ИСТИНА

Background: Latent inflammation in adipose tissue promoting systemic inflammation, insulin resistance and cardiovascular risk is the main hazard of metabolic syndrome. Insulin signal transduction induces phosphorylation of Akt kinase which in turn results in a spectrum of physiological effects including translocation of GLUT4 to cell membrane and glucose utilization. This process is downregulated in inflammation, suggesting the involvement of paracrine factors secreted by leukocytes. However, mechanisms participating in this modulation are yet not studied definitely. Purpose: We studied the role of Akt kinase regulation by polarized macrophages (Mphs) and free fatty acids (FFA) in development of insulin resistance in 3T3L1 adipocytes. Methods: RAW 264.7 macrophages (Mphs) were polarized into M1 and M2 types by LPS/INF-γ and by IL-4, respectively. Mph M1/M2 polarization was characterized by real time PCR, ELISA and by estimating reactive oxygen species (ROS) production. Insulin resistance in 3T3L1 adipocytes was evaluated as a decrease of membrane GLUT4 content measured by flow cytometry after short-term cell treatment by insulin. Akt phosphorylation on Thr-308/Ser-473 was estimated by Western-blotting. Results: M1 Mphs expressed predominantly inflammatory markers (TNF-α, CXCL9, CXCL11), with abundant generation of ROS. On a contrary, M2 Mphs produced predominantly anti-inflammatory markers (ALOX-15, IGF-1, MCP-4) and had lesser ROS generation than M1. GLUT4 exposure on 3T3L1 adipocyte membrane was decreased after treatment by FFA, M1-cell conditioned media (CM) and M2-cell CM, and was significantly affected by M0-cell CM. In proinflammatory conditions, pAkt-Thr308 decreased while pAkt-Ser473 increased, in contrast to normal cells. We have found that M1-cell CM exert a decrease of pAkt-Thr308 while increasing pAkt-Ser473 in 3T3L1 adipocytes; these effects were not observed if M2 cell CM was used for adipocyte treatment. On the contrary, M2 Mph CM induced the increase of pAkt-Thr308 and decrease in pAkt-Ser473. The decline of Thr-308 phosphorylation in inflammatory conditions may be due to regulatory inhibitory Ser/Thr phosphorylation of IRS-1 by IKK. Conclusion: We have demonstrated the possibility of adipocyte insulin sensitivity regulation via modulation of inflammatory status of macrophages. The proposed mechanism of pAkt-Ser473 increase in inflammatory conditions may be associated with IKK-dependent activation of mTORC2, the possibility of which is shown in some studies. Thus, possible mechanism mediating the influence of M1/M2-polarized Mphs on insulin-dependent Akt-phosphorylation may be associated with divergent regulation of IKK-kinase in target cells. It may be realized in latent inflammation, insulin resistance and metabolic syndrome.