蛋白激酶Cδ通过抑制自噬促进TRAIL诱导前列腺癌细胞凋亡
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摘要
目的探讨蛋白激酶Cδ在去势抵抗性前列腺癌细胞中对自噬的影响,并研究其相关机制。方法使用去势抵抗性前列腺癌细胞C4-2和CWR22Rv1作为研究对象,将细胞进行分组,分别使用DMSO对照、BAF-A1、TRAIL、PKCδ激活剂PMA和PKCδ抑制剂rottlerin对细胞进行处理。用MTT检测各组药物处理后细胞存活状况;用Western blot检测各组药物处理后mTOR通路、自噬相关蛋白和凋亡相关蛋白的变化;用GFP-LC3荧光蛋白标记检测细胞内自噬作用的变化。结果 TRAIL诱导去势抵抗性前列腺癌细胞C4-2和CWR22Rv1自噬作用增强。PMA可以通过激活mTOR通路抑制自噬并提高细胞对TRAIL的凋亡敏感性。PKCδ在此过程中是抑制自噬作用的关键分子。结论本研究证实在去势抵抗性前列腺癌细胞C4-2和CWR22Rv1中,PKCδ/AKT/mTOR通路对细胞自噬的负向调节作用,激活该通路可以促进TRAIL诱导的细胞凋亡现象。
Objective To investigate the effects of protein kinase(PK)Cδon autophagy in castration-resistant prostate cancer cells and to explore the possible mechanism.Methods Castration-resistant prostate cancer cell lines C4-2 and CWR22 Rv1 were used as cell models,cultured and divided into DMSO control,BAF-A1,TRAIL,PMA and rottlerin groups.The cell survival was detected with MTT assay.The changes of mTOR pathway,autophagy-related protein markers and apoptosis-related protein markers were assessed with Western blot.The change of autophagy was determined with GFP-LC3.Results The autophagy of C4-2 and CWR22 Rv1 cells was induced by TRAIL.PMA inhibited the autophagy by activating the mTOR signaling pathway and increasing the apoptosis sensitivity to TRAIL.PKCδwas a key molecule inhibiting autophagy in this process.Conclusion The PKCδ/AKT/mTOR pathway negatively regulates autophagy of C4-2 and CWR22 Rv1 cells,and activation of this pathway promotes TRAIL-induced apoptosis.
Objective To investigate the effects of protein kinase(PK)Cδon autophagy in castration-resistant prostate cancer cells and to explore the possible mechanism.Methods Castration-resistant prostate cancer cell lines C4-2 and CWR22 Rv1 were used as cell models,cultured and divided into DMSO control,BAF-A1,TRAIL,PMA and rottlerin groups.The cell survival was detected with MTT assay.The changes of mTOR pathway,autophagy-related protein markers and apoptosis-related protein markers were assessed with Western blot.The change of autophagy was determined with GFP-LC3.Results The autophagy of C4-2 and CWR22 Rv1 cells was induced by TRAIL.PMA inhibited the autophagy by activating the mTOR signaling pathway and increasing the apoptosis sensitivity to TRAIL.PKCδwas a key molecule inhibiting autophagy in this process.Conclusion The PKCδ/AKT/mTOR pathway negatively regulates autophagy of C4-2 and CWR22 Rv1 cells,and activation of this pathway promotes TRAIL-induced apoptosis.
引文
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[24]CHEN JL,LIN HH,KIM KJ,et al.PKC delta signaling:a dual role in regulating hypoxic stress-induced autophagy and apoptosis[J].Autophagy,2009,5(2):244-246.
[25]SINGH BN,KUMAR D,SHANKAR S,et al.Rottlerin induces autophagy which leads to apoptotic cell death through inhibition of PI3K/Akt/mTOR pathway in human pancreatic cancer stem cells[J].Biochem Pharmacol,2012,84(9):1154-1163.
[2]ABDULGHANI J,EL-DEIRY WS.TRAIL receptor signaling and therapeutics[J].Expert Opin Ther Targets,2010,14(10):1091-1108.
[3]WU Z,CHANG PC,YANG JC,et al.Autophagy blockade sensitizes prostate cancer cells towards src family kinase inhibitors[J].Genes&cancer,2010,1(1):40-49.
[4]JIN Z,LI Y,PITTI R,et al.Cullin3-based polyubiquitination and p62-dependent aggregation of caspase-8mediate extrinsic apoptosis signaling[J].Cell,2009,137(4):721-735.
[5]SINGH K,SHARMA A,MIR MC,et al.Autophagic flux determines cell death and survival in response to Apo2L/TRAIL(dulanermin)[J].Mol Cancer,2014,13:70.
[6]SHI Q,JIA J,HUI K,et al.KLF5promotes apoptosis induced by phorbol ester as an effector of the autocrine factor TNFalpha in LNCaP prostate cancer cells[J].Oncol Lett,2017,14(2):1847-1854.
[7]ALTMAN A,KONG KF.Protein Kinase C Enzymes in the Hematopoietic and immune systems[J].Annu Rev Immunol,2016,34:511-538.
[8]ISAKOV N.Protein kinase C(PKC)isoforms in cancer,tumor promotion and tumor suppression[J].Semin Cancer Biol,2018,48:36-52.
[9]GASPARIAN ME,BYCHKOV ML,YAGOLOVICH AV,et al.The effect of cisplatin on cytotoxicity of anticancer cytokine TRAIL and its receptor-selective mutant variant DR5-B(1)[J].Dokl Biochem Biophys,2017,477(1):385-388.
[10]WEI B,LIANG J,HU J,et al.TRAF2is a valuable prognostic biomarker in patients with prostate cancer[J].Med Sci Monit,2017,23:4192-4204.
[11]WEI B,RUAN J,MI Y,et al.Knockdown of TNF receptor-associated factor 2(TRAF2)modulates in vitro growth of TRAIL-treated prostate cancer cells[J].Biomed Pharmacother,2017,93:462-469.
[12]KIM YC,GUAN K L.mTOR:apharmacologic target for autophagy regulation[J].J Clin Invest,2015,125(1):25-32.
[13]OCANA A,VERA-BADILLO F,AL-MUBARAK M,et al.Activation of the PI3K/mTOR/AKT pathway and survival in solid tumors:systematic review and meta-analysis[J].PLoS One,2014,9(4):e95219.
[14]NORMAN LL,BRUGUES J,SENGUPTA K,et al.Cell blebbing and membrane area homeostasis in spreading and retracting cells[J].Biophysic J,2010,99(6):1726-1733.
[15]DICKSTEIN RJ,NITTI G,DINNEY CP,et al.Autophagy limits the cytotoxic effects of the AKT inhibitor AZ7328in human bladder cancer cells[J].Cancer Biol Ther,2012,13(13):1325-1338.
[16]COOKE M,MAGIMAIDAS A,CASADO-MEDRANO V,et al.Protein kinase C in cancer:The top five unanswered questions[J].Mol Carcinog,2017,56(6):1531-1542.
[17]ZHOU X,SHEN L,PARRIS T,et al.Regulation of the viability of Nf1deficient cells by PKC isoforms[J].Oncotarget,2014,5(21):10709-10717.
[18]KIM H,NA YR,KIM SY,et al.Protein Kinase C Isoforms Differentially Regulate Hypoxia-Inducible Factor-1alpha Accumulation in Cancer Cells[J].J Cell Biochem,2016,117(3):647-658.
[19]KAWAKAMI Y,KITAURA J,YAO L,et al.A Ras activation pathway dependent on Syk phosphorylation of protein kinase C[J].Proceed Nation Acad Sci USA,2003,100(16):9470-9475.
[20]CONLEY-LACOMB M K,SALIGANAN A,KANDAGATLAP,et al.PTEN loss mediated Akt activation promotes prostate tumor growth and metastasis via CXCL12/CXCR4signaling[J].Mol Cancer,2013,12(1):85.
[21]BERARDI D E,FLUMIAN C,RODRIGUEZ C E,et al.PKCdelta inhibition impairs mammary cancer proliferative capacity but selects cancer stem cells,involving autophagy[J].J Cell Biochem,2016,117(3):730-740.
[22]HAFEEZ A,ELMADHOUN O,PENG C,et al.Reduced Apoptosis by Ethanol and Its Association with PKC-delta and Akt Signaling in Ischemic Stroke[J].Aging Dis,2014,5(6):366-372.
[23]MARQUINA-SANCHEZ B,GONZALEZ-JORGE J,HANS-BERG-PASTOR V,et al.The interplay between intracellular progesterone receptor and PKC plays a key role in migration and invasion of human glioblastoma cells[J].J Steroid Biochem Mol Biol,2017,172:198-206.
[24]CHEN JL,LIN HH,KIM KJ,et al.PKC delta signaling:a dual role in regulating hypoxic stress-induced autophagy and apoptosis[J].Autophagy,2009,5(2):244-246.
[25]SINGH BN,KUMAR D,SHANKAR S,et al.Rottlerin induces autophagy which leads to apoptotic cell death through inhibition of PI3K/Akt/mTOR pathway in human pancreatic cancer stem cells[J].Biochem Pharmacol,2012,84(9):1154-1163.
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