中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2014年
5期
1004-1006
,共3页
张莉娟%黄月红%陈治新%陈运新%郑伟达%王小众
張莉娟%黃月紅%陳治新%陳運新%鄭偉達%王小衆
장리연%황월홍%진치신%진운신%정위체%왕소음
白细胞介素-10%肿瘤坏死因子-α%肝星状细胞%信号转导和转录活化因子-1
白細胞介素-10%腫瘤壞死因子-α%肝星狀細胞%信號轉導和轉錄活化因子-1
백세포개소-10%종류배사인자-α%간성상세포%신호전도화전록활화인자-1
Interleukin-10%Tumor necrosis factor-α%Hepatic stellate cell%Signal transducer and activator of transcription-1
目的 观察腺病毒介导的白细胞介素-10(IL-10)基因(Ad-IL-10)转染肝星状细胞(HSC)对肿瘤坏死因子-α(TNF-α)诱导的细胞凋亡及信号转导和转录活化因子-1(STAT-1)表达的影响.方法 以293细胞包装构建重组腺病毒载体系统Ad5-hIL10-IRES-增强型绿色荧光蛋白(EGFP).将体外培养的人HSC分为5组:空白对照组、空载组、TNF-α(10 μg/L)组、空载组+TNF-α(10 μg/L)组、Ad-IL-10+ TNF-α(10 μg/L)组.流式细胞仪检测HSC细胞的凋亡比例;Western blot检测HSC细胞STAT-1及其磷酸化蛋白水平.结果 成功构建Ad5-hIL-10-IRES-EGFP;流式细胞仪检测显示TNF-α诱导使HSC凋亡率显著增加,Ad-IL-10干预可使HSC凋亡率下降(P＜0.05);其凋亡率分别为:空白组0.010±0.001,TNF-α组0.230±0.090,空载组0.210±0.080,IL-10组0.070±0.030.Westernblot检测显示,TNF-α诱导后STAT-1磷酸化蛋白水平显著增加,而Ad-IL-10干预可使两者水平下降(P＜0.01);各组STAT-P/STAT-1吸光度值分别为:空白组0.028±0.006,空载组0.025±0.007,TNF-α组0.261±0.085,空载对照组0.254±0.065,IL-10组0.029±0.009.结论 IL-10能抑制TNF-α诱导HSC细胞发生凋亡,其信号途径与STAT-1有关.
目的 觀察腺病毒介導的白細胞介素-10(IL-10)基因(Ad-IL-10)轉染肝星狀細胞(HSC)對腫瘤壞死因子-α(TNF-α)誘導的細胞凋亡及信號轉導和轉錄活化因子-1(STAT-1)錶達的影響.方法 以293細胞包裝構建重組腺病毒載體繫統Ad5-hIL10-IRES-增彊型綠色熒光蛋白(EGFP).將體外培養的人HSC分為5組:空白對照組、空載組、TNF-α(10 μg/L)組、空載組+TNF-α(10 μg/L)組、Ad-IL-10+ TNF-α(10 μg/L)組.流式細胞儀檢測HSC細胞的凋亡比例;Western blot檢測HSC細胞STAT-1及其燐痠化蛋白水平.結果 成功構建Ad5-hIL-10-IRES-EGFP;流式細胞儀檢測顯示TNF-α誘導使HSC凋亡率顯著增加,Ad-IL-10榦預可使HSC凋亡率下降(P＜0.05);其凋亡率分彆為:空白組0.010±0.001,TNF-α組0.230±0.090,空載組0.210±0.080,IL-10組0.070±0.030.Westernblot檢測顯示,TNF-α誘導後STAT-1燐痠化蛋白水平顯著增加,而Ad-IL-10榦預可使兩者水平下降(P＜0.01);各組STAT-P/STAT-1吸光度值分彆為:空白組0.028±0.006,空載組0.025±0.007,TNF-α組0.261±0.085,空載對照組0.254±0.065,IL-10組0.029±0.009.結論 IL-10能抑製TNF-α誘導HSC細胞髮生凋亡,其信號途徑與STAT-1有關.
목적 관찰선병독개도적백세포개소-10(IL-10)기인(Ad-IL-10)전염간성상세포(HSC)대종류배사인자-α(TNF-α)유도적세포조망급신호전도화전록활화인자-1(STAT-1)표체적영향.방법 이293세포포장구건중조선병독재체계통Ad5-hIL10-IRES-증강형록색형광단백(EGFP).장체외배양적인HSC분위5조:공백대조조、공재조、TNF-α(10 μg/L)조、공재조+TNF-α(10 μg/L)조、Ad-IL-10+ TNF-α(10 μg/L)조.류식세포의검측HSC세포적조망비례;Western blot검측HSC세포STAT-1급기린산화단백수평.결과 성공구건Ad5-hIL-10-IRES-EGFP;류식세포의검측현시TNF-α유도사HSC조망솔현저증가,Ad-IL-10간예가사HSC조망솔하강(P＜0.05);기조망솔분별위:공백조0.010±0.001,TNF-α조0.230±0.090,공재조0.210±0.080,IL-10조0.070±0.030.Westernblot검측현시,TNF-α유도후STAT-1린산화단백수평현저증가,이Ad-IL-10간예가사량자수평하강(P＜0.01);각조STAT-P/STAT-1흡광도치분별위:공백조0.028±0.006,공재조0.025±0.007,TNF-α조0.261±0.085,공재대조조0.254±0.065,IL-10조0.029±0.009.결론 IL-10능억제TNF-α유도HSC세포발생조망,기신호도경여STAT-1유관.
Objective To study the effects of adenovirus-mediated mterleukin-10 (IL-10) gene on apoptosis of hepatic stellate cells (HSCs) induced by tumor necrosis factor-α (TNF-α) and the expres-sion of signal transducer and activator of transcription-1 (STAT-1),and to investigate the relation of IL-10 and apoptosis of HSCs.Methods The recombinant adenovirus vector carrying IL-10 marked enhanced green fluourescent protein [Ad5-hIL-10-IRES-enhanced green fluorescent protein(EGFP)] was constructed.Cultured human HSCs were divided into five groups:control group,empty vector control group,TNF-α group,empty vector + TNF-α group and Ad-IL-10 + TNF-α group.The apoptosis rate of HSCs was assayed by flow cytometry.The levels of STAT-1 and STAT-1 phosphorylation were assayed by Western blotting.Results Ad5-hIL-10-IRES-EGFP was established successfully.Flow cytometry showed that apoptosis rate of HSCs induced by TNF-α was increased significantly,and Ad-IL-10 intervention decreased ap-optosis rate of HSCs (P ＜0.05).The apoptosis rate was as follows:control group 0.010 ±0.001,TNF-αgroup 0.230 ± 0.090,empty vector group 0.210 ± 0.080,Ad-IL-10 + TNF-α group 0.070 ± 0.030,respectively.Western blotting analysis showed that STAT-1 and STAT-1 phosphorylation protein levels were increased significantly by TNF-α,and Ad-IL-10 interventions could reduce the STAT-1 levels (P ＜0.01).The STAT-P/STAT-1 Awas as follows:control group 0.028 ±0.006,empty vector group 0.025 ±0.007,TNF-α group 0.261 ± 0.085,empty vector + TNF-α group 0.254 ± 0.065,Ad-IL-10 + TNF-α group 0.029 ± 0.009,respectively.Conclusion IL-10 could inhibit TNF-α-induced HSCs apoptosis and its signaling pathways involved in STAT-1.
