Zhou Hui, Fan Yan-ying, Wu Chang-you
Department of Clinical Laboratory, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou 510080, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2009 Nov;25(11):976-9.
To study the effect and mechanism of BCG on human nature killer cells.
PBMC or purified NK cells were isolated from normal human peripheral blood with negative anti-Mycobacterium tuberculosis antibody and cultured with BCG, IL-12, BCG plus IL-12 and BCG plus anti-IL-12R beta 1 mAb (2B10), respectively. The levels of IFN-gamma and IL-12p40 in the culture supernatants were measured by ELISA. The frequency of IFN-gamma and granzyme B producing cells were analyzed by ELISpot. The cytolytic activity was detected by MTT reduction assay. The surface expression of IL-12R beta 1 on NK cells was detected by flow cytometry.
BCG significantly induced IFN-gamma production by PBMC in a dose-dependent manner. When PBMC was stimulated with BCG, the frequency of granzyme B producing cells was higher than that in unstimulated PBMC (P<0.05). BCG enhanced the cytotoxic activity of PBMC. BCG alone didn't induce IFN-gamma production by purified NK cells, but it can augment IL-12-induced IFN-gamma production by purified NK cells. The cytotoxic activities of BCG-stimulated and unstimulated purified NK cells were not significantly different (P>0.05). BCG induced IL-12 production by PBMC in a dose-dependent manner and enhanced IL-12R beta 1 expression on different subsets of NK cells. Blocking the effect of IL-12 by anti-IL-12R beta 1 mAb (2B10) inhibited BCG-induced IFN-gamma production and granzyme B releasing by PBMC.
BCG can indirectly promote biologic activity of NK cells and the production of endogenous IL-12 combined with up-regulation IL-12R beta 1 expression on the surface of NK cells is a part of the mechanisms of IL-12 on human NK cells.
研究卡介苗对人自然杀伤细胞的作用及机制。
用抗结核分枝杆菌阴性抗体从正常人外周血中分离出外周血单个核细胞（PBMC）或纯化的自然杀伤细胞（NK细胞），分别与卡介苗、白细胞介素-12（IL-12）、卡介苗加IL-12以及卡介苗加抗IL-12Rβ1单克隆抗体（2B10）共同培养。采用酶联免疫吸附测定（ELISA）法检测培养上清液中γ-干扰素（IFN-γ）和IL-12p40的水平。采用酶联免疫斑点法（ELISpot）分析产生IFN-γ和颗粒酶B的细胞频率。采用MTT比色法检测细胞溶解活性。采用流式细胞术检测NK细胞表面IL-12Rβ1的表达。
卡介苗以剂量依赖方式显著诱导PBMC产生IFN-γ。当用卡介苗刺激PBMC时，产生颗粒酶B的细胞频率高于未刺激的PBMC（P<0.05）。卡介苗增强了PBMC的细胞毒性活性。单独的卡介苗不能诱导纯化的NK细胞产生IFN-γ，但它可以增强IL-12诱导的纯化NK细胞产生IFN-γ。卡介苗刺激和未刺激的纯化NK细胞的细胞毒性活性无显著差异（P>0.05）。卡介苗以剂量依赖方式诱导PBMC产生IL-12，并增强NK细胞不同亚群表面IL-12Rβ1的表达。用抗IL-12Rβ1单克隆抗体（2B10）阻断IL-12的作用可抑制卡介苗诱导的PBMC产生IFN-γ和释放颗粒酶B。
卡介苗可间接促进NK细胞的生物学活性，内源性IL-12的产生以及NK细胞表面IL-12Rβ1表达上调是IL-12作用于人类NK细胞机制的一部分。
