Gene Regulation Of Immun-inflammatory Reponses In NOD1/RIPK2/NF-?B Signaling By Lnc TINCR

The underlying mechanism of type 2 diabetes remains unclear,however,insulin resistance is closely linked to the pathogenesis of type 2 diabetes.We investigated the molecular mechanisms of insulin resistance to offer exciting opportunities for the development of new therapeutic strategies for the treatment of type 2 diabetes.Recent studies have confirmed that several lncRNAs are involved in the onset and development of diabetes.Chronic low-grade inflammation of adipose tissue is one of the main causes of obesity associated insulin resistance and diabetes.lncRNA is aberrantly expressed in inflammation-related diseases and is involved in the expression of various proinflammatory mediators and activation of signaling pathways.Innate immune receptors(e.g.NOD1)play critical roles in innate immune responses and inflammation.Recent literature proposes that lncRNA Mirt2 Inhibits activation of NF-?B and MAPK pathways and thus acts as a negative feedback regulator of excessive inflammation.We therefore hypothesized that lncRNA may be mediate inflammatory response and effect the process of insulin resistance in adipocytes.In the present study,3T3L1 preadipocytes were differentiated to adipocytes.Then adipocytes were pretreated with NOD1 synthetic ligands Tri-DAP to induce chronic inflammation.We performed RNA-Seq to identify the expression of the lncRNAs in Tri-DAP stimulated adipocytes to examined the role of lncRNAs in insulin resistance.Finally,we knockdowned lncRNA which was up-related with transfection experiments to explore the effect of lncRNA in inflammatory response.Objective: Investigating the expression of the lncRNAs in Tri-DAP stimulated mouse mature adipocytes.Addressing the mechanism underlying the regulation of inflammation response by lnc TINCR in insulin resistant adipocytes.Methods: 3T3-L1 preadipocytes were differentiated into adipocytes by 10?g/ml insulin,1?M dexamethasone,and 0.5 mM 3-isobutyl-1-methylxanthine.Oil red O staining was applied to observe the mature adipocytes.Mature adipocytes were randomized to normal control group and Tri-DAP(NOD1 ligand)group.After the establishment of insulin resistance,RNA sequencing(RNA-Seq)was applied to identify lncRNAs that are regulated during NOD1 activation in mouse adipocytes.Gene Ontology and KEGGPathway analysiswere used to explore the potential molecular function of the differentially expressed mRNAs.Real-time PCR was used to analyze the expression of lnc TINCR.,proinflammatory TNF-? ? IL-6 ?CXCL1 and RIPK2 in the presence or absence of Tri-DAP(10 g/ml).Whether knockdown of lnc TINCR could regulate the expression of proinflammatory cytokines IL-6?TNF-??CXCL1 and RIPK2 stimulation with Tri-DAP are determined using Real-time PCR and ELISA.Results: Deep RNA sequencing identified 81 lncRNAs and 167 mRNAs were obviously up-related(P<0.05)whereas 78 lncRNAs and 82 mRNAs were obviously down-related(P<0.05)more than twofold in NOD1 activation in adipocytes.We discovered that lncRNA NONMMUT038418.2,termed lnc TINCR(Tri-DAP-inducible non-protein coding RNA)is greatly upregulated in Tri-DAP activated adipocytes.Moreover,knockdown of lncTINCR with siRNA dampens the proinflammatory IL-6 ?TNF-? ?CXCL1 and RIPK2(P<0.05).Conclusions: LncRNA expression between normal and Tri-DAP stimulated 3T3-L1 adipocytes changed significantly.lnc TINCR is a positive regulator of adipocytes inflammation presumably via activation of inflammatory chemokine,TNF-?,IL-6,and CXCL1 as well as NOD1/RIPK2/ NF-?B signaling pathways.lnc TINCR maybe a regulator of inflammation-induced insulin resistance through acivation of NOD1 signaling pathways.