| Objective:Obesity, dyslipidemia, fatty liver and other metabolic diseases has become commonly and frequently encountered disease, has seriously impact on human health, but the pathogenesis is not fully understood. low abundance of metabolic inflammation which triggered by nutrient and metabolite is considered the major risk factors for the development of metabolic disease. The class B scavenger receptor CD36, not only can mediate transmembrane transport of long-chain fatty acids, can also identify a number of proinflammatory endogenous metabolites, is closely associated with metabolic inflammation. This study use C57BL/6J mice, wild-type (widetype, WT)/CD36 knockout (CD36 knockout, CD36KO) mice and 3T3-L1 adipocytes, investigating the effect of high-fat diet(HFD) on the expression of CD36 and inflammatory responses in adipose tissue, and furthermore to explore the role of CD36 in the metabolism inflammation of adipose tissue.Methods:C57BL/6J mice were fed with a normal-chow diet (NCD) or a high-fat diet (HFD) for 14 weeks. The contents of free fatty acid (FFA) in the serum were measured by ELISA. The expression of CD36, cytokines and chemokines at mRNA and protein levels in adipose tissues was determined by real-time polymerase chain reaction and Western blotting. Immunohistochemical staining was used to examine the macrophages infiltration in the adipose tissues. And futher, the inflammatory responses in CD36 knockout mice and widetype mice with high-fat diet were analyzed use the same method.3T3-L1 preadipocyte, via differentiation induction into adipocytes, were incubated for 24 hours in medium containing 0.2% BSA (Control, CTL) or 0.2% BSA plus different concentrations of palmitic acid treatment group (Palmitic acid, PA). Lipid droplet formation was visulised using oil red O staining. The expression of CD36, cytokines (IL-1β, IL-6, TNF-α) and chemokines (MCP-1, MIP-1) at mRNA and protein levels in 3T3-L1 adipocytes was determined by real-time polymerase chain reaction and Western blotting. NF-κB activity was detected by dual luciferase reporter. phosphorylated and total protein levels of Jun N-terminal kinase was also detected by Western blotting, extracting lipid rafts and caveolin-1 was measured by western blotting. CD36/TLR4 copolymer formation was investigated using Co-immunoprecipitation.Results:The levels of FAT/CD36 were higher in HFD group than that in NCD group (P<0.05). HFD feeding also enhanced the mRNA and protein expression of IL-1, IL-6, TNF-a, MCP-1 and MIP-1 (P<0.05), as well as promoting macrophage infiltration in the adipose tissues. Interestingly, as fed with HFD, the expression of cytokines/chemokines and macrophage infiltration were significantly reduced in adipose tissues of the CD36 knockout mice, compared with the widetype mice. The results in vitro experiments showed that palmitate increased the mRNA and protein expression of CD36, inflammatory cytokines (IL-1β, IL-6, TNF-α), and chemokines (MCP-1, MIP-1), extracting 3T3-L1 adipocytes lipid rafts, we found that the proportion of CD36 in lipid rafts and total CD36 protein levels increased in PA-treated group (P<0.05). Immunoprecipitation indicated that PA enhanced CD36/TLR4 copolymer formation (P<0.05). Meanwhile, Western Blotting further revealed that the phosphorylation of Jun N-terminal kinase raised in PA-treated group (P<0.05). Dual luciferase reporter showed that the activity of NF-κB was significantly increased (P<0.05).Conclusions:High fat or Palmitic acid can be significantly increases the levels of CD36 in adipose tissue and adipocyte, prompting the formation of CD36/TLR4 copolymer, and then activate JNK/NF-κB inflammatory signaling pathways, involved in the generation of metabolism inflammation. |
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