人枯否细胞在同种肝移植免疫中作用机制初步探讨

Role and mechanism of human Kupffer cells in liver allo-geneic transplantation

目的 探讨肝脏枯否细胞（KC）在肝移植后早期免疫反应中的可能作用.方法 将KC和（或）异体PBMC共培养,收集细胞上清液,培养结束时分别收获培养的KC和PBMC.检测HLA-G在细胞表面的表达;测定上清液中NO、IFN-γ、IL-10和TGF-β1的浓度;MTT试验观察KC对淋巴细胞增殖的影响.结果 实验组及对照组中KC和PBMC表面均未检测到HLA-G的表达.与不含KC实验组相比,含KC实验组中,NO、IL-10和TGF-β1的产量显著升高,而IFN-γ呈相对偏低趋势;对照组中未能检测到IL-10和IFN-γ的分泌,仅含KC的对照组中含少量NO及TGF-β1,且显著低于实验组.MTT实验发现,不含KC实验组OD值显著高于含KC实验组及对照组.结论 体外KC接触异体PBMC后早期,各细胞膜表面均无HLA-G表达,但参与了NO及Th2/Th3样细胞因子的分泌调节,并能抑制淋巴细胞增殖反应,可能促进肝脏移植早期免疫耐受的形成.

Objective To explore the possible role and mechanism of the Kupffer cells （KCs） in liver allo-geneic transplantation at the early stage. Methods In vitro cell contact coculture system was established. Culture supernatants were collected respectively on the 1st, 2nd, 4th, 6th d after cocul-ture and the KCs and PBMCs were harvested on the 6th day after culture. The expression of HLA-G on the membrane of the KCs and PBMCs was detected with immunochemistry. Nitrate reduction test was used to determine the concentration of nitric oxide. IFN-γ, IL-10, TGF-β1 cytokine levels in the supernatants were also measured with ELISA. The proliferation of lymphocytes was evaluated with MTT. Results six days later, no HLA-G molecules were detected on the membrane of the KCs and PBMCs. In the experimental group containing KCs, the levels of NO, IL-10 and TGF-β1 was signifi-cantly increased（P〈0. 05）, while the levels of IFN-γ was relatively lower（P〈0. 05） as compared to the experimental group without KCs. No IL-10 and IFN-γ were detected in the control group, and on-ly few NO and TGF-β1 was found in the control group with KCs. MTT test showed that the value of optical density was lower in the experimental group with KCs than that in any other group（P〈0. 05）.Conclusion No HLA-G is expressed on the membrane of KCs and PBMCs after contact coculture.KCs may participate in regulating production of NO and Th2/Th3-like cytokines and suppressing the proliferation of lymphocytes, through which KCs probably take part in inducing immunotolerance of liver transplantation in early stage.