目的:探讨肝脏中lamp2a对plin5表达及其功能的影响。方法:利用Western blot及免疫组化检测正常人和脂肪肝患者肝脏中lamp2a与plin5表达相关性。利用HepG2细胞构建lamp2a敲减的细胞系HepG2-L2A,通过给予油酸刺激、溶酶体抑制剂处理等进...目的:探讨肝脏中lamp2a对plin5表达及其功能的影响。方法:利用Western blot及免疫组化检测正常人和脂肪肝患者肝脏中lamp2a与plin5表达相关性。利用HepG2细胞构建lamp2a敲减的细胞系HepG2-L2A,通过给予油酸刺激、溶酶体抑制剂处理等进一步在细胞水平证实其相关性。结果:Western-Blot及免疫组化检测显示脂肪肝患者肝组织中lamp2a的表达明显下降,而plin5表达显著升高。Western-Blot试验提示正常培养的HepG2-L2A细胞中plin5表达较HepG2细胞中升高,Bodipy染色提示HepG2-L2A细胞中脂滴数量也明显增加(124±15.3 vs 273±19.1)。在分解试验中,HepG2细胞中plin5明显降低,脂滴数量也明显减少(282±24.1 vs 192±17.5);而HepG2-L2A细胞中plin5仍然处于高水平,脂滴数量仍然较多(325±24.0 vs 286±28.7)。当给予溶酶体抑制剂处理时,HepG2细胞中plin5降解明显抑制,而HepG2-L2A中plin5水平未见明显改变。结论:肝脏中lamp2a的异常表达影响plin5的表达,进而影响了脂质的分解。展开更多
脂滴包被蛋白5(perilipin5,Plin5)在脂肪酸氧化程度较高的组织中表达,虽然在细胞和动物模型中均发现Plin5具有促进脂滴堆积、提高线粒体功能和缓解脂毒性的作用,但Plin5如何发挥其功能特点的具体机制尚未得到分析整合。通过分析相关文...脂滴包被蛋白5(perilipin5,Plin5)在脂肪酸氧化程度较高的组织中表达,虽然在细胞和动物模型中均发现Plin5具有促进脂滴堆积、提高线粒体功能和缓解脂毒性的作用,但Plin5如何发挥其功能特点的具体机制尚未得到分析整合。通过分析相关文献发现:Plin5可以通过和CGI-58(comparative gene identification-58)结合来调节关键脂解酶脂肪甘油三酯水解酶(adiposetiglyceridelipase,ATGL)以调控脂肪的代谢过程;通过核转位与沉默信息调节因子2相关酶1(silent mating type information regulator 2homolog 1,SIRT1)和过氧化物酶体增殖物激活受体γ辅激活因子1α(PPARγcoactivator-1α,PGC-1α)结合促进线粒体功能基因的表达,提高线粒体的氧化能力和呼吸功能;通过增加围脂滴线粒体的含量促使脂滴含量增加并增强线粒体功能,这揭示了Plin5参与调节脂代谢和线粒体功能的具体机制。而耐力运动锻炼会显著增加Plin5的含量,表明Plin5是解释"运动员悖论"现象的关键因子,这为Plin5作为新的2型糖尿病治疗靶点提供了理论依据。展开更多
Objective:To investigate the regulatory mechanism in liver fibrosis progression by nuclear receptor of farnesoid X receptor(FXR)and the lipid droplet-associated protein of perilipin 5(PLIN5).Methods:FXR response eleme...Objective:To investigate the regulatory mechanism in liver fibrosis progression by nuclear receptor of farnesoid X receptor(FXR)and the lipid droplet-associated protein of perilipin 5(PLIN5).Methods:FXR response element(FXRE)upstream of PLIN5 gene was found by bioinformatics,and confirmed by a dual luciferase reporter gene system;a hepatic fibrosis model based on human hepatic stellate cell LX-2 was established by induction of transforming growth factor-β1(TGF-β1);mRNA and protein levels ofα-smooth muscle actin(α-SMA)and collagen栺were measured by qPCR and Western blot after transient overexpression of FXR or PLIN5;Oil red O staining was used to study the formation of lipid droplets.Results:The promoter region of the PLIN5 gene contained a known reverse repeats-1(IR-1);the gene expression of PLIN5 in LX-2 cells was up-regulated after FXR activation(P<0.01);overexpression of PLIN5 promoted the formation of lipid droplets and significantly reduced the TGF-β1 induced fibrosis gene expression(P<0.05);FXR activation showed no effects on the inhibition of LX-2 cells activation.Conclusion:Overexpression of PLIN5 promotes the formation of lipid droplets and inhibits activation of LX-2 cells.FXR might bind to the FXRE site upstream of PLIN5 gene and regulate its gene expression.In summary,FXR may prevent liver fibrosis progression partially by regulating lipid droplet-associated protein of PLIN5.展开更多
文摘目的:探讨肝脏中lamp2a对plin5表达及其功能的影响。方法:利用Western blot及免疫组化检测正常人和脂肪肝患者肝脏中lamp2a与plin5表达相关性。利用HepG2细胞构建lamp2a敲减的细胞系HepG2-L2A,通过给予油酸刺激、溶酶体抑制剂处理等进一步在细胞水平证实其相关性。结果:Western-Blot及免疫组化检测显示脂肪肝患者肝组织中lamp2a的表达明显下降,而plin5表达显著升高。Western-Blot试验提示正常培养的HepG2-L2A细胞中plin5表达较HepG2细胞中升高,Bodipy染色提示HepG2-L2A细胞中脂滴数量也明显增加(124±15.3 vs 273±19.1)。在分解试验中,HepG2细胞中plin5明显降低,脂滴数量也明显减少(282±24.1 vs 192±17.5);而HepG2-L2A细胞中plin5仍然处于高水平,脂滴数量仍然较多(325±24.0 vs 286±28.7)。当给予溶酶体抑制剂处理时,HepG2细胞中plin5降解明显抑制,而HepG2-L2A中plin5水平未见明显改变。结论:肝脏中lamp2a的异常表达影响plin5的表达,进而影响了脂质的分解。
文摘脂滴包被蛋白5(perilipin5,Plin5)在脂肪酸氧化程度较高的组织中表达,虽然在细胞和动物模型中均发现Plin5具有促进脂滴堆积、提高线粒体功能和缓解脂毒性的作用,但Plin5如何发挥其功能特点的具体机制尚未得到分析整合。通过分析相关文献发现:Plin5可以通过和CGI-58(comparative gene identification-58)结合来调节关键脂解酶脂肪甘油三酯水解酶(adiposetiglyceridelipase,ATGL)以调控脂肪的代谢过程;通过核转位与沉默信息调节因子2相关酶1(silent mating type information regulator 2homolog 1,SIRT1)和过氧化物酶体增殖物激活受体γ辅激活因子1α(PPARγcoactivator-1α,PGC-1α)结合促进线粒体功能基因的表达,提高线粒体的氧化能力和呼吸功能;通过增加围脂滴线粒体的含量促使脂滴含量增加并增强线粒体功能,这揭示了Plin5参与调节脂代谢和线粒体功能的具体机制。而耐力运动锻炼会显著增加Plin5的含量,表明Plin5是解释"运动员悖论"现象的关键因子,这为Plin5作为新的2型糖尿病治疗靶点提供了理论依据。
基金National Natural Science Foundation of China(81973376)。
文摘Objective:To investigate the regulatory mechanism in liver fibrosis progression by nuclear receptor of farnesoid X receptor(FXR)and the lipid droplet-associated protein of perilipin 5(PLIN5).Methods:FXR response element(FXRE)upstream of PLIN5 gene was found by bioinformatics,and confirmed by a dual luciferase reporter gene system;a hepatic fibrosis model based on human hepatic stellate cell LX-2 was established by induction of transforming growth factor-β1(TGF-β1);mRNA and protein levels ofα-smooth muscle actin(α-SMA)and collagen栺were measured by qPCR and Western blot after transient overexpression of FXR or PLIN5;Oil red O staining was used to study the formation of lipid droplets.Results:The promoter region of the PLIN5 gene contained a known reverse repeats-1(IR-1);the gene expression of PLIN5 in LX-2 cells was up-regulated after FXR activation(P<0.01);overexpression of PLIN5 promoted the formation of lipid droplets and significantly reduced the TGF-β1 induced fibrosis gene expression(P<0.05);FXR activation showed no effects on the inhibition of LX-2 cells activation.Conclusion:Overexpression of PLIN5 promotes the formation of lipid droplets and inhibits activation of LX-2 cells.FXR might bind to the FXRE site upstream of PLIN5 gene and regulate its gene expression.In summary,FXR may prevent liver fibrosis progression partially by regulating lipid droplet-associated protein of PLIN5.
