Reversine对肝星状细胞凋亡及相关蛋白表达的影响  

Influence of Reversine on Hepatic Stellate Cell Apoptosis and Relevant Protein Expression

作　　者：黄迪[1,2,3] 黄宇 翁杰锋[1,2,3] 黄子圣[1] 麦振豪 张强 张汉意 张帅[1,2,3] 古维立 HUANG Di;HUANG Yu;WENG Jiefeng;HUANG Zisheng;MAI Zhenhao;ZHANG Qiang;ZHANG Hanyi;ZHANG Shuai;GU Weili(Guangzhou First People＇ s Hospital,Guangzhou Medical University,Guangzhou 510180,China)

机构地区：[1]广州医科大学附属广州市第一人民医院,广东广州510180 [2]广州消化疾病中心,广东广州510180 [3]华南理工大学第二附属医院,广东广州510180

出　　处：《现代医院》2018年第8期1181-1184,共4页Modern Hospitals

基　　金：广东省科技计划项目(20118061300024;201313021800057);广东省自然基金项目(10151006001000013);广东省医学科研基金(A2016106);广州市医药卫生科技一般引导项目(20171A011025;20181A011002);广州市中医药科技项目(20172A011004)

摘　　要：目的研究Reversine对肝星状细胞增殖、凋亡及相关蛋白表达的影响,进一步探讨Reversine防治肝纤维化的机制。方法体外培养HSCs细胞,分别给予不同浓度Reversine(5、10、20、40μg/mL)处理24 h后,采用CCK-8法检测Reversine对HSCs细胞增殖抑制率;流式细胞术检测Reversine对HSCs凋亡的影响;细胞免疫荧光法检测TGF-β1、caspase-3、Aurora B、SMAD2/3、Desmin、p16INK4a、SMP-30、p66SHC等蛋白的表达情况。结果不同浓度Reversine(5、10、20、40μg/mL)明显抑制HSCs的增殖(38.03±9.05、44.97±5.85、55.33±2.40、64.19±3.66,F=27.356,P=0.000),且促进HSCs凋亡(13.14±1.87、15.85±1.00、20.57±0.90、21.24±1.07,F=298.75,P=0.000)。荧光显微镜显示Reversine作用HSCs后出现典型的细胞凋亡形态学改变,细胞体积变小,细胞质浓缩,细胞核固缩、碎裂,出现多倍体等,荧光定性检测发现TGF-β_1、caspase-3、Aurora B、SMAD2/3等蛋白有表达,Desmin、p16INK4a、SMP-30、p66SHC等蛋白未见表达。结论 Reversine能抑制肝星状细胞的增殖,诱导其凋亡,并且影响肝星状细胞TGF-β_1、caspase-3、Aurora B、SMAD2/3等蛋白质的表达。Objective This study aimed to determine the influence of reversine on hepatic stellate cell（ HSC） proliferation and apoptosis and relevant protein expression and evaluate its applicability in the treatment of liver fibrosis. Methods HSCs were bred in vitro,and different concentrations of reversine（ 5,10,20,and 40 μg/m L） were used. The CCK-8 method was used to determine the proliferation inhibition rate of reversine on HSCs. Flow cytometry was adopted to observe the effects of reversine on HSC apoptosis. Cell immunofluorescence method was used to detect the expression levels of TGF-β1,caspase-3,Aurora B,SMAD2/3,Desmin,p16 INK4 a,SMP-30,and p66 SHC proteins. Results Different concentrations of reversine（ 5,10,20,and 40 μg/mL） significantly inhibited HSC proliferation（ 38. 03 ± 9. 05,44. 97 ±5. 85,55. 33 ± 2. 40,and 64. 19 ± 3. 66,respectively,F = 27. 356,P = 0. 000） and improved HSC apoptosis（ 13. 14 ±1. 87,15. 85 ± 1. 00,20. 57 ± 0. 90,and 21. 24 ± 1. 07,respectively,F = 298. 75,P = 0. 000）. Fluorescence microscopic images showed that HSCs appear to undergo typical morphological changes of apoptosis after reversine treatment due to the appearance of small cells,cytoplasm concentration,cell nuclear pyknosis and fragmentation,and polyploids. Qualitative fluorescence detection discovered the expression of TGF-β1,caspase-3,Aurora B,and SMAD2/3 proteins but not for Desmin,p16 INK4 a,SMP-30,and p66 SHC. Conclusion Reversine can inhibit HSC proliferation,induce HSC apoptosis,and affect the expression of TGF-β1,caspase-3,Aurora B,collagen-I,and SMAD2/3 proteins in HSC.

关 键 词：Reversine 肝星状细胞 增殖 凋亡 肝纤维化 

分 类 号：R329.25[医药卫生—人体解剖和组织胚胎学]