Analysis of induction and activation mechanism of protective CD8 T cells against pulmonary tuberculosis and application for the development of anti-tuberculosis vaccine

虽然在CD8缺失或CD8T细胞缺陷的小鼠身上已经报道了CD8T细胞在预防肺结核中的重要性，但在结核分枝杆菌(Mtb)感染的肺中，分枝杆菌抗原特异性T细胞的迁移和激活是如何调节的还不清楚。为了解决这个问题，我们产生了MHC H-2D^b和分枝杆菌Ag Mtb32a表位复合体(D^&lt；b-&gt；Mtb32a 5mer)的五聚体。利用该试剂，我们从结核分枝杆菌感染后第21天的肺组织和第28天的纵隔淋巴结中鉴定出分枝杆菌Mtb32a特异性CD8T细胞。相比之下，卡介苗疫苗株也表达与结核分枝杆菌相同的Mtb32a表位，且BCG皮下感染后第7天即可诱导出D^b-Mtb32a 5聚体染色的CD8 T细胞，早于结核分枝杆菌肺部感染后的观察者。卡介苗接种小鼠肺内感染Mtb后30天，肺内Mtb32a特异性CD8 T细胞的诱导速度加快，从第7天开始明显检测到Mtb。此外，Mtb32a特异性T细胞的增加与肺对Mtb32a多肽反应产生的干扰素-γ的增加平行。结果提示，结核分枝杆菌特异性CD8T细胞在结核分枝杆菌感染肺中的诱导延迟，卡介苗可加速干扰素-γ的诱导。

Although importance of CD8+T cells in protection against pulmonary tuberculosis has been reported using CD8-depleted or CD8+T cell-deficient mice, it is not clear how migration and activation of the mycobacterial Ag-specific T cells are regulated in the Mycobacterium tuberculosis (Mtb)-infected lung. To address the issue, we generated pentamer of MHC H-2D^b and mycobacterial Ag Mtb32a epitope complex (D^<b->Mtb32a 5mer). Using the reagent we identified mycobacterial Mtb32a-specific CD8+T cells from day 21 in the lung and from day 28 in the mediastinal lymph node after Mtb lung infection. In contrast, Mycpbacterium bovis BCG, a vaccine strain, also express Mtb32a epitope identical to that of Mtb, and BCG subcutaneous infection induced D^b-Mtb32a 5mer-stained CD8+T cells from day 7 which is earlier than that observer after Mtb lung infection. When BCG vaccinated mice were infected with Mtb in the lung 30 days after the vaccination, induction of Mtb32a-specific CD8+T cells in the lung accelerated and clearly detected from day 7 after the infection. Furthermore, the increase of the Mtb32a-specificT cells parallel to increase of interferon-gamma production in the lung in response to Mtb32a peptide. The results suggest that induction of Mtb-specific CD8+T cells delays in the lung of Mtb-infected lung, and BCG vaccination accelerate the induction with interferon-gamma production.