CAJ | 학술논문

目的:研究卡介苗(BCG)对人自然杀伤细胞(NK)功能的作用及其机制.方法:分离抗结核抗体阴性志愿者外周血PBMC、纯化NK细胞, 分别与BCG、 IL-12、 BCG+IL-12、 BCG+抗IL-12Rβ1 mAb(2B10)培养.利用ELISA方法检测培养上清液IFN-γ、 IL-12p40含量;利用ELISpot方法检测IFN-γ、颗粒酶B产生细胞的频率;利用四甲基偶氮唑盐(MTT)比色法测定杀伤功能.利用流式细胞术检测NK细胞IL-12Rβ1的表达.结果:BCG呈剂量依赖的方式诱导PBMC产生IFN-γ.在BCG刺激条件下, PBMC颗粒酶B分泌细胞数明显高于不加任何刺激剂组(P<0.05).BCG增强PBMC杀伤活性.BCG不能诱导纯化NK细胞产生IFN-γ, 但与IL-12同时刺激则表现出协同作用.纯化NK细胞经BCG刺激后杀伤活性与未刺激相比差异无统计学意义.BCG呈剂量依赖方式诱导PBMC产生IL-12、并促进NK细胞不同亚群表达IL-12Rβ1.2B10抗体抑制BCG对PBMC产生IFN-γ和分泌颗粒酶B的诱导作用.结论:BCG间接地促进NK细胞的生物学活性, 其部分机制是通过诱导单核细胞产生内源性IL-12、并上调NK细胞表达IL-12R.
목적:연구잡개묘(BCG)대인자연살상세포(NK)공능적작용급기궤제.방법:분리항결핵항체음성지원자외주혈PBMC、순화NK세포, 분별여BCG、 IL-12、 BCG+IL-12、 BCG+항IL-12Rβ1 mAb(2B10)배양.이용ELISA방법검측배양상청액IFN-γ、 IL-12p40함량;이용ELISpot방법검측IFN-γ、과립매B산생세포적빈솔;이용사갑기우담서염(MTT)비색법측정살상공능.이용류식세포술검측NK세포IL-12Rβ1적표체.결과:BCG정제량의뢰적방식유도PBMC산생IFN-γ.재BCG자격조건하, PBMC과립매B분비세포수명현고우불가임하자격제조(P<0.05).BCG증강PBMC살상활성.BCG불능유도순화NK세포산생IFN-γ, 단여IL-12동시자격칙표현출협동작용.순화NK세포경BCG자격후살상활성여미자격상비차이무통계학의의.BCG정제량의뢰방식유도PBMC산생IL-12、병촉진NK세포불동아군표체IL-12Rβ1.2B10항체억제BCG대PBMC산생IFN-γ화분비과립매B적유도작용.결론:BCG간접지촉진NK세포적생물학활성, 기부분궤제시통과유도단핵세포산생내원성IL-12、병상조NK세포표체IL-12R.
AIM: To study the effect and mechanism of BCG on human nature killer cells. METHODS: 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-12Rβ1 mAb (2B10), respectively. The levels of IFN-γ and IL-12p40 in the culture supernatants were measured by ELISA. The frequency of IFN-γ and granzyme B producing cells were analyzed by ELISpot. The cytolytic activity was detected by MTT reduction assay. The surface expression of IL-12Rβ1 on NK cells was detected by flow cytometry. RESULTS: BCG significantly induced IFN-γ 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-γ production by purified NK cells, but it can augment IL-12-induced IFN-γ 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-12Rβ1 expression on different subsets of NK cells. Blocking the effect of IL-12 by anti-IL-12Rβ1 mAb (2B10) inhibited BCG-induced IFN-γ production and granzyme B releasing by PBMC. CONCLUSION: BCG can indirectly promote biologic activity of NK cells and the production of endogenous IL-12 combined with up-regulation IL-12Rβ1 expression on the surface of NK cells is a part of the mechanisms of IL-12 on human NK cells.