NRF-DST Centre of Excellence for Biomedical Tuberculosis Research; South African Medical Research Council Centre for Tuberculosis Research; Division of Molecular Biology and Human Genetics, Faculty of Medicine and Health Sciences, Stellenbosch University, Cape Town, South Africa.
South African Tuberculosis Bioinformatics Initiative (SATBBI), Faculty of Medicine and Health Sciences, Stellenbosch University, Cape Town, South Africa.
Virulence. 2020 Dec;11(1):170-182. doi: 10.1080/21505594.2020.1726561.
The comparison of the host immune response when challenged with pathogenic and nonpathogenic species of mycobacteria can provide answers to the unresolved question of how pathogens subvert or inhibit an effective response. We infected human monocyte derived macrophages (hMDMs) with different species of mycobacteria, in increasing order of pathogenicity, i.e. BCG, and R179 that had been cultured in the absence of detergents. RNA was isolated post-infection and transcriptomic analysis using amplicons (Ampliseq) revealed 274 differentially expressed genes (DEGs) across three species, out of which we selected 19 DEGs for further validation. We used qRT-PCR to confirm the differential expression of 19 DEGs. We studied biological network through Ingenuity Pathway Analysis® (IPA) which revealed up-regulated pathways of the interferon and interleukin family related to the killing of . Apart from interferon and interleukin family, we found one up-regulated ( and two down-regulated ( and ) genes as unique potential targets found by Ampliseq and qRT-PCR which may be involved in the intracellular mycobacterial killing. The roles of these genes have not previously been described in tuberculosis. Multiplex ELISA of culture supernatants showed increased host immune response toward as compared to BCG and R179. These results enhance our understanding of host immune response against infection.
当宿主受到致病性和非致病性分枝杆菌物种的挑战时，比较宿主免疫反应可以为尚未解决的问题提供答案，即病原体如何颠覆或抑制有效的反应。我们用不同的分枝杆菌物种（递增的致病性，即卡介苗和 R179）感染人单核细胞衍生的巨噬细胞（hMDMs），这些分枝杆菌是在没有去污剂的情况下培养的。感染后分离 RNA，并使用扩增子（Ampliseq）进行转录组分析，揭示了三种物种中 274 个差异表达基因（DEGs），我们从中选择了 19 个 DEGs 进行进一步验证。我们使用 qRT-PCR 来验证 19 个 DEGs 的差异表达。我们通过 Ingenuity Pathway Analysis®（IPA）研究了生物网络，结果显示干扰素和白细胞介素家族的上调途径与 的杀伤有关。除了干扰素和白细胞介素家族，我们还发现了一个上调（和两个下调（和）基因，这是 Ampliseq 和 qRT-PCR 发现的独特的潜在靶点，可能参与细胞内分枝杆菌的杀伤。这些基因在结核病中的作用以前没有被描述过。培养上清液的多重 ELISA 显示，与卡介苗和 R179 相比，宿主对 的免疫反应增强。这些结果增强了我们对宿主对 感染的免疫反应的理解。
