CYP2J2 And Its Metabolites EETs Attenuate Insulin Resistance Via Regulation Of Adipose Tissue Macrophage Polarization

Background and ObjectiveInsulin resistance (IR) is the condition in which a cell, tissue, or organism fails to respond appropriately to a given dose of insulin. Obesity is the most common cause of insulin resistance, and obesity induced low-grade chronic inflammation in adipose tissue play an important role in the development of IR. Recent studies have found that macrophages infiltration and polarization in adipose tissue are closely related with the chronic inflammation and insulin resistance in mice. Regulation of adipose tissue macrophages infiltration and polarization can provide new targets for clinical prevention and treatment of insulin resistance.Cytochrome P450 epoxygenase metabolize arachidonic acid (AA) to four different epoxyeicosatrienoic acids (5,6-EET,8,9-EET,11,12-EET and 14,15-EET). Soluble epoxide hydrolase (sEH) hydrolyzes EETs to dihydroxyeicosatrienoic acids (DHET), which exhibits a weaker biologically activity. CYP450-EETs-sEH system plays an important role in the regulation of various cellular and physiologic processes in various organs and tissues, especially in cardiovascular and renal system.Preliminary studies demonstrated that endogenous epoxygenases are immuno-modulators regulating monocyte/macrophage activation depending on the underlying activation state, regulate macrophages polarization and prevent LPS-induced cardiac dysfunction. Macrophages act as the main important part of the immune system, has the characteristics of morphological plasticity and function diversity, and play an important role in the development of inflammation process. Obesity induced by high fat diet (HFD) showed a continuous state of chronic low-grade inflammation, adipose tissue macrophages accumulation promoted inflammation progress. Whether CYP2J2-EETs-sEH-DHET metabolic pathway regulate macrophage diversity and function to attenuate inflammation, improve insulin sensitivity and the related mechanism need further research. In this study, we tested the hypothesis that CYP2J2-EETs-sEH metabolic pathways prevent obesity-induced insulin resistance by regulating adipose tissue macrophages infiltration and polarization.Methods and ResultsFirstly, insulin resistance mice model was established by high fat diet (60 Kcal% Fat) for 12 weeks. Continuous infusion of EETs (11,12-EET or 14,15-EET) by application of mini-pump for further 4 weeks and observed the effects of EETs on insulin resistance. Results demonstrated that EETs directly attenuated HFD induced obesity; reduced the levels of serum glucose, insulin, triglyceride and cholesterol; decreased the secretion of pro-inflammatory cytokines (IL-1?, IL-6 and MCP-1) and significantly improved the glucose tolerance and insulin sensitivity. EETs also significantly reduced the adipose tissue mass, attenuated HFD-induced MAPK and NF-?B inflammatory signaling pathways and improved the insulin downstream P-AKT expression. Furthermore, over-expression of CYP2J2 or inhibited the activity of sEH by TUPS significantly increased the level of EETs indirectly. We further explore the mechanisms of EETs on insulin resistance. Results showed that CYP2J2 or TUPS significantly elevated EETs level; decreased body weight gain; improved the metabolism of glucose and lipid; improved the glucose tolerance and insulin sensitivity; decreased the levels of serum inflammatory cytokines and pro-inflammatory monocytes in circulation. The mechanism of which were related with regulating the balance of M1/M2 macrophages polarization and the related membrane molecules expression, and finally attenuated the HFD induced inflammation and insulin resistance in adipose tissue. Finally, our in vitro experiments aimed to explore the effects of exogenous EETs on FFA-induced macrophages polarization and the potential mechanism. Both LPS and FFA significantly induced macrophage migration, and the adipocytes-derived condition medium further increased the ability of macrophage migration.11,12-EET and TUPS inhibited these effects. In addition,11,12-EET significantly inhibited the M1 macrophage polarization induced by LPS or FFA, and promoted IL-4 induced M2 polarization. Microarray analysis of gene expression profiles of bone-morrow derived macrophages (BMDM) and the verification results by real-time PCR and western blot indicated cAMP-EPAC signaling pathway may play important role in macrophage polarization. Using EPAC specific cAMP agonist revealed that increased the level of EPAC by 8-CPT-2'-O-Me-cAMP mimiced the effect of FFA on the regulation of MAPK and NF-?B signaling pathway, EETs inhibited the expression of EPAC, thus attenuated the downstream inflammatory signaling pathway.Conclusion1. EETs significantly attenuated HFD induced obesity and insulin resistance.2. EETs prevented HFD-induced insulin resistance by regulating adipose tissue macrophage accumulation and polarization and subsequently attenuated adipose tissue and systemic inflammation.3. EETs regulated macrophage polarization and inflammation may be through the cAMP-EPAC signaling pathway.