Chatterjee Bhowmick Diti, Ahn Miwon, Bhattacharya Supriyo, Aslamy Arianne, Thurmond Debbie C
bioRxiv. 2024 Dec 23:2024.12.22.629540. doi: 10.1101/2024.12.22.629540.
Type 1 diabetic human islet β-cells are deficient in double C 2 like domain beta (DOC2b) protein. Further, DOC2b protects against cytokine-induced pancreatic islet β-cell stress and apoptosis. However, the mechanisms underpinning the protective effects of DOC2b remain unknown.
Biochemical studies, qPCR, proteomics, and immuno-confocal microscopy were conducted to determine the underlying protective mechanisms of DOC2b in β-cells. DOC2b- enriched or-depleted primary islets (human and mouse) and β-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.
A significant elevation of stress-induced CXCL10 mRNA was observed in DOC2b- depleted β-cells and primary mouse islets. Further, DOC2b enrichment markedly attenuated cytokine-induced CXCL10 levels in primary non-diabetic human islets and β-cells. DOC2b enrichment also reduced total-NF-κB p65 protein levels in human islets challenged with T1D mimicking proinflammatory cytokines. IKKβ, NF-κB p65, and STAT-1 are capable of associating with DOC2b in cytokine-challenged β-cells. DOC2b enrichment in cytokine-stressed human islets and β-cells corresponded with a significant reduction in activated and total IKKβ protein levels. Total IκBβ 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-IKKβ and STAT-1 crosstalk in the rat INS1-832/13 β-cell line.
The mechanisms underpinning the protective effects of DOC2b involve attenuation of IKKβ-NF-κB p65 and STAT-1 signaling, and reduced CXCL10 expression.
1型糖尿病患者的胰岛β细胞中双C2样结构域β（DOC2b）蛋白缺乏。此外，DOC2b可保护胰岛β细胞免受细胞因子诱导的应激和凋亡。然而，DOC2b发挥保护作用的机制尚不清楚。
进行了生化研究、定量聚合酶链反应（qPCR）、蛋白质组学和免疫共聚焦显微镜检查，以确定DOC2b在β细胞中的潜在保护机制。这些研究使用了富含或缺乏DOC2b的原代胰岛（人和小鼠）以及β细胞系，分别用或不用促炎细胞因子进行刺激，还使用了经多次低剂量链脲佐菌素（MLD-STZ）处理的全球DOC2b杂合敲除小鼠。
在缺乏DOC2b的β细胞和原代小鼠胰岛中，观察到应激诱导的CXCL10 mRNA显著升高。此外，在原代非糖尿病人类胰岛和β细胞中，富集DOC2b可显著降低细胞因子诱导的CXCL10水平。在受到模拟1型糖尿病的促炎细胞因子刺激的人类胰岛中，富集DOC2b还可降低总NF-κB p65蛋白水平。在受到细胞因子刺激的β细胞中，IKKβ、NF-κB p65和STAT-1能够与DOC2b结合。在受到细胞因子应激的人类胰岛和β细胞中富集DOC2b，与活化的和总的IKKβ蛋白水平显著降低相对应。在受到急性细胞因子刺激的富含DOC2b的人类胰岛中，总IκBβ蛋白增加。在富含DOC2b的人类胰岛中，细胞因子诱导的活化的和总的STAT-1蛋白及mRNA水平显著降低。有趣的是，DOC2b还可防止大鼠INS1-832/13β细胞系中内质网应激-IKKβ和STAT-1的相互作用。
DOC2b发挥保护作用的机制包括减弱IKKβ-NF-κB p65和STAT-1信号传导，以及降低CXCL10表达。
