氯化钆诱导大鼠腹腔巨噬细胞内质网应激和凋亡的研究

Study of endoplasmic reticulum stress and apoptosis of peritoneal macrophage in ratinduced by Gadolinium chloride

目的探讨氯化钆对大鼠腹腔巨噬细胞内质网应激和凋亡的作用。方法分离和培养大鼠腹腔巨噬细胞(PMΦ),分别经氯化钆(Gd)和衣霉素(Tunicamycin)处理后,采用细胞免疫组化S-P法检测ARMET的表达,免疫荧光双标染色检测ARMET和CHOP的表达,PI染色观察对细胞凋亡的影响。结果氯化钆处理组细胞数目减少,胞体变小变圆;ARMET在氯化钆组(5×10-6mol/L)的阳性细胞百分数为(42.1±6.6)%,在正常对照组的阳性率为(6.1±1.6)%,差异有统计学意义(P<0.05);而其在阳性对照Tunicamycin组的阳性率为(43.4±5.9)%,两者差异无统计学意义(P>0.05);免疫荧光双标染色提示氯化钆组ARMET和CHOP均高表达;PI染色可见氯化钆组细胞被深染、胞核断裂等凋亡特征。结论氯化钆可诱导大鼠腹腔巨噬细胞内质网应激和凋亡,其作用机制可能与胞内钙超载引起的非折叠蛋白反应有关。

Objective To investigate the effect of endoplasmic reticulum stress and apoptosis of peritoneal macrophage in rat induced by Gadolinium chloride.Methods Rat peritoneal macrophage（PMΦ）were isolated and cultured.After PMΦ treated with Gadolinium chloride and Tunicamycin,the expression of ARMET was determined by immunohistochemical method,immunoflurorescent double staining were employed to detect the expression of CHOP and ARMET,PI stain was used to observe PMΦs apoptosis.Results The amount of cells were greatly reduced and the cell body were shrinked to round in GdCl3 group.The percentage of cell ARMET positive-expressed were（42.1±6.6）% in GdCl3 group（5×10-6 mol/L）,while the value of control group were（6.1±1.6）%,the difference between the two groups was significant（P0.01）.But compared with the value of GdCl3 group were（43.4±5.9）%,there was no significant statistical difference in ARMET expression.The expression of ARMET and CHOP were markedly high in GdCl3 group by immunofluorescence double labeled stain.PI stain revealed the apoptosis feature that cells were deeply red-stained and nuclear fragment.Conclusion GdCl3 can induce endoplasmic reticulum stress and apoptosis of peritoneal macrophage in rat,the potential meachnism is related to endoplasmic reticulum Ca2＋ efflux producing unfolded protein response（UPR）.