Majumder Nivedita, Dey Ranadhir, Mathur Ram Kumar, Datta Sriparna, Maitra Madhumita, Ghosh Sanjukta, Saha Bhaskar, Majumdar Subrata
Department of Microbiology, Bose Institute, P1/12 CIT Scheme VII M, Kolkata, India.
Glycoconj J. 2006 Dec;23(9):675-86. doi: 10.1007/s10719-006-9017-9.
Various species of Mycobacteria produce a major cell wall-associated lipoglycan, called Lipoarabinomannan (LAM), which is involved in the virulence of Mycobacterial species. In this study, we tried to establish the role of the increased IL-10 secretion under Arabinosylated-LAM (Ara-LAM) treatment, the LAM that induces apoptosis in host macrophages or PBMC. We have studied the survival and apoptotic factors by western blotting, and estimated nitrite generation by Griess reaction, quantified iNOS mRNA by semi-quantitative RT-PCR, and ultimately the fate of the cells were studied by Flow Cytometric Analysis of AnnexinV-FITC binding. As per our observations, neutralization of released IL-10 in C57BL/6 peritoneal macrophages prior to Ara-LAM treatment, as well as macrophages from IL-10 knockout (KO) mice treated with Ara-LAM, showed significant down regulation of pro-apoptotic factors and up regulation of survival factors. These effects were strikingly similar to those when peritoneal macrophages were subjected to TNF-alpha and IL-12 neutralization followed by Ara-LAM-treatment. However, under similar conditions virulent Mannosylated-LAM (from Mycobacterium tuberculosis) treatment of macrophages clearly depicts the importance of IL-10 in the maintenance of pathogenesis, proving its usual immunosuppressive role. Thus, from our detailed investigations we point out an unusual pro-inflammatory action of IL-10 in Ara-LAM treated macrophages, where it behaves in a similar manner as the known Th1 cytokines TNF- alpha and IL-12.
多种分枝杆菌会产生一种主要的与细胞壁相关的脂多糖，称为脂阿拉伯甘露聚糖（LAM），它与分枝杆菌的毒力有关。在本研究中，我们试图确定在阿拉伯糖基化LAM（Ara-LAM）处理下IL-10分泌增加所起的作用，Ara-LAM是一种能诱导宿主巨噬细胞或外周血单核细胞凋亡的LAM。我们通过蛋白质印迹法研究了生存和凋亡因子，通过格里斯反应估计亚硝酸盐的产生，通过半定量逆转录聚合酶链反应（RT-PCR）对诱导型一氧化氮合酶（iNOS）mRNA进行定量，最终通过膜联蛋白V-异硫氰酸荧光素（AnnexinV-FITC）结合的流式细胞术分析研究细胞的命运。根据我们的观察，在Ara-LAM处理之前中和C57BL/6腹膜巨噬细胞中释放的IL-10，以及用Ara-LAM处理的IL-10基因敲除（KO）小鼠的巨噬细胞，均显示促凋亡因子显著下调，生存因子上调。这些效应与腹膜巨噬细胞先接受肿瘤坏死因子-α（TNF-α）和白细胞介素-12（IL-12）中和然后进行Ara-LAM处理时的效应极为相似。然而，在类似条件下，用有毒的甘露糖基化LAM（来自结核分枝杆菌）处理巨噬细胞清楚地表明了IL-10在维持发病机制中的重要性，证明了其通常的免疫抑制作用。因此，通过我们的详细研究，我们指出IL-10在Ara-LAM处理的巨噬细胞中具有一种不寻常的促炎作用，其表现与已知的Th1细胞因子TNF-α和IL-12相似。
