MicroRNA-29a Alleviate Lipid Deposition Mediated Metabolic Inflammation Via Downregulation Of CD36

【Backgrounds & Aims】 Metabolic inflammation is a chronic low-grade inflammation and one of the core pathogenesis of metabolic disorders in obesity.Macrophages are key to the generation and persistence of metabolic inflammation.Persistent metabolic inflammation is closely associated with the development of diseases such as type 2 Diabetes Mellitus(T2DM),metabolic associated fatty liver disease(MAFLD)and cardiovascular disease.Previous studies have confirmed that MicroRNA-29a(miR-29a)is associated with macrophage proinflammatory polarization,but there are few studies on its association with obesity-related metabolic disorders.In the current study,we explored the effect and mechanism of miR-29 a on obesity and its related metabolic disorders.【Methods】 The expression levels of miR-29 a in liver,muscle and adipose tissues of obese mice induced by high fat diet(HFD)were detected by quantitative real-time PCR(q PCR).miR-29 a overexpression(miR-29a)mice were constructed,and wild-type(WT)and miR-29 a mice were fed with HFD,and the changes of body weight and food intake were recorded.Fasting lipid profile,blood glucose and insulin were detected,and homeostatic model assessment for insulin resistance(HOMA-IR)was calculated,and glucose tolerance test(GTT)and insulin tolerance test(ITT)were performed.Alanine aminotransferase(ALT),aspartate aminotransferase(AST)and lactate dehydrogenase(LDH)in serum of mice were detected to evaluate liver function.The weights of liver and white adipose tissues were recorded,and their indexes were calculated,and the expression levels of miR-29 a were detected.The pathological changes of liver tissues and white adipose tissues were observed by HE and immunohistochemistry staining,and the lipid deposition in liver tissues of mice was observed by oil red staining.The expression of CD36 was detected by q PCR and western blot(WB).WB was used to detect the protein levels of AMP activated protein kinase(AMPK)and phosphorylated AMPK.The protein levels of Toll-like receptor 4(TLR4),myeloid differentiation factor 88(My D88),nuclear factor-κB(NF-κB),p38-mitogen-activated protein kinase(p38),c-Jun N-terminal kinase(JNK),extracellular signa 1/2 regulated kinase(ERK1/2)and nod-like receptor protein 3(NLRP3)signaling pathways in liver tissues were detected.The expression of CD36,fatty acid synthase(FASN)and inflammatory factors in epididymal white adipose tissues were detected by q PCR.Liver metabolic inflammation model induced by methionine and choline deficient diet(MCD)was established,and the changes of body weight of mice were recorded,and the blood lipids,blood glucose,insulin,ALT,AST and LDH of mice were detected.The lipid deposition and inflammation in liver tissues were observed by HE and immunohistochemistry.The expression of CD36 in liver tissues was detected by q PCR and WB,and the activation of AMPK and TLR4 and its downstream My D88,NF-κB,P38,JNK,ERK1/2 and NLRP3 signaling pathways were detected by WB.【Results】 The expression levels of miR-29 a in liver,muscle and interscapular brown adipose tissues of HFD mice were not changed,but the expression levels of miR-29 a in epididymal white adipose tissue and inguinal white adipose tissues were decreased.miR-29a-HFD mice had no change in body weight compared with WT-HFD mice,but their blood glucose was decreased,and HOMA-IR,GTT and ITT were improved;liver weight and lipid deposition were decreased,and liver function and inflammatory cell infiltration were improved;the weight of epididymis white adipose tissues was increased,and inflammatory cell infiltration was decreased.Compared with WT-HFD mice,the expression of CD36 in liver tissues and epididymal white adipose tissue of miR-29a-HFD mice was decreased;the activation of AMPK in liver tissue was increased,and the expression of FASN was decreased,and the TLR4-My D88 pathway was inhibited,and the activation of NF-κB,P38,ERK1/2 signaling pathway and NLRP3 inflammasome were decreased.While the expression of FASN in epididymal white adipose tissues was increased,the expression levels of TLR4,C-C motif chemokine ligand 2(CCL2),tumor necrosis factor alpha(TNF-α)and other inflammatory factors decreased.Compared with WT-MCD mice,miR-29 a overexpression mice with MCD fed(miR-29a-MCD mice)showed no difference in liver lipid deposition,but alleviated liver inflammation and fibrosis.The expression of CD36 was decreased and the activation of AMPK was increased in liver tissues of miR-29a-MCD mice,but the expression of FASN was not changed.miR-29 a overexpression inhibited the activation of p38 and NLRP3 signaling pathways,the downstream of TLR4,in liver tissues of MCD diet mice.【Conclusions】 HFD inhibited the expression of miR-29 a in white adipose tissues of mice.Overexpression of miR-29 a did not change the body weight of HFD mice,but could reduce fasting blood glucose and improve lipid metabolism and insulin resistance,and reduce lipid deposition in liver,and reduce inflammatory cell infiltration in liver and white adipose tissue,and alleviated metabolic inflammation in liver and adipose tissue.Further studies found that these mechanisms may be closely related to alleviated metabolic inflammation mediated by lipid deposition by miR-29 a via downregulation of CD36: the overexpression of miR-29 a inhibited lipid synthesis in liver tissues of HFD mice,but had no effect on MCD mice,and also increased lipid synthesis in epididymal white adipose tissues of HFD mice;the overexpression of miR-29 a inhibited TLR4-My D88 signaling pathway in liver tissues of HFD and MCD mice,reduced the activation of inflammatory signaling pathway of p38 and NLRP3,and alleviated the metabolic inflammation mediated by lipid deposition,and also inhibited the expression of TLR4 and its downstream inflammatory factors in epididymal white adipose tissue of HFD mice,and alleviated the metabolic inflammation in white adipose tissue.