Yoneda T, Ellner J J
Department of Medicine, Case Western Reserve University, and University Hospitals, Cleveland, Ohio, USA.
Am J Respir Crit Care Med. 1998 Aug;158(2):395-403. doi: 10.1164/ajrccm.158.2.9707102.
We established an in vitro model of the phagocytosis of Mycobacterium tuberculosis by human peripheral blood monocytes to evaluate the subsequent inhibition of intracellular replication of the organism. Highly purified T cells (94% CD3(+)/CD16(-)) or natural killer (NK) cells (96% CD16(+)/CD3(-)) isolated by Percoll discontinuous density gradient of peripheral blood mononuclear cells were incubated with M. tuberculosis-infected monocyte monolayers. Monocytes were lysed immediately and at 4, 7, and 10 d after infection for quantification of intracellular replication, which was assessed by quantitative plating techniques as colony-forming units (CFU). Whereas control monocytes permitted intracellular replication, T cells activated monocytes to kill 77% (p < 0.01) of intracellular M. tuberculosis compared with control monocytes by 10 d after infection. NK cells activated monocytes to kill 84% (p < 0.01) of M. tuberculosis in comparison with control monocytes. Lymphokine (IL-2)-activated-killer (LAK) cells were capable of activating monocytes to kill 97% (p < 0.01) of the intracellular organisms compared with control monocytes. In purified protein derivative (PPD)-positive donors, PPD-specific-CD4(+) lymphocytes stimulated monocytes to kill intracellular M. tuberculosis in a Class II major histocompatibility complex-restricted manner. In contrast, in PPD-negative donors, CD4(-) lymphocytes activated monocytes in a genetically unrestricted manner. Both T cell supernatant and NK cell supernatant generated from cocultivation with M. tuberculosis-infected monocytes also activated monocytes to augment mycobactericidal function. In conclusion, T cells, NK cells, LAK cells, and their supernatants activated mycobactericidal function of monocytes, although these pathways of activation differed in terms of antigenic specificity and genetic restriction.
我们建立了人外周血单核细胞吞噬结核分枝杆菌的体外模型，以评估该菌随后细胞内复制的抑制情况。通过外周血单个核细胞的Percoll不连续密度梯度分离出的高度纯化的T细胞（94% CD3(+)/CD16(-)）或自然杀伤（NK）细胞（96% CD16(+)/CD3(-)）与感染结核分枝杆菌的单核细胞单层共同孵育。感染后立即以及在4、7和10天时裂解单核细胞以定量细胞内复制情况，通过定量平板技术将其评估为菌落形成单位（CFU）。对照单核细胞允许细胞内复制，而T细胞激活单核细胞，与对照单核细胞相比，感染后10天时杀死了77%（p < 0.01）的细胞内结核分枝杆菌。与对照单核细胞相比，NK细胞激活单核细胞杀死了84%（p < 0.01）的结核分枝杆菌。与对照单核细胞相比，淋巴因子（IL-2）激活的杀伤（LAK）细胞能够激活单核细胞杀死97%（p < 0.01）的细胞内菌。在纯化蛋白衍生物（PPD）阳性供体中，PPD特异性CD4(+)淋巴细胞以II类主要组织相容性复合体限制的方式刺激单核细胞杀死细胞内结核分枝杆菌。相反，在PPD阴性供体中，CD4(-)淋巴细胞以非基因限制的方式激活单核细胞。与感染结核分枝杆菌的单核细胞共培养产生的T细胞上清液和NK细胞上清液也激活单核细胞以增强杀分枝杆菌功能。总之，T细胞、NK细胞、LAK细胞及其上清液激活了单核细胞的杀分枝杆菌功能，尽管这些激活途径在抗原特异性和基因限制方面有所不同。
