Department of Molecular Biology, Rajendra Memorial Research Institute of Medical Sciences, Agamkuan, Patna, Bihar, India.
Department of Microbiology, All India Institute of Medical Sciences, Phulwarisharif, Patna, Bihar, India.
Sci Rep. 2020 Feb 26;10(1):3523. doi: 10.1038/s41598-020-60435-2.
Visceral leishmaniasis is characterized by mixed production of Th1/2 cytokines and the disease is established by an enhanced level of Th2 cytokine. CD4+ T cells are main cell type which produces Th1/2 cytokine in the host upon Leishmania infection. However, the regulatory mechanism for Th1/2 production is not well understood. In this study, we co-cultured mice CD4+ T cells with Leishmania donovani infected and uninfected macrophage for the identification of dysregulated miRNAs in CD4+ T cells by next-generation sequencing. Here, we identified 604 and 613 known miRNAs in CD4+ T cells in control and infected samples respectively and a total of only 503 miRNAs were common in both groups. The expression analysis revealed that 112 miRNAs were up and 96 were down-regulated in infected groups, compared to uninfected control. Nineteen up-regulated and 17 down-regulated miRNAs were statistically significant (p < 0.05), which were validated by qPCR. Further, using insilco approach, we identified the gene targets of significant miRNAs on the basis of CD4+ T cell biology. Eleven up-regulated miRNAs and 9 down-regulated miRNAs were associated with the cellular immune responses and Th1/2 dichotomy upon Leishmania donovani infection. The up-regulated miRNAs targeted transcription factors that promote differentiation of CD4+ T cells towards Th1 phenotype. While down-regulated miRNAs targeted the transcription factors that facilitate differentiation of CD4+ T cells towards Th2 populations. The GO and pathway enrichment analysis also showed that the identified miRNAs target the pathway and genes related to CD4+ T cell biology which plays important role in Leishmania donovani infection.
内脏利什曼病的特征是 Th1/2 细胞因子的混合产生，并且疾病是通过 Th2 细胞因子水平的增强而建立的。CD4+T 细胞是主要的细胞类型，在利什曼原虫感染宿主时产生 Th1/2 细胞因子。然而，Th1/2 产生的调节机制尚不清楚。在这项研究中，我们将小鼠 CD4+T 细胞与感染和未感染巨噬细胞的利什曼原虫共培养，通过下一代测序鉴定 CD4+T 细胞中失调的 miRNA。在这里，我们分别在对照和感染样本中鉴定出 CD4+T 细胞中的 604 和 613 个已知 miRNA，而两组共有总共只有 503 个 miRNA 是共同的。表达分析显示，与未感染对照相比，感染组中有 112 个 miRNA 上调，96 个下调。与未感染对照组相比，感染组中有 19 个上调和 17 个下调的 miRNA 具有统计学意义（p<0.05），并通过 qPCR 进行了验证。此外，通过 insilco 方法，我们根据 CD4+T 细胞生物学，确定了显著 miRNA 的基因靶标。11 个上调 miRNA 和 9 个下调 miRNA 与利什曼原虫感染时的细胞免疫反应和 Th1/2 二分法有关。上调的 miRNA 靶向促进 CD4+T 细胞向 Th1 表型分化的转录因子。而下调的 miRNA 靶向有利于 CD4+T 细胞向 Th2 群体分化的转录因子。GO 和途径富集分析还表明，鉴定出的 miRNA 靶向与 CD4+T 细胞生物学相关的途径和基因，这些基因在利什曼原虫感染中发挥重要作用。
