柳氮磺胺吡啶诱导HSC-T6肝星状细胞凋亡及其机制

Apoptosis of hepatic stellate cells induced by sulfasalazine

目的观察柳氮磺胺吡啶及其分解产物对肝星状细胞株HSC-T6增殖和凋亡的影响,并对其分子机制作初步探讨。方法用CCK-8比色法检测细胞增殖;流式细胞术AnnexinV-FITC/PI双染检测HSC-T6细胞凋亡率;AO/EB染色法观察细胞凋亡形态,Western blot检测NF-κBP65、P-IKK、P-IκB蛋白的表达,激光共聚焦显微镜观察P65核转位。结果柳氮磺胺吡啶能剂量依赖性的抑制HSC-T6肝星状细胞的增殖;AO/EB染色法、AnnexinV/PI染色结果证明柳氮磺胺吡啶能诱导HSC-T6肝星状细胞的凋亡;而5-氨基水杨酸和磺胺吡啶对HSC-T6的增殖和凋亡无影响。Western blot结果表明柳氮磺胺吡啶不是5-氨基水杨酸和磺胺吡啶抑制IKK、IκB的磷酸化、细胞核NF-κBP65的表达(P<0.05),激光共聚焦显微镜下观察到,柳氮磺胺吡啶组P65核转位被抑制。结论柳氮磺胺吡啶可抑制HSC-T6的增殖并诱导凋亡,其机制可能与其抑制Rel/NF-κB/IκB/IKK信号通路有关。

Objective To determine whether sulfasalazine stimulates hepatic stellate cell （HSC-T6） apoptosis and its possible mechanism. Methods CCK-8 assay, acridine orange/ethidium bromide （AO/EB） and Annexin V FITC/PI were used to determine cell growth and cell apoptosis. The expression of NF-κB P65, phospho-IKK and phospho-IKB was detected by Western blotting. The nuclear translocation of HSC-T6 P65 was observed with laser confocal microscopy. Results Sulfasalazine displayed a strong growth inhibition and promoting apoptosis effect on HSC-T6 cells in a dose and time-dependent manner. Sulfasalazine, but not 5-amin- osalicylic acid or sulfapyridine, inhibited the activation of NF-κB by down-regulating the expressions of P-IKK, P-IκB and the nuclear translocation of P65. Conclusion Sulfasalazine can inhibit NF-KB activity and promote apoptosis in HSC-T6 cells, where the Rel/NF-κB/IKB/IKK pathway plays an important role in HSC survival.