外源性硫化氢对人肾癌786-O细胞增殖及凋亡的影响
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摘要
目的探讨外源性硫化氢对人肾癌786-0细胞增殖和凋亡作用的影响及机制。方法 2017年6月一12月于武汉大学人民医院中心实验室进行实验,将不同浓度(0、10、25、50μmol/L)的硫化氢作用于体外培养的肾癌786-0细胞。采用MTT法检测硫化氢对细胞的增殖抑制作用;流式细胞术检测硫化氢对细胞凋亡的影响;Western blot法检测硫化氢对786-0细胞内Caspase-3和Bax蛋白表达的影响;RT-PCR法检测凋亡相关基因Caspase-3和Bax mRNA的表达水平。结果随着硫化氢浓度的增加,786-0细胞增殖率显著降低(F=20.24,P=0.001),凋亡率增加(F=7.66,P=0.004);细胞内Caspase-3和Bax蛋白表达水平增加(F=137.64,P=0.001),凋亡相关基因Caspase-3和BaxmRNA表达均显著增强(F=28.76、63.22,P均=0.001),具有显著的剂量依赖性。结论外源性硫化氢对人肾癌786-0细胞可显著抑制增殖和促进其凋亡,有望成为治疗和预防肾癌的有效药物。
Objective To investigate the effects and mechanism of hydrogen sulfide on proliferation and apoptosis in human renal cell carcinoma 786-0 cells. Methods The experiment was carried out in the Central Laboratory of Renmin Hospital of uhan University from June 2017 to December in 2017. The renal cell carcinoma 786-0 cells was exposed to different concentrations of hydrogen sulfide in virto. MTT and flow cytometry assays were used to measure the function of hydrogen sulfide on proliferation and apoptosis, respectively. The apoptosis related genes caspase 3 and Bax protein and mRNA levels were detected by western blot and qRT PCR, respectively. Results With increasing of hydrogen sulfide concentration, the proliferation rate of 786-0 cells decreased and the apoptosis increased gradually. Caspase 3 and Bax were significantly increased in the level of transcription and translation. Conclusion Hydrogen sulfide can significantly inhibit cell proliferation and promote cell apoptosis in human renal cell carcinoma 786-0 cells. It is expected to be an effective drug for the treatment and prevention of renal cell carcinoma.
Objective To investigate the effects and mechanism of hydrogen sulfide on proliferation and apoptosis in human renal cell carcinoma 786-0 cells. Methods The experiment was carried out in the Central Laboratory of Renmin Hospital of uhan University from June 2017 to December in 2017. The renal cell carcinoma 786-0 cells was exposed to different concentrations of hydrogen sulfide in virto. MTT and flow cytometry assays were used to measure the function of hydrogen sulfide on proliferation and apoptosis, respectively. The apoptosis related genes caspase 3 and Bax protein and mRNA levels were detected by western blot and qRT PCR, respectively. Results With increasing of hydrogen sulfide concentration, the proliferation rate of 786-0 cells decreased and the apoptosis increased gradually. Caspase 3 and Bax were significantly increased in the level of transcription and translation. Conclusion Hydrogen sulfide can significantly inhibit cell proliferation and promote cell apoptosis in human renal cell carcinoma 786-0 cells. It is expected to be an effective drug for the treatment and prevention of renal cell carcinoma.
引文
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[2]Siegel R,Ward E,Brawley 0,et al.Cancer statistics,2011:the impact of eliminating socioeconomic and racial disparities on premature cancer deaths[J].CA Cancer J Clin,2011,61(4):212-236.DOI:10.3322/caac.20121.
[3]Langrand-Escure J,Vallard A,Rivoirard R,et al.Safety assessment of molecular targeted therapies in association with radiotherapy in metastatic renal cell carcinoma:a real-life report[J].Anticancer Drugs,2016,27(5):427-432.DOI:10.1097/CAD.0000000000000349.
[4]Ivanyi P,Beutel G,Drewes N,et al.Therapy of treatment-related hypertension in metastatic renal-cell cancer patients receiving sunitinib[J].Clin Genitourin Cancer,2017,15(2):280-290.DOI:10.1016/j.clgc.2016.10.004.
[5]Yang CT,Chen L,Xu S,et al.Recent development of hydrogen sulfide releasing/stimulating reagents and their potential applications in cancer and glycometabolic disorderes[J].Front Pharmacol,2017,8:664.DOI:10.3389/fphar.2017.00664.
[6]Humphries LS,Baluch DA,Nystrom LM,et al.Interfascicular renal cell carcinoma metastasis to the ulnar nerve:A case report[J].Hand(NY),2016,11(2):1-4.DOI:10.1177/1558944715627620.
[7]Ljungberg B,Campbell SC,Choi HY,et al.The epidemiology of renal cell carcinoma[J].Eur Urol,2011,60(4):615-621.DOI:10.1016/j.eururo.2011.06.049.
[8]Win AZ.Renal cell carcinoma metastasis to the gallbladder detected by FDG-PET/CT[J].J Clin Med Res,2014.6(6):482-486.DOI:10.14740/jocmr2146e.
[9]Wu SW,Chen PN,Lin CY,et al.Everolimus suppresses invasion and migration of renal cell carcinoma by inhibiting FAK activity and reversing epithelial to mesenchymal transition in vitro and in vivo[J].Environ Toxicol,2017,32(7):1888-1898.DOI:10.1002/tox.22411.
[10]Lu M,Liu YH,Goh HS,et al.Hydrogen sulfide inhibits plasma renin activity[J].J Am Soc Nephrol,2010,21(6):993-1002.DOI:10.1681/ASN.2009090949.
[11]Bozlu M,Acar D,Cayan S,et al.Protective effect of trapidil on long-term histologic damage in a rat model of testicular ischemia-reperfusion injury[J].World J Urol,2009,27(I):117-122.DOI:10.1007/s00345-008-0323-7.
[12]Szabo C,Papapetropoulos A.International union of basic and clinical pharmacology.CII:pharmacological modulation of H_2S levels:H_2S donors and H_2S biosynthesis inhibitors[J].Pharmacol Rev,2017,69(4):497-564.DOI:10.1124/pr.117.014050.
[13]Nussbaum BL,Vogt J,Wachter U,et al.Metabolic,cardiac,and renal effects of the slow hydrogen sulfide-releasing molecule GYY4137 during resuscitated septic shock in swine with pre-existing coronary artery disease[J].Shock,2017,48(2):175-184.DOI:10.1097/SHK.0000000000000834.
[14]Wu D,Luo N,Wang L,et al.Hydrogen sulfide ameliorates chronic renal failure in rats by inhibiting apoptosis and inflammation through ROS/MAPK and NF-kappaB signaling pathways[J].Sci Rep,2017,7(1):455.DOI:10.1038/s41598-017-00557-2.
[15]Wang SS,Chen YH,Chen N,et al.Hydrogen sulfide promotes autophagy of hepatocellular carcinoma cells through the PI3K/Akt/mTOR signaling pathway[J].Cell Death Dis,2017,8(3):e2688.DOI:10.1038/cddis.2017.18.
[16]Olah G,Modis K,Toro G,et al.Role of endogenous and exogenous nitric oxide,carbon monoxide and hydrogen sulfide in HCT116 colon cancer cell proliferation[J].Biochem Pharmacol.2017.DOI:10.1016/j.bcp.2017.10.011.
[17]Feng W,Teo XY,Novera W,et al.Discovery of new H_2S releasing phosphordithioates and 2,3-Dihydro-2-phenyl-2-sulfanylenebenzo[d][1,3,2]oxazaphospholes with improved antiproliferative activity[J].J Med Chem,2015,58(16):6456-6480.DOI:10.1021/acs.jmedchem.5b00848.
[18]Jiang D,Wu D,Zhang Y,et al.Protective effects of hydrogen rich saline solution on experimental testicular ischemia-reperfusion injury in rats[J].J Urol,2012,187(6):2249-2253.DOI:10.1016/j.juro.2012.01.029.
[19]Mcgreal E P,Miller J L,Cordon S.Ligand recognition by antigenpresenting cell C-type lectin receptors[J].Curr Opin Immunol,2005.17(1):18-24.DOI:10.1016/j.coi.2004.12.001.
[20]赵丽敏,赵帅,毕仁杰,等.乳腺癌组织中caspase-3及caspase-8的表达及意义[J].中国老年学杂志,2017,37(14):3486-3488.DOI:10.3969/j.issn.1005-9202.2017.14.047.
[21]饶韬,陈迪诗,曾飞.Caspase-3和Caspase-6在胃癌组织中的表达及意义[J].南昌大学学报:医学版,2011,51(1):45-48,52.DOI:10.3969/j.issn.1000-2294.2011.01.015.
[22]Cai SP,Cheng XY,Chen PJ,et al.Transmembrane protein 88 attenuates liver fibrosis by promoting apoptosis and reversion of activated hepatic stellate cells[J].Mol Immunol,2016,80:58-67.DOI:10.1016/j.molimm.2016.11.002.
[23]Pu H,Wang X,Su L,et al.Heroin activates ATF3 and CytC via cJun N-terminal kinase pathways to mediate neuronal apoptosis[J].Med Sci Monit Basic Res,2015,21:53-62.DOI:10.12659/MSMBR.893827.
[24]Amoult D,Parone P,Martinou J C,et al.Mitochondrial release of apoptosis-inducing factor occurs downstream of cytochrome c release in response to several proapoptotic stimuli[J].J Cell Biol,2002,159(6):923-929.DOI:10.1083/jcb.200207071.