The chronic systemic inflammation in type I diabetes mellitus (T1DM), which is driven by signaling through the interleukin-1 (IL-1) 1 receptor (IL1R) and the adaptor protein myeloid differentiation factor 88 (MyD88), may be associated with the enhanced susceptibility of diabetics to systemic bacterial infection (sepsis). We hypothesized that low insulin concentrations trigger the enzyme 5-lipoxygenase (5-LO) to produce the lipid mediator leukotriene B4 (LTB4), serving as a trigger of systemic inflammation and increased susceptibility to polymicrobial sepsis in T1DM. In support of this hypothesis, we found that the abundance of MyD88 and its direct transcriptional regulator, STAT-1 were higher in peritoneal macrophages from two mouse models of T1DM compared to nondiabetic mice. Expression of Alox5, synthesis of LTB4, and concentrations of the proinflammatory cytokine IL-1β were also increased in peritoneal macrophages and serum from T1DM mice. Insulin treatment restored LTB4 concentrations and Myd88 and Stat1 expression in T1DM mice. T1DM mice lacking Alox5 or treated with a 5-LO inhibitor showed reduced Myd88 and Il1b mRNA expression and increased IL-1 receptor antagonist concentration. The transcription factor cJun drove LTB4-dependent transcription of Stat1 in macrophages from T1DM mice. Compared to wild-type or untreated diabetic mice, T1DM mice lacking 5-LO or treated with a 5-LO inhibitor survived polymicrobial sepsis and showed reduced production of proinflammatory cytokines and decreased bacterial counts, suggesting that high LTB4 concentrations contribute to enhanced susceptibility to sepsis in T1DM. These results uncover a role for LTB4 in promoting sterile inflammation in diabetes and enhanced susceptibility to sepsis in T1DM.

MicroRNAs are known to control Toll like receptor activation in phagocytes. We have shown that leukotriene (LT) B4 (LTB4) positively regulates macrophage MyD88 expression by decreasing suppressor of cytokine signaling-1 (SOCS-1) mRNA stability. Here, we investigated the possibility that LTB4 control of MyD88 expression involves the generation of microRNAs. Our data show that LTB4, via its receptor B leukotriene receptor 1 (BLT1) and Gαi signaling, increased macrophage expression of inflammatory microRNAs, including miR-155, miR-146b, and miR-125b. LTB4-mediated miR-155 generation was attributable to AP-1 activation. Furthermore, macrophage transfection with antagomirs against miR-155 and miR-146b prevented both the LTB4-mediated decrease in SOCS-1 and increase in MyD88. Transfection with miR-155 and miR-146b mimics decreased SOCS-1 levels, increased MyD88 expression, and restored TLR4 responsiveness in both WT and LT-deficient macrophages. Together, our data unveil a heretofore unrecognized role for the GPCR BLT1 in controlling expression of microRNAs which regulate MyD88-dependent activation of macrophages.

Polymicrobial sepsis induces organ failure and is accompanied by overwhelming inflammatory response and impairment of microbial killing. Peroxisome proliferator-activated receptor-gamma (PPAR-γ) is a nuclear receptor with pleiotropic effects on lipid metabolism, inflammation, and cell proliferation. The insulin-sensitizing drugs thiazolidinediones (TZDs) are specific PPAR-γ agonists. TZDs exert anti-inflammatory actions in different disease models, including polymicrobial sepsis. The TZD pioglitazone, which is an FDA-approved drug, improves sepsis outcome; however, the molecular programs that mediate the effect of pioglitazone have not been determined. In a murine model of sepsis, we now show that pioglitazone treatment acts to improve microbial clearance and to enhance neutrophil recruitment to the site of infection. We also observed reduced pro-inflammatory cytokine production and high IL-10 levels in pioglitazone-treated mice. These effects were associated with a decrease in STAT-1-dependent expression of myeloid differentiation factor-88 (MyD88) in vivo and in vitro. IL-10R blockage abolished PPAR-γ-mediated inhibition of MyD88 expression. These data demonstrate that the primary mechanism by which pioglitazone protects against polymicrobial sepsis is by impairing MyD88 responses. This appears to represent a novel regulatory program. In this regard, pioglitazone provides advantages as a therapeutic tool, since it improves different aspects of host defense during sepsis, ultimately enhancing survival.