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Certain hexavalent chromium ( Cr(VI) ) compounds are well established occupational respiratory tract carcinogens . However , despite extensive studies , the cellular and molecular mechanisms underlying Cr(VI)-induced lung cancer remain poorly understood . In fact , the models used were often suboptimal and yielded conflicting results that were heavily dependent upon the system and experimental conditions employed . Here , we investigated the effects of chronic subcytotoxic and mildly cytotoxic ( 0.1-2 microM ) Cr(VI) exposures on cultures of human bronchial epithelial cells , the main targets of Cr(VI) carcinogenicity . Our studies with the nontumorigenic BEAS-2B cell line suggest that relatively short exposures ( h ) to sublethal Cr(VI) doses ( 0.1-1 microM ) may render these cells less sensitive to contact inhibition . We have also observed a reduced sensitivity to Cr(VI)-induced apoptosis shortly after the beginning of exposure to a mildly cytotoxic Cr(VI) dose ( 2 microM ) . Further studies are needed to determine whether these two phenotypes are involved in the Cr(VI)-induced carcinogenic process . Additionally , evidence gathered in this study strongly points to a Cr(VI) interference with cell adhesion to the substratum and with cell-cell interactions . Finally , by chronically exposing BEAS-2B cells to mildly cytotoxic Cr(VI) doses ( 1 and 2 microM ) , we were able to induce changes in cell morphology and pattern of growth characteristic of an early phase of pre-malignant progression .
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20336777
|
A large number of human cancers display alterations in the Ink4a/cyclin D/Cdk4 genetic pathway , suggesting that activation of Cdk4 plays an important role in oncogenesis . Here we report that Cdk4-null mouse embryonic fibroblasts are resistant to transformation in response to Ras activation with dominant-negative ( DN ) p53 expression or in the Ink4a/Arf-null background , judged by foci formation , anchorage-independent growth , and tumorigenesis in athymic mice . Cdk4-null fibroblasts proliferate at normal rates during early passages . Whereas Cdk4(+/+)Ink4a/Arf(-/-) cells are immortal in culture , Cdk4(-/-)Ink4a/Arf(-/-) cells undergo senescence during continuous culture , as do wild-type cells . Activated Ras also induces premature senescence in Cdk4(-/-)Ink4a/Arf(-/-) cells and Cdk4(-/-) cells with DNp53 expression . Thus , Cdk4 deficiency causes senescence in a unique Arf/p53-independent manner , which accounts for the loss of transformation potential . Cdk4-null cells express high levels of p21(Cip1/Waf1) with increased protein stability . Suppression of p21(Cip1/Waf1) by small interfering RNA ( siRNA ) , as well as expression of HPV-E7 oncoprotein , restores immortalization and Ras-mediated transformation in Cdk4(-/-)Ink4a/Arf(-/-) cells and Cdk4(-/-) cells with DNp53 expression . Therefore , Cdk4 is essential for immortalization , and suppression of Cdk4 could be a prospective strategy to recruit cells with inactive Arf/p53 pathway to senescence .
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12435633
|
Archaeal and eukaryotic B-family DNA polymerases ( pols ) mainly replicate chromosomal DNA but stall at lesions , which are often bypassed with Y-family pols . In this study , a B-family pol Vent ( exo(-) ) from the euryarchaeon Thermococcus litoralis was studied with three types of DNA lesions-N(2)-alkylG , O(6)-alkylG , and an abasic ( AP ) site-in comparison with a model Y-family pol Dpo4 from Sulfolobus solfataricus , to better understand the effects of various DNA modifications on binding , bypass efficiency , and fidelity of pols . Vent ( exo(-) ) readily bypassed N(2)-methyl(Me)G and O(6)-MeG , but was strongly blocked at O(6)-benzyl(Bz)G and N(2)-BzG , whereas Dpo4 efficiently bypassed N(2)-MeG and N(2)-BzG and partially bypassed O(6)-MeG and O(6)-BzG . Vent ( exo(-) ) bypassed an AP site to an extent greater than Dpo4 , corresponding with steady-state kinetic data . Vent ( exo(-) ) showed 180- , and 300-fold decreases in catalytic efficiency ( k(cat)/K(m) ) for nucleotide insertion opposite an AP site , N(2)-MeG , and O(6)-MeG but and 5000-fold decreases opposite O(6)-BzG and N(2)-BzG , respectively , as compared to G , whereas Dpo4 showed little or only decreases opposite N(2)-MeG and N(2)-BzG but decreases opposite O(6)-MeG , O(6)-BzG , and the AP site . Vent ( exo(-) ) preferentially misinserted G opposite N(2)-MeG , T opposite O(6)-MeG , and A opposite an AP site and N(2)-BzG , while Dpo4 favored correct C insertion opposite those lesions . Vent ( exo(-) ) and Dpo4 both bound modified DNAs with affinities similar to unmodified DNA . Our results indicate that Vent ( exo(-) ) is as or more efficient as Dpo4 in synthesis opposite O(6)-MeG and AP lesions , whereas Dpo4 is much or more efficient opposite ( only ) N(2)-alkylGs than Vent ( exo(-) ) , irrespective of DNA-binding affinity . Our data also suggest that Vent ( exo(-) ) accepts nonbulky DNA lesions ( e.g. , N(2)- or O(6)-MeG and an AP site ) as manageable substrates despite causing error-prone synthesis , whereas Dpo4 strongly favors minor-groove N(2)-alkylG lesions over major-groove or noninstructive lesions .
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22793782
|
OBJECTIVE Chronic generation of inflammatory cytokines and reactive oxygen species are implicated in atherosclerosis , aging , cancers , and other chronic diseases . We hypothesized that zinc induces A20 in premonocytic , endothelial , and cancer cells , and A20 binds to tumor necrosis factor ( TNF)-receptor associated factor , and inhibits Iκ kinase-α ( IKK-α)/nuclear factor-κB ( NF-κB ) , resulting in downregulation of TNF-α and interleukin-1β ( IL-1β ) . METHODS To test this hypothesis , we used HL-60 , human umbilical vein endothelial cells , and SW480 cell lines under zinc-deficient and zinc-sufficient conditions in this study . We measured oxidative stress markers , inflammatory cytokines , A20 protein and mRNA , A20-FRAF-1 complex , and IKK-α/NF-κB signaling in stimulated zinc-deficient and zinc sufficient cells . We also conducted antisense A20 and siRNA studies to investigate the regulatory role of zinc in TNF-α and IL-1β via A20 . RESULTS We found that zinc increased A20 and A20-tumor necrosis factor-receptor associated factor-1 complex , decreased the IKK-α/NF-κB signaling pathway , oxidative stress markers , and inflammatory cytokines in these cells compared with zinc-deficient cells . We confirmed that zinc-induced A20 contributes to downregulation of TNF-α and IL-1β by antisense and short interfering RNA A20 studies . CONCLUSION Our studies suggest that zinc suppresses generation of NF-κB-regulated inflammatory cytokines by induction of A20 .
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1
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21035309
|
Monoclonal antibodies ( mAbs ) to HER2 are currently used to treat breast cancer , but low clinical efficacy , along with primary and acquired resistance to therapy , commonly limit clinical applications . We previously reported that combinations of antibodies directed at non-overlapping epitopes of HER2 are endowed with enhanced antitumor effects , probably due to accelerated receptor degradation . Here , we extend these observations to three-dimensional mammary cell models , and compare the effects of single mAbs with the effects of antibody combinations . Collectively , our in vitro assays and computational image analyses indicate that combining mAbs against different epitopes of HER2 better inhibits invasive growth . Importantly , while growth factors are able to reduce intraluminal apoptosis and induce an invasive phenotype , combinations of mAbs better than single mAbs can reverse the growth factor-induced phenotypes of HER2-overexpressing spheroids . In conclusion , our studies propose that mAb combinations negate the biological effects of growth factors on invasive growth of HER2-overexpressing cells . Hence , combining mAbs offers a therapeutic strategy , potentially able to enhance clinical efficacy of existing antireceptor immunotherapeutics .
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1,
0
] |
21132012
|
In order to determine the in vivo immune response in glioblastoma , monoclonal and polyclonal antibodies specific for inflammatory leukocytes and immunoregulatory products were utilized to stain tissue from four surgical specimens . The more activated the inflammatory cells , the more activated the tumors appeared to be . In the tumor with the largest infiltration ( Case 3 ) , inflammatory cells were stained for interferon-gamma , interleukin-2 , interleukin-1 beta , lymphotoxin , tumor necrosis factor-alpha , and transforming growth factor-beta . The tumor cells also expressed interleukin-1 beta , interleukin-6 , transforming growth factor-beta , tumor necrosis factor-alpha , and prostaglandin E. In contrast , in the tumor with the least inflammatory response ( Case 1 ) , the tumor cells did not express any cytokines . Expression of cytokines by glioma cells was modest in the two cases with modest inflammatory responses . Cellular inflammation , primarily consisting of T cells and macrophages with few or no B cells or natural killer cells , was two- to 15-fold greater outside the tumor than within . In contrast to leukocytes outside the tumor , which were activated and expressing class II major histocompatibility antigens , leukocytes within the tumor parenchyma or at the tumor's edge were negative for these antigens . In the four specimens studied here , the tumor cells themselves were also negative for class II major histocompatibility antigens . These findings , although preliminary , suggest that inflammatory cells within gliomas are inactivated and that glioma cells may increase the expression of immunosuppressive cytokines in response to an increased lymphocyte infiltrate . This observation , if corroborated by more extensive studies , may help to explain the failure of immune treatments in glioblastoma multiforme .
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1318961
|
1,25-Dihydroxyvitamin D(3) ( 1,25D ) , the hormonal form of vitamin D , and several analogs have failed as monotherapies for cancer because of poor efficacy or acquired resistance . However , 1,25D analogs are amenable to bifunctionalization . Preclinical studies have revealed combinatorial effects of 1,25D analogs and histone deacetylase inhibitors ( HDACi ) . Secosteroidal hybrid molecules combining vitamin D receptor ( VDR ) agonism with HDACi displayed enhanced efficacy but are laborious to synthesize . Here , we have developed easily assembled , fully integrated , non-secosteroidal VDR agonist/HDACi hybrids . The most promising are full VDR agonists with lower potency than 1,25D . Structure/function studies revealed that antiproliferative activity against 1,25D-resistant squamous carcinoma cells required VDR agonism and HDACi . Remarkably , modeling and X-ray crystallography reveal non-secosteroidal hybrids bind in the VDR ligand binding domain in the opposite orientation of their secosteroidal counterparts .
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22921063
|
Activated lymphocytes and malignant lymphoma cells derived from them ( Ki-1 positive lymphoma cells ) share similar mechanisms of proliferation . To further examine the inhibitory role of endogenous transforming growth factor beta ( TGF beta ) in Ki-1 positive lymphoma cells , the authors studied anti-TGF beta antibodies and measured their effect on proliferation . A monoclonal antibody ( T1A5 ) prepared against a unique antigenic epitope of high molecular weight Hodgkin's TGF beta and a polyclonal rabbit antibody prepared against highly purified 25,000 D porcine platelet TGF beta 1 were used . Both antibodies are shown here to inhibit the biological activity of Hodgkin's TGF beta and to crossreact with their respective antigens in immunoblotting . DNA synthesis by Ki-1 lymphoma cells was increased 138-fold by anti-TGF beta 1 antibody and 262-fold by anti-Hodgkin's TGF beta . Exogenous TGF beta 1 suppression was completely reversed by anti-TGF beta 1 antibody and IL-2-induced proliferation was markedly potentiated ( 41 fold ) . L-428 Reed-Sternberg cells secrete physiologically active TGF beta but have fewer than 500 TGF beta receptor sites per cell ; no significant proliferative response was measured for either anti-TGF beta 1 or anti-Hodgkin's TGF beta . These results show the suppressive effect of exogenous TGF beta 1 on indolent Ki-1 lymphoma cells and suggest that the endogenous secretion of high molecular weight physiologically active TGF beta is important in maintaining the indolent nature of this low-grade Ki-1 positive lymphoma .
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1,
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] |
1312308
|
In response to DNA double strand breaks , the histone variant H2AX at the break site is phosphorylated at serine 139 by DNA damage sensor kinases such as ataxia telangiectasia-mutated , forming gamma-H2AX . This phosphorylation event is critical for sustained recruitment of other proteins to repair the break . After repair , restoration of the cell to a prestress state is associated with gamma-H2AX dephosphorylation and dissolution of gamma-H2AX-associated damage foci . The phosphatases PP2A and PP4 have previously been shown to dephosphorylate gamma-H2AX . Here , we demonstrate that the wild-type p53-induced phosphatase 1 ( WIP1 ) also dephosphorylates gamma-H2AX at serine 139 in vitro and in vivo . Overexpression of WIP1 reduces formation of gamma-H2AX foci in response to ionizing and ultraviolet radiation and blocks recruitment of MDC1 ( mediator of DNA damage checkpoint 1 ) and 53BP1 ( p53 binding protein 1 ) to DNA damage foci . Finally , these inhibitory effects of WIP1 on gamma-H2AX are accompanied by WIP1 suppression of DNA double strand break repair . Thus , WIP1 has a homeostatic role in reversing the effects of ataxia telangiectasia-mutated phosphorylation of H2AX .
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0,
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20118229
|
Thellungiella salsuginea , a close relative of Arabidopsis , represents an extremophile model for abiotic stress tolerance studies . We present the draft sequence of the T. salsuginea genome , assembled based on coverage to seven chromosomes with a coding capacity of at least 28,457 genes . This genome provides resources and evidence about the nature of defense mechanisms constituting the genetic basis underlying plant abiotic stress tolerance . Comparative genomics and experimental analyses identified genes related to cation transport , abscisic acid signaling , and wax production prominent in T. salsuginea as possible contributors to its success in stressful environments .
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22778405
|
In the present study , the question was addressed whether anthocyanins interfere with the topoisomerase I poison irinotecan in vivo . In vivo complexes of enzyme to DNA bioassay was used to detect irinotecan-induced stabilization of topoisomerase I/DNA complexes and single cell gel electrophoresis to determine DNA-strand-break induction in the colon of male Wistar rats . Furthermore , analysis of anthocyanin concentrations in rat plasma and rat colon was included in the testing , demonstrating that anthocyanins reach the colon and the concentrations do not differ between rats that only received anthocyanins and the anthocyanin/irinotecan group . Blackberry extract was found to significantly reduce irinotecan-mediated topoisomerase I/DNA cleavable complex formation . Overall , anthocyanins did not notably increase cleavable complex formation . However , a significant increase of DNA damage was shown after a single dose of irinotecan as well as the single compounds cyanidin ( cy ) and cyanidin-3-glucoside ( cy-3-g ) . Furthermore , a significant reduction of irinotecan-induced DNA-strand breaks after a pretreatment with cy , cy-3-g and blackberry extract was observed . Thus , the question arises whether anthocyanin-rich preparations might interfere with chemotherapy or whether , due to low systemic bioavailability , the preparations might provide protective potential in the gastrointestinal tract .
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23275152
|
MicroRNAs ( miRNAs ) are 19 nt non-coding RNA molecules that regulate expression of target genes at the post-transcriptional level . Evidence indicates that miRNAs play an essential role in physiological and pathological conditions including pulmonary development , inflammation , fibrosis and cancer . The aim of the present study is to investigate the altered miRNAs expression profile in rats with experimental silicosis . We duplicated silicosis rat model , and identify the miRNA expression pattern of silicosis rat with miRNA microarray . Compared with normal lung tissue , fourteen miRNAs were found significantly up-regulated while the other twenty-five down-regulated in silicosis samples . The differential expression of two selected miRNAs was confirmed by stem-loop real-time reverse transcription-polymerase chain reaction . Our results indicate that the 39 altered miRNAs may be involved in lung fibrosis of rats that were exposed to silica dust . Furthermore , the microarray results provide a solid basis for further validation , such as identification of other miRNAs that may be related to inflammation and fibrosis . The findings are paving way for silicosis early prevention , prognosis and possible therapy .
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23208435
|
Metastasis , commonly to the lung , is the major cause of mortality from testicular cancer . The aim of the present study was to examine the effect of a novel nutrient mixture ( NM ) containing ascorbic acid , amino acids and green tea extract on the inhibition of melanoma growth and metastasis using a model of intratesticular inoculation of B16FO cells into nude mice . Male athymic mice ( n=12 ) , 10-12 weeks of age , were inoculated with 5�10(5) B16FO melanoma cells in 100 μl of PBS into the right testis , while the left testis was left untreated . Following inoculation , the mice were randomly divided into two groups . The control group ( n=6 ) was fed a regular mouse chow diet and the NM 1% group ( n=6 ) the same diet , but supplemented with 1% NM . Four weeks later the mice were sacrificed and the abdominal cavity was opened . Mice in the control group exhibited extensive metastasis in the peritoneal cavity and severely enlarged right testes and necrotic seminiferous tubules . By contrast , in the NM 1% fed group there was no evidence of peritoneal metastasis in 50% of the animals and mild metastasis in the remaining 50% . The right testes were enlarged and seminiferous tubules in the area of invasion showed evidence of degeneration . No metastasis to the liver , kidney or spleen were evident in either group . However , severe lung metastasis was observed in 2 of 6 mice in the control group and mild metastasis in 2 of 6 mice in the NM 1% group . In conclusion , these results confirm earlier studies and verify the anti-metastatic potential of NM .
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1,
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] |
23226724
|
OBJECTIVES : Cigarette smoking is a major risk factor for pancreatic cancer ( PaCa ) . However , the mechanisms of smoking-induced PaCa remain unknown . Here we investigated the effect of smoking compounds on cell death pathways in pancreatic ductal cells , precursors of PaCa . METHODS : Human pancreatic ductal cells ( HPDE6-c7 ) were cultured with cigarette smoke extract ( CSE ) or smoking compound 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone ( NNK ) . Apoptosis and autophagy were assessed by DNA fragmentation and immunofluorescence , respectively . RESULTS : Exposure to CSE or NNK decreased DNA fragmentation and up-regulated BclXL . Akt kinase was activated by smoking compounds through reactive oxygen species-dependent mechanism . Specifically , Akt activation was prevented by inhibition of nicotinamide adenine dinucleotide oxidase . Molecular or pharmacologic inhibitions of Akt prevented the antiapoptotic effect of smoking compounds . Smoking compounds stimulated rapid ( 1 hour ) and transient activation of 5'-adenosine monophosphate-activated protein kinase and formation of autophagic vacuoles , indicating stimulation of autophagy . Repeated exposure to CSE/NNK ( 48 hours or longer ) abolished the early activation of autophagic markers . Inhibition of Akt prevented the antiautophagic effect of long exposure to smoking compounds , indicating that smoking-induced late activation of Akt prevents autophagy . CONCLUSIONS : Long exposure of pancreatic ductal cells to smoking compounds inhibited apoptosis and autophagy . The results revealed a central role for Akt kinase in mediating key procarcinogenic effects of smoking compounds .
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23271397
|
BACKGROUND : To quantify tumour angiogenesis , microvessel density ( MVD ) has been widely used . We here present a novel angiogenesis marker , microvessel proliferation ( MVP ) , based on dual immunohistochemical staining of nestin and Ki-67 . Immature endothelial cells express nestin , and when co-expressed with the proliferation marker Ki-67 , the number of proliferating immature blood vessels can be measured . MATERIALS AND METHODS : Microvessel proliferation was evaluated in 178 breast cancer samples and estimated by vascular proliferation index ( VPI ) , the ratio between the number of vessels containing proliferating endothelial cells and the total number of immature vessels . RESULTS : High VPI was strongly associated with several markers of aggressive breast cancer , such as negative oestrogen receptor ( ER ) status ( p=0.003 ) , high tumour cell proliferation by Ki-67 ( p=0.004 ) , high p53 expression ( p=0.001 ) , and five profiles for the basal-like phenotype ( odds ratios ( OR ) ; range 3.4-6.3 ) . Also , high VPI was significantly associated with interval detected breast cancer compared with screening detected lesions ( p<0.0005 ) , and adverse outcome in univariate and multivariate survival analysis ( p=0.034 and p=0.022 , respectively ) . CONCLUSION : Microvessel proliferation is a novel marker of ongoing angiogenesis and was associated with aggressive tumour features , basal-like phenotypes , interval presentation , and prognosis in this series of breast cancer .
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22840462
|
Key cellular decisions , such as proliferation or growth arrest , typically occur at spatially defined locations within tissues . Loss of this spatial control is a hallmark of many diseases , including cancer . Yet , how these patterns are established is incompletely understood . Here , we report that physical and architectural features of a multicellular sheet inform cells about their proliferative capacity through mechanical regulation of YAP and TAZ , known mediators of Hippo signaling and organ growth . YAP/TAZ activity is confined to cells exposed to mechanical stresses , such as stretching , location at edges/curvatures contouring an epithelial sheet , or stiffness of the surrounding extracellular matrix . Weidentify the F-actin-capping/severing proteins Cofilin , CapZ , and Gelsolin as essential gatekeepers that limit YAP/TAZ activity in cells experiencing low mechanical stresses , including contact inhibition of proliferation . We propose that mechanical forces are overarching regulators of YAP/TAZ in multicellular contexts , setting responsiveness to Hippo , WNT , and GPCR signaling .
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23954413
|
This study aims to identify significant predictors of survival in pediatric and adolescent colorectal carcinoma . We retrospectively analyzed our experience with 29 histologically verified cases , of which 20 were resected for cure . Variables analyzed as predictors of survival included : ( 1 ) resectability , ( 2 ) regional nodal involvement , ( 3 ) depth of invasion , ( 4 ) grade , and ( 5 ) interval from symptom onset to diagnosis . Signet ring or anaplastic lesions were considered high grade . Survival curves were generated on both the overall group and those resected for cure . Multivariate analysis was performed on the overall group . The median age at diagnosis was 19 years ( range , 10 to 21 ) . Median follow-up in survivors was 4.7 years . Signet ring tumors occurred in 45% and another 24% were poorly differentiated . Seventy-six percent presented with regional lymph node metastases . The median survival for the overall group was 16 months , whereas that for those undergoing complete resection was 33 months . In patients undergoing resection for cure , grade ( P = .005 ) , regional nodal involvement ( P = .007 ) , and depth of invasion ( P = .03 ) were significant predictors of outcome in univariate analysis . In the overall group these variables as well as resectability and distant metastases were significant in univariate analysis . In multivariate analysis high-grade lesions and lymph node involvement were highly correlated , as were resectability and metastases . Thus , either variable ( but not both ) of each pair added information to the multivariate model . In patients resected for cure , positive nodes or high histological grade became the only significant predictors of survival.(ABSTRACT TRUNCATED AT 250 WORDS )
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1403541
|
The generation and detection of mechanical forces is a ubiquitous aspect of cell physiology , with direct relevance to cancer metastasis(1) , atherogenesis(2) and wound healing(3) . In each of these examples , cells both exert force on their surroundings and simultaneously enzymatically remodel the extracellular matrix ( ECM ) . The effect of forces on ECM has thus become an area of considerable interest due to its likely biological and medical importance(4-7) . Single molecule techniques such as optical trapping(8) , atomic force microscopy(9) , and magnetic tweezers(10,11) allow researchers to probe the function of enzymes at a molecular level by exerting forces on individual proteins . Of these techniques , magnetic tweezers ( MT ) are notable for their low cost and high throughput . MT exert forces in the range of pN and can provide millisecond temporal resolution , qualities that are well matched to the study of enzyme mechanism at the single-molecule level(12) . Here we report a highly parallelizable MT assay to study the effect of force on the proteolysis of single protein molecules . We present the specific example of the proteolysis of a trimeric collagen peptide by matrix metalloproteinase 1 ( MMP-1 ) ; however , this assay can be easily adapted to study other substrates and proteases .
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22871786
|
OBJECTIVES Despite the well-defined histological types of non-small cell lung cancer ( NSCLC ) , a given stage is often associated with wide-ranging survival rates and treatment outcomes . This disparity has led to an increased demand for the discovery and identification of new informative biomarkers . METHODS In the current study , we screened 81 NSCLC samples using Illumina whole-genome gene expression microarrays in an effort to identify differentially expressed genes and new NSCLC biomarkers . RESULTS We identified novel genes whose expression was upregulated in NSCLC , including SPAG5 , POLH , KIF23 , and RAD54L , which are associated with mitotic spindle formation , DNA repair , chromosome segregation , and dsDNA break repair , respectively . We also identified several novel genes whose expression was downregulated in NSCLC , including SGCG , NLRC4 , MMRN1 , and SFTPD , which are involved in extracellular matrix formation , apoptosis , blood vessel leakage , and inflammation , respectively . We found a significant correlation between RNA degradation and survival in adenocarcinoma cases . CONCLUSIONS Even though the follow-up time was too limited to draw final conclusions , we were able to show better prediction p values in a group selection based on molecular profiles compared to histology . The current study also uncovered new candidate biomarker genes that are likely to be involved in diverse processes associated with NSCLC development .
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21412013
|
Prostate cancer is responsible for major deaths globally after lung cancer . However , etiology of prostate cancer is still unknown . Individual risk and incidence of prostate cancer may result from the interaction of genetic susceptibility with exposure to environmental factors such as infectious agents , tobacco , occupational exposure , dietary carcinogens , and/or hormonal imbalances leading to injury of the prostate and to the development of chronic inflammation . About 30% of all human cancers are caused by tobacco smoking and inhaled pollutants . Inflammation is now regarded as an important hallmark of cancer . The present study has been aimed to explore the pro-inflammatory levels in prostate carcinoma patients by examining the serum levels of novel cytokine interleukin-18 ( IL-18 ) expression in tobacco exposed population . A total of 578 ( n = 284 biopsy proven prostate cancer patients , n = 294 controls with and without tobacco exposed population ) were recruited . Serum IL-18 ( Interleukin-18 ) level was done by ELISA . The IL-18 levels between cancer patients and controls within same mode tobacco exposure as tobacco smoking ( overall ) showed significant difference ( P < 0.0001 ) and further we compared within stratified group , it significantly differ ( P < 0.0001 ) in bidi and cigarette smoking than control non users . Furthermore , IL-18 levels in tobacco chewers ( overall ) with gutkha and khaini chewers showed significant difference ( P < 0.01 ) than controls non users . Moreover , the IL-18 levels between cancer patients and controls with in of combined mode chewers smokers and alcohol ( CSA ) , smokers with alcohol showed significant difference ( P < 0.01 ) than controls . The IL-18 levels also differed significantly ( P < 0.05 ) with smokers and chewers in higher stages of III and IV , and showed non significant with in lower stages . Tobacco exposure enhance the inflammation in prostate carcinoma patients in stratified group as it have been represented as a risk factors in various cancers , but this study provide further its role that seems to influence inflammation especially in prostate carcinoma .
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23293472
|
Colorectal cancer represents one of the most challenging diseases . Increasing evidence indicates that aberrant expression of microRNAs ( miRNAs ) is related to pathogenesis of colorectal cancer . Cancer cells reprogram metabolic pathways to sustain higher proliferation rates . Whether mechanisms underlying the role of miRNA in colorectal cancer are involved in metabolic reprogramming and the mechanisms through which miRNAs alter cancer metabolism are as yet unknown . Herein , we show that miR-124 , miR-137 and miR-340 are associated with poor prognosis of colorectal cancer . Expression of these miRNAs inhibits the growth of colorectal cancer cells . PKM ( pyruvate kinase isozyme ) alternative splicing proteins ( PTB1/hnRNAPA1/hnRNAPA2 ) , which control the inclusion of exon 9 ( PKM1 ) or exon 10 ( PKM2 ) , are targeted by miR-124 , miR-137 and miR-340 . Consequently , miR-124 , miR-137 and miR-340 switch PKM gene expression from PKM2 to PKM1 . High ratios of PKM1/PKM2 inhibit the glycolysis rate , but elevate the glucose flux into oxidative phosphorylation . These results demonstrate that miRNAs ( miR-124 , miR-137 and miR-340 ) impair colorectal cancer growth by counteracting the Warburg effect due to regulating alternative splicing of the PKM gene .
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22895557
|
DNA damage checkpoints arrest cell cycle progression to facilitate DNA repair . The ability to survive genotoxic insults depends not only on the initiation of cell cycle checkpoints but also on checkpoint maintenance . While activation of DNA damage checkpoints has been studied extensively , molecular mechanisms involved in sustaining and ultimately inactivating cell cycle checkpoints are largely unknown . Here , we explored feedback mechanisms that control the maintenance and termination of checkpoint function by computationally identifying an evolutionary conserved mitotic phosphorylation network within the DNA damage response . We demonstrate that the non-enzymatic checkpoint adaptor protein 53BP1 is an in vivo target of the cell cycle kinases Cyclin-dependent kinase-1 and Polo-like kinase-1 ( Plk1 ) . We show that Plk1 binds 53BP1 during mitosis and that this interaction is required for proper inactivation of the DNA damage checkpoint. 53BP1 mutants that are unable to bind Plk1 fail to restart the cell cycle after ionizing radiation-mediated cell cycle arrest . Importantly , we show that Plk1 also phosphorylates the 53BP1-binding checkpoint kinase Chk2 to inactivate its FHA domain and inhibit its kinase activity in mammalian cells . Thus , a mitotic kinase-mediated negative feedback loop regulates the ATM-Chk2 branch of the DNA damage signaling network by phosphorylating conserved sites in 53BP1 and Chk2 to inactivate checkpoint signaling and control checkpoint duration .
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0,
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20126263
|
OBJECTIVE This study describes a new method used in the clinical laboratory at Hospital Israelita Albert Einstein to detect mutations in exons 8 and 17 of the KIT gene in patients with acute myeloid leukemia . METHODS Genomic DNA extraction was performed on 54 samples of peripheral blood or bone marrow from patients with acute myeloid leukemia . The extracted DNA was amplified by polymerase chain reaction and sequenced , and the fragments were analyzed . RESULTS Within the analyzed samples , we detected four mutations in exon 8 , two mutations in exon 17 , and mutations or a double mutation in one sample . CONCLUSION The tests detecting mutations in exon 8 and 17 on the KIT gene were successfully standardized . The test is now included among the routine diagnostics employed for patients at Hospital Israelita Albert Einstein clinical laboratory .
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23386005
|
A number of cancer chemotherapeutic drugs designed to have cytotoxic actions on tumor cells have recently been shown to also have antiangiogenic activities . Endothelial cell migration and proliferation are key components of tumor angiogenesis , and agents that target the microtubule cytoskeleton can interfere with these processes . In this study , the effect on endothelial cell functions of the microtubule-stabilizing drugs Taxotere and Taxol were evaluated in three in vitro assays : a chemokinetic migration assay , an angiogenesis factor-mediated chemotactic migration assay , and a three-dimensional Matrigel tubule formation assay , using rat fat pad endothelial cells ( RFPECs ) and/or human umbilical vein endothelial cells ( HUVECs ) . Taxotere was active in all three assays at concentrations that were not cytotoxic and did not inhibit endothelial cell proliferation . In the RFPEC chemokinetic migration and in vitro tubule formation assays , the IC50 values were approximately 10(-9) M for both Taxotere and Taxol . HUVEC migration , however , was more sensitive to Taxotere , with an observed IC50 of 10(-12) M in a chemokinetic assay . In a Boyden chamber assay , HUVEC chemotaxis stimulated by either of two angiogenic factors , thymidine phosphorylase or vascular endothelial growth factor , was inhibited by Taxotere with an IC50 of 10(-11) M and was ablated at 10(-9) M. Taxotere was also up to 1000-fold more potent than Taxol in inhibiting either chemokinetic or chemotactic migration . When the microtubule cytoskeleton was visualized using immunofluorescence staining of alpha-tubulin , there were no gross morphological changes observed in HUVECs or RFPECs treated with Taxotere at concentrations that inhibited endothelial cell migration but not proliferation . The effects of Taxotere on migration were associated with a reduction in the reorientation of the cell's centrosome , at concentrations that did not affect gross microtubule morphology or proliferation . Reorientation of the centrosome , which acts as the microtubule organizing center , in the intended direction of movement is a critical early step in the stabilization of directed cell migration . These data indicate that endothelial cell migration correlates more closely with changes in microtubule plasticity than with microtubule gross structure . The antiangiogenic activity of Taxotere in vivo was assessed in a Matrigel plug assay . In this assay , the angiogenic response to fibroblast growth factor 2 was inhibited in vivo by Taxotere with an ID50 of 5.4 mg/kg when injected twice weekly over a 14-day period , and angiogenesis was completely blocked in mice that received 10 mg/kg Taxotere . The in vivo data further suggested that Taxotere had selectivity for endothelial cell migration and/or microvessel formation because infiltration of inflammatory cells into the Matrigel plug was much less sensitive to inhibition by Taxotere . In conclusion , Taxotere is a potent and potentially specific inhibitor of endothelial cell migration in vitro and angiogenesis in vitro and in vivo .
|
[
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1,
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0,
1
] |
12479700
|
Acetyl-11-keto-beta-boswellic acid ( AKBA ) is a naturally occurring pentacyclic triterpene isolated from the gum resin exudate from the stem of the tree Boswellia serrata ( frankincense ) . AKBA has been recently identified as a novel , orally active , non-redox and non-competitive 5-lipoxygenase inhibitor that also inhibits topisomerase I and II in vitro . Because natural pentacyclic triterpenes have an antiproliferative effect against different tumor types , we investigated the effects of AKBA on the proliferation of 11 primary cell cultures established from human surgical specimens of meningiomas , common central nervous system tumors . Treatment of meningioma cells by AKBA revealed a potent cytotoxic activity with half-maximal inhibitory concentrations in the range of 2-8 microM . At similar , physiologically achievable concentrations , AKBA rapidly ( within minutes ) and potently inhibited the phosphorylation of extracellular signal-regulated kinase 1 and 2 ( Erk-1 and Erk-2 ) in meningioma cells stimulated with platelet-derived growth factor BB . High expression level of 5-LO was detected in primary meningioma cells and surgical specimens by immunoblotting analysis , suggesting the possible role of 5-LO in meningioma tumorigenesis . Considering the critical importance of the Erk-1/2 signal transduction pathway not only in meningiomas but in other human neoplasms , the interruption of signaling through this evolutionarily conserved pathway might be one of the mechanisms by which AKBA induces suppression of proliferation and apoptosis of different tumor types .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
12664615
|
Tumor necrosis factor related apoptosis-inducing ligand ( TRAIL ) and agonistic anti-DR4/TRAIL-R1 and anti-DR5/TRAIL-R2 antibodies are currently under clinical investigation for treatment of different malignancies . TRAIL activates DR4 and DR5 and thereby triggers apoptotic and non-apoptotic signaling pathways , but possible different roles of DR4 or DR5 in these responses has poorly been addressed so far . In the present work , we analyzed cell viability , DISC formation as well as IL-8 and NF-kappaB activation side by side in responses to TRAIL and agonistic antibodies against DR4 ( mapatumumab ) and against DR5 ( lexatumumab ) in pancreatic ductal adenocarcinoma cells . We found that all three reagents are able to activate cell death and pro-inflammatory signaling . Death-inducing signaling complex ( DISC ) analysis revealed that mapatumumab and lexatumumab induce formation of homocomplexes of either DR4 or DR5 , whereas TRAIL additionally stimulated the formation of heterocomplexes of both receptors . Notably , blocking of receptors using DR4- and DR5-specific Fab fragments indicated that TRAIL exerted its function predominantly via DR4 . Interestingly , inhibition of PKC by Goe6983 enabled DR5 to trigger apoptotic signaling in response to TRAIL and also strongly enhanced lexatumumab-mediated cell death . Our results suggest the existence of mechanisms that silence DR5 for TRAIL- but not for agonistic-antibody treatment .
|
[
0,
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1,
0,
1
] |
20354842
|
UNLABELLED ABC transporters like P-glycoprotein ( P-gp/ABCB1 ) are membrane proteins responsible for the transport of toxic compounds out of non-malignant cells and tumor tissue . AIM To investigate the effect of glycolysis and the tissue redox state on P-gp expression in multicellular tumor spheroids derived from prostate adenocarcinoma cells ( DU-145 ) , glioma cells ( Gli36 ) , and the human cervix carcinoma cell line KB-3-1 transfected with a P-gp-EGFP fusion gene that allows monitoring of P-gp expression in living cells . During cell culture of DU-145 , Gli36 , and KB-3-1 tumor spheroids P-gp expression was observed as well as increased lactate and decreased pyruvate levels and expression of glycolytic enzymes . Inhibition of glycolysis for 24 h by either iodoacetate ( IA ) or 2-deoxy-D-glucose ( 2-DDG ) downregulated P-gp expression which was reversed upon coincubation with the radical scavenger ebselen as shown by semi-quantitative immunohistochemisty in DU-145 and Gli36 tumor spheroids , and by EGFP fluorescence in KB-3-1 tumor spheroids . Consequently endogenous ROS generation in DU-145 tumor spheroids was increased in the presence of either IA or 2-DDG , which was abolished upon coincubation with ebselen . Exogenous addition of pyruvate significantly reduced ROS generation , increased P-gp expression as well as efflux of the P-gp substrate doxorubicin . Doxorubicin transport was significantly blunted by 2-DDG and IA , indicating that inhibition of glycolysis reversed the multidrug resistance phenotype . In summary our data demonstrate that P-gp expression in tumor spheroids is closely related to the glycolytic metabolism of tumor cells and can be downregulated by glycolysis inhibitors via mechanisms that involve changes in the cellular redox state .
|
[
0,
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1,
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0,
0
] |
19950199
|
The aryl hydrocarbon receptor ( AhR ) is a ligand-activated transcription factor . Recent studies have reported the anti-tumor effects of the AhR in breast cancer . In this study , we investigated the anti-tumor effect of AhR activation based on the cancer stem cell hypothesis . We show that AhR activation suppressed mammosphere formation of MCF-7 cells and decreased the proportion of cells with high ALDH-1 ( aldehyde dehydrogenase 1 ) activity . In addition , we also demonstrate that AhR activation regulates self-renewal signaling by down-regulating Wnt/β-catenin and Notch .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
22123295
|
Here , we provide the necessary proof of concept , that it is possible to metabolically create a non-permissive or " hostile " stromal microenvironment , which actively prevents tumor engraftment in vivo . We developed a novel genetically engineered fibroblast cell line that completely prevents tumor formation in mice , with a 100% protection rate . No host side effects were apparent.This could represent a viable cellular strategy for preventing and treating a variety of human cancers . More specifically , we examined the autocrine and paracrine effects of the cellular delivery of TNF-alpha on breast cancer tumor growth and cancer metabolism . For this purpose , we recombinantly overexpressed TNF-alpha in human breast cancer cells ( MDA-MB-231 ) or human immortalized fibroblasts ( hTERT-BJ1 ) . Our results directly show that TNF-alpha functions as a potent tumor suppressor . Remarkably , TNF-alpha-expressing breast cancer cells were viable , without any significant increases in their basal apoptotic rate . However , after 4 weeks post-implantation , TNF-alpha-expressing breast cancer cells failed to form any tumors in xenografted mice ( 0 tumors/10 injections ) , ultimately conferring 100% protection against tumorigenesis . Similarly , TNF-alpha-overexpressing fibroblasts were also viable , without any increases in apoptosis . Significantly , complete tumor suppression was obtained by co-injecting TNF-alpha expressing stromal fibroblasts with human breast cancer cells , indicating that paracrine cell-mediated delivery of TNF-alpha can also prevent tumor engraftment and growth ( 0 tumors/10 injections ) . Mechanistically , TNF-alpha induced autophagy and mitochondrial dysfunction in both epithelial cancer cells and stromal fibroblasts , preventing energy transfer from the tumor microenvironment , likely " starving " the cancer cells to death . In addition , via qRT-PCR analysis of MDA-MB-231 cells , we observed that TNF-alpha mediated the up-regulation of gene transcripts associated with inflammation and senescence [ IL-1-beta , IL-6 , IL-8 , MCP-1 , COX-2 , p21(WAF1/CIP1) ] and downregulated known tumor-promoting genes ( collagen VI and MMP2 ) . Recombinant overexpression of TNF-alpha receptor(s) in MDA-MB-231 cells also significantly reduced tumor growth , but was not as effective as the TNF-alpha ligand itself in preventing tumor growth . Thus , we propose that stromal cell-mediated delivery of TNF-alpha to human tumors [ using transfected fibroblasts or mesenchymal stem cells ( hMSCs) ] may be a novel and effective strategy for the prevention and treatment of human cancers .
|
[
0,
0,
0,
1,
0,
0,
0,
1,
0,
1
] |
23292149
|
Diallyl disulfide ( DADS ) is a major organo-sulfur compound derived from garlic ( Allium sativum ) , which inhibits the proliferation of various types of cancer cells . In this study we investigated the effect of DADS on the induction of apoptosis , as well as its regulatory effect on the activation of transcription factors in B16F-10 melanoma cells . Treatment of B16F-10 cells with nontoxic concentrations of DADS resulted in the presence of apoptotic bodies and induced DNA fragmentation in a dose-dependent manner . Cell-cycle analysis revealed that the occurrence of the sub-G1 peak was significantly elevated in DADS-treated cells . DADS treatment also down-reguated Bcl-2 expression and up-regulated p53 , caspase-9 , and caspase-3 expression in B16F-10 melanoma cells . The study also reveals that DADS inhibited the activation and nuclear translocation of p65 , p50 , and c-Rel subunits of nuclear factor ( NF)-B and other transcription factors , such as c-fos , activated transcription factor-2 , and cyclic adenosine monophosphate response element-binding protein , in B16F-10 melanoma cells The pro-inflammatory cytokine production and gene expression of tumor necrosis factor ( TNF)-α , interleukin ( IL)-1β , IL-6 , and granulocyte-macrophage colony-stimulating factor ( GM-CSF ) were down-regulated in DADS-treated cells compared with control B16F-10 metastatic melanoma cells . DADS induces caspase-dependent apoptosis through a mitochondria-mediated intrinsic pathway in B16F-10 melanoma cells by activating p53 and caspase-3 gene expression and suppressing pro-inflammatory cytokines and NF-B-mediated Bcl-2 activation .
|
[
0,
0,
0,
0,
0,
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0,
1,
0,
1
] |
20932246
|
Piceatannol is a stilbenoid , a metabolite of resveratrol found in red wine . Piceatannol and sera from rats orally given piceatannol were found to dose-dependently suppress both the proliferation and invasion of AH109A hepatoma cells in culture . Its antiproliferative effect was based on cell cycle arrest at lower concentration ( 25 μM ) and on apoptosis induction at higher concentration ( 100 μM ) . Piceatannol suppressed reactive oxygen species-potentiated invasive capacity by scavenging the intracellular reactive oxygen species . These results suggest that piceatannol , unlike resveratrol , has a potential to suppress the hepatoma proliferation by inducing cell cycle arrest and apoptosis induction . They also suggest that the antioxidative property of piceatannol , like resveratrol , may be involved in its anti-invasive action . Subsequently , piceatannol was found to suppress the growth of solid tumor and metastasis in hepatoma-bearing rats . Thus , piceatannol may be a useful anticancer natural product .
|
[
0,
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0,
0,
0,
0,
0,
0,
0,
0
] |
22496613
|
Sunitinib is an oral multitargeted receptor tyrosine kinase inhibitor with antiangiogenic and antitumor activity that mainly targets vascular endothelial growth factor receptors , and recently , it has been shown to be an active agent for the treatment of malignant pheochromocytomas . Previously , we demonstrated that sunitinib directly inhibited mTORC1 signaling in rat pheochromocytoma PC12 cells . Although autophagy is a highly regulated cellular process , its relevance to cancer seems to be complicated . It is of note that inhibition of mTORC1 is a prerequisite for autophagy induction . Indeed , direct mTORC1 inhibition initiates ULK1/2 autophosphorylation and subsequent Atg13 and FIP200 phosphorylation , inducing autophagy . Here , we demonstrated that sunitinib significantly increased the levels of LC3-II , concomitant with a decrease of p62 in PC12 cells . Following sunitinib treatment , immunofluorescent imaging revealed a marked increased punctate LC3-II distribution . Furthermore , Atg13 knockdown significantly reduced its protein level , which in turn abolished sunitinib-induced autophagy . Moreover , inhibition of autophagy by siRNAs targeting Atg13 or by pharmacological inhibition with ammonium chloride , enhanced both sunitinib-induced apoptosis and anti-proliferation . Thus , sunitinib-induced autophagy is dependent on the suppression of mTORC1 signaling and the formation of ULK1/2-Atg13-FIP200 complexes . Inhibition of autophagy may be a promising therapeutic option for improving the anti-tumor effect of sunitinib .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
0
] |
23269235
|
Keratinocyte growth factor ( KGF , fibroblast growth factor-7 ) is a fibroblast-derived mitogen , which stimulates proliferation of epithelial cells . The expression of KGF by dermal fibroblasts is induced following injury and it promotes wound repair . However , the role of KGF in cutaneous carcinogenesis and cancer progression is not known . We have examined the role of KGF in progression of squamous cell carcinoma ( SCC ) of the skin . The expression of KGF receptor ( KGFR ) mRNA was lower in cutaneous SCCs ( n = 6 ) than in normal skin samples ( n = 6 ) . Expression of KGFR mRNA was detected in 6 out of 8 cutaneous SCC cell lines and the levels were downregulated by 24-h treatment with KGF . KGF did not stimulate SCC cell proliferation , but it reduced invasion of SCC cells through collagen . Gene expression profiling of three cutaneous SCC cell lines treated with KGF for 24 h revealed a specific gene expression signature characterized by upregulation of a set of genes specifically downregulated in SCC cells compared to normal epidermal keratinocytes , including genes with tumor suppressing properties ( SPRY4 , DUSP4 , DUSP6 , LRIG1 , PHLDA1 ) . KGF also induced downregulation of a set of genes specifically upregulated in SCC cells compared to normal keratinocytes , including genes associated with tumor progression ( MMP13 , MATN2 , CXCL10 , and IGFBP3 ) . Downregulation of MMP-13 and KGFR expression in SCC cells and HaCaT cells was mediated via ERK1/2 . Activation of ERK1/2 in HaCaT cells and tumorigenic Ha-ras-transformed HaCaT cells resulted in downregulation of MMP-13 and KGFR expression . These results provide evidence , that KGF does not promote progression of cutaneous SCC , but rather suppresses the malignant phenotype of cutaneous SCC cells by regulating the expression of several genes differentially expressed in SCC cells , as compared to normal keratinocytes .
|
[
1,
0,
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0,
0,
0,
0,
0,
1,
0
] |
22427941
|
Exosomes are small-membrane vesicles secreted by hematopoietic and malignant epithelial cells as well as trophoblasts . The composition of cancerous exosomes has been proven to play pivotal roles in the maintenance of the microenvironment that is beneficial for the progression of cancer , such as Fas-ligand-triggered lymphocyte apoptosis . We supposed that the immunosuppressive effect of cancerous exosomes might be helpful in the treatment of diseases characterized by overactivation of the immune system and subsequent tissue injury . The aim of this study was to evaluate the protective effect of tumor-derived exosomes in the mice model of lipopolysaccharide ( LPS)-induced inflammation . Tetrazolium ( MTT ) and DNA electrophoresis were used to measure the cytotoxicity of exosomes on lymphocytes . Pathologic observation of tissue sections , serologic analysis of aspartate aminotransferase/alanine aminotransferase ( AST/ALT ) , and urinary analysis of protein were used to assess the protection effect of exosomes in LPS-induced multiorgan damage . In vitro outcomes of MTT and DNA electrophoresis showed the cytotoxicity of exosomes on lymphocytes . Together with the alleviation of organ damages evaluated by urine protein , serum AST/ALT , and pathologic analysis , we confirmed the possibility that pretreatment of mice with exosomes , produced by H22 hepatic tumor cells , resulted in protection against LPS-induced tissue damage , which is caused by overactivation of the immune system and inflammation response . This therapeutic strategy will raise an interesting way to search new therapeutics in pairs of diseases with complementarities in etiology and pathology , namely , a strategy of taking advantage of the mutual complementarities between diseases .
|
[
0,
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0,
0,
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0,
0,
0,
1
] |
22984881
|
Although some types of carbon nanotubes ( CNTs ) have been described to induce mesothelioma in rodents and genotoxic effects in various cell systems , there are few previous studies on the genotoxicity of CNTs in mesothelial cells . Here , we examined in vitro DNA damage induction by short multi-wall CNTs ( MWCNTs ; 10-30 nm × 1-2 \u03bcm ) and single-wall CNTs ( SWCNTs ; >50% SWCNTs , other CNTs ; <2 nm × 1-5 \u03bcm ) in human mesothelial ( MeT-5A ) cells and bronchial epithelial ( BEAS 2B ) cells , using the single cell gel electrophoresis ( comet ) assay and the immunoslot blot assay for the detection of malondialdehyde ( M1dG ) DNA adducts . In BEAS 2B cells , we also studied the induction of micronuclei ( MN ) by the CNTs using the cytokinesis-block method . The cells were exposed to the CNTs ( 5-200 \u03bcg/cm(2) , corresponding to 19-760 \u03bcg/ml ) for 24 and 48h in the comet assay and for 48 and 72 h in the MN and M1dG assays . Transmission electron microscopy ( TEM ) showed more MWCNT fibres and SWCNT clusters in BEAS 2B than MeT-5A cells , but no significant differences were seen in intracellular dose expressed as area of SWCNT clusters between TEM sections of the cell lines . In MeT-5A cells , both CNTs caused a dose-dependent induction of DNA damage ( % DNA in comet tail ) in the 48-h treatment and SWCNTs additionally in the 24-h treatment , with a statistically significant increase at 40 \u03bcg/cm(2) of SWCNTs and ( after 48 h ) 80 \u03bcg/cm(2) of both CNTs . SWCNTs also elevated the level of M1dG DNA adducts at 1 , 5 , 10 and 40 \u03bcg/cm(2) after the 48-h treatment , but both CNTs decreased M1dG adduct level at several doses after the 72-h treatment . In BEAS 2B cells , SWCNTs induced a statistically significant increase in DNA damage at 80 and 120 \u03bcg/cm(2) after the 24-h treatment and in M1dG adduct level at 5 \u03bcg/cm(2) after 48 h and 10 and 40 \u03bcg/cm(2) after 72 h ; MWCNTs did not affect the level of DNA damage but produced a decrease in M1dG adducts in the 72-h treatment . The CNTs did not affect the level of MN . In conclusion , MWCNTs and SWCNTs induced DNA damage in MeT-5A cells but showed a lower ( SWCNTs ) or no ( MWCNTs ) effect in BEAS 2B cells , suggesting that MeT-5A cells were more sensitive to the DNA-damaging effect of CNTs than BEAS 2B cells , despite the fact that more CNT fibres or clusters were seen in BEAS 2B than MeT-5A cells . M1dG DNA adducts were induced by SWCNTs but decreased after a 3-day exposure to MWCNTs and ( in MeT-5A cells ) SWCNTs , indicating that CNTs may lead to alterations in oxidative effects within the cells . Neither of the CNTs was able to produce chromosomal damage ( MN ) .
|
[
0,
0,
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0,
0,
0,
0,
0,
0,
0
] |
23266321
|
In both mitotic and meiotic processes , cellular surveillance of the integrity of genetic information transmission from parental cells to their subsequent generations is carried out by a network of proteins primarily involved in cell-cycle regulation , DNA replication , DNA repair , and chromosome segregation . Within this context , the mammalian MRE11 represents an essential multifunctional protein that promotes repair of DNA double-strand breaks and plays a role in the signaling of DNA damage response . Mutations in human hMRE11 gene could contribute to the rare " AT-like " disorder . However , at present time the functional roles of hMRE11 in these cellular processes are elusive . In the current study , we provide evidence that hMRE11 interacts physically with the mismatch repair protein hMLH1 through yeast two-hybrid analysis . In addition , we show that recombinant hMRE11 and hMLH1 proteins interact when these two proteins are coexpressed in bacterial cells , and both proteins can be co-immunoprecipitated from human cell extracts . Furthermore , hMRE11 and hMLH1 display similar expression patterns when examined with a human normal/tumor DNA array . Together , these data suggest that hMRE11 and hMLH1 might act in a co-operative fashion during DNA damage detection , signaling , and repair .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
12509276
|
Abstract The use of immunotherapeutics in melanoma has received much attention , and recent advances to further characterize the regulatory components of the immune system and the importance of co-stimulatory molecules have opened a new area for clinical investigation . Cytotoxic T lymphocyte-associated antigen 4 ( CTLA-4 ) serves as a negative regulator of immunity . Recent trials administering fully human anti-CTLA-4 monoclonal antibodies to melanoma patients have demonstrated clinically meaningful responses . Treatment with CTLA-4 blocking antibodies , however , is not without potential toxicities . Autoimmune side-effects , the most common being colitis-associated diarrhea , are frequently associated with clinical responses . In efforts to build upon prior vaccination efforts as well as attempt to offer patients clinically meaningful immune responses with a CTLA-4 blockade but without significant toxicities , we conducted a clinical trial in patients who previously received autologous tumor cells engineered to secrete granulocyte-macrophage colony stimulating factor ( GVAX ; Cell Genesys , South San Francisco , CA , USA ) with periodic infusions of CTLA-4 blocking antibodies . This sequential treatment resulted in clinically significant anti-tumor immunity without grade 3 or 4 toxicity in most patients . Pathological analyses following treatment of pre-existing tumors revealed a linear correlation between tumor necrosis and the ratio of intra-tumoral CD8+ effector cells to FoxP3+ regulatory cells ( T(regs) ) . Effective anti-tumor immunity and serious autoimmunity can be disassociated . Further targeting of anti-tumor T(regs)in combinatorial therapy approaches may be a rich avenue of future investigation .
|
[
0,
1,
0,
0,
0,
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1,
0,
0
] |
20482528
|
Environmental carcinogen exposure may play an important role in the incidence of cancer in children . In addition to environmental pollutants , maternal smoking during pregnancy may be a contributing factor . Major carcinogenic components of cigarette smoke and other combustion by-products in the environment include polycyclic aromatic hydrocarbons ( PAH ) . Mouse offspring exposed during midpregnancy to the PAH , benzo[a]pyrene ( B[a]P ) , show significant deficiencies in their immune functions , observed in late gestation which persist for at least 18 months . Tumor incidences in these progeny are 8 to 10-fold higher than in controls . We have demonstrated a significant reduction in thymocytes ( CD4+ CD8+ , CD4+ CD8+ Vbeta8+ , CD4+ CD8+ Vgamma2+ ) from newborn and splenocytes ( CD4+ CD8+ ) from 1-week-old mouse progeny exposed to B[a]P in utero . To investigate possible causes of the observed T cell reduction , we analyzed the thymocytes and splenocytes from progeny and maternal tissues for the presence of B[a]P-DNA adducts . Adducts were detected in maternal , placental and offspring lymphoid tissues at day 19 of gestation , at birth and 1-wk after birth . The presence of B[a]P-DNA adducts in immature T cells may , in part , explain the previously observed T cell immunosuppression and tumor susceptibility in mice exposed to B[a]P in utero . The effects of DNA lesions on progeny T cells may include interference with normal T-cell development . These results provide a possible explanation for the relationship between maternal smoking during pregnancy and childhood carcinogenesis .
|
[
0,
1,
0,
0,
0,
1,
0,
0,
0,
0
] |
12375734
|
Imiquimod is a synthetic compound with antitumor properties ; a 5% cream formulation is successfully used to treat skin tumors . The antitumor effect of imiquimod is multifactorial , although its ability to modulate immune responses by triggering TLR7/8 is thought to be key . Among the immune cells suggested to be involved are plasmacytoid DCs ( pDCs ) . However , a direct contribution of pDCs to tumor killing in vivo and the mechanism of their recruitment to imiquimod-treated sites have never been demonstrated . Using a mouse model of melanoma , we have now demonstrated that pDCs can directly clear tumors without the need for the adaptive immune system . Topical imiquimod treatment led to TLR7-dependent and IFN-α/β receptor 1-dependent ( IFNAR1-dependent ) upregulation of expression of the chemokine CCL2 in mast cells . This was essential to induce skin inflammation and for the recruitment of pDCs to the skin . The recruited pDCs were CD8α+ and induced tumor regression in a TLR7/MyD88- and IFNAR1-dependent manner . Lack of TLR7 and IFNAR1 or depletion of pDCs or CD8α+ cells from tumor-bearing mice completely abolished the effect of imiquimod . TLR7 was essential for imiquimod-stimulated pDCs to produce IFN-α/β , which led to TRAIL and granzyme B secretion by pDCs via IFNAR1 signaling . Blocking these cytolytic molecules impaired pDC-mediated tumor killing . Our results demonstrate that imiquimod treatment leads to CCL2-dependent recruitment of pDCs and their transformation into a subset of killer DCs able to directly eliminate tumor cells .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
1
] |
22251703
|
Dendritic cells ( DCs ) and natural killer ( NK ) cells are central components of innate immunity for controlling tumor growth . The therapeutic effects of certain anti-myeloma drugs are partially mediated by targeting the innate immune response . In addition , novel types of natural compounds have been developed that efficiently modulate the activity of both the cellular and humoral compartments of immunity . MGN-3 is known as an activator of natural killer cells , inducer of apoptosis and cytokine production , and modulator of dendritic cell maturation and differentiation in vitro . We have performed a randomized , placebo-controlled study to examine the effects of MGN-3 on innate immune system parameters in 48 multiple myeloma patients . We performed immunophenotypic analysis of peripheral blood samples , determined NK cell activity , and assessed the cytokine profiles of plasma before and during 3months of treatment . The results demonstrate a clear increase in NK activity in MGN-3-treated patients compared to the placebo group , an increased level of myeloid DCs in peripheral blood , and augmented concentrations of T helper cell type 1-related cytokines . The present study suggests that MGN-3 may represent an immunologically relevant product for activating innate immunity in multiple myeloma patients and warrants further testing to demonstrate clinical efficacy .
|
[
0,
1,
0,
0,
0,
0,
0,
0,
0,
0
] |
22941038
|
Genetic defects in breast cancer ( BC ) susceptibility genes , most importantly BRCA1 and BRCA2 , account for of hereditary BC and ovarian cancer ( OC ) . Little is known about the contribution of constitutive ( soma-wide ) epimutations to the remaining cases . We developed bisulfite pyrosequencing assays to screen >600 affected BRCA1/BRCA2 mutation-negative patients from the German Consortium for Hereditary Breast and Ovarian Cancer for constitutive hypermethylation of ATM , BRCA1 , BRCA2 , RAD51C , PTEN and TP53 in blood cells . In a second step , patients with \u22656% promoter methylation were analyzed by bisulfite plasmid sequencing to demonstrate the presence of hypermethylated alleles ( epimutations ) , indicative of epigenetic gene silencing . Altogether we identified nine ( 1.4% ) patients with constitutive BRCA1 and three ( 0.5% ) with RAD51C hypermethylation . Epimutations were found in both sporadic cases , in particular in 2 ( 5.5% ) of 37 patients with early-onset BC , and familial cases , in particular 4 ( 10% ) of 39 patients with OC . Hypermethylation was always confined to one of the two parental alleles in a subset ( 12-40% ) of the analyzed cells . Because epimutations occurred in cell types from different embryonal layers , they most likely originated in single cells during early somatic development . We propose that analogous to germline genetic mutations constitutive epimutations may serve as the first hit of tumor development . Because the role of constitutive epimutations in cancer development is likely to be largely underestimated , future strategies for effective testing of susceptibility to BC and OC should include an epimutation screen .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
22843497
|
We studied the effect of oxidative stress induced by hyperoxia , hydrogen peroxide , or menadione on mouse leukemia P388 cells at early ( 4 days ) and late ( 7 days ) stages of tumor growth . Oxidative stress proved to inhibit cell division and to induce apoptosis . Seven-day leukemia cells feature lower proliferative potential and higher sensitivity to oxidative stress and platidiam .
|
[
0,
0,
0,
0,
0,
0,
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1,
0,
1
] |
12561326
|
Oncostatin M ( OSM ) , an interleukin-6 type cytokine , acts via the gp130 signaling receptor to inhibit proliferation and induce differentiation of breast cancer cells . EGF , a mitogen for breast cells , signals via EGFR/ErbB tyrosine kinase receptors which are implicated in breast cancer pathogenesis . Here we show paradoxically that EGF enhanced the OSM-induced inhibition of proliferation and induction of cellular differentiation in both estrogen receptor positive and negative breast cancer cells . This functional synergism was also seen with heregulin but not SCF , PDGF or IGF-1 , indicating that it was specific to EGF-related growth factors . Immunoprecipitation experiments revealed that gp130 was constitutively associated with ErbB-2 and ErbB-3 . There was a similar association between the OSMRbeta and ErbB-2 . Furthermore , EGF unexpectedly induced tyrosine phosphorylation of gp130 . We show that OSM induced phosphorylation of STAT3 . Both OSM and EGF activated the p42/44 MAP kinases , but while the MEK inhibitor , PD98059 , ablated the OSM-induced inhibition , it only partially ablated the inhibitory effects of OSM plus EGF . Thus , we have demonstrated that the receptors and signalling pathways of two apparently unrelated growth factors were intimately linked , resulting in an unexpected biological effect . This provides a new mechanism for generating signalling diversity and has potential clinical implications in breast cancer .
|
[
0,
0,
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0,
0,
0,
0,
0,
1,
0
] |
11821958
|
In order to elucidate the mechanisms by which tumour-specific CD4+ T-cell responses are impaired during tumour development , an attempt was made to identify factors which impair CD4+ T-cell responses at a late tumour-bearing stage . Plasma from mice bearing B16 melanoma for 30 days ( plasma d30 ) showed a more profound immunosuppressive effect on the in vitro proliferation of unrelated antigen-specific CD4+ T cells in the presence of both antigen and antigen-presenting cells ( APC ) than plasma from na�ve mice . The level of plasma transforming growth factor ( TGF)- was elevated in mice bearing B16 melanoma for 30 days compared with na�ve mice , and the suppressive effect of plasma d30 was partially diminished by the neutralization of TGF- . Interestingly , immunoglobulin ( IgG)-bound TGF- , but not IgG-unbound TGF- , in plasma d30 was suggested to be responsible for the immunosuppressive activity . In addition , no suppressive effect of plasma d30 was observed when antigen was added as a class II peptide , thus suggesting that the impaired proliferation of CD4+ T cells in the presence of plasma d30 was due to a dysfunction of antigen uptake/processing by APC . Furthermore , dissociation between IgG and TGF- resulted in a loss of the suppressive activity of plasma d30 . Taken together , these results suggest that circulating IgG-bound TGF- is , at least in part , responsible for the impaired responses of CD4+ T cells at the late tumour-bearing stage by suppressing antigen uptake/ processing by APC .
|
[
0,
1,
0,
0,
0,
0,
0,
0,
1,
0
] |
12041507
|
cDNA microarray-based gene expression analysis has been successfully employed to explore the action mechanism and to validate the targets of several drugs . In the present study , we evaluated anti-angiogenic activity of demethoxycurcumin ( DC ) , a structural analog of curcumin , isolated from Curcuma aromatica , and investigated the effect of DC on genetic reprogramming in cultured human umbilical vein endothelial cells ( HUVECs ) using cDNA microarray analysis . Of 1024 human cancer-focused genes arrayed , 187 genes were up-regulated and 72 genes were down-regulated at least 2-fold by DC . Interestingly , 9 angiogenesis-related genes were down-regulated over 5-fold in response to DC , suggesting that the genetic reprogramming was crucially involved in anti-angiogenesis by the compound . To verify the results obtained from cDNA microarray analysis , matrix metalloproteinase-9 ( MMP-9 ) , the product of one of the angiogenesis-related genes down-regulated over 5-fold by DC , was investigated using gelatin zymography . DC potently inhibited the expression of MMP-9 , yet showed no direct effect on its activity . These data show that gene expressional change of MMP-9 is a major mediator for angiogenesis inhibition by DC . All genes identified and microarray data are available on the web at http://dasan.sejong.ac.kr/
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
12495478
|
Whereas estrogen-estrogen receptor α ( ER ) signaling plays an important role in breast cancer growth , it is also necessary for the differentiation of normal breast epithelial cells . How this functional conversion occurs , however , remains unknown . Based on a genome-wide sequencing study that identified mutations in several breast cancer genes , we examined some of the genes for mutations , expression levels , and functional effects on cell proliferation and tumorigenesis . We present the data for C1orf64 or ER-related factor ( ERRF ) from 31 cell lines and 367 primary breast cancer tumors . Whereas mutation of ERRF was infrequent ( 1 of 79 or 1.3% ) , its expression was up-regulated in breast cancer , and the up-regulation was more common in lower-stage tumors . In addition , increased ERRF expression was significantly associated with ER and/or progesterone receptor ( PR ) positivity , which was still valid in human epidermal growth factor receptor 2 ( HER2)-negative tumors . In ER-positive tumors , ERRF expression was inversely correlated with HER2 status . Furthermore , higher ERRF protein expression was significantly associated with better disease-free survival and overall survival , particularly in ER- and/or PR-positive and HER2-negative tumors ( luminal A subtype ) . Functionally , knockdown of ERRF in two ER-positive breast cancer cell lines , T-47D and MDA-MB-361 , suppressed cell growth in vitro and tumorigenesis in xenograft models . These results suggest that ERRF plays a role in estrogen-ER-mediated growth of breast cancer cells and could , thus , be a potential therapeutic target .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
22341523
|
Proliferating cells consume more glucose to cope with the bioenergetics and biosynthetic demands of rapidly dividing cells as well as to counter a shift in cellular redox environment . This study investigates the hypothesis that manganese superoxide dismutase ( MnSOD ) regulates cellular redox flux and glucose consumption during the cell cycle . A direct correlation was observed between glucose consumption and percentage of S-phase cells in MnSOD wild-type fibroblasts , which was absent in MnSOD homozygous knockout fibroblasts . Results from electron paramagnetic resonance spectroscopy and flow cytometric assays showed a significant increase in cellular superoxide levels in S-phase cells , which was associated with an increase in glucose and oxygen consumption , and a decrease in MnSOD activity . Mass spectrometry results showed a complex pattern of MnSOD-methylation at both lysine ( 68 , 89 , 122 , and 202 ) and arginine ( 197 and 216 ) residues . MnSOD protein carrying a K89A mutation had significantly lower activity compared with wild-type MnSOD . Computational-based simulations indicate that lysine and arginine methylation of MnSOD during quiescence would allow greater accessibility to the enzyme active site as well as increase the positive electrostatic potential around and within the active site . Methylation-dependent changes in the MnSOD conformation and subsequent changes in the electrostatic potential around the active site during quiescence versus proliferation could increase the accessibility of superoxide , a negatively charged substrate . These results support the hypothesis that MnSOD regulates a " metabolic switch " during progression from quiescent through the proliferative cycle . We propose MnSOD as a new molecular player contributing to the Warburg effect .
|
[
0,
0,
1,
0,
0,
1,
0,
0,
1,
0
] |
22710435
|
PURPOSE Idarubicin is a synthetic anthracycline anticancer drug widely used in the treatment of some hematological malignancies . The studies in our laboratory have clearly demonstrated that idarubicin can undergo reductive bioactivation by NADPH-cytochrome P450 reductase to free radicals with resulting formation of DNA strand breaks , which can potentially contribute to its genotoxic effects [ Celik , H. , Arinç , E. , Bioreduction of idarubicin and formation of ROS responsible for DNA cleavage by NADPH-cytochrome P450 reductase and its potential role in the antitumor effect . J Pharm Pharm Sci , 11(4):68-82 , 2008 ] . In the current study , our aim was to investigate the possible protective effects of several phenolic antioxidants , quercetin , rutin , naringenin , resveratrol and trolox , against the DNA-damaging effect of idarubicin originating from its P450 reductase-catalyzed bioactivation . METHODS DNA damage was measured by detecting single-strand breaks in plasmid pBR322 DNA using a cell-free agarose gel method . RESULTS Our results indicated that , among the compounds tested , quercetin was the most potent antioxidant in preventing DNA damage . Quercetin significantly decreased the extent of DNA strand breaks in a dose-dependent manner ; 100 microM of quercetin almost completely inhibited the DNA strand breakage . Unlike quercetin , its glycosidated conjugate rutin , failed to provide any significant protection against idarubicin-induced DNA strand breaks except at the highest concentration tested ( 2 mM ) . The protective effects of other antioxidants were significantly less than that of quercetin even at high concentrations . Quercetin was found to be also an effective protector against DNA damage induced by mitomycin C. CONCLUSION We conclude that quercetin , one of the most abundant flavonoids in the human diet , is highly effective in reducing the DNA damage caused by the antitumor agents , idarubicin and mitomycin C , following bioactivation by P450 reductase .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
20816008
|
Colon cancer is a leading cause of morbidity and mortality in Western countries . Basic fibroblast growth factor ( bFGF ) was up-regulated in patients with colon cancer and was considered as a potential therapeutic target . In this study , we first demonstrated that a novel bFGF-binding peptide ( named P7 ) inhibited proliferation of several colon cancer cell lines including HT-29 , LoVo , and Caco2 cells stimulated by bFGF . Further investigations with HT-29 cells indicated that P7 arrested the cell cycle at the G0/G1 phase of bFGF-stimulated cells , reduced the levels of phospho-Erk1/Erk2 induced by bFGF , and caused significant changes in the expression of proteins related to proliferation , cell cycle , and cancer . Our results suggested that the bFGF-binding peptide has a potential antitumor effect on colon cancer .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
20331620
|
SMG1 is a member of the phosphoinositide kinase-like kinase family of proteins that includes ATM , ATR , and DNA-PK , proteins with known roles in DNA damage and cellular stress responses . SMG1 has a well-characterized role in nonsense-mediated decay as well as suggested roles in the DNA damage response , resistance to oxidative stress , regulation of hypoxic responses , and apoptosis . To understand the roles of SMG1 further , we generated a Genetrap Smg1 mouse model . Smg1 homozygous KO mice were early embryonic lethal , but Smg1 heterozygous mice showed a predisposition to a range of cancers , particularly lung and hematopoietic malignancies , as well as development of chronic inflammation . These mice did not display deficiencies in known roles of SMG1 , including nonsense-mediated decay . However , they showed elevated basal tissue and serum cytokine levels , indicating low-level inflammation before the development of tumors . Smg1 heterozygous mice also showed evidence of oxidative damage in tissues . These data suggest that the inflammation observed in Smg1 haploinsufficiency contributes to susceptibility to cancer and that Smg1-deficient animals represent a model of inflammation-enhanced cancer development .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
1
] |
23277562
|
Extracellular respiration of solid-phase electron acceptors in some microorganisms requires a complex chain of multiheme c-type cytochromes that span the inner and outer membranes . In Shewanella species , MtrA , an periplasmic decaheme c-type cytochrome , is an essential component for extracellular respiration of iron(III) . The exact mechanism of electron transport has not yet been resolved , but the arrangement of the polypeptide chain may have a strong influence on the capability of the MtrA cytochrome to transport electrons . The iron hemes of MtrA are bound to its polypeptide chain via proximal ( CXXCH ) and distal histidine residues . In this study , we show the effects of mutating histidine residues of MtrA to arginine on protein expression and extracellular respiration using Shewanella sp. strain ANA-3 as a model organism . Individual mutations to six out of nine proximal histidines in CXXCH of MtrA led to decreased protein expression . However , distal histidine mutations resulted in various degrees of protein expression . In addition , the effects of histidine mutations on extracellular respiration were tested using ferrihydrite and current production in microbial fuel cells . These results show that proximal histidine mutants were unable to reduce ferrihydrite . Mutations to the distal histidine residues resulted in various degrees of ferrihydrite reduction . These findings indicate that mutations to the proximal histidine residues affect MtrA expression , leading to loss of extracellular respiration ability . In contrast , mutations to the distal histidine residues are less detrimental to protein expression , and extracellular respiration can proceed .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
22923588
|
BACKGROUND Astaxanthin modulates immune response , inhibits cancer cell growth , reduces bacterial load and gastric inflammation , and protects against UVA-induced oxidative stress in in vitro and rodent models . Similar clinical studies in humans are unavailable . Our objective is to study the action of dietary astaxanthin in modulating immune response , oxidative status and inflammation in young healthy adult female human subjects . METHODS Participants ( averaged 21.5 yr ) received 0 , 2 , or 8 mg astaxanthin ( n = 14/diet ) daily for 8 wk in a randomized double-blind , placebo-controlled study . Immune response was assessed on wk 0 , 4 and 8 , and tuberculin test performed on wk 8 . RESULTS Plasma astaxanthin increased ( P < 0.01 ) dose-dependently after 4 or 8 wk of supplementation . Astaxanthin decreased a DNA damage biomarker after 4 wk but did not affect lipid peroxidation . Plasma C-reactive protein concentration was lower ( P < 0.05 ) on wk 8 in subjects given 2 mg astaxanthin . Dietary astaxanthin stimulated mitogen-induced lymphoproliferation , increased natural killer cell cytotoxic activity , and increased total T and B cell subpopulations , but did not influence populations of Thelper , Tcytotoxic or natural killer cells . A higher percentage of leukocytes expressed the LFA-1 marker in subjects given 2 mg astaxanthin on wk 8 . Subjects fed 2 mg astaxanthin had a higher tuberculin response than unsupplemented subjects . There was no difference in TNF and IL-2 concentrations , but plasma IFN-gamma and IL-6 increased on wk 8 in subjects given 8 mg astaxanthin . CONCLUSION Therefore , dietary astaxanthin decreases a DNA damage biomarker and acute phase protein , and enhances immune response in young healthy females .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
1
] |
20205737
|
The suppression of MAPK and oxidative stress could be an important anti-inflammatory mechanism . Fucoidan regulates MAPK activity in several cell lines . However , the mechanism for the anti-inflammatory and oxidative stress effect of low molecular weight fucoidan ( LMWF ) is poorly understood in RAW264.7 cells . The objective of this study was to examine the critical role of LMWF during LPS-induced inflammation in RAW264.7 cells . To determine the potential role of LMWF , we analysed pro-inflammatory cytokine , transcription factor , inflammation-related and oxidative stress related protein expression in vitro during LPS-induced inflammation in RAW264.7 cells . In this study , we demonstrated the anti-inflammatory effects of LMWF on LPS-induced inflammation in macrophages through the regulation of signalling pathways , including its effect on the attenuation of inflammatory cytokines , such as IL-1β , IL-1 and TNF-α , and the degradation of phosphorylated p38 MAPK , ERK1/2 and JNK . This study also demonstrates that LMWF might block NO as well as the expression of reactive oxygen species ( ROS ) , which subsequently inhibits the iNOS and COX-2 expression induced by LPS . Based on these findings , we suggest that LMWF might have great potential as an external pathogen prevention and intervention agent for inflammatory diseases .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
1
] |
23006539
|
Although prostate cancer ( CaP ) is the most frequently diagnosed malignant tumor in American men , the mechanisms underlying the development and progression of CaP remain largely unknown . Recent studies have shown that downregulation of the microRNA miR-124 occurs in several types of human cancer , suggesting a tumor suppressive function of miR-124 . Until now , however , it has been unclear whether miR-124 is associated with CaP . In the present study , we completed a series of experiments to understand the functional role of miR-124 in CaP . We detected the expression level of miR-124 in clinical CaP tissues , evaluated the influence of miR-124 on the growth of CaP cells and investigated the mechanism underlying the dysregulation of miR-124 . We found that ( i ) miR-124 directly targets the androgen receptor ( AR ) and subsequently induces an upregulation of p53 ; ( ii ) miR-124 is significantly downregulated in malignant prostatic cells compared to benign cells , and DNA methylation causes the reduced expression of miR-124 ; and ( iii ) miR-124 can inhibit the growth of CaP cells in vitro and in vivo . Data from this study revealed that loss of miR-124 expression is a common event in CaP , which may contribute to the pathogenesis of CaP . Our studies also suggest that miR-124 is a potential tumor suppressive gene in CaP , and restoration of miR-124 expression may represent a novel strategy for CaP therapy.Oncogene advance online publication , 15 October 2012 ; doi:10.1038/onc.2012.425 .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
23069658
|
BACKGROUND Oxidative stress and inflammation are important steps in the pathogenesis of atherosclerosis . We postulated that therapeutic concentrations of aspirin and pravastatin , especially in combination , may suppress oxidative stress and inflammation in endothelial cells , and this concept was examined in human coronary artery endothelial cells ( HCAECs ) . METHODS Human coronary artery endothelial cells were cultured and treated with oxidized-low density lipoprotein ( ox-LDL , 60 microg/ml for 24 hours ) alone , or pre-treated with aspirin ( 1 , 2 or 5 mmol/L ) , pravastatin ( 1 , 5 or 10 micromol/L ) or their combination ( 1 mmol/L aspirin and 5 micromol/L pravastatin ) , followed by ox-LDL treatment . After respective treatment , superoxide anion production , p38 mitogen activated protein kinase and transcription factor NF-kappaB activation , protein expression of lectin-like ox-LDL receptor-1 ( LOX-1 ) and adhesion molecules , and monocyte adhesion were measured . RESULTS Ox-LDL treatment greatly elicited its receptor LOX-1 expression , superoxide anion production and inflammatory response , which were minimally affected by low concentration of aspirin ( 1 mmol/L ) or pravastatin ( 5 micromol/L ) , but were markedly decreased by their combination . Activation of p38 mitogen activated protein kinase and NF-kappaB , the expression of intercellular adhesion molecule-1 and monocyte chemotactic protein-1 , which were only mildly affected by aspirin or pravastatin alone , were significantly attenuated by their combination . As a consequence , monocyte adhesion to endothelial cells was markedly attenuated by the combination of the two agents . Well-known anti-oxidants alpha-tocopherol and gamma-tocopherol had similar inhibitory effects on ox-LDL-mediated oxidative stress and LOX-1 expression as well as monocyte adhesion as did the combination of aspirin and pravastatin . CONCLUSIONS These studies point to a positive interaction between aspirin and pravastatin with regard to endothelial biology . Anti-oxidant and subsequent anti-inflammatory effect may be one of the potential underling mechanisms .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
1
] |
20819511
|
BACKGROUND Modulation of the expression of retinoic acid receptors ( RAR ) alpha and gamma in adult rat prostate by testosterone ( T ) suggests that RAR signaling events might mediate some of the androgen effects on prostate cells . METHOD In this study , we examined the interactions between T and retinoic acid ( RA ) in cell growth of human prostate carcinoma cells , LNCaP , and their relationship with the expression of RAR and epidermal growth factor receptor ( EGF-R ) . RESULTS Both T and RA , when administered alone , stimulated 3H-thymidine incorporation in LNCaP cells in a dose-dependent manner ; the effect of each agent was reciprocally attenuated by the other agent . Testosterone treatment of LNCaP cells also resulted in dose dependent , biphasic increases in RAR alpha and gamma mRNAs ; increases paralleled that of 3H-thymidine incorporation and were attenuated by the presence of 100 nM RA . These results suggest a link between RAR signaling and the effect of T on LNCaP cell growth . Gel electrophoretic mobility shift assays revealed the presence of putative androgen responsive element ( ARE ) in the promoter region of RAR alpha gene , suggesting that a direct AR-DNA interaction might mediate the effects of T on RAR alpha gene . Furthermore , treatment of LNCaP cells with 20 nM T resulted in an increase in EGF-R . In contrast , EGF-R was suppressed by 100 nM RA that also suppressed the effect of T. CONCLUSIONS Current results demonstrate interactions between T and RA in the expression of RARs and cell growth in LNCaP cells . The presence of putative ARE in the promoter of the RAR alpha gene suggests that AR-DNA interaction might mediate the effects of T on RAR alpha gene . The opposite effects of T and RA on the expression of RAR and EGF-R suggest that signal events of these receptors might be involved in the interaction between T and RA in the control of LNCaP cell growth .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
12069693
|
The current study was undertaken to study the effect of a macerated extract of Nigella sativa seeds in normal as well as in tumour bearing mice against gamma radiation-induced cellular damage to normal tissues . This was done to mimic the clinical setting where in , normal tissues of cancer patients undergoing radiotherapy are exposed to the deleterious effects of radiation . The protection of cellular DNA was analysed in peripheral blood leucocytes of whole body irradiated mice following pretreatment with macerated extract of Nigella sativa seeds ( 100 mg/kg ) , using alkaline comet assay , and also estimating biochemical and blood parameters such as levels of antioxidant enzymes superoxide dismutase and catalase , thiobarbituric acid reactive substances and protein oxidation in organs such as spleen , liver , brain and intestine haemoglobin and total leucocyte count , respectively . The results showed that the macerated extract of Nigella sativa seeds protected the liver , spleen , brain and intestines both in normal as well as tumour bearing mice . This study concludes that macerated extract of Nigella sativa seeds has protective effects against radiation-induced damage and biochemical alterations which could be attributed to the ability to scavenge free radicals and its antioxidant properties . Hence macerated extract of Nigella sativa seeds , could be used in combination with radiation to protect against oxidative stress in normal tissues and improving the quality of life of cancer patients by mitigating unwanted side effects of radiation in normal tissues .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
1
] |
23716868
|
BACKGROUND/AIMS Adenocarcinoma of the gallbladder is a highly malignant neoplasm. p16 is a tumor suppressor gene protein , which is a cyclin-dependent kinase inhibitor that regulates the G1-S phase of the cell cycle . The purpose of the present study was to investigate the expression of p16 in gallbladder carcinoma and its precancerous conditions and to examine the relationship between p16 expression and clinicopathological parameters . METHODOLOGY Formalin-fixed , paraffin-embedded tissue sections from 20 cases of normal gallbladder , 20 cases of chronic cholecystitis , 20 cases of gallbladder adenoma , 20 cases of dysplasia , and 58 cases of adenocarcinoma were examined . The expression of p16 was evaluated by immunohistochemical analysis . RESULTS In normal gallbladder , no expression of p16 was found . In chronic cholecystitis , expression of p16 was not found . In gallbladder adenomas , expression of p16 was found in 20% ( 4/20 ) . In low grade dyspalsias , expression of p16 was not found . In high grade dysplasias , p16 expression was present in 45.0% ( 9/20 ) . In gallbladder adenocarcinomas , p16 expression was found in 27.6% ( 16/58 ) . Expression of p16 correlated significantly with histologic grade ( p < 0.05 ) . No correlation was found between p16 expression and age , gender , tumor size , gross type , location , vascular invasion , lymph node metastasis , and TNM stage , respectively . CONCLUSIONS P16 protein overexpression is an early and relatively common event in carcinogenesis of gallbladder carcinoma . Expression of p16 protein is absent in normal or chronic cholecystitis . Expression of p16 may be an ancillary diagnostic marker of gallbladder carcinoma and its precancerous conditions .
|
[
0,
0,
0,
0,
1,
0,
0,
0,
0,
0
] |
20422865
|
BACKGROUND Luteolin is a 3',4',5,7-tetrahydroxyflavone found in various fruits and vegetables . We have shown previously that luteolin reduces HT-29 cell growth by inducing apoptosis and cell cycle arrest . The objective of this study was to examine whether luteolin downregulates the insulin-like growth factor-I receptor ( IGF-IR ) signaling pathway in HT-29 cells . METHODS In order to assess the effects of luteolin and/or IGF-I on the IGF-IR signaling pathway , cells were cultured with or without 60 μmol/L luteolin and/or 10 nmol/L IGF-I . Cell proliferation , DNA synthesis , and IGF-IR mRNA levels were evaluated by a cell viability assay , [ 3H]thymidine incorporation assays , and real-time polymerase chain reaction , respectively . Western blot analyses , immunoprecipitation , and in vitro kinase assays were conducted to evaluate the secretion of IGF-II , the protein expression and activation of IGF-IR , and the association of the p85 subunit of phophatidylinositol-3 kinase ( PI3K ) with IGF-IR , the phosphorylation of Akt and extracellular signal-regulated kinase ( ERK)1/2 , and cell division cycle 25c ( CDC25c ) , and PI3K activity . RESULTS Luteolin ( 0 - 60 μmol/L ) dose-dependently reduced the IGF-II secretion of HT-29 cells . IGF-I stimulated HT-29 cell growth but did not abrogate luteolin-induced growth inhibition . Luteolin reduced the levels of the IGF-IR precursor protein and IGF-IR transcripts . Luteolin reduced the IGF-I-induced tyrosine phosphorylation of IGF-IR and the association of p85 with IGF-IR . Additionally , luteolin inhibited the activity of PI3K activity as well as the phosphorylation of Akt , ERK1/2 , and CDC25c in the presence and absence of IGF-I stimulation . CONCLUSIONS The present results demonstrate that luteolin downregulates the activation of the PI3K/Akt and ERK1/2 pathways via a reduction in IGF-IR signaling in HT-29 cells ; this may be one of the mechanisms responsible for the observed luteolin-induced apoptosis and cell cycle arrest .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
1,
0
] |
22269172
|
Ataxia-telangiectasia mutated ( ATM ) is a cellular damage sensor that coordinates the cell cycle with damage-response checkpoints and DNA repair to preserve genomic integrity . However , ATM also has been implicated in metabolic regulation , and ATM deficiency is associated with elevated reactive oxygen species ( ROS ) . ROS has a central role in many physiological and pathophysiological processes including inflammation and chronic diseases such as atherosclerosis and cancer , underscoring the importance of cellular pathways involved in redox homeostasis . We have identified a cytoplasmic function for ATM that participates in the cellular damage response to ROS . We show that in response to elevated ROS , ATM activates the TSC2 tumor suppressor via the LKB1/AMPK metabolic pathway in the cytoplasm to repress mTORC1 and induce autophagy . Importantly , elevated ROS and dysregulation of mTORC1 in ATM-deficient cells is inhibited by rapamycin , which also rescues lymphomagenesis in Atm-deficient mice . Our results identify a cytoplasmic pathway for ROS-induced ATM activation of TSC2 to regulate mTORC1 signaling and autophagy , identifying an integration node for the cellular damage response with key pathways involved in metabolism , protein synthesis , and cell survival .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
1
] |
20160076
|
Ubiquitylation is a reversible post-translational modification that has emerged as a key regulator of most complex cellular processes . It may rival phosphorylation in scope and exceed it in complexity . The dynamic nature of ubiquitylation events is important for governing protein stability , maintaining ubiquitin homeostasis and controlling ubiquitin-dependent signalling pathways . The human genome encodes active deubiquitylating enzymes ( DUBs , also referred to as deubiquitinases ) , which exhibit distinct specificity profiles towards the various ubiquitin chain topologies . As a result of their ability to reverse ubiquitylation , these enzymes control a broad range of key cellular processes . In this Commentary we discuss the cellular functions of DUBs , such as their role in governing membrane traffic and protein quality control . We highlight two key signalling pathways--the Wnt and transforming growth factor \u03b2 ( TGF-\u03b2 ) pathways , for which dynamic ubiquitylation has emerged as a key regulator . We also discuss the roles of DUBs in the nucleus , where they govern transcriptional activity and DNA repair pathways .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
22357969
|
BACKGROUND Nucleolin is one of the major proteins of the nucleolus , but it is also expressed on the cell surface where is serves as a binding protein for variety of ligands implicated in tumorigenesis and angiogenesis . Emerging evidence suggests that the cell-surface expressed nucleolin is a strategic target for an effective and nontoxic cancer therapy . METHODOLOGY/PRINCIPAL FINDINGS By monitoring the expression of nucleolin mRNA , and by measuring the level of nucleolin protein recovered from the surface and nucleus of cells , here we show that the presence of nucleolin at the cell surface is dependent on the constant induction of nucleolin mRNA . Indeed , inhibitors of RNA transcription or translation block expression of surface nucleolin while no apparent effect is observed on the level of nucleolin in the nucleus . The estimated half-life of surface nucleolin is less than one hour , whereas that of nuclear nucleolin is more than 8 hours . Nucleolin mRNA induction is reduced markedly in normal fibroblasts that reach confluence , while it occurs continuously even in post-confluent epithelial tumor cells consistent with their capacity to proliferate without contact inhibition . Interestingly , cold and heat shock induce nucleolin mRNA concomitantly to enhanced mRNA expression of the heat shock protein 70 , thus suggesting that surface nucleolin induction also occurs in response to an environmental insult . At the cell surface , one of the main functions of nucleolin is to shuttle specific extracellular ligands by an active transport mechanism , which we show here to be calcium dependent . CONCLUSION/SIGNIFICANCE Our results demonstrate that the expression of surface nucleolin is an early metabolic event coupled with tumor cell proliferation and stress response . The fact that surface nucleolin is constantly and abundantly expressed on the surface of tumor cells , makes them a preferential target for the inhibitory action of anticancer agents that target surface nucleolin .
|
[
0,
0,
0,
0,
1,
0,
0,
0,
1,
0
] |
21203423
|
The aim of the present study was to investigate the apoptosis of human ovarian cancer cell lines , A2780 and CP70 , induced by a novel curcumin analogue , B19 . The proliferation of cells was detected with methyl thiazolyl tetrazolium ( MTT ) assay and apoptosis was examined by flow cytometry . Reactive oxygen species ( ROS ) were assessed by the fluorescent indicator DCF-DA . The protein expression of the endoplasmic reticulum ( ER ) stress pathways , GRP78 , XBP-1 , ATF-4 and CHOP , was examined with Western blotting . A growth inhibitory effect was observed after treatment with B19 in a dose-dependent manner and with more potential than curcumin . At 20 �M , B19 induced significant apoptosis in CP70 cells . Furthermore , B19 induced the ER stress response , while curcumin had no effect on ER stress . These results suggest that B19 has more effective antitumor properties than curcumin , and is associated with the activation of ER stress and ROS in ovarian cancer cells .
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[
0,
0,
0,
0,
0,
0,
0,
1,
0,
0
] |
22159410
|
OBJECTIVE To evaluate the Expression and correlation of cyclooxygenase-2 ( COX-2 ) and vascular endothelial growth factor receptor ( VEGF ) in nasopharyngeal carcinoma . METHOD In this study , expression levels of COX-2 , VEGF were examined in 58 patients with nasopharyngeal carcinoma and 38 patients with inflammation in nasopharyngeal mucosa by immunohistochemistry method . RESULT The expression of COX-2 , VEGF were higher in nasopharyngeal carcinoma than those in nasopharyngeal mucosa ( P < 0.05 ) , and they had some correlation with the invasion and lymphatic metastasis and with the clinical stage of nasopharyngeal carcinoma ( P < 0.05 ) . The expression of COX-2 was positively correlated with that of VEGF ( P < 0.05 ) . CONCLUSION The coexpression of COX-2 and VEGF may play animportant role in the carcinogenesis and development of nasopharyngeal carcinoma , and they may prom ( see text ) lymph node metastasis of nasopharyngeal carcinoma .
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[
1,
0,
0,
0,
0,
0,
1,
0,
1,
1
] |
22803408
|
In normal tissues , strict control of tissue size is achieved by regulating cell numbers . The mechanism that controls total cell number is known as contact inhibition of growth and it depends on the NF2/Merlin pathway . Negative regulation of this pathway by deleterious mutations or by oncogenes results in cell transformation and tumor progression . Here we provide evidence that the CD43 sialomucin cooperates with oncogenic signals to promote cell transformation by abrogating the contact inhibition of growth through a molecular mechanism that involves AKT-dependent Merlin phosphorylation and degradation . Accordingly , inhibition of endogenous CD43 expression by RNA interference in lung , cervix and colon human cancer cells impaired tumor growth in vivo . These data underscore a previously unidentified role for CD43 in non-hematopoietic tumor progression .
|
[
0,
0,
0,
0,
1,
0,
0,
0,
0,
0
] |
24260485
|
The calcium-calmodulin activated kinase CaMKII mediates many forms of learning and memory . Activity-regulated translocation of CaMKII to synapses is important for its functions in synaptic plasticity . Here , we tested the role of the NMDA receptor subunit GluN2B in recruiting CaMKII\u03b1 to synapses with different paradigms : global bath stimulation of NMDA receptors , a chemical long term potentiation ( cLTP ) protocol that selectively activates synaptic NMDA receptors , or local stimulation of NMDA receptors at a contiguous set of synapses that triggers a propagating synaptic accumulation of CaMKII . Global or cLTP-induced synaptic accumulation of CaMKII\u03b1 occurred in wild-type but not sister GluN2B -/- cultured mouse hippocampal neurons . Expression of YFP-GluN2B , but not a similar level of YFP-GluN2A , rescued global and cLTP-induced CaMKII\u03b1 translocation . Using chimeric constructs , the pore-forming extracellular and membrane domains of GluN2A combined with the cytoplasmic tail of GluN2B were sufficient to rescue CaMKII\u03b1 translocation , whereas the reverse chimera was ineffective . Furthermore , the dual point mutation R1300Q,S1303D in GluN2B that blocks interaction of this high affinity site with CaMKII abolished rescue . Thus , CaMKII binding to GluN2B is required for global and cLTP-induced synaptic accumulation of CaMKII\u03b1 . However , surprisingly , locally induced propagating synaptic accumulation of CaMKII\u03b1 occurred normally in GluN2B -/- neurons , indistinguishably from wild-type . Thus , synaptic trapping of CaMKII during locally induced propagating translocation occurs by different mechanisms and molecular partners compared with global stimulation and cLTP paradigms . These findings underscore the complex regulatory properties and molecular interactions of CaMKII\u03b1 , a key player in synaptic plasticity .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
22902837
|
Certain components of apples have been shown to prevent cancer growth and impede cancer progression . We hypothesized that extracted apple polysaccharides ( APs ) might , therefore , have anticancer effects , through a mechanism involving the induction of apoptosis in cancer cells , partly via the NF-κB pathway . Two human colorectal cancer ( CRC ) cell lines , HT-29 and SW620 , were exposed to different concentrations of APs ( 0.01 , 0.1 or 1 mg/ml ) . Cell apoptosis was measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay by flow cytometry and incorporation of 5'-bromodeoxyuridine ( BrdU ) into DNA to identify the proliferating cell fraction , using fluorescence microscopy in vitro . The protein levels of NF-κB/p65 , I-κBα , pI-κBα , Bax , Bcl-xl and Bcl-2 were evaluated by western blotting . The target sites of APs on CRC cells were assessed by flow cytometry . At concentrations of 0.1 and 1 mg/ml , APs showed apoptosis-inducing effects , increased expressions of Bax , nuclear p65 and cytoplasmic pI-κBα , and decreased expressions of Bcl-2 , Bcl-xl and cytoplasmic I-κBα . APs induced apoptosis by slightly activating the NF-κB pathway ; the AP target site could be the Toll-like receptor 4 on the cell membrane . These results demonstrate the potential of APs as agents for clinical prevention and treatment of CRC .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
0
] |
22470124
|
Autophagy is a catabolic process by which a cell degrades its intracellular materials to replenish itself . Induction of autophagy under various cellular stress stimuli can lead to either cell survival or cell death via apoptotic and/or autophagic ( nonapoptotic ) pathways . The NSAID sulindac sulfide induces apoptosis in colon cancer cells . Here , we show that inhibition of autophagy under serum-deprived conditions resulted in significant reductions of sulindac sulfide-induced apoptosis in HT-29 colon cancer cells . In contrast , inhibition of autophagy under conditions where serum is available significantly increased sulindac sulfide-induced apoptosis in HT-29 cells . We previously showed that the apoptosis inhibitor , survivin , plays a role in regulating NSAID-induced apoptosis and autophagic cell death . Here , we show that survivin protein half-life is increased in the presence of autophagy inhibitors under serum-deprived conditions , but not under conditions when serum is available . Thus , the increased levels of survivin may be a factor contributing to inhibition of sulindac sulfide-induced apoptosis under serum-deprived conditions . These results suggest that whether a cell lives or dies due to autophagy induction depends on the balance of factors that regulate both autophagic and apoptotic processes .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
0
] |
23431290
|
Histopathological features of the lymph node involvement were studied in 104 patients with thoracic esophageal cancer who underwent subtotal esophagectomy combined with extended radical lymph adenectomy in cervicothoracoabdominal region . Metastatic involvement was found in a total number of 503 lymph nodes from 73 patients by histologic examination . The mean of long and short diameter was found to be less than 5mm in 125 ( 24.9% ) of these 503 nodes . The involved area on the section was less than one third in 149 nodes ( 29.6% ) , and was significantly smaller in mediastinal lymph nodes than those in cervical or abdominal ones . Sixty-seven ( 13.3% ) of 503 nodes were partially invaded by micrometastasis of 1mm or less in diameter . Micrometastasis also more frequently occurred in mediastinal nodes with a statistically significant difference . Extranodal proliferation ( ENP ) of cancer cells was found in 106 nodes ( 21.1% ) , and extranodal lymphatic and/or blood vessel invasion ( ENly , v ) was also recognized in 60 nodes ( 11.9% ) . Micrometastasis and ENP with or without ENly , v were found in 24 ( 32.9% ) and 29 ( 39.7% ) of 73 patients with positive lymph node metastasis , respectively . Postoperative survival rate in patients with micrometastasis and/or ENP with or without ENly , v was inferior to that in patients with neither of them .
|
[
1,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
1435691
|
Lung cancer is primarily caused by exposure to tobacco smoke . Tobacco smoke contains numerous carcinogens , including polycyclic aromatic hydrocarbons ( PAH ) . The most common PAH studied is benzo[a]pyrene ( B[a]P ) . B[a]P is metabolically activated through multiple routes , one of which is catalyzed by aldo-keto reductase ( AKR ) to B[a]P-7,8-dione ( BPQ ) . BPQ undergoes a futile redox cycle in the presence of NADPH to generate reactive oxygen species ( ROS ) . ROS , in turn , damages DNA . Studies with a yeast p53 mutagenesis system found that the generation of ROS by PAH o-quinones may contribute to lung carcinogenesis because of similarities between the patterns ( types of mutations ) and spectra ( location of mutations ) and those seen in lung cancer . The patterns were dominated by G to T transversions , and the spectra in the experimental system have mutations at lung cancer hotspots . To address repair mechanisms that are responsible for BPQ induced damage we observed the effect of mutating two DNA repair genes OGG1 and APE1 ( APN1 in yeast ) and tested them in a yeast reporter system for p53 mutagenesis . There was an increase in both the mutant frequency and the number of G:C/T:A transversions in p53 treated with BPQ in ogg1 yeast but not in apn1 yeast . Knocking out APN2 increased mutagenesis in the apn1 cells . In addition , we did not find a strand bias on p53 treated with BPQ in ogg1 yeast . These studies suggest that Ogg1 is involved in repairing the oxidative damage caused by BPQ , Apn1 and Apn2 have redundant functions and that the stand bias seen in lung cancer may not be due to impaired repair of oxidative lesions .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
23117049
|
Ataxia telangiectasia mutated ( ATM ) kinase is a central player in cellular response to DNA damage . Phosphorylation of the histone H2AX by ATM is required for the accumulation of repair proteins at the sites of double-strand breaks . Recently , it was reported that the histone acetyltransferase Tat interactive protein-60 ( IPP60 ) is required to acetylate ATM prior to its activation . The RuvB-like proteins TIP48 and TIP49 are known to be necessary for the assembly and functional activity of the TIP60 acetyltransferase complex . In the present communication , we investigated the requirements of IIP48 and IIP49 for ATM activation by monitoring the cell cycle distribution and H2AX phosphorylation after irradiation of IIP48- and IIP49-depleted cells . We found that neither the cell cycle norgammay-H2AX were affected in IIP48- and IIP49-silenced cells , suggesting that the IIP60 chromatin modification complex is not engaged in DNA damage signaling upstream of ATM .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
20355335
|
Estrogens play essential roles in the progression of mammary and prostatic diseases . The transcriptional effects of estrogens are transduced by two estrogen receptors , ERα and ERβ , which elicit opposing roles in regulating proliferation : ERα is proliferative while ERβ is anti-proliferative . Exogenous expression of ERβ in ERα-positive cancer cell lines inhibits cell proliferation in response to estrogen and reduces xenografted tumor growth in vivo , suggesting that ERβ might oppose ERα's proliferative effects via formation of ERα/β heterodimers . Despite biochemical and cellular evidence of ERα/β heterodimer formation in cells co-expressing both receptors , the biological roles of the ERα/β heterodimer remain to be elucidated . Here we report the identification of two phytoestrogens that selectively activate ERα/β heterodimers at specific concentrations using a cell-based , two-step high throughput small molecule screen for ER transcriptional activity and ER dimer selectivity . Using ERα/β heterodimer-selective ligands at defined concentrations , we demonstrate that ERα/β heterodimers are growth inhibitory in breast and prostate cells which co-express the two ER isoforms . Furthermore , using Automated Quantitative Analysis ( AQUA ) to examine nuclear expression of ERα and ERβ in human breast tissue microarrays , we demonstrate that ERα and ERβ are co-expressed in the same cells in breast tumors . The co-expression of ERα and ERβ in the same cells supports the possibility of ERα/β heterodimer formation at physio- and pathological conditions , further suggesting that targeting ERα/β heterodimers might be a novel therapeutic approach to the treatment of cancers which co-express ERα and ERβ .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
22347418
|
The ARF locus is frequently inactivated in human cancer . The oncosuppressor ARF has indeed been described as a general sensor for different situation of cellular stress . We have previously demonstrated that ARF deficiency severely impairs inflammatory responses in vitro and in vivo , establishing a role for ARF in the regulation of innate immunity . The aim of the present work was to get further insights into the immune functions of ARF and to evaluate its possible contribution to the polarization of macrophages toward the M1 or M2 phenotype . Our results demonstrate that resting Arf(-/-) macrophages express high levels of Ym1 and Fizz-1 , two typical markers of alternatively-activated macrophages ( M2 ) . Additionally , Arf(-/-) peritoneal macrophages showed an impaired response to lipopolysaccharide ( a classical inducer of M1 polaryzation ) and a reduced production of pro-inflammatory cytokines/chemokines . Moreover , upon stimulation with interleukin-4 ( IL-4 ) , an inducer of the M2 phenotype , well established M2 markers such as Fizz-1 , Ym1 and arginase-1 were upregulated in Arf(-/-) as compared with wild type macrophages . Accordingly , the cytokine and chemokine profile associated with the M2 phenotype was significantly overexpressed in Arf(-/-) macrophages responding to IL-4 . In addition , multiple pro-angiogenic factors such as VEGF and MMP-9 were also increased . In summary , these results indicate that ARF contributes to the polarization and functional plasticity of macrophages .
|
[
0,
0,
0,
0,
0,
0,
1,
0,
0,
1
] |
23243586
|
Normal human lung fibroblast diploid cells , WI-38 , become senescent after a definite number of divisions . VA-13 is a line of immortalized cells established by transformation of WI-38 cells by SV40 virus . To determine whether SV40 large T ( SV40-T ) antigen is essential for this immortalization of WI-38 cells we introduced an antisense gene for T antigen into VA-13 . Two morphologically different types of antisense transformant ( VA-AS5-8 and VA-AS37-8 ) were obtained . In both antisense transformants the expression of T antigen was reduced by more than 70% as compared to that in the parent cells . The morphology of the antisense transformants indicated a partial conversion to the senescent phenotype of WI-38 . The relative number of cells in the S phase of the antisense transformants was decreased as compared to that in cultures of VA-13 and about 50% of cells were at G1/0 . The doubling time of the transformants was prolonged to close to the doubling time of WI-38 . The level of expression of retinoblastoma protein ( pRB ) complexed with SV40-T antigen of the antisense transformants was significantly decreased although the level of total pRB was much higher than that in VA-13 . The pRB was present exclusively in the underphosphorylated form . Thus , the decreased level of formation of the complex between SV40-T and pRB or the underphosphorylation of pRB may explain the suppression of growth of antisense transformants . Together , these results show that an antisense gene for SV40-T antigen can efficiently block the cell proliferation and the cell immortalization of VA-13 cells .
|
[
0,
0,
0,
1,
0,
0,
0,
0,
1,
0
] |
1338304
|
BACKGROUND Though an increased efficacy of carmustine and temozolomide ( TMZ ) has been demonstrated by inactivation of O(6)-methylguanine-DNA methyltransferase ( MGMT ) with O(6)-benzyl-guanine ( BG ) in human pancreatic tumors refractive to alkylating agents , the regulatory mechanisms have not been explored . METHODS The effects of TMZ and BG on apoptosis , cell growth , the mitotic index , cell cycle distribution , and protein expression were studied by TUNEL , cell counting , flow cytometry , and Western blot analysis , respectively . RESULTS The wt-p53 human pancreatic tumor cell line Capan-2 and p53-efficient mouse embryonic fibroblasts ( MEFs ) were more responsive to treatment with TMZ + BG than mutant p53 Capan-1 and p53-null MEFs . S phase delay with a subsequent G2/M arrest was observed in Capans in response to BG + TMZ . The G1-to-S transition delay in Capan-2 was associated with p53-dependent apoptosis and was distinctly different from the presumed mismatch repair ( MMR ) killing operative during the G2/M arrest . The effect of p53 on BG + TMZ toxicity was supported by a marked change in apoptosis when p53 function was restored/inactivated . There was an early induction of MMR proteins in p53-efficient lines . CONCLUSION p53 provokes a classic proapoptotic response by delaying G1-to-S progression , but it may also facilitate cell killing by enhancing MMR-related cell cycle arrest and cell death .
|
[
0,
0,
0,
0,
1,
1,
0,
1,
1,
0
] |
20980775
|
OBJECTIVE In addition to its effects on cholesterol levels , apoE3 has lipid-independent effects that contribute to cardiovascular protection ; one of these effects is the ability to inhibit cell cycling in VSMCs . The goal of this study was to identify and characterize cell cycle-regulatory mechanisms responsible for the anti-mitogenic effect of apoE . METHODS AND RESULTS Primary VSMCs were stimulated with serum in the absence or presence of apoE3. apoE3 upregulated expression of the cdk inhibitor , p27(kip1) , in primary VSMCs , and this effect required Cox2 and activation of PGI(2)-IP signaling . The microRNA family , miR221/222 has recently been identified as a post-translational regulator of p27 , and apoE3 inhibited miR221/222 expression in a Cox2- and PGI(2)/IP-dependent manner . Moreover , reconstituted miR222 expression was sufficient to override the effects of apoE on p27 expression and S phase entry . The ability to repress expression of miR221/222 is shared by apoE3-containing HDL but is absent from apoA-1 , LDL and apoE-depleted HDL . All three apoE isoforms regulate miR221/222 , and the effect is independent of the C-terminal lipid-binding domain. miR221/222 levels are increased in the aortae of apoE3-null mice and reduced when apoE3 expression is reconstituted by adeno-associated virus infection . Thus , regulation of miR221/222 by apoE3 occurs invivo as well as invitro . CONCLUSIONS ApoE inhibits VSMC proliferation by regulating p27 through miR221/222 . Control of cell cycle-regulatory microRNAs adds a new dimension to the spectrum of cardiovascular protective effects afforded by apoE and apoE-HDL .
|
[
0,
0,
0,
0,
1,
0,
0,
0,
0,
0
] |
23294923
|
Chromatographic , peptide mapping and mass spectrometric analysis were used to examine hemoglobin ( Hb ) from heavy drinkers and abstainers for alcohol consumption-related modifications . Heavy drinker and abstainer hemoglobin samples contained similar amounts of glycosylated Hb and significantly different ( p < 0.05 ) amounts of " fast " hemoglobin . The presence of higher amounts of " fast " Hb in heavy drinker relative to abstainer samples suggested the presence of alcohol-consumption related modifications . To further examine Hb for modifications , tryptic peptides of the " fast " hemoglobin HbA1c were isolated and analyzed by plasma desorption mass spectrometry ( PDMS). [ 14C]acetaldehyde ( AcH)-Hb was synthesized in vivo for use as a standard . Specific peptides were chosen based on co-migration with radiolabeled peptides from a tryptic digest of the [ 14C]acetaldehyde-Hb . The masses obtained by PDMS for two heavy drinker peptides were identical to two radiolabeled peptides ; the two pairs of peptides co-migrated on HPLC . A comparison of the observed mass for the peptides with the theoretical masses for acetaldehyde-modified Hb peptides suggested that the peptides were AcH-modified alpha and beta chain N-termini of Hb . The modified peptides were found in five of six heavy drinker samples . This is the first description of site-specific AcH-Hb adducts occurring in vivo . The routine detection of such adducts has potential for characterizing usual alcohol intake .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
1471764
|
DNA damage signaling and repair take place in a chromatin context . Consequently , chromatin-modifying enzymes , including adenosine triphosphate-dependent chromatin remodeling enzymes , play an important role in the management of DNA double-strand breaks ( DSBs ) . Here , we show that the p400 ATPase is required for DNA repair by homologous recombination ( HR ) . Indeed , although p400 is not required for DNA damage signaling , DNA DSB repair is defective in the absence of p400 . We demonstrate that p400 is important for HR-dependent processes , such as recruitment of Rad51 to DSB ( a key component of HR ) , homology-directed repair , and survival after DNA damage . Strikingly , p400 and Rad51 are present in the same complex and both favor chromatin remodeling around DSBs . Altogether , our data provide a direct molecular link between Rad51 and a chromatin remodeling enzyme involved in chromatin decompaction around DNA DSBs .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
23266955
|
Altered p53 protein is prevalently associated with the pathologic class of triple-negative breast cancers and loss of p53 function has recently been linked to the induction of an epithelial-mesenchymal transition ( EMT ) and acquisition of stemness properties . We explored the association between TP53 mutational status and expression of some genes involved in the canonical TGF-\u03b2 signaling pathway ( the most potent EMT inducer ) and in two early EMT associated events : loss of cell polarity and acquisition of stemness-associated features . We used a publicly accessible microarray dataset consisting of 251 p53-sequenced primary breast cancers . Statistical analysis indicated that mutant p53 tumors ( especially those harboring a severe mutation ) were consistent with the aggressive class of triple-negative cancers and that , differently from cell cultures , surgical tumors underexpressed some TGF-\u03b2 related transcription factors known as involved in EMT ( ID1 , ID4 , SMAD3 , SMAD4 , SMAD5 , ZEB1 ) . These unexpected findings suggest an interesting relationship between p53 mutation , mammary cell dedifferentiation , and the concomitant acquisition of stemlike properties ( as indicated by the overexpression of PROM1 and NOTCH1 genes ) , which improve tumor cells aggressiveness as indicated by the overexpression of genes associated with cell proliferation ( CDK4 , CDK6 , MKI67 ) and migration ( CXCR4 , MMP1 ) .
|
[
1,
0,
0,
0,
0,
1,
0,
0,
1,
0
] |
22899882
|
BACKGROUND Taxol is one of the most effective chemotherapeutic agents for the treatment of patients with breast cancer . Despite impressive clinical responses initially , the majority of patients eventually develop resistance to Taxol . Lactate dehydrogenase-A ( LDH-A ) is one of the predominant isoforms of LDH expressed in breast tissue , which controls the conversion of pyruvate to lactate and plays an important role in glucose metabolism . In this study we investigated the role of LDH-A in mediating Taxol resistance in human breast cancer cells . RESULTS Taxol-resistant subclones , derived from the cancer cell line MDA-MB-435 , sustained continuous growth in high concentrations of Taxol while the Taxol-sensitive cells could not . The increased expression and activity of LDH-A were detected in Taxol-resistant cells when compared with their parental cells . The downregulation of LDH-A by siRNA significantly increased the sensitivity of Taxol-resistant cells to Taxol . A higher sensitivity to the specific LDH inhibitor , oxamate , was found in the Taxol-resistant cells . Furthermore , treating cells with the combination of Taxol and oxamate showed a synergistical inhibitory effect on Taxol-resistant breast cancer cells by promoting apoptosis in these cells . CONCLUSION LDH-A plays an important role in Taxol resistance and inhibition of LDH-A re-sensitizes Taxol-resistant cells to Taxol . This supports that Warburg effect is a property of Taxol resistant cancer cells and may play an important role in the development of Taxol resistance . To our knowledge , this is the first report showing that the increased expression of LDH-A plays an important role in Taxol resistance of human breast cancer cells . This study provides valuable information for the future development and use of targeted therapies , such as oxamate , for the treatment of patients with Taxol-resistant breast cancer .
|
[
0,
0,
1,
0,
0,
0,
0,
0,
0,
0
] |
20144215
|
Here , we investigated the compartment-specific role of cell cycle arrest and senescence in breast cancer tumor growth . For this purpose , we generated a number of hTERT-immortalized senescent fibroblast cell lines overexpressing CDK inhibitors , such as p16(INK4A) , p19(ARF) or p21(WAF1/CIP1) . Interestingly , all these senescent fibroblast cell lines showed evidence of increased susceptibility toward the induction of autophagy ( either at baseline or after starvation ) , as well as significant mitochondrial dysfunction . Most importantly , these senescent fibroblasts also dramatically promoted tumor growth ( up to without any comparable increases in tumor angiogenesis . Conversely , we generated human breast cancer cells ( MDA-MB-231 cells ) overexpressing CDK inhibitors , namely p16(INK4A) or p21(WAF1/CIP1) . Senescent MDA-MB-231 cells also showed increased expression of markers of cell cycle arrest and autophagy , including β-galactosidase , as predicted . Senescent MDA-MB-231 cells had retarded tumor growth , with up to a near 2-fold reduction in tumor volume . Thus , the effects of CDK inhibitors are compartment-specific and are related to their metabolic effects , which results in the induction of autophagy and mitochondrial dysfunction . Finally , induction of cell cycle arrest with specific inhibitors ( PD0332991 ) or cellular stressors [ hydrogen peroxide ( H(2)O(2) ) or starvation ] indicated that the onset of autophagy and senescence are inextricably linked biological processes . The compartment-specific induction of senescence ( and hence autophagy ) may be a new therapeutic target that could be exploited for the successful treatment of human breast cancer patients .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
22935696
|
PURPOSE Genomic complexity is present in approximately 15% to 30% of all chronic lymphocytic leukemia ( CLL ) and has emerged as a strong independent predictor of rapid disease progression and short remission duration in CLL . We conducted this study to advance our understanding of the causes of genomic complexity in CLL . EXPERIMENTAL DESIGN We have obtained quantitative measurements of radiation-induced apoptosis and radiation-induced ATM autophosphorylation in purified CLL cells from 158 and 140 patients , respectively , and have used multivariate analysis to identify independent contributions of various biological variables on genomic complexity in CLL . RESULTS Here , we identify a strong independent effect of radiation resistance on elevated genomic complexity in CLL and describe radiation resistance as a predictor for shortened CLL survival . Furthermore , using multivariate analysis , we identify del17p/p53 aberrations , del11q , del13q14 type II ( invariably resulting in Rb loss ) , and CD38 expression as independent predictors of genomic complexity in CLL , with aberrant p53 as a predictor of approximately 50% of genomic complexity in CLL . Focusing on del11q , we determined that normalized ATM activity was a modest predictor of genomic complexity but was not independent of del11q . Through single nucleotide polymorphism array-based fine mapping of del11q , we identified frequent monoallelic loss of Mre11 and H2AFX in addition to ATM , indicative of compound del11q-resident gene defects in the DNA double-strand break response . CONCLUSIONS Our quantitative analysis links multiple molecular defects , including for the first time del11q and large 13q14 deletions ( type II ) , to elevated genomic complexity in CLL , thereby suggesting mechanisms for the observed clinical aggressiveness of CLL in patients with unstable genomes .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
20086003
|
Here , we identified the milk protein α-casein as a novel suppressor of tumor growth and metastasis . Briefly , Met-1 mammary tumor cells expressing α-casein showed a reduction in tumor growth and a near 10-fold decrease in experimental metastasis . To identify the molecular mechanism(s) , we performed genome-wide transcriptional profiling . Interestingly , our results show that α-casein upregulates gene transcripts associated with interferon/STAT1 signaling and downregulates genes associated with " stemness. " These findings were validated by immunoblot and FACS analysis , which showed the upregulation and hyperactivation of STAT1 and a decrease in the number of CD44(+) " cancer stem cells. " These gene signatures were also able to predict clinical outcome in human breast cancer patients . Thus , we conclude that a lactation-based therapeutic strategy using recombinant α-casein would provide a more natural and non-toxic approach to the development of novel anticancer therapies .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
23047602
|
BACKGROUND Cellular stress responses trigger signaling cascades that inhibit proliferation and protein translation to help alleviate the stress or if the stress cannot be overcome induce apoptosis . In recent studies , we demonstrated the ability of lovastatin , an inhibitor of mevalonate synthesis , to induce the Integrated Stress Response as well as inhibiting epidermal growth factor receptor ( EGFR ) activation . METHODOLOGY/PRINCIPAL FINDINGS In this study , we evaluated the effects of lovastatin on the activity of the LKB1/AMPK pathway that is activated upon cellular energy shortage and can interact with the above pathways . In the squamous cell carcinoma ( SCC ) cell lines SCC9 and SCC25 , lovastatin treatment ( 1-25 �M , 24 hrs ) induced LKB1 and AMPK activation similar to metformin ( 1-10 mM , 24 hrs ) , a known inducer of this pathway . Lovastatin treatment impaired mitochondrial function and also decreased cellular ADP/ATP ratios , common triggers of LKB1/AMPK activation . The cytotoxic effects of lovastatin were attenuated in LKB1 null MEFs indicating a role for this pathway in regulating lovastatin-induced cytotoxicity . Of clinical relevance , lovastatin induces synergistic cytotoxicity in combination with the EGFR inhibitor gefitinib . In LKB1 deficient ( A549 , HeLa ) and expressing ( SCC9 , SCC25 ) cell lines , metformin enhanced gefitinib cytotoxicity only in LKB1 expressing cell lines while both groups showed synergistic cytotoxic effects with lovastatin treatments . Furthermore , the combination of lovastatin with gefitinib induced a potent apoptotic response without significant induction of autophagy that is often induced during metabolic stress inhibiting cell death . CONCLUSION/SIGNIFICANCE Thus , targeting multiple metabolic stress pathways including the LKB1/AMPK pathway enhances lovastatin's ability to synergize with gefitinib in SCC cells .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
1,
0
] |
23029387
|
Accumulating evidence from epidemiological studies indicates that chronic inflammation and oxidative stress play critical roles in neoplastic development . The aim of this study was to investigate the anti-inflammatory , anti-oxidative stress activities , and differential regulation of Nrf2-mediated genes by tea Chrysanthemum zawadskii ( CZ ) and licorice Glycyrrhiza uralensis ( LE ) extracts . The anti-inflammatory and anti-oxidative stress activities of hexane/ethanol extracts of CZ and LE were investigated using in vitro and in vivo approaches , including quantitative real-time PCR ( qPCR ) and microarray . Additionally , the role of the transcriptional factor Nrf2 ( nuclear erythroid-related factor 2 ) signaling pathways was examined . Our results show that CZ and LE extracts exhibited potent anti-inflammatory activities by suppressing the mRNA and protein expression levels of pro-inflammatory biomarkers IL-1β , IL-6 , COX-2 and iNOS in LPS-stimulated murine RAW 264.7 macrophage cells . CZ and LE also significantly suppressed the NO production of LPS-stimulated RAW 264.7 cells . Additionally , CZ and LE suppressed the NF-κB luciferase activity in human HT-29 colon cancer cells . Both extracts also showed strong Nrf2-mediated antioxidant/Phase II detoxifying enzymes induction . CZ and LE induced NQO1 , Nrf2 , and UGT and antioxidant response element ( ARE)-luciferase activity in human hepatoma HepG2 C8 cells . Using Nrf2 knockout [ Nrf2 ( -/-) ] and Nrf2 wild-type ( +/+ ) mice , LE and CZ showed Nrf2-dependent transactivation of Nrf2-mediated antioxidant and phase II detoxifying genes . In summary , CZ and LE possess strong inhibitory effects against NF-κB-mediated inflammatory as well as strong activation of the Nrf2-ARE-anti-oxidative stress signaling pathways , which would contribute to their overall health promoting pharmacological effects against diseases including cancer .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
1
] |
20967519
|
BACKGROUND Cellular and clinical sensitivity to ionizing radiation ( IR ) is determined by DNA double-strand breaks ( DSB ) repair . Here , we investigate the molecular mechanism underlying the extreme response of a head and neck tumor case ( SKX ) to standard radiotherapy . METHODS Immunofluorescence ( IF ) was used for the assessment of DSB repair , Western blot and real-time PCR for protein and mRNA expression , respectively . RESULTS SKX cells exhibited a pronounced radiosensitivity associated with numerous residual \u03b3-H2AX foci after IR . This was not associated with lacking canonical repair proteins . SKX cells did not express any ATM protein . Accordingly , immunoblotting revealed no ATM kinase activity toward substrates such as p-SMC1 , p-CHK2 and p-KAP1 . Sequencing of all 66 exons of ATM showed no mutation . ATM mRNA level was moderately reduced , which could be reverted by 5'-Aza-C treatment but without restoring protein levels . Importantly , we demonstrated a post-transcriptional regulation in SKX cells via 6-fold enhanced levels of miR-421 , which targets the 3'-UTR of ATM mRNA . Transfection of SKX cells with either anti-miR-421 inhibitor or a microRNA-insensitive ATM vector recovered ATM expression and abrogated the hyper-radiosensitivity . CONCLUSION This is the first report describing microRNA-mediated down-regulation of ATM leading to clinically manifest tumor radiosensitivity .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
23199656
|
The number of renal cancers has increased over the last ten years and patient survival in advanced stages remains very poor . Therefore , new therapeutic approaches for renal cancer are essential . Englerin A is a natural product with a very potent and selective cytotoxicity against renal cancer cells . This makes it a promising drug candidate that may improve current treatment standards for patients with renal cancers in all stages . However , little is known about englerin A's mode of action in targeting specifically renal cancer cells . Our study is the first to investigate the biological mechanism of englerin A action in detail . We report that englerin A is specific for renal tumor cells and does not affect normal kidney cells . We find that englerin A treatment induces necrotic cell death in renal cancer cells but not in normal kidney cells . We further show that autophagic and pyroptotic proteins are unaffected by the compound and that necrotic signaling in these cells coincided with production of reactive oxygen species and calcium influx into the cytoplasm . As the first study to analyze the biological effects of englerin A , our work provides an important basis for the evaluation and validation of the compound's use as an anti-tumor drug . It also provides a context in which to identify the specific target or targets of englerin A in renal cancer cells .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
1
] |
23144724
|
Hepatocellular carcinoma ( HCC ) is one of the most common malignancies worldwide ; however , the prognosis of HCC patients remains poor . This poor prognosis is mainly attributed to the high rate of intrahepatic and distant metastasis . HCC often occurs in a hypoxic environment and hypoxia can activate metastatic programs , ultimately leading to tumor recurrence or metastasis . Thus , the discovery and subsequent development of novel agents to block HCC invasion and migration are the primary objectives of hepatic cancer research . The Notch1 signaling pathway might be involved in hypoxia-induced carcinoma metastasis . However , the mechanisms by which Notch1 mediates cell metastasis , particularly in hepatocellular carcinoma , are not yet entirely clear . The results of the present study show that hypoxia increases the invasion and migration capacities of different HCC cells . Activation of the Notch1 signaling pathway contributes to hypoxia-induced invasion and migration in HCC cells . The activated Notch1 signaling pathway can regulate Snail/E-cadherin through cyclooxygenase-2 ( COX-2 ) under hypoxic conditions . The above results suggest that the Notch1/COX-2/Snail/E-cadherin pathway is possibly associated with hypoxia-induced invasion and migration in HCC cells . Thus , targeting Notch1 may be useful for devising novel preventive and therapeutic strategies for HCC .
|
[
1,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
23124652
|
We investigated the direct effects of LH-releasing hormone ( LH-RH ) antagonist , Cetrorelix , on the growth of HTOA human epithelial ovarian cancer cell line . RT-PCR revealed the expression of mRNA for LH-RH and its receptor in HTOA cells . Cetrorelix , at concentrations between 10(-9) and 10(-5) M , exerted a dose-dependent antiproliferative action on HTOA cells , as measured by 5-bromo-2'-deoxyuridine incorporation into DNA . Flow cytometric analysis indicated that Cetrorelix , at 10(-5) M , arrested cell cycle in HTOA cells , at G1 phase , after 24 h of treatment . Western blot analysis of cell cycle-regulatory proteins demonstrated that treatment with Cetrorelix ( 10(-5) M ) for 24 h did not change the steady-state levels of cyclin D1 , cyclin E , and cyclin-dependent kinase ( Cdk)4 but decreased the levels of cyclin A and Cdk2 . The protein levels of p21 ( a Cdk inhibitor ) and p53 ( a suppressor of tumor cell growth and a positive regulator for p21 expression ) were increased by Cetrorelix , but the levels of p27 ( a Cdk inhibitor ) did not change significantly . Flow cytometric analysis and terminal deoxynucleotidyltransferase-mediated deoxyuridine 5-triphosphate nick end labeling staining demonstrated that Cetrorelix ( 10(-5) M ) induced apoptosis in HTOA cells . In conclusion , Cetrorelix directly inhibits the proliferation of human epithelial ovarian cancer cells through mechanisms mediated by LH-RH receptor and involving multiple events in cell cycle progression , including G1 phase cell cycle arrest coupled with down-regulation of cyclin A-Cdk2 complex levels , presumably attributable to an up-regulation of p53 and p21 protein levels and apoptosis .
|
[
0,
0,
0,
0,
1,
0,
0,
1,
1,
0
] |
12161501
|
Nuclear factor kappa-B ( NF-κB ) activates multiple genes with overlapping roles in cell proliferation , inflammation and cancer . Using an unbiased approach we identified human CDK6 as a novel kinase phosphorylating NF-κB p65 at serine 536 . Purified and reconstituted CDK6/cyclin complexes phosphorylated p65 in vitro and in transfected cells . The physiological role of CDK6 for basal as well as cytokine-induced p65 phosphorylation or NF-κB activation was revealed upon RNAi-mediated suppression of CDK6 . Inhibition of CDK6 catalytic activity by PD332991 suppressed activation of NF-κB and TNF-induced gene expression . In complex with a constitutively active viral cyclin CDK6 stimulated NF-κB p65-mediated transcription in a target gene specific manner and this effect was partially dependent on its ability to phosphorylate p65 at serine 536 . Tumor formation in thymi and spleens of v-cyclin transgenic mice correlated with increased levels of p65 Ser536 phosphorylation , increased expression of CDK6 and upregulaton of the NF-κB target cyclin D3 . These results suggest that aberrant CDK6 expression or activation that is frequently observed in human tumors can contribute through NF-κB to chronic inflammation and neoplasia .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
0,
1
] |
23300567
|
Topoisomerase 1 ( Top1)-DNA cleavage complexes induced by camptothecin ( CPT ) cause DNA strand breaks during DNA replication or transcription . Although the cellular responses to replication-mediated DNA double-strand breaks have been well studied , the responses to transcription-mediated DNA strand breaks have not . Here , we show that poly ( ADP-ribose ) polymerase ( PARP ) and cockayne syndrome group B protein ( CSB ) modulate the CPT-induced formation of discrete p53-binding protein 1 ( 53BP1 ) nuclear foci at sites of transcription-mediated DNA strand breaks . Inhibition of PARP activity enhanced the formation of these foci , while knockdown of essential components of the base excision repair ( BER ) pathway did not . These findings suggest that PARP suppresses transcription-mediated 53BP1 foci formation , but that this does not occur through the BER pathway . In addition , knockdown of CSB , one of the key factors of transcription-coupled repair , slowed the kinetics of 53BP1 foci formation . These data suggest that PARP and CSB modulate the formation of 53BP1 foci during the processing of transcription-mediated DNA strand breaks .
|
[
0,
0,
0,
0,
0,
1,
0,
0,
0,
0
] |
23229313
|
The Warburg effect describes a heightened propensity of tumor cells to produce lactic acid in the presence or absence of O(2) . A generally held notion is that the Warburg effect is related to energy . Using whole-genome , proteomic MALDI-TOF-MS and metabolite analysis , we investigated the Warburg effect in malignant neuroblastoma N2a cells . The findings show that the Warburg effect serves a functional role in regulating acidic pericellular pH ( pHe ) , which is mediated by metabolic inversion or a fluctuating dominance between glycolytic-rate substrate level phosphorylation ( SLP ) and mitochondrial ( mt ) oxidative phosphorylation ( OXPHOS ) to control lactic acid production . The results also show that an alkaline pHe caused an elevation in SLP/OXPHOS ratio ( approximately 98% SLP/OXPHOS ) ; while the ratio was approximately 56% at neutral pHe and approximately 93% in acidic pHe . Acidic pHe paralleled greater expression of mitochondrial biogenesis and OXPHOS genes , such as complex III-V ( Uqcr10 , Atp5 and Cox7c ) , mt Fmc1 , Romo1 , Tmem 173 , Tomm6 , aldehyde dehydrogenase , mt Sod2 mt biogenesis component PPAR-\u03b3 co-activator 1 adjunct to loss of mt fission ( Mff ) . Moreover , acidic pHe corresponded to metabolic efficiency evidenced by a rise in mTOR nutrient sensor G\u03b2L , its downstream target ( Eif4ebp1 ) , insulin modulators ( Trib3 and Fetub ) and loss of catabolic ( Hadhb , Bdh1 and Pygl)/glycolytic processes ( aldolase C , pyruvate kinase , Nampt and aldose-reductase ) . In contrast , alkaline pHe initiated loss of mitofusin 2 , complex II-IV ( Sdhaf1 , Uqcrq , Cox4i2 and Aldh1l2 ) , aconitase , mitochondrial carrier triple repeat 1 and mt biosynthetic ( Coq2 , Coq5 and Coq9 ) . In conclusion , the Warburg effect might serve as a negative feedback loop that regulates the pHe toward a broad acidic range by altering lactic acid production through inversion of metabolic systems . These effects were independent of changes in O(2) concentration or glucose supply .
|
[
0,
0,
1,
0,
0,
0,
0,
0,
0,
0
] |
22320183
|
We compared gadolinium diethylenetriaminepentaacetic acid ( Gd-DTPA ) enhanced T1-weighted images ( T1-Gd ) with the histopathological findings in 13 patients with bone or soft tissue sarcomas . Signal intensity of the viable tumor tissue was increased in T1-Gd in 92% of the patients . The necrotic or cystic areas in the tumor were not enhanced , rendering them distinctly . The degree of enhancement of the edematous area around the tumor was similar to or more marked than that of the tumor in 54% of the patients . Area showing inflammatory cells infiltration and edematous areas in the tumor tissue were also enhanced . Thus , the effect of preoperative chemotherapy in tumor tissues other than necrotic and cystic areas tended to be underestimated in T1-Gd . Its effect should be comprehensively evaluated based on not only T1-Gd but also T2-weighted images and findings of other imaging techniques .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
1
] |
1485542
|
Vascular integrity is fundamental to the formation of mature blood vessels and depends on a functional , quiescent endothelial monolayer . However , how endothelial cells enter and maintain quiescence in the presence of angiogenic factors is still poorly understood . Here we identify the fibroblast growth factor ( FGF ) antagonist Sprouty2 ( Spry2 ) as a key player in mediating endothelial quiescence and barrier integrity in mouse aortic endothelial cells ( MAECs ) : Spry2 knockout MAECs show spindle-like shapes and are incapable of forming a functional , impermeable endothelial monolayer in the presence of FGF2 . Whereas dense wild type cells exhibit contact inhibition and stop to proliferate , Spry2 knockout MAECs remain responsive to FGF2 and continue to proliferate even at high cell densities . Importantly , the anti-proliferative effect of Spry2 is absent in sparsely plated cells . This cell density-dependent Spry2 function correlates with highly increased Spry2 expression in confluent wild type MAECs . Spry2 protein expression is barely detectable in single cells but steadily increases in cells growing to high cell densities , with hypoxia being one contributing factor . At confluence , Spry2 expression correlates with intact cell-cell contacts , whereas disruption of cell-cell contacts by EGTA , TNFα and thrombin decreases Spry2 protein expression . In confluent cells , high Spry2 levels correlate with decreased extracellular signal-regulated kinase 1/2 ( Erk1/2 ) phosphorylation . In contrast , dense Spry2 knockout MAECs exhibit enhanced signaling by Erk1/2 . Moreover , inhibiting Erk1/2 activity in Spry2 knockout cells restores wild type cobblestone monolayer morphology . This study thus reveals a novel Spry2 function , which mediates endothelial contact inhibition and barrier integrity .
|
[
0,
0,
0,
0,
1,
0,
0,
0,
0,
0
] |
23232625
|
A number of growth factors have been implicated in the control of the proliferation of breast cancer cells and some have been reported to mediate the proliferative effects of oestradiol . MCF-7 cells were treated with growth factors in the presence and absence of oestradiol . Oestradiol increased the response of cells to the proliferative effects of epidermal growth factor ( EGF ) , transforming growth factor alpha ( TGF-alpha ) and basic fibroblast growth factor ( bFGF ) . Platelet derived growth factor ( PDGF ) and cathepsin D had no effect in the presence or absence of oestradiol while TGF-beta slightly reduced the stimulation by oestradiol . In the absence of oestradiol , there was little effect of combinations of growth factors although the effects of bFGF and IGF-I were additive . In the presence of oestradiol , the effects of bFGF and TGF-alpha were additive whereas bFGF acted as an IGF-I antagonist . Overall , bFGF had the greatest effect on cell proliferation although this was less marked than the previously described effect of the IGFs and insulin . The effects of oestradiol on the sensitivity of cells to the proliferative effects of bFGF did not appear to result from regulation of bFGF receptor expression .
|
[
0,
0,
0,
0,
0,
0,
0,
0,
1,
0
] |
1419600
|
Cav-1 ( -/- ) deficient stromal cells are a new genetic model for myofibroblasts and cancer-associated fibroblasts . Using an unbiased informatics analysis of the transcriptional profile of Cav-1 ( -/- ) deficient mesenchymal stromal cells , we have now identified many of the major signaling pathways that are activated by a loss of Cav-1 , under conditions of metabolic restriction ( with low glucose media ) . Our informatics analysis suggests that a loss of Cav-1 induces oxidative stress , which mimics a constitutive pseudo-hypoxic state , leading to ( 1 ) aerobic glycolysis and ( 2 ) inflammation in the tumor stromal microenvironment . This occurs via the activation of two major transcription factors , namely HIF ( aerobic glycolysis ) and NFκB ( inflammation ) in Cav-1 ( -/- ) stromal fibroblastic cells . Experimentally , we show that Cav-1 deficient stromal cells may possess defective mitochondria , due to the over-production of nitric oxide ( NO ) , resulting in the tyrosine nitration of the mitochondrial respiratory chain components ( such as complex I ) . Elevated levels of nitro-tyrosine were observed both in Cav-1 ( -/- ) stromal cells , and via acute knock-down with siRNA targeting Cav-1 . Finally , metabolic restriction with mitochondrial ( complex I ) and glycolysis inhibitors was synthetically lethal with a Cav-1 ( -/- ) deficiency in mice . As such , Cav-1 deficient mice show a dramatically reduced mitochondrial reserve capacity . Thus , a mitochondrial defect in Cav-1 deficient stromal cells could drive oxidative stress , leading to aerobic glycolysis , and inflammation , in the tumor microenvironment . These stromal alterations may underlie the molecular basis of the " reverse Warburg effect " , and could provide the key to targeted anti-cancer therapies using metabolic inhibitors . In direct support of these findings , the transcriptional profile of Cav-1 ( -/- ) stromal cells overlaps significantly with Alzheimer disease , which is characterized by oxidative stress , NO over-production ( peroxynitrite formation ) , inflammation , hypoxia and mitochondrial dysfunction . We conclude that Cav-1 ( -/- ) deficient mice are a new whole-body animal model for an activated lethal tumor microenvironment , i.e. , " tumor stroma " without the tumor . Since Cav-1 ( -/- ) mice are also an established animal model for profibrotic disease , our current results may have implications for understanding the pathogenesis of scleroderma ( systemic sclerosis ) and pulmonary fibrosis , which are also related to abnormal mesenchymal stem cell function .
|
[
0,
0,
1,
0,
0,
0,
0,
0,
0,
1
] |
20519932
|
Transforming growth factor-beta1 ( TGF-beta1 ) exerts potent immunosuppressive effects . In this study , we investigated the potential role of TGF-beta1 produced by hepatocellular carcinoma ( HCC ) cell lines in immunosuppression mechanisms . Using the Mv1Lu cell-growth inhibition assay and an enzyme-linked immunosorbent assay ( ELISA ) , we detected optimal levels of TGF-beta1 in the culture supernatants conditioned by the HCC cell lines PLC/PRF/5 , Hep3B , and HepG2 . To determine the biological activity of TGF-beta1 in the supernatants , we examined the effects of the culture supernatants on the production of interferon ( IFN)-gamma induced during the culture of peripheral blood mononuclear cells ( PBMCs ) stimulated with interleukin ( IL)-12 . IFN-gamma production of IL-12-stimulated PBMCs in the 1:1 dilution of the acid-activated conditioned medium of PLC/PRF/5 , Hep3B , and HepG2 reduced to 14.7 +/- 0.8 , 17.3 +/- 9.0 , and 35.9 +/- 14.6% , respectively , compared with the value in the culture with control medium ( complete culture medium ) . These results suggest that HCC cells producing TGF-beta1 may reduce the generation or activation of cytotoxic T lymphocytes ( CTL ) and natural killer ( NK ) cells , and thus could enhance their ability to escape immune-mediated surveillance .
|
[
0,
1,
0,
0,
0,
0,
0,
0,
0,
0
] |
12685860
|
IL-16 is a ligand and chemotactic factor for CD4+ T cells . IL-16 inhibits the CD3 mediated lymphocyte activation and proliferation . The effects of IL-16 on the target cells are dependent on the cell type , the presence of co-activators etc . To understand the regulation function and mechanism of IL-16 on target cells , we used a 130 a.a. recombinant IL-16 to study its effects on the growth of Jurkat T leukemia cells in vitro . We found that the rIL-16 stimulated the proliferation of Jurkat cells at low dose ( 10(-9)M ) , but inhibited the growth of the cells at higher concentration ( 10(-5)M ) . Results showed that 10(-5) M of rIL-16 treatment induced an enhanced apoptosis in Jurkat cells . The treatment blocked the expression of FasL , but up-regulated the c-myc and Bid expression in the cells . Pre-treatment of PKC inhibitor or MEK1 inhibitor markedly increased or decreased the rIL-16 induced growth-inhibiting effects on Jurkat cells , respectively . The results suggested that the rIL-16 might be a regulator for the growth or apoptosis of Jurkat cells at a dose-dependent manner . The growth-inhibiting effects of rIL-16 might be Fas/FasL independent , but , associated with the activation of PKC , up-regulated expression of c-Myc and Bid , and the participation of the ERK signal pathway in Jurkat cells .
|
[
0,
0,
0,
0,
0,
0,
0,
1,
0,
0
] |
12528894
|
We report a case of recurrent gastric cancer successfully treated by a combination of CPT-11 and CDDP as the third- line chemotherapy . A 78-year-old man with advanced gastric cancer underwent a curative distal gastrectomy . Three years later , the tumor marker level began to rise and computed tomography ( CT ) revealed lymph node metastasis invading the pancreas resulting in pancreatic duct dilatation . S-1 treatment was initiated but was discontinued because of a systemic exanthem . Paclitaxel was administrated as secondary chemotherapy . But , after 2 courses , a further increase in the tumor marker level and portal vein invasion were observed . Combination therapy of CPT-11 and CDDP was administered as the third-line chemotherapy . After 3 courses , the tumor marker level normalized , tumor size decreased , and the invasion was eliminated . Third-line chemotherapy against recurrent gastric cancer should be considered if patient performance status ( PS ) is maintained .
|
[
1,
0,
0,
0,
0,
0,
0,
0,
0,
0
] |
23268075
|
CDA-2 ( cell differentiation agent 2 ) , a urinary preparation , has potent anti- proliferative and pro-apoptotic properties in cancer cells . However , the mechanisms of tumor inhibitory action of CDA-2 are far from clear , and especially there was no report on lung cancer . Here we demonstrate that CDA-2 and its main component phenylacetylglutamine ( PG ) reduce the metastatic lung tumor growth , and increases survival time after inoculation with Lewis lung carcinoma ( LLC ) cells in a dose-dependent manner in C57BL6 mice . Proliferative program analysis in cancer cells revealed a fundamental impact of CDA-2 and PG on proliferation and apoptosis , including Bcl-2 , Bcl-XL , cIAP1 , Survivin , PCNA , Ki-67 proteins and TUNEL assays . CDA-2 and PG significantly reduced NF-κB DNA-binding activity in lung cancer cells and in alveolar macrophages of tumor bearing mice and especially decreased the release of inflammatory factors including TNFα , IL-6 , and KC . Furthermore , CDA-2 and PG decrease the expressions of TLR2 , TLR6 , and CD14 , but not TLR1 , TLR3 , TLR4 , and TLR9 in bone-marrow-derived macrophages ( BMDM ) of mice stimulated by LLC-conditioned medium ( LLC-CM ) . Over-expressing TLR2 in BMDM prevented CDA-2 and PG from inhibiting NF-κB activation , as well as induction of TNFα and IL-6 . TLR2:TLR6 complexes mediate the effect of NF-κB inactivation by CDA-2 . In conclusion , CDA-2 potently inhibits lung tumor development by reduction of the inflammation in lung through suppression of NF-κB activation in myeloid cells , associating with modulation of TLR2 signaling .
|
[
1,
0,
0,
0,
0,
0,
0,
1,
0,
1
] |
23284890
|
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