LukS-PV通过C5aR诱导急性髓系白血病细胞THP-1凋亡的研究

Study on LukS-PV regulated apoptosis by C5aR in human acute myeloid leukemia THP-1 cells

目的探讨金黄色葡萄球菌杀白细胞毒素亚组分(LukS-PV)是否通过C5aR发挥其诱导急性髓系白血病细胞THP-1凋亡的作用。方法将THP-1细胞株分为5组:未处理细胞为对照组,使用不同浓度的C5aR拮抗剂(0、50、100、150 nmol/L)预先作用于THP-1细胞后,再使用1μmol/L LukS-PV刺激细胞为处理组。24 h后通过Annexin V/PI染色法,线粒体膜电位法检测细胞凋亡,qRT-PCR法检测凋亡相关基因,Western blot法检测凋亡相关蛋白,酶标仪法检测Caspase-3活性。结果经过不同浓度C5aR拮抗剂干预后,LukS-PV诱导THP-1细胞凋亡的比率逐渐下降,促凋亡基因Bax、Bak、Bim和促凋亡蛋白cleaved-Caspase-3、Bak、Bax的表达量也逐渐减低,抑凋亡基因Bcl-2、Bcl-w、Bcl-x和抑凋亡蛋白Bcl-2的表达量逐渐升高,Caspase-3活性逐渐降低。结论LukS-PV可能通过结合C5aR从而激活下游线粒体通路,发挥其诱导人急性髓系白血病细胞THP-1凋亡的作用。

Objective To study the role of CSaR on the apoptosis induced by subunit of Panton-Valentine leukocidin （LukS-PV） in the acute myeloid leukemia THP-1 cells. Methods THP-1 ceils were divided into five groups： the control group for untreated ceils ,the use of 1 μmol/L LukS-PV to stimulate THP-1 cells which had been treated with different concentrations （0,50,100,150 nmoL/L） of CSaR antagonists for processing cell group. After 24 h, the rates of cells apoptosis were detected by Annexin V/PI staining and mitochondrial membrane potential method. The expression of apoptosis genes were detected by qRT-PCR. The expression of apoptosis proteins were detected by Western blot and the Caspase-3 activity was also examined. Results Because of the intervention of different concentrations of CSaR antagonist, the ratio of cells apoptosis induced by LukS-PV gradually decreased; the ex- pression of pro-apoptosis genes Bax, Bak, Bim and pro-apoptosis proteins cleaved-Caspase-3, Bak, Bax gradually reduced ; the expression of anti-apoptosis genes Bcl-2, Bcl-w, Bcl-x and anti-apoptosis proteins Bcl-2 increased and the Caspase-3 activity also gradually decreased. Conclusion LukS-PV perhaps regulates apoptosis in human acute myeloid leukemia THP-1 cells by CSaR/mitochondrial pathway.