Adipose tissue-resident macrophages are an important contributor to systemic chronic inflammation and cardiometabolic abnormalities associated with obesity. However, the etiological factors that alter the number and polarization of adipose-resident macrophages remain poorly characterized. As our previous clinical study has identified microRNA (miR)-34a as one of the most significantly elevated genes in visceral fat of obese patients, we investigated the roles of miR-34a in obesity-related adipose tissue inflammation and metabolic disorders by generated adipose tissue-specific miR-34a knockout (KO) mice. After feeding with high fat diet (HFD), adipose miR-34a KO mice exhibited significant attenuation of HFD-induced glucose intolerance, insulin resistance, hyperlipidemia, steatohepatitis and systemic inflammation compared to HFD-fed wild type (WT) controls. These benefits of miR-34a deficiency were mainly attributed to the reduced number of total macrophages and polarization of adipose macrophages towards the pro-inflammatory M1 phenotype. Mechanistically, we found that miR-34a directly repressed the expression of the transcription factor Klf4, thereby enhancing macrophage polarization to the M1 phenotype. Silencing of Klf4 in the adipose-specific miR-34a KO mice suppressed M2 macrophage polarization. In visceral fat from obese patients, the expression of Klf4 was significantly decreased and negatively correlated with miR-34a level. These data demonstrated that increased miR-34a in adipose tissues exacerbates obesity-induced adipose tissue inflammation by suppressing the expression of Klf4, thereby leading to increased accumulation of pro-inflammatory M1 macrophages. Disclosure Y. Pan: None. H. Hui: None. R. Hoo: None. T. Feng: None. K.S. Lam: None. A. Xu: None.

Background: Type 1 diabetes is an autoimmune disease resulted from self-destruction of insulin-producing pancreatic beta-cells. However, the pathological pathways that trigger the autoimmune destruction remain poorly understood. Our previous studies demonstrated that increased circulating neutrophil elastase (NE) is closely associated with beta-cell autoimmunity in patients with T1D. Here we investigate the role and mechanism whereby NE participates in the pathogenesis of T1D, and to explore the therapeutic potential of the pharmacological NE inhibitors for autoimmune diabetes. Method: Biochemical and immunological analysis were adopted to determine the dynamic change in the infiltration and activation of neutrophils in pancreas of NOD mice. NE-specific endogenous (pre-elafin/elafin) or pharmaceutical (sivelestat) inhibitors were supplemented into NOD mice to evaluate their effects on diabetes incidence and insultitis. Flow cytometry was employed to explore the effects of NE in mediating the crosstalk of immune cells. Results: A dynamic change in the infiltration of neutrophils was observed in pancreatic islets of NOD mice, leading to a transient elevation of local NE activity. Supplementation of NE-specific inhibitors rectified the elevated pancreatic NE activity, alleviated insulitis and reduced the development of T1D in NOD mice. Inhibition of NE activity significantly attenuated the infiltration of pro-inflammatory macrophages into pancreas, reduced the production of inflammatory cytokines. In vitro studies showed that NE directly induced inflammatory responses in both min6 beta-cells and RAW264.7 macrophages through the activation of TLR4-IκB-NFκB pathway. Discussion: NE-mediated crosstalk between beta-cell and macrophages forms a vicious feedback loop, resulting in the exaggerated beta-cell damage in NOD mice. Pharmacological inhibition of NE may represent a promising therapeutic strategy for treatment of autoimmune diabetes. Disclosure S. Lingling: None. L. Zhong: None. R. Hoo: None. A. Xu: None.

Liver fibrosis is the result of chronic liver injury and the first step toward the development of liver cirrhosis and hepatic carcinoma. Under the stress of damage factors, quiescent hepatic stellate cells (qHSCs) trans-differentiate into myofibroblast-like cells, and take the main role of extracellular matrix secretion contributing to the development of liver fibrosis. In recent clinical study, circulating level of A-FABP, was found to positively correlate with the stages of liver fibrosis and liver cirrhosis. Here, we investigate the role of A-FABP in the development of liver fibrosis. A-FABP knockout (A-FABP KO) mice and their wild type (WT) littermates were subjected to bile duct ligation (BDL) for two weeks to induce liver fibrosis. Plasma and hepatic A-FABP were significantly elevated in BDL treated WT mice. Liver sinusoidal endothelial cell (LSEC) was identified as the major cellular source of hepatic A-FABP in response to BDL. In the BDL treated groups, comparing to the WT mice, A-FABP KO mice showed significantly reduced collagen formation and HSC activation which were accompanied by an attenuated induction of hepatic expression of transforming growth factor beta 1 (TGFβ1), a central regulator in hepatic fibrogenesis. As the LSECs and HSCs are closely attached to each other, we hypothesize that LSECs-derived A-FABP may act in a paracrine manner to stimulate the expression of TGFβ1 in HSCs. Our in vitro study demonstrated that treatment of recombinant A-FABP protein (rA-FABP) significantly induced the expression of TGFβ1 in primary HSCs. Mechanistically, BDL induces the release of A-FABP from LSECs which stimulates the TGFβ1 gene transcription through enhancing the activator protein-1 (AP-1) activity on its promoter via upregulating the phosphorylation of c-Jun, a component of AP-1. In conclusion, A-FABP contributes to the development of liver fibrosis via enhancing the expression of TGFβ1 in HSCs. Disclosure X. Wu: None. L. Shu: None. K.S. Lam: None. A. Xu: None. R. Hoo: None.

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