CAJ | 학술논문

目的评价七氟醚和异氟醚对大鼠大脑皮质神经干细胞增殖的影响.方法取孕15 d的SD大鼠,分离皮层进行神经干细胞原代培养,以(1～2)×106个/ml的细胞密度稳定传代到第3代后,接种至多聚赖氨酸包被的6孔培养板中.采用随机数字表法,将其分为3组(n=24):对照组(C组)、七氟醚组(S组)和异氟醚组(Ⅰ组).将培养板细胞置于暴露箱,S组通入4.9％七氟醚和5％CO2-95％O2的混合气体;Ⅰ组通入2.8％异氟醚和5％ CO2-95％O2的混合气体;C组通人5％CO2-95％O2的混合气体.暴露6h后,取出培养板并于37℃细胞培养箱中继续培养2h.采用免疫细胞化学法和酶标法测定细胞增殖情况,采用实时荧光定量PCR法检测增殖相关基因:信号转导和转录激活子3(STAT3)、Sox2、Notch1和P21的mRNA表达水平.采用Western blot法测定总STAT3蛋白和磷酸化STAT3(p-STAT3)蛋白的表达水平.结果与C组比较,S组和Ⅰ组神经干细胞增殖率降低,p-STAT3蛋白表达下调,Ⅰ组STAT3 mRNA表达下调(P＜0.05),S组STAT3 mRNA表达差异无统计学意义(P＞0.05).3组间Sox2 mRNA、Notch1 mRNA、P21 mRNA及总STAT3蛋白表达差异无统计学意义(P＞0.05).结论异氟醚和七氟醚均可抑制大鼠大脑皮质神经干细胞的增殖,其机制可能为七氟醚与抑制STAT3蛋白激活有关,而异氟醚不仅抑制了STAT3蛋白的激活,而且抑制了STAT3基因的转录.
목적 평개칠불미화이불미대대서대뇌피질신경간세포증식적영향.방법 취잉15 d적SD대서,분리피층진행신경간세포원대배양,이(1～2)×106개/ml적세포밀도은정전대도제3대후,접충지다취뢰안산포피적6공배양판중.채용수궤수자표법,장기분위3조(n=24):대조조(C조)、칠불미조(S조)화이불미조(Ⅰ조).장배양판세포치우폭로상,S조통입4.9％칠불미화5％CO2-95％O2적혼합기체;Ⅰ조통입2.8％이불미화5％ CO2-95％O2적혼합기체;C조통인5％CO2-95％O2적혼합기체.폭로6h후,취출배양판병우37℃세포배양상중계속배양2h.채용면역세포화학법화매표법측정세포증식정황,채용실시형광정량PCR법검측증식상관기인:신호전도화전록격활자3(STAT3)、Sox2、Notch1화P21적mRNA표체수평.채용Western blot법측정총STAT3단백화린산화STAT3(p-STAT3)단백적표체수평.결과 여C조비교,S조화Ⅰ조신경간세포증식솔강저,p-STAT3단백표체하조,Ⅰ조STAT3 mRNA표체하조(P＜0.05),S조STAT3 mRNA표체차이무통계학의의(P＞0.05).3조간Sox2 mRNA、Notch1 mRNA、P21 mRNA급총STAT3단백표체차이무통계학의의(P＞0.05).결론 이불미화칠불미균가억제대서대뇌피질신경간세포적증식,기궤제가능위칠불미여억제STAT3단백격활유관,이이불미불부억제료STAT3단백적격활,이차억제료STAT3기인적전록.
Objective To evaluate the effects of sevoflurane and isoflurane on the proliferation of neural stem cells in rat cerebral cortex.Methods The neural stem cells were isolated from Sprague-Dawley rats at 15 days of gestation and cultured at a density of (1-2) × 106 cells/ml.The cells of 3rd generation were seeded in 6-well plates coated with poly-lysine and randomly divided into 3 groups (n =24 wells each) using a random number table:control group (group C),sevoflurane group (group S) and isoflurane group (group Ⅰ).In S and Ⅰ groups,the cells were exposed to 4.9％ sevoflurane and 2.8％ isoflurane in a mixture of 5％ CO2-95％ O2 for 6 h,respectively.The cells were exposed to a mixture of 5 ％ CO2-95 ％ O2 for 6 h in group C.After 6 h of exposure,the plates were removed and the cells were continuously incubated for 2 h in an incubator at 37 ℃.The proliferation of cells was detected by immunocytochemistry and microplate method.The expression of proliferation-related genes such as signal transducers and activators of transcription 3 (STAT3),Sox2,Notchl and P21 mRNA was detected using quantitative real-time PCR.The expression of total STAT3 protein and phosphorylated STAT3 protein (p-STAT3) was determined using Western blot.Results Compared with C group,the rate of proliferation was significantly decreased,and the expression of p-STAT3 was down-regulated in I and S groups,and the expression of STAT3 mRNA was down-regulated in Ⅰ group (P ＜ 0.05),and no significant change was found in the expression of STAT3 mRNA in S group (P ＞ 0.05).There was no significant difference in the expression of Sox2,Notch1 and P21 mRNA and total STAT3 protein between the three groups (P ＞ 0.05).Conclusion Sevoflurane and isoflurane both can inhibit the proliferation of neural stem cells in the rat cerebral cortex,and the mechanism may be that sevoflurane inhibits activation of STAT3 protein,however,isoflurane not only inhibits the activation of STAT3 protein,but also inhibit the transcription of STAT3 gene.