Insulin-Attenuated Inflammatory Response of Dendritic Cells in Diabetes by Regulating RAGE-PKCβ1-IRS1-NF-κB Signal Pathway: A Study on the Anti-Inflammatory Mechanism of Insulin in Diabetes

Figure 6

The role of IR and IRS1 in AGEs-BSA-stimulated BMDCs immune maturation. (a) Flow cytometric analysis was performed for cell surface CD83, CD86, and MHC-II expression examinations. IRS1 siRNA attenuated AGEs-BSA-induced CD83 and CD86 upregulation; (b) the concentrations of TNF-α, IFN-γ, IL-6, IL-1b, and CCL2 in the supernatants of the culture were measured by ELISA; (c, d) the expressions of cytokines and chemokines mRNA in BMDCs were analyzed by real-time quantitative RT-PCR. IRS1 siRNA attenuated AGEs-BSA-induced the expressions of cytokines and chemokines and upregulated the expression of IL-10; (e) the expressions of NF-κB and IκB were determined by western blot with the samples treated with IRS1 specific siRNA or scramble siRNA. The phosphorylation of NF-κB and IκB was downregulated after IRS1 silencing; (f) the expressions of NF-κB and IκB were determined by western blot in the presence of IR neutralizing antibody or insulin or IRS1 siRNA. The results showed that the IR-neutralizing antibody had no impact on the suppressing effect of insulin. Insulin treatment and IRS1 silencing showed no significant impact on the suppression of phosphorylation of NF-κB and IκB. Data are expressed as ; ; vs. control group; ^# vs. AGEs-BSA group; ^Δ vs.. AGEs-BSA: advanced glycosylation end-bovine serum albumin; BMDCs: bone marrow-derived dendritic cells; IRS1: insulin receptor subunit-1; IR: insulin receptor; TNF-α: tumor necrosis factor-α; IFN-γ: interferon-γ; MHC-II: major histocompatibility complex-II; NF-κB: nuclear factor-κB.

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