Abstract：
Whether naive CD4+ T cells are dysregulated and associated with the overactivation of CD4+ T cells in primary SS (pSS) remains unclear. We aimed to explore the role and underlying mechanism of naive CD4+ T cells in pSS.
We examined the activation, proliferation and differentiation of naive CD4+ T cells from pSS patients and healthy controls. Differentially expressed genes were identified using RNA sequencing, and were overexpressed or silenced to determine the gene regulating follicular helper T (Tfh) cells. Assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) with chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) was performed to explore the epigenetic mechanism. Naive CD4+ T cells were treated with pSS-related cytokines to explore the upstream signalling pathway.
pSS naive CD4+ T cells had higher potentials of activation, proliferation and differentiation towards Tfh cells. Thymocyte selection-associated high mobility group box protein (TOX) was upregulated in pSS naive CD4+ T cells and promoted T cell activation and Tfh cell polarization. TOX silencing in pSS naive CD4+ T cells downregulated B cell lymphoma 6 (BCL6) expression and altered levels of multiple Tfh-associated genes. ChIP-seq analysis implied that TOX bound to the BCL6 locus, where there were accessible regions found by ATAC-seq. IFN-α induced TOX overexpression, which was attenuated by Janus kinase (JAK) and signal transducer and activator of transcription 1 (STAT1) inhibitors.
Our data suggest that TOX in pSS naive CD4+ T cells is upregulated, which facilitates Tfh cell differentiation. Mechanistically, IFN-α induces TOX overexpression in naive CD4+ T cells through JAK-STAT1 signalling and TOX regulates BCL6 expression. Therefore, IFN-α-JAK-STAT1 signalling and TOX might be potential therapeutic targets in pSS.
We examined the activation, proliferation and differentiation of naive CD4+ T cells from pSS patients and healthy controls. Differentially expressed genes were identified using RNA sequencing, and were overexpressed or silenced to determine the gene regulating follicular helper T (Tfh) cells. Assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) with chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) was performed to explore the epigenetic mechanism. Naive CD4+ T cells were treated with pSS-related cytokines to explore the upstream signalling pathway.
pSS naive CD4+ T cells had higher potentials of activation, proliferation and differentiation towards Tfh cells. Thymocyte selection-associated high mobility group box protein (TOX) was upregulated in pSS naive CD4+ T cells and promoted T cell activation and Tfh cell polarization. TOX silencing in pSS naive CD4+ T cells downregulated B cell lymphoma 6 (BCL6) expression and altered levels of multiple Tfh-associated genes. ChIP-seq analysis implied that TOX bound to the BCL6 locus, where there were accessible regions found by ATAC-seq. IFN-α induced TOX overexpression, which was attenuated by Janus kinase (JAK) and signal transducer and activator of transcription 1 (STAT1) inhibitors.
Our data suggest that TOX in pSS naive CD4+ T cells is upregulated, which facilitates Tfh cell differentiation. Mechanistically, IFN-α induces TOX overexpression in naive CD4+ T cells through JAK-STAT1 signalling and TOX regulates BCL6 expression. Therefore, IFN-α-JAK-STAT1 signalling and TOX might be potential therapeutic targets in pSS.
摘要：
目的：在原发性SS（pSS）中，初始CD4+T细胞是否失调以及是否与CD4+T细胞过度活化有关，目前尚不清楚。我们旨在探讨幼稚CD4T细胞在pSS中的作用和潜在机制。
方法：我们检查了激活，来自pSS患者和健康对照的幼稚CD4+T细胞的增殖和分化。使用RNA测序鉴定差异表达的基因，并过表达或沉默以确定调节滤泡辅助性T（Tfh）细胞的基因。通过高通量测序（ATAC-seq）和染色质免疫沉淀（ChIP-seq）进行转座酶可接近的染色质测定，以探索表观遗传机制。用pSS相关细胞因子处理幼稚CD4+T细胞以探索上游信号通路。
结果：pSS幼稚CD4+T细胞具有较高的活化潜能，向Tfh细胞增殖和分化。胸腺细胞选择相关的高迁移率族框蛋白（TOX）在pSS幼稚CD4T细胞中上调，并促进T细胞活化和Tfh细胞极化。pSS幼稚CD4+T细胞中的TOX沉默下调B细胞淋巴瘤6(BCL6)表达和多个Tfh相关基因水平的改变。ChIP-seq分析暗示TOX与BCL6基因座结合，在那里有ATAC-seq发现的可访问区域。IFN-α诱导的TOX过表达,Janus激酶（JAK）和信号转导和转录激活因子1（STAT1）抑制剂减弱。
结论：我们的数据表明pSS初始CD4+T细胞中的TOX上调，促进Tfh细胞分化。机械上,IFN-α通过JAK-STAT1信号传导在幼稚CD4+T细胞中诱导TOX过表达，TOX调节BCL6表达。因此,IFN-α-JAK-STAT1信号传导和TOX可能是pSS的潜在治疗靶点。
方法：我们检查了激活，来自pSS患者和健康对照的幼稚CD4+T细胞的增殖和分化。使用RNA测序鉴定差异表达的基因，并过表达或沉默以确定调节滤泡辅助性T（Tfh）细胞的基因。通过高通量测序（ATAC-seq）和染色质免疫沉淀（ChIP-seq）进行转座酶可接近的染色质测定，以探索表观遗传机制。用pSS相关细胞因子处理幼稚CD4+T细胞以探索上游信号通路。
结果：pSS幼稚CD4+T细胞具有较高的活化潜能，向Tfh细胞增殖和分化。胸腺细胞选择相关的高迁移率族框蛋白（TOX）在pSS幼稚CD4T细胞中上调，并促进T细胞活化和Tfh细胞极化。pSS幼稚CD4+T细胞中的TOX沉默下调B细胞淋巴瘤6(BCL6)表达和多个Tfh相关基因水平的改变。ChIP-seq分析暗示TOX与BCL6基因座结合，在那里有ATAC-seq发现的可访问区域。IFN-α诱导的TOX过表达,Janus激酶（JAK）和信号转导和转录激活因子1（STAT1）抑制剂减弱。
结论：我们的数据表明pSS初始CD4+T细胞中的TOX上调，促进Tfh细胞分化。机械上,IFN-α通过JAK-STAT1信号传导在幼稚CD4+T细胞中诱导TOX过表达，TOX调节BCL6表达。因此,IFN-α-JAK-STAT1信号传导和TOX可能是pSS的潜在治疗靶点。
