CAJ | 학술논문

目的评价异氟醚对转染APPsw基因SH-SY5Y细胞凋亡的影响和1,4,5-三磷酸肌醇(IP3)受体在其中的作用.方法将转染APPsw基因的SH-SY5Y细胞以1.2×104个/cm2密度接种于培养瓶中,采用随机数字表法,将其分为4组(n=6):对照组(C组)、IP3受体拮抗剂组(Ⅹ组)、异氟醚组(Ⅰ组)和异氟醚+ IP3受体拮抗剂组(Ⅰ+Ⅹ组).继续培养24h细胞贴壁后,C组常规培养；Ⅹ组和Ⅰ+Ⅹ组加入IP3受体拮抗剂Xestospongin C 100 nmol/L; 30 min后Ⅰ组和Ⅰ+Ⅹ组给予1.2％异氟醚处理8h.收集细胞,电镜下观察超微结构,采用流式细胞术检测细胞凋亡率和胞浆内游离钙离子浓度([Ca2+]i),采用Western blot法测定IP3受体蛋白表达.结果与C组比较,Ⅹ组细胞凋亡率、[Ca2+]i和IP3受体蛋白表达差异无统计学意义(P＞0.05),Ⅰ组和Ⅰ+Ⅹ组细胞凋亡率增加,[Ca2+]i升高,IP3受体蛋白表达上调(P ＜0.05或0.01)；与Ⅰ组比较,Ⅰ+Ⅹ组细胞凋亡率和[Ca2+]i降低,IP3受体蛋白表达下调(P＜0.01).Ⅰ组和Ⅰ+Ⅹ组细胞出现病理学损伤,Ⅰ组损伤重于Ⅰ+Ⅹ组.结论异氟醚可通过升高胞浆内游离Ca2+浓度、上调IP3受体蛋白表达,引起转染APPsw基因的SH-SY5Y细胞凋亡.
목적 평개이불미대전염APPsw기인SH-SY5Y세포조망적영향화1,4,5-삼린산기순(IP3)수체재기중적작용.방법 장전염APPsw기인적SH-SY5Y세포이1.2×104개/cm2밀도접충우배양병중,채용수궤수자표법,장기분위4조(n=6):대조조(C조)、IP3수체길항제조(Ⅹ조)、이불미조(Ⅰ조)화이불미+ IP3수체길항제조(Ⅰ+Ⅹ조).계속배양24h세포첩벽후,C조상규배양；Ⅹ조화Ⅰ+Ⅹ조가입IP3수체길항제Xestospongin C 100 nmol/L; 30 min후Ⅰ조화Ⅰ+Ⅹ조급여1.2％이불미처리8h.수집세포,전경하관찰초미결구,채용류식세포술검측세포조망솔화포장내유리개리자농도([Ca2+]i),채용Western blot법측정IP3수체단백표체.결과 여C조비교,Ⅹ조세포조망솔、[Ca2+]i화IP3수체단백표체차이무통계학의의(P＞0.05),Ⅰ조화Ⅰ+Ⅹ조세포조망솔증가,[Ca2+]i승고,IP3수체단백표체상조(P ＜0.05혹0.01)；여Ⅰ조비교,Ⅰ+Ⅹ조세포조망솔화[Ca2+]i강저,IP3수체단백표체하조(P＜0.01).Ⅰ조화Ⅰ+Ⅹ조세포출현병이학손상,Ⅰ조손상중우Ⅰ+Ⅹ조.결론 이불미가통과승고포장내유리Ca2+농도、상조IP3수체단백표체,인기전염APPsw기인적SH-SY5Y세포조망.
Objective To evaluate the effect of isoflurane on the apoptosis of SH-SYSY cells transfected with APPsw gene and the role of inositol 1,4,5-triphosphate (IP3) recepters.Methods The SH-SYSY ceils transfected with APPsw gene were seeded in culture flasks with the density of 1.2 × 104/cm2.The cells were randomly divided into 4 groups (n =6 each):control group (group C),IP3 receptor antagonist group (group Ⅹ),isoflurane group (group Ⅰ) and isoflurane + IP3 receptor antagonist group (group Ⅰ + Ⅹ).After the cells were cultured for 24 h and attached to the wall,the cells were cultured routinely in group C,and Xestospongin C 100 nmol/L (IP3 receptor antagonist) was added to DMEM culture medium in groups X and Ⅰ + X,and 30 min later the cells were exposed to 1.2 ％ sevoflurane for 8 h in groups Ⅰ and Ⅰ + X.The cells were collected for examination of the ultrastructure and for determination of cell apoptosis,intracellular free calcium ion concentration [Ca2 +] i (by flow cytometry) and expression of IP3 receptor protein (by Western blot).The apoptosis rate was calculated.Results Compared with group C,there was no significant change in the apoptosis rate,[Ca2 +]i or IP3 receptor protein expression in group Ⅹ (P ＞ 0.05),while the cell apoptosis rate and [Ca2 +] i were significantly increased and IP3 receptor protein expression was up-regulated in groups I and Ⅰ + Ⅹ (P ＜ 0.05 or 0.01).Compared with group Ⅰ,cell apoptosis rate and [Ca2+]i were significantly decreased and IP3 receptor protein expression was down-regulated in group Ⅰ + Ⅹ (P ＜ 0.01).The pathological changes of the cells happened in groups Ⅰ and Ⅰ + Ⅹ,and the pathological changes were severer in group Ⅰ than in group Ⅰ + Ⅹ.Conclusion Isoflurane can induce apoptosis of SH-SY5Y cells transfected with APPsw gene through increasing [Ca2+]i and up-regulating IP3 receptor protein expression.