中波段紫外线对角质形成细胞凋亡的诱导和钙信号的影响

目的　观察中波段紫外线 (UVB)照射诱导人永生化角质形成细胞 (HaCaT细胞 )的凋亡和对细胞静息Ca2 + 及thapsigargin和adrenaline引起的Ca2 + 运动影响。方法　采用四甲基偶氮唑盐 (MTT)染色法分析受UVB照射后细胞存活率的变化 ,用流式细胞术、琼脂糖凝胶电泳和Hoechst332 5 8核染色法观察UVB对HaCaT细胞凋亡的影响 ,采用Fura 2 /AM荧光分光光度法测定胞浆游离Ca2 + 浓度的变化。结果　在 1 2 5～ 6 2 5J·m-2 的UVB照射剂量之间和照射后培养时间 2 4h内的条件下 ,UVB照射以剂量和时间依赖的方式抑制细胞的存活率 ;在照射后培养时间 1 8h内以剂量依赖的方式诱导细胞凋亡 ;2 5 0J·m-2 UVB照射后 ,细胞的静息钙迅速下降 ,在照射后第 6h静息钙降至最低 ,且thapsigargin(1 0μmol·L-1 )和adrenaline(1 0 μmol·L-1 )所引起的钙释放及由此产生的钙内流也随照射后培养时间的延长而逐渐减少。结论　UVB照射可诱导HaCaT细胞的凋亡及引起细胞钙稳态的失衡。

Effect of UVB-irradiation on induction of apoptosis and changes of Ca2+ signal in human keratinocyte HaCaT cells

AIM To investigate the effect of UVB irradiation on the induction of apoptosis, the changes of basal Ca 2+ and mobilization induced by thapsigargin and adrenaline in human keratinocyte HaCaT cells. METHODS MTT assay was performed to determine cell viability. Apoptosis was evaluated by Hoechst33258 staining, agarose gel electrophoresis and flow cytometry. [Ca 2+ ] i was measured using Fura 2/AM spectrophoto fluometry. RESULTS Within 24 h incubation time, 125 to 625 J·m -2 UVB irradiation caused a gradual decrease in cell viability in a dose and time dependent manner. Within 18 h incubation time,125 to 625 J·m -2 UVB irradiation resulted in apoptotic cell death in a dose dependent manner. The basal [Ca 2+ ] i decreased immediately after 250 J·m -2 UVB irradiation. Thapsigargin and adrenaline induced Ca 2+ release and influx also decreased in a time dependent manner. CONCLUSION UVB irradiation could induce apoptotic cell death and interrupt the homeostasis of Ca 2+ in human keratinocyte HaCaT cells.