DOC2b enrichment mitigates proinflammatory cytokine-induced CXCL10 expression by attenuating IKKβ and STAT-1 signaling in human islets

Abstract

IntroductionType 1 diabetic human islet {beta}-cells are deficient in double C 2 like domain beta (DOC2b) protein. Further, DOC2b protects against cytokine-induced pancreatic islet {beta}-cell stress and apoptosis. However, the mechanisms underpinning the protective effects of DOC2b remain unknown. MethodsBiochemical studies, qPCR, proteomics, and immuno-confocal microscopy were conducted to determine the underlying protective mechanisms of DOC2b in {beta}-cells. DOC2b- enriched or-depleted primary islets (human and mouse) and {beta}-cell lines challenged with or without proinflammatory cytokines, global DOC2b heterozygous knockout mice subjected to multiple-low-dose-streptozotocin (MLD-STZ), were used for these studies. ResultsA significant elevation of stress-induced CXCL10 mRNA was observed in DOC2b- depleted {beta}-cells and primary mouse islets. Further, DOC2b enrichment markedly attenuated cytokine-induced CXCL10 levels in primary non-diabetic human islets and {beta}-cells. DOC2b enrichment also reduced total-NF-{kappa}B p65 protein levels in human islets challenged with T1D mimicking proinflammatory cytokines. IKK{beta}, NF-{kappa}B p65, and STAT-1 are capable of associating with DOC2b in cytokine-challenged {beta}-cells. DOC2b enrichment in cytokine-stressed human islets and {beta}-cells corresponded with a significant reduction in activated and total IKK{beta} protein levels. Total I{kappa}B{beta} protein was increased in DOC2b-enriched human islets subjected to acute cytokine challenge. Cytokine-induced activated and total STAT-1 protein and mRNA levels were markedly reduced in DOC2b-enriched human islets. Intriguingly, DOC2b also prevents ER-stress-IKK{beta} and STAT-1 crosstalk in the rat INS1-832/13 {beta}-cell line. ConclusionThe mechanisms underpinning the protective effects of DOC2b involve attenuation of IKK{beta}-NF-{kappa}B p65 and STAT-1 signaling, and reduced CXCL10 expression. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=179 HEIGHT=200 SRC="FIGDIR/small/629540v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1b9d0acorg.highwire.dtl.DTLVardef@32db73org.highwire.dtl.DTLVardef@e5645aorg.highwire.dtl.DTLVardef@1f7619_HPS_FORMAT_FIGEXP M_FIG C_FIG