Zhang Liting, Xie Zehui, Yu Hongmiao, Du Haoxuan, Wang Xuqiao, Cai Jiazheng, Qiu Yingfei, Chen Rui, Jiang Xiaofeng, Liu Zelin, Li Yi, Chen Tuo
State Key Laboratory of Cryospheric Science, Northwest Institute of Eco-Environment and Resources, Chinese Academy of Sciences, Lanzhou, China.
University of Chinese Academy of Sciences, Beijing, China.
Ann Transl Med. 2021 Sep;9(18):1429. doi: 10.21037/atm-21-4012.
Gut microbiome dysbiosis is related to the pathogenesis of nonalcoholic fatty liver disease (NAFLD), and the role of toll-like receptor 2 (TLR2) in its molecular mechanism is controversial. Here, we investigated the effects and mechanisms of -derived lipopolysaccharide (LPS) on lipid accumulation and lipotoxicity in palmitic acid (PA)-treated L02 cell as an NAFLD cell model, and the role of TLR2 in this process.
Oil red O staining assay and free fatty acid (FFA) content test were performed to determine the effects of LPS on lipid accumulation in a PA-induced NAFLD cell model with or without TLR2 inhibition. The levels of IL-6 and TNF-α were measured to investigate inflammation conditions. Hoechst 33342 staining assay and Caspase-3 activity assay were used to test cell apoptosis, and the expression levels of proteins in the IRS1/PI3K/AKT signaling pathway, TLR2/MyD88/IKKα/NF-κB signaling pathway, and mitochondrion-dependent apoptotic signaling pathway were detected using Western blot.
Lipid accumulation, pro-inflammatory cytokine release, and cell apoptosis with high levels were observed in the PA-induced NAFLD cell model, and LPS aggravated these processes. Whereas TLR2 inhibition could significantly ameliorate PA-induced and LPS-amplified lipid accumulation, inflammatory, and cell apoptosis, it had no significant effect on L02 cells treated with LPS alone.
These results were confirmed by activation or inhibition of the IRS1/PI3K/AKT signaling pathway, TLR2/MyD88/IKKα/NF-κB signaling pathway, and mitochondrion-dependent apoptotic signaling pathway, and were reflected by changes on their proteins expression. TLR2 is involved in PA-induced lipid accumulation and lipotoxicity in L02 cells, which could be aggravated by LPS, although LPS-induced amplification might not be through direct interaction with TLR2.
肠道微生物群失调与非酒精性脂肪性肝病（NAFLD）的发病机制有关，而Toll样受体2（TLR2）在其分子机制中的作用存在争议。在此，我们研究了细菌衍生的脂多糖（LPS）对棕榈酸（PA）处理的L02细胞（作为NAFLD细胞模型）中脂质积累和脂毒性的影响及机制，以及TLR2在此过程中的作用。
进行油红O染色试验和游离脂肪酸（FFA）含量检测，以确定LPS对有或无TLR2抑制的PA诱导的NAFLD细胞模型中脂质积累的影响。检测白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）水平以研究炎症情况。采用Hoechst 33342染色试验和半胱天冬酶-3（Caspase-3）活性检测来检测细胞凋亡，并使用蛋白质印迹法检测胰岛素受体底物1（IRS1）/磷脂酰肌醇-3激酶（PI3K）/蛋白激酶B（AKT）信号通路、TLR2/髓样分化因子88（MyD88）/核因子κB抑制蛋白激酶α（IKKα）/核因子κB（NF-κB）信号通路以及线粒体依赖性凋亡信号通路中蛋白质的表达水平。
在PA诱导的NAFLD细胞模型中观察到脂质积累、促炎细胞因子释放以及高水平的细胞凋亡，且LPS加剧了这些过程。而TLR2抑制可显著改善PA诱导的以及LPS增强的脂质积累、炎症反应和细胞凋亡，对单独用LPS处理的L02细胞无显著影响。
这些结果通过激活或抑制IRS1/PI3K/AKT信号通路、TLR2/MyD88/IKKα/NF-κB信号通路以及线粒体依赖性凋亡信号通路得到证实，并通过其蛋白质表达的变化得以体现。TLR2参与了PA诱导的L02细胞脂质积累和脂毒性，LPS可使其加重，尽管LPS诱导的增强作用可能不是通过与TLR2直接相互作用实现的。
