CAJ | 학술논문

目的：观察自噬抑制剂3-甲基腺嘌呤（3-MA）对乙醇诱导的肝星状细胞（HSC）活化作用，并探讨其作用机制。方法体外培养大鼠肝星状细胞株HSC-T6，设立空白对照组、阳性对照组（乙醇刺激组）、低剂量组（5 mmol/L 3-MA+100 mmol/L乙醇）、高剂量组（10 mmol/L 3-MA+100 mmol/L乙醇）。RT-PCR分析各组HSC活化标志蛋白α-平滑肌肌动蛋白(α-SMA)及Ⅰ型胶原基因表达，Western blot法检测各组HSC中自噬水平标志蛋白LC3Ⅱ、α-SMA、Ⅰ型胶原表达；MTT法检测3-MA对乙醇诱导的HSC增殖的影响。结果与空白对照组比较，阳性对照组中α-SMA、Ⅰ型胶原mRNA和蛋白表达及LC3Ⅱ表达量明显增加（P＜0.05），高剂量组却显著减少（P＜0.01）；与阳性对照组比较，低剂量组和高剂量组中α-SMA、Ⅰ型胶原mRNA和蛋白表达及LC3Ⅱ表达量逐渐减少（P＜0.05）；与低剂量组比较，高剂量组中α-SMA、Ⅰ型胶原mRNA和蛋白表达及LC3Ⅱ表达减少（P＜0.05）。与阳性对照组比较，加入3-MA处理后的HSC增殖显著减少（P＜0.05）。结论3-MA可抑制乙醇诱导的HSC-T6细胞中LC3Ⅱ蛋白的表达、α-SMA、Ⅰ型胶原mRNA及蛋白的表达，抑制HSC增殖，且高剂量的作用更明显。
목적：관찰자서억제제3-갑기선표령（3-MA）대을순유도적간성상세포（HSC）활화작용，병탐토기작용궤제。방법체외배양대서간성상세포주HSC-T6，설립공백대조조、양성대조조（을순자격조）、저제량조（5 mmol/L 3-MA+100 mmol/L을순）、고제량조（10 mmol/L 3-MA+100 mmol/L을순）。RT-PCR분석각조HSC활화표지단백α-평활기기동단백(α-SMA)급Ⅰ형효원기인표체，Western blot법검측각조HSC중자서수평표지단백LC3Ⅱ、α-SMA、Ⅰ형효원표체；MTT법검측3-MA대을순유도적HSC증식적영향。결과여공백대조조비교，양성대조조중α-SMA、Ⅰ형효원mRNA화단백표체급LC3Ⅱ표체량명현증가（P＜0.05），고제량조각현저감소（P＜0.01）；여양성대조조비교，저제량조화고제량조중α-SMA、Ⅰ형효원mRNA화단백표체급LC3Ⅱ표체량축점감소（P＜0.05）；여저제량조비교，고제량조중α-SMA、Ⅰ형효원mRNA화단백표체급LC3Ⅱ표체감소（P＜0.05）。여양성대조조비교，가입3-MA처리후적HSC증식현저감소（P＜0.05）。결론3-MA가억제을순유도적HSC-T6세포중LC3Ⅱ단백적표체、α-SMA、Ⅰ형효원mRNA급단백적표체，억제HSC증식，차고제량적작용경명현。
Objective To observe the effect of autophagy inhibitor on the activation of alcohol induced hepatic stel-late cells, and the mechanisms thereof. Methods HSC-T6 cells were cultured in vitro and divided into four groups, includ-ing blank control group, alcohol group, 5 mmol/L 3-MA+alcohol group (low alcohol group) and 10 mmol/L 3-MA+alcohol group (high alcohol group). RT-PCR was used to detect the expression levels ofα-smooth muscle actin (α-SMA) and typeⅠcollagen. The levels of LC3Ⅱ,α-SMA and typeⅠcollagen were detected by Western blot assay. The cell viability of HSC-T6 was detected by MTT assay. Results The mRNA expressions ofα-SMA, typeⅠcollagen and the protein of expressionsα-SMA, typeⅠcollagen and LC3Ⅱwere significantly up-regulated in alcohol group compared with those of control group (P<0.05), while the expressions of those parameters were significantly down-regulated in 10 mmol/L 3-MA+alcohol group (P<0.01). The mRNA and protein levels ofα-SMA and typeⅠcollagen were significantly decreased in two 3-MA-treated groups compared with those in alcohol group (P<0.05). Meanwhile, compared with the 5 mmol/L 3-MA+alcohol group,the protein expressions ofα-SMA, typeⅠcollagen and LC3Ⅱwere significantly decreased in10 mmol/L 3-MA+alcohol group (P < 0.05 ). Compared with the alcohol group,there was significantly lower proliferation activity in all two 3-MA-treated groups (P<0.05). Conclusion 3-MA can inhibit the protein expression of LC3Ⅱ,α-SMA and typeⅠcollagen induced by alcohol in HSC-T6 cells, and inhibit the proliferation of HSC cells.