LPS induced CXCL10 chemokine production via IFN- α receptor and toll-like receptor 4 signalling

CXCL10 is a chemokine induced by IFN-γ and implicated in the pathogenesis of type 1 diabetes (T1D). It attracts IFN-γ-producing T cells to the pancreatic islets, forming a positive feed-forward loop where T cells induce CXCL10 and attract themselves, but it is unclear what attracts the first IFN-γ-producing T cells in islets. Lipopolysaccharide induced CXCL10 production in cultured islets of nonobese diabetic (NOD) mice, and the lack of its inhibition in islets from NOD mice deficient for RAG1 or IFN-g receptor revealed its dissociation from the requirement of T cells and IFN-γ. To further investigate the mechanism of LPS induced CXCL10 production, the gene expression profile of pancreatic beta cells after culture of whole islets in LPS was analysed. Overall a gene expression signature of cytokine signalling, particularly by type I IFNs and TLR4 was upregulated after culture of islets in LPS. Islets were isolated from 6-8 week old male NOD mice and cultured for 48 hours with 50ng/ml lipopolysaccharide. Islets were then dispersed to single cells with trypsin and beta cells were sorted using FACS. RNA was isolated and subjected to RNA sequencing. There were n=5 independent samples per group, untreated and LPS-treated.

趋化因子CXCL10是由干扰素γ（Interferon-γ, IFN-γ）诱导产生的趋化因子，参与1型糖尿病（type 1 diabetes, T1D）的发病机制。其可将分泌干扰素γ的T细胞招募至胰岛，形成正前馈环路：T细胞诱导CXCL10表达并进一步招募自身细胞群，但目前尚不明确是什么因素首次将分泌干扰素γ的T细胞招募至胰岛内。研究发现，脂多糖（Lipopolysaccharide, LPS）可在体外培养的非肥胖糖尿病（nonobese diabetic, NOD）小鼠胰岛中诱导CXCL10产生；而在重组激活基因1（Recombination Activating Gene 1, RAG1）或干扰素γ受体缺陷的NOD小鼠胰岛中，该诱导作用并未受到抑制，这表明CXCL10的产生并不依赖于T细胞与干扰素γ。为进一步探究脂多糖诱导CXCL10产生的分子机制，本研究对经脂多糖体外培养的完整胰岛来源的胰腺β细胞进行了基因表达谱分析。整体而言，经脂多糖培养的胰岛中，细胞因子信号通路（尤其是I型干扰素与Toll样受体4（Toll-like receptor 4, TLR4）相关通路）的基因表达特征显著上调。实验所用胰岛分离自6-8周龄的雄性NOD小鼠，经含有50ng/ml脂多糖的培养基体外培养48小时；随后将胰岛用胰酶分散为单个细胞，通过荧光激活细胞分选术（Fluorescence-Activated Cell Sorting, FACS）分选出胰腺β细胞，提取总RNA并进行RNA测序。本研究设置未处理组与LPS处理组，每组均包含5个独立生物学重复样本。