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1.
Dongqiang Song Yuanmin Li Jiatian Cao Zhihua Han Lin Gao Zuojun Xu Zhaofang Yin Guifang Wang Yuqi Fan Changqian Wang 《PloS one》2013,8(6)
Aim
Iron deficiency is a common comorbidity in chronic heart failure (CHF) which may exacerbate CHF. The c-kit+ cardiac stem cells (CSCs) play a vital role in cardiac function repair. However, much is unknown regarding the role of iron deficiency in regulating c-kit+ CSCs function. In this study, we investigated whether iron deficiency regulates c-kit+ CSCs proliferation, migration, apoptosis, and differentiation in vitro.Method
All c-kit+ CSCs were isolated from adult C57BL/6 mice. The c-kit+ CSCs were cultured with deferoxamine (DFO, an iron chelator), mimosine (MIM, another iron chelator), or a complex of DFO and iron (Fe(III)), respectively. Cell migration was assayed using a 48-well chamber system. Proliferation, cell cycle, and apoptosis of c-kit+ CSCs were analyzed with BrdU labeling, population doubling time assay, CCK-8 assay, and flow cytometry. Caspase-3 protein level and activity were examined with Western blotting and spectrophotometric detection. The changes in the expression of cardiac-specific proteins (GATA-4,TNI, and β-MHC) and cell cycle-related proteins (cyclin D1, RB, and pRB) were detected with Western blotting.Result
DFO and MIM suppressed c-kit+ CSCs proliferation and differentiation. They also modulated cell cycle and cardiac-specific protein expression. Iron chelators down-regulated the expression and phosphorylation of cell cycle-related proteins. Iron reversed those suppressive effects of DFO. DFO and MIM didn’t affect c-kit+ CSCs migration and apoptosis.Conclusion
Iron deficiency suppressed proliferation and differentiation of c-kit+ CSCs. This may partly explain how iron deficiency affects CHF prognosis. 相似文献2.
Wei Lin Lixia Jin Hua Chen Qingjun Wu Yunyun Fei Wenjie Zheng Qian Wang Ping Li Yongzhe Li Wen Zhang Yan Zhao Xiaofeng Zeng Fengchun Zhang 《Arthritis research & therapy》2014,16(3):R118
Introduction
IgG4-related disease (IgG4-RD) is a multisystem-involved autoimmune disease. Abnormally activated and differentiated B cells may play important roles. Regulatory B cells (Breg) are newly defined B cell subgroups with immunosuppressive functions. In this study, we investigated the differences of B cell subsets, the expressions of co-stimulatory molecules on B cells, and the function of Breg cells in patients with IgG4-RD, primary Sjögren’s syndrome (pSS) as well as in healthy controls (HC).Methods
Newly diagnosed IgG4-RD patients (n = 48) were enrolled, 38 untreated pSS patients and 30 healthy volunteers were recruited as disease and healthy controls. To analyze B cell subsets and B cell activity, PBMCs were surface stained and detected by flow cytometry. The function of Breg cells was tested by coculturing isolated CD19 + CD24hiCD38hi Breg cells with purified CD4 + CD25- T cells. Serum cytokines were measured by ELISA and cytometric bead array. Relationship between clinical data and laboratory findings were analyzed as well.Results
Compared with pSS patients and HC, IgG4-RD patients had a lower frequency of peripheral Breg cells. Interestingly, CD19 + CD24-CD38hi B cell subsets were significantly higher in peripheral B cells from IgG4-RD patients than in pSS patients and HC, which correlated with serum IgG4 levels. The expression of BAFF-R and CD40 on B cells was significantly lower in IgG4-RD patients compared with those in pSS patients and HC. Unlike HC, Breg cells from pSS patients lacked suppressive functions.Conclusions
B cells in patients with IgG4-RD and pSS display a variety of abnormalities, including disturbed B cell subpopulations, abnormal expression of key signaling molecules, co-stimulatory molecules, and inflammatory cytokines. In addition, a significantly increased B cell subset, CD19 + CD24-CD38hi B cells, may play an important role in the pathogenesis of IgG4-RD. 相似文献3.
Ilaria Nichele Alberto Zamò Anna Bertolaso Francesco Bifari Martina Tinelli Marta Franchini Roberta Stradoni Fiorenza Aprili Giovanni Pizzolo Mauro Krampera 《PloS one》2012,7(12)
Background
small B-cell neoplasms can show plasmacytic differentiation and may potentially progress to aggressive lymphoma (DLBCL). Epstein-Barr virus (EBV) infection may cause the transformation of malignant cells in vitro.Design and Method
we established VR09 cell line with plasmacytic differentiation, obtained from a case of atypical, non-CLL B-cell chronic lymphoproliferative disease with plasmacytic features. We used flow cytometry, immunohistochemistry, polymerase chain reaction, cytogenetic analysis and florescence in situ hybridization in the attempt at thoroughly characterizing the cell line. We showed VR09 tumorigenic potential in vivo, leading to the development of activated DLBCL with plasmacytic features.Results
VR09 cells displayed plasmacytic appearance and grew as spherical tumors when inoculated subcutaneously into immunodeficient Rag2−/− γ-chain−/− mice. VR09 cell line and tumors displayed the phenotype of activated stage of B cell maturation, with secretory differentiation (CD19+ CD20+ CD79a+ CD79b+/− CD138+ cyclin D1- Ki67 80% IgM+ IgD+ MUM1+ MNDA+ CD10- CD22+ CD23+ CD43+ K+, λ- Bcl2+ Bcl6-) and they presented episomal EBV genome, chromosome 12 trisomy, lack of c-MYC rearrangement and Myd88 gene mutation, presence of somatic hypermutation in the VH region, and wild-type p53.Conclusion
This new EBV-positive cell line may be useful to further characterize in vivo activated DLBCL with plasmacytic features. 相似文献4.
Fen Li Xuan Yu Claudia K. Szynkarski Cong Meng Beiyan Zhou Rola Barhoumi Richard E. White Cristine L. Heaps John N. Stallone Guichun Han 《PloS one》2013,8(6)
Background
Vascular pathology and dysfunction are direct life-threatening outcomes resulting from atherosclerosis or vascular injury, which are primarily attributed to contractile smooth muscle cells (SMCs) dedifferentiation and proliferation by re-entering cell cycle. Increasing evidence suggests potent protective effects of G-protein coupled estrogen receptor 1 (GPER) activation against cardiovascular diseases. However, the mechanism underlying GPER function remains poorly understood, especially if it plays a potential role in modulating coronary artery smooth muscle cells (CASMCs).Methodology/Principal Findings
The objective of our study was to understand the functional role of GPER in CASMC proliferation and differentiation in coronary arteries using from humans and swine models. We found that the GPER agonist, G-1, inhibited both human and porcine CASMC proliferation in a concentration- (10−8 to 10−5 M) and time-dependent manner. Flow cytometry revealed that treatment with G-1 significantly decreased the proportion of S-phase and G2/M cells in the growing cell population, suggesting that G-1 inhibits cell proliferation by slowing progression of the cell cycle. Further, G-1-induced cell cycle retardation was associated with decreased expression of cyclin B, up-regulation of cyclin D1, and concomitant induction of p21, and partially mediated by suppressed ERK1/2 and Akt pathways. In addition, G-1 induces SMC differentiation evidenced by increased α-smooth muscle actin (α-actin) and smooth muscle protein 22α (SM22α) protein expressions and inhibits CASMC migration induced by growth medium.Conclusion
GPER activation inhibits CASMC proliferation by suppressing cell cycle progression via inhibition of ERK1/2 and Akt phosphorylation. GPER may constitute a novel mechanism to suppress intimal migration and/or synthetic phenotype of VSMC. 相似文献5.
6.
7.
Objective
To evaluate changes of nuclear factor-kappa B (NF-κB) during radioiodine 131 (131I) therapy and whether NF-κB inhibition could enhance 131I-induced apoptosis in differentiated thyroid cancer (DTC) cells in a synergistic manner.Methods
Three human DTC cell lines were used. NF-κB inhibition was achieved by using a NF-κB inhibitor (Bay 11-7082) or by p65 siRNA transfection. Methyl-thiazolyl-tetrazolium assay was performed for cell viability assessment. DNA-binding assay, luciferase reporter assay, and Western blot were adopted to determine function and expression changes of NF-κB. Then NF-κB regulated anti-apoptotic factors XIAP, cIAP1, and Bcl-xL were measured. Apoptosis was analyzed by Western blot for caspase 3 and PARP, and by flow cytometry as well. An iodide uptake assay was performed to determine whether NF-κB inhibition could influence radioactive iodide uptake.Results
The methyl-thiazolyl-tetrazolium assay showed significant decrease of viable cells by combination therapy than by mono-therapies. The DNA-binding assay and luciferase reporter assay showed enhanced NF-κB function and reporter gene activities due to 131I, yet significant suppression was achieved by NF-κB inhibition. Western blot proved 131I could increase nuclear NF-κB concentration, while NF-κB inhibition reduced NF-κB concentration. Western blot also demonstrated significant up-regulation of XIAP, cIAP1, and Bcl-xL after 131I therapy. And inhibition of NF-κB could significantly down-regulate these factors. Finally, synergism induced by combined therapy was displayed by significant enhancements of cleaved caspase 3 and PARP from Western blot, and of Annexin V positively staining from flow cytometry. The iodine uptake assay did not show significant changes when NF-κB was inhibited.Conclusion
We demonstrated that 131I could induce NF-κB activation, which would attenuate 131I efficacy in DTC cells. NF-κB inhibition by Bay 11-7082 or by p65 siRNA transfection was effective in suppressing NF-κB regulated anti-apoptotic changes and in combined regimen apoptosis was achieved synergistically. 相似文献8.
Rosa H. Jimenez Joan M. Boylan Ju-Seog Lee Mirko Francesconi Gastone Castellani Jennifer A. Sanders Philip A. Gruppuso 《PloS one》2009,4(10)
Background
The mTOR inhibitor rapamycin has anti-tumor activity across a variety of human cancers, including hepatocellular carcinoma. However, resistance to its growth inhibitory effects is common. We hypothesized that hepatic cell lines with varying rapamycin responsiveness would show common characteristics accounting for resistance to the drug.Methodology/Principal Findings
We profiled a total of 13 cell lines for rapamycin-induced growth inhibition. The non-tumorigenic rat liver epithelial cell line WB-F344 was highly sensitive while the tumorigenic WB311 cell line, originally derived from the WB-F344 line, was highly resistant. The other 11 cell lines showed a wide range of sensitivities. Rapamycin induced inhibition of cyclin E–dependent kinase activity in some cell lines, but the ability to do so did not correlate with sensitivity. Inhibition of cyclin E–dependent kinase activity was related to incorporation of p27Kip1 into cyclin E–containing complexes in some but not all cell lines. Similarly, sensitivity of global protein synthesis to rapamycin did not correlate with its anti-proliferative effect. However, rapamycin potently inhibited phosphorylation of two key substrates, ribosomal protein S6 and 4E-BP1, in all cases, indicating that the locus of rapamycin resistance was downstream from inhibition of mTOR Complex 1. Microarray analysis did not disclose a unifying mechanism for rapamycin resistance, although the glycolytic pathway was downregulated in all four cell lines studied.Conclusions/Significance
We conclude that the mechanisms of rapamycin resistance in hepatic cells involve alterations of signaling downstream from mTOR and that the mechanisms are highly heterogeneous, thus predicting that maintaining or promoting sensitivity will be highly challenging. 相似文献9.
Liang Ye Zhongyang Wen Yanqun Li Bingni Chen Ting Yu Lanying Liu Jinshun Zhang Yanmei Ma Shuying Xiao Liping Ding Li Li Zhong Huang 《Arthritis research & therapy》2014,16(2):R96
Introduction
Our objective in the present study was to determine the signaling pathway of interleukin 10 (IL-10) for modulating IL-17 expression in macrophages and the importance of this mediation in collagen-induced arthritis (CIA).Methods
IL-10-knockout (IL-10−/−) mice and wild-type (WT) mice were immunized with chicken type II collagen (CII) to induce arthritis. The expression levels of IL-17 and retinoid-related orphan receptor γt (RORγt) in macrophages and joint tissues of IL-10−/− and WT mice were analyzed by enzyme-linked immunosorbent assay, quantitative RT-PCR (qRT-PCR) and Western blotting. The F4/80 macrophages and positive IL-17-producing macrophages in synovial tissues of the mice were determined by immunohistochemistry. The populations of classically activated macrophage (M1) and alternatively activated macrophage (M2) phenotypes were analyzed by flow cytometry. The expression of genes associated with M1 and M2 markers was analyzed by qRT-PCR.Results
Compared to WT mice, IL-10−/− mice had exacerbated CIA development, which was associated with increased production of T helper 17 cell (Th17)/Th1 proinflammatory cytokines and CII-specific immunoglobulin G2a antibody after CII immunization. Macrophages in IL-10−/− mice had increased amounts of IL-17 and RORγt compared with the amounts in WT mice with CIA. Immunofluorescence microscopy showed that the number of IL-17-producing macrophages in synovial tissues was significantly higher in IL-10−/− mice than in WT mice. IL-10 deficiency might promote macrophage polarization toward the proinflammatory M1 phenotype, which contributes to the rheumatoid arthritis inflammation response.Conclusion
IL-10 inhibits IL-17 and RORγt expression in macrophages and suppresses macrophages toward the proinflammatory M1 phenotype, which is important for the role of IL-10 in mediating the pathogenesis of CIA. 相似文献10.
Wunchana Seubwai Chaisiri Wongkham Anucha Puapairoj Narong Khuntikeo Ake Pugkhem Chariya Hahnvajanawong Jariya Chaiyagool Kazuo Umezawa Seiji Okada Sopit Wongkham 《PloS one》2014,9(8)
Background
Up-regulation and association of nuclear factor kappa B (NF-κB) with carcinogenesis and tumor progression has been reported in several malignancies. In the current study, expression of NF-κB in cholangiocarcinoma (CCA) patient tissues and its clinical significance were determined. The possibility of using NF-κB as the therapeutic target of CCA was demonstrated.Methodology
Expression of NF-κB in CCA patient tissues was determined using immunohistochemistry. Dehydroxymethylepoxyquinomicin (DHMEQ), a specific NF-κB inhibitor, was used to inhibit NF-κB action. Cell growth was determined using an MTT assay, and cell apoptosis was shown by DNA fragmentation, flow cytometry and immunocytofluorescent staining. Effects of DHMEQ on growth and apoptosis were demonstrated in CCA cell lines and CCA-inoculated mice. DHMEQ-induced apoptosis in patient tissues using a histoculture drug response assay was quantified by TUNEL assay.Principal Findings
Normal bile duct epithelia rarely expressed NF-κB (subunits p50, p52 and p65), whereas all CCA patient tissues (n = 48) over-expressed all NF-κB subunits. Inhibiting NF-κB action by DHMEQ significantly inhibited growth of human CCA cell lines in a dose- and time-dependent manner. DHMEQ increased cell apoptosis by decreasing the anti-apoptotic protein expressions–Bcl-2, XIAP–and activating caspase pathway. DHMEQ effectively reduced tumor size in CCA-inoculated mice and induced cell apoptosis in primary histocultures of CCA patient tissues.Conclusions
NF-κB was over-expressed in CCA tissues. Inhibition of NF-κB action significantly reduced cell growth and enhanced cell apoptosis. This study highlights NF-κB as a molecular target for CCA therapy. 相似文献11.
Purpose
To examine whether 99mTc(V)-DMSA could be used as a non-invasive measure of cancer cell proliferation.Methods
Human breast cancer MCF-7, MDA-MB-231 and pII, and prostate cancer PC-3 cell lines were grown to 30, 50 and 100% confluency and pulsed with 99mTc(V)-DMSA in media for 60 min at 37°C. DNA synthesis was analysed by quantification of the S phase using flow cytometry, [methyl-3H]thymidine incorporation and expression of proliferation markers PCNA and Ki-67 using realtime PCR. One way ANOVA was used to compare groups.Results
In all cell lines rates of 99mTc(V)-DMSA uptake were inversely related to cell density. This was paralleled by similar trends in S phase proportions, [methyl-3H]thymidine incorporation and expression of PCNA and Ki-67.Conclusion
Rates of 99mTc(V)-DMSA uptake into different types of tumour cells correlate well with cell density that is useful as a non-invasive measure of tumour cellular proliferation in vivo. 相似文献12.
Geng Guo Dong Kuai Sang Cai Naizhao Xue Yueting Liu Jiehe Hao Yimin Fan Ji Jin Xinggang Mao Bolin Liu Chengliang Zhong Xiang Zhang Yi Yue Xiaodong Liu Ning Ma Yuhong Guo 《PloS one》2013,8(4)
Background
FRAT1 positively regulates the Wnt/β-catenin signaling pathway by inhibiting GSK-3-mediated phosphorylation of β-catenin. It was originally characterized as a protein frequently rearranged in advanced T cell lymphoma, but has recently also been identified as a proto-oncogene involved in tumorigenesis. Our previous studies showed that FRAT1 was dramatically overexpressed in gliomas and its expression level was significantly increased along with clinicopathological grades.Methods
In the current study, we used RT-PCR and Western blotting to assess the mRNA and protein levels of FRAT1 in three glioma cell lines. In addition, to evaluate its functional role in gliomas, we examined the effects of FRAT1 knockdown on proliferation, migration and invasion in vitro and tumor growth in vivo using glioblastoma U251 cells and RNAi.Results
FRAT1 was highly expressed in all three glioma cell lines. RNAi-mediated down-regulation of endogenous FRAT1 in human glioblastoma U251 cells resulted in suppression of cell proliferation, arrest of cell cycle, inhibition of cell migration and invasion in vitro. Moreover, FRAT1 depletion significantly impaired tumor xenograft growth in nude mice.Conclusions
Our results highlight the potential role of FRAT1 in tumorigenesis and progression of glioblastoma. These findings provide a biological basis for FRAT1 as a potential molecular marker for improved pathological grading and as a novel candidate therapeutic target for glioblastoma management. 相似文献13.
Cécilia Szatkowski Judith Vallet Mojdeh Dormishian Nadia Messaddeq Phillippe Valet Mounia Boulberdaa Daniel Metzger Pierre Chambon Canan G. Nebigil 《PloS one》2013,8(12)
Background
Adipocyte renewal from preadipocytes occurs throughout the lifetime and contributes to obesity. To date, little is known about the mechanisms that control preadipocyte proliferation and differentiation. Prokineticin-2 is an angiogenic and anorexigenic hormone that activate two G protein-coupled receptors (GPCRs): PKR1 and PKR2. Prokineticin-2 regulates food intake and energy metabolism via central mechanisms (PKR2). The peripheral effect of prokineticin-2 on adipocytes/preadipocytes has not been studied yet.Methodology/Principal Findings
Since adipocytes and preadipocytes express mainly prokineticin receptor-1 (PKR1), here, we explored the role of PKR1 in adipose tissue expansion, generating PKR1-null (PKR1−/−) and adipocyte-specific (PKR1ad−/−) mutant mice, and using murine and human preadipocyte cell lines. Both PKR1−/− and PKR1ad−/− had excessive abdominal adipose tissue, but only PKR1−/− mice showed severe obesity and diabetes-like syndrome. PKR1ad−/−) mice had increased proliferating preadipocytes and newly formed adipocyte levels, leading to expansion of adipose tissue. Using PKR1-knockdown in 3T3-L1 preadipocytes, we show that PKR1 directly inhibits preadipocyte proliferation and differentiation. These PKR1 cell autonomous actions appear targeted at preadipocyte cell cycle regulatory pathways, through reducing cyclin D, E, cdk2, c-Myc levels.Conclusions/Significance
These results suggest PKR1 to be a crucial player in the preadipocyte proliferation and differentiation. Our data should facilitate studies of both the pathogenesis and therapy of obesity in humans. 相似文献14.
Background
Currently available methods for diagnosis and staging of prostate cancer lack the sensitivity to distinguish between patients with indolent prostate cancer and those requiring radical treatment. Alterations in key adherens (AJ) and tight junction (TJ) components have been hailed as potential biomarkers for prostate cancer progression but the majority of research has been carried out on individual molecules.Objective
To elucidate a panel of biomarkers that may help distinguish dormant prostate cancer from aggressive metastatic disease.Methods
We analysed the expression of 7 well known AJ and TJ components in cell lines derived from normal prostate epithelial tissue (PNT2), non-invasive (CAHPV-10) and invasive prostate cancer (LNCaP, DU145, PC-3) using gene expression, western blotting and immunofluorescence techniques.Results
Claudin 7, α –catenin and β-catenin protein expression were not significantly different between CAHPV-10 cells and PNT2 cells. However, in PC-3 cells, protein levels for claudin 7, α –catenin were significantly down regulated (−1.5 fold, p = <.001) or undetectable respectively. Immunofluoresence showed β-catenin localisation in PC-3 cells to be cytoplasmic as opposed to membraneous.Conclusion
These results suggest aberrant Claudin 7, α – and β-catenin expression and/or localisation patterns may be putative markers for distinguishing localised prostate cancer from aggressive metastatic disease when used collectively. 相似文献15.
Background
The sympathetic neurotransmitter Norepinephrine (NE) contributes to tumorigenesis and cancer progression. This study aims to investigate the role of NE in modulating the immune phenotype and allowing pancreatic carcinoma (PC) cells to escape the immune response.Methods
Varied concentrations of NE and interferon-gamma (IFN-γ) were administrated to MIA PaCa-2 and BxPC-3 cell lines for 48 hours. Proliferation and invasion were then investigated using an MTT assay and a membrane invasion culture system respectively. MHC-I, B7-1, IDO and B7-H1 expression were measured using real-time quantitative RT-PCR, western blotting and immunocytochemistry. The synergistic and time-dependent effects of NE/IFN-γ were also investigated. Adrenergic antagonists were used to identify the relevant target receptor of NE.Results
The results showed that NE had dose-dependent and time-dependent effects on cell biological processes as well as on the expression of MHC-I, B7-1, IDO and B7-H1. These effects occurred mainly via the β2-adrenergic receptor. Long-term NE treatment was able to antagonize some of the effects of IFN-γ (after 2 weeks of treatment), but NE and IFN-γ had significant synergistic stimulatory effects on IDO and B7-H1 expression. The residual effects on biological activities lasted for 2 weeks, while the immunophenotypic changes decreased at early time points after treatment.Conclusions
NE plays important roles in modulating PC cell biological activities and affecting MHC-I, B7-1, IDO and B7-H1 expression in vitro, mainly via the β2-adrenergic receptor (β2-AR) in a time- and dose-dependent fashion. Only at extended treatment durations could NE affect PC cell progression and immune evasion. 相似文献16.
Sreelatha Gopinath Rama Rao Malla Christopher S. Gondi Kiranmai Alapati Daniel Fassett Jeffrey D. Klopfenstein Dzung H. Dinh Meena Gujrati Jasti S. Rao 《PloS one》2010,5(7)
Background
Cathepsin B and urokinase plasminogen activator receptor (uPAR) are both known to be overexpressed in gliomas. Our previous work and that of others strongly suggest a relationship between the infiltrative phenotype of glioma and the expression of cathepsin B and uPAR. Though their role in migration and adhesion are well studied the effect of these molecules on cell cycle progression has not been thoroughly examined.Methodology/Principal Findings
Cathespin B and uPAR single and bicistronic siRNA plasmids were used to downregulate these molecules in SNB19 and U251 glioma cells. FACS analysis and BrdU incorporation assay demonstrated G0/G1 arrest and decreased proliferation with the treatments, respectively. Immunoblot and immunocyto analysis demonstrated increased expression of p27Kip1 and its nuclear localization with the knockdown of cathepsin B and uPAR. These effects could be mediated by αVβ3/PI3K/AKT/FOXO pathway as observed by the decreased αVβ3 expression, PI3K and AKT phosphorylation accompanied by elevated FOXO3a levels. These results were further confirmed with the increased expression of p27Kip1 and FOXO3a when treated with (10 µM) and increased luciferase expression with the siRNA and Ly294002 treatments when the FOXO binding promoter region of p27Kip1 was used. Our treatment also reduced the expression of cyclin D1, cyclin D2, p-Rb and cyclin E while the expression of Cdk2 was unaffected. Of note, the Cdk2-cyclin E complex formation was reduced significantly. Ly294002Conclusion/Significance
Our study indicates that cathepsin B and uPAR knockdown induces G0/G1 arrest by modulating the PI3K/AKT signaling pathway and further increases expression of p27Kip1 accompanied by the binding of FOXO3a to its promoter. Taken together, our findings provide molecular mechanism for the G0/G1 arrest induced by the downregulation of cathepsin B and uPAR in SNB19 and U251 glioma cells. 相似文献17.
Deepika Kanojia Manoj Garg Shikha Saini Sumit Agarwal Deepak Parashar Nirmala Jagadish Amlesh Seth Amar Bhatnagar Anju Gupta Rajive Kumar Nirmal Kumar Lohiya Anil Suri 《PloS one》2013,8(12)
Background
Majority of bladder cancer deaths are caused due to transitional cell carcinoma (TCC) which is the most prevalent and chemoresistant malignancy of urinary bladder. Therefore, we analyzed the role of Sperm associated antigen 9 (SPAG9) in bladder TCC.Methodology and Findings
We examined SPAG9 expression and humoral response in 125 bladder TCC patients. Four bladder cancer cell lines were assessed for SPAG9 expression. In addition, we investigated the effect of SPAG9 ablation on cellular proliferation, cell cycle, migration and invasion in UM-UC-3 bladder cancer cells by employing gene silencing approach. Our SPAG9 gene and protein expression analysis revealed SPAG9 expression in 81% of bladder TCC tissue specimens. High SPAG9 expression (>60% SPAG9 positive cells) was found to be significantly associated with superficial non-muscle invasive stage (P = 0.042) and low grade tumors (P = 0.002) suggesting SPAG9 putative role in early spread and tumorigenesis. Humoral response against SPAG9 was observed in 95% of patients found positive for SPAG9 expression. All four bladder cancer cell lines revealed SPAG9 expression. In addition, SPAG9 gene silencing in UM-UC-3 cells resulted in induction of G0–G1 arrest characterized by up-regulation of p16 and p21 and consequent down-regulation of cyclin E, cyclin D and cyclin B, CDK4 and CDK1. Further, SPAG9 gene silencing also resulted in reduction in cellular growth, and migration and invasion ability of cancer cells in vitro.Conclusions
Collectively, our data in clinical specimens indicated that SPAG9 is potential biomarker and therapeutic target for bladder TCC. 相似文献18.
Objective
Mitochondrial oxidative stress is the basis for pancreatic β-cell apoptosis and a common pathway for numerous types of damage, including glucotoxicity and lipotoxicity. We cultivated mice pancreatic β-cell tumor Min6 cell lines in vitro and observed pancreatic β-cell apoptosis and changes in mitochondrial function before and after the addition of Exendin-4. Based on these observations, we discuss the protective role of Exendin-4 against mitochondrial oxidative damage and its relationship with Ca2+-independent phospholipase A2.Methods
We established a pancreatic β-cell oxidative stress damage model using Min6 cell lines cultured in vitro with tert-buty1 hydroperoxide and hydrogen peroxide. We then added Exendin-4 to observe changes in the rate of cell apoptosis (Annexin-V-FITC-PI staining flow cytometry and DNA ladder). We detected the activity of the caspase 3 and 8 apoptotic factors, measured the mitochondrial membrane potential losses and reactive oxygen species production levels, and detected the expression of cytochrome c and Smac/DLAMO in the cytosol and mitochondria, mitochondrial Ca2-independent phospholipase A2 and Ca2+-independent phospholipase A2 mRNA.Results
The time-concentration curve showed that different percentages of apoptosis occurred at different time-concentrations in tert-buty1 hydroperoxide- and hydrogen peroxide-induced Min6 cells. Incubation with 100 µmol/l of Exendin-4 for 48 hours reduced the Min6 cell apoptosis rate (p<0.05). The mitochondrial membrane potential loss and total reactive oxygen species levels decreased (p<0.05), and the release of cytochrome c and Smac/DLAMO from the mitochondria was reduced. The study also showed that Ca2+-independent phospholipase A2 activity was positively related to Exendin-4 activity.Conclusion
Exendin-4 reduces Min6 cell oxidative damage and the cell apoptosis rate, which may be related to Ca2-independent phospholipase A2. 相似文献19.
Avignat S. Patel Jin Woo Song Sarah G. Chu Kenji Mizumura Juan C. Osorio Ying Shi Souheil El-Chemaly Chun Geun Lee Ivan O. Rosas Jack A. Elias Augustine M. K. Choi Danielle Morse 《PloS one》2015,10(3)
Background
Epithelial cell death is a major contributor to fibrogenesis in the lung. In this study, we sought to determine the function of mitochondria and their clearance (mitophagy) in alveolar epithelial cell death and fibrosis.Methods
We studied markers of mitochondrial injury and the mitophagy marker, PTEN-induced putative kinase 1 (PINK1), in IPF lung tissues by Western blotting, transmission electron microscopy (TEM), and immunofluorescence. In vitro experiments were carried out in lung epithelial cells stimulated with transforming growth factor-β1 (TGF-β1). Changes in cell function were measured by Western blotting, flow cytometry and immunofluorescence. In vivo experiments were performed using the murine bleomycin model of lung fibrosis.Results
Evaluation of IPF lung tissue demonstrated increased PINK1 expression by Western blotting and immunofluorescence and increased numbers of damaged mitochondria by TEM. In lung epithelial cells, TGF-β1 induced mitochondrial depolarization, mitochondrial ROS, and PINK1 expression; all were abrogated by mitochondrial ROS scavenging. Finally, Pink1 -/- mice were more susceptible than control mice to bleomycin induced lung fibrosis.Conclusion
TGF-β1 induces lung epithelial cell mitochondrial ROS and depolarization and stabilizes the key mitophagy initiating protein, PINK1. PINK1 ameliorates epithelial cell death and may be necessary to limit fibrogenesis. 相似文献20.