摘要
目的:探讨泛素蛋白酶系统(ubiquitin-proteasome system,UPS)在体外脂肪细胞分化中的作用。方法:常规方法诱导前脂肪细胞分化为脂肪细胞,Western blot检测蛋白质表达,免疫共沉淀检查蛋白质间结合,油红O染色检测脂肪细胞中的脂质,RT-PCR检测mRNA表达。结果:UPS抑制剂硼替佐米可抑制3T3-L1前脂肪细胞分化为脂肪细胞,表现为细胞内脂质含量降低以及脂肪细胞标志蛋白mRNA表达的降低。蛋白激酶G激动剂西地那非可激活UPS,并增强脂肪细胞的分化。抑制UPS可降低热休克蛋白90(HSP90)和过氧化物酶体增殖物活化受体γ(PPARγ)间的结合;同时降低细胞可溶性组分中HSP90和PPARγ的表达,增强其在不可溶性组分中的表达。HSP90特异性的N末端抑制剂格尔德霉素可抑制西地那非促进的PPARγ表达和脂肪细胞的分化。结论:泛素蛋白酶系统通过HSP90调节PPARγ的表达,从而影响脂肪细胞的分化。
AIM： To investigate the role of ubiquitin-proteasome system （ UPS） in adipocyte differentiation. METHODS：Differentiation of 3T3-L1 preadipocytes into adipocytes was induced by treatment with insulin, 3-isobutyl-1-methylxanthine and dexamethasone.Western blot and immunoprecipitation were performed to detect the protein abundances and association, respectively.Oil red O staining was used to determine the intracellular lipid of 3T3-L1 adipocytes.The levels of mRNA were measured by reverse transcription polymerase chain reaction （ RT-PCR） .RESULTS：UPS inhibitor bortezomib （BZM） suppressed the differentiation of 3T3-L1 pre-adipocytes, evidenced by reduced intracellular content of triglyceride, and decreased mRNA expression of adipogenic marker proteins such as adiponectin and adipocyte protein 2.In contrast, administration of sildenafil （SDN）, an activator of protein kinase G which was also found to activate UPS, promo-ted adipocyte differentiation.In addition, BZM treatment decreased the expression of heat shock protein 90 （HSP90） and peroxisome proliferator-activated receptor gamma （ PPARγ） in the soluble fraction and reduced association of HSP90 and PPARγ.Furthermore, HSP90-specific N-terminal inhibitor geldanamic mitigated SDN-induced increase in PPARγlevel and 3T3-L1 cell differentiation.CONCLUSION：UPS modulates HSP90-dependent PPARγstability, thus leading to pro-motion of adipocyte differentiation.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2015年第5期888-893,共6页
Chinese Journal of Pathophysiology
基金
国家自然科学基金资助项目(No.30770758/H0178)
