SIVmac239对猕猴B淋巴细胞增殖的抑制机制

Inhibition mechanism of SIVmac239 in macques B lymphocyte(MM133) proliferation in vitro

目的:探讨SIVmac239感染CD+4T淋巴细胞后,能否产生某些可抑制B淋巴细胞生长的活性因子,及其抑制作用的机制,为HIV/AIDS进一步的病原学研究和控制提供有益的线索。方法:用MTT实验观察不同时间收集的SIVmac239感染的CD4+T淋巴细胞上清和未感染SIVmac239的CD4+T淋巴细胞的上清对猕猴B淋巴细胞生长的抑制作用,用3H thymidineuptake assay判断这些上清是否抑制B淋巴细胞的DNA合成,用凝胶电泳方法检测DNA合成被抑制是否为凋亡的机制,用Western blot方法检测Cyclin D1的表达情况。结果:SIVmac239感染的CD4+T淋巴细胞上清含有抑制猕猴B淋巴细胞生长的因子,并且,其抑制效果随着作用时间的延长而增加。这些因子可较强的抑制CD+4T淋巴细胞的DNA合成,最高抑制率可达81%。凝胶试验结果未见DNA梯状条带。Western blot试验发现Cyclin D1表达被抑制。结论:SIVmac239感染CD4+T淋巴细胞可产生抑制B淋巴细胞生长的因子。此因子可较强的抑制猕猴B淋巴细胞DNA的合成,但并不引起B淋巴细胞凋亡。其主要机制为抑制猕猴B淋巴细胞的细胞周期蛋白D1的产生,使B淋巴细胞停止在G1期,即抑制作用的主要的机制为细胞被捕获在细胞周期的G1期。本研究结果发现一个新的现象,即猕猴B淋巴细胞虽然不被SIVmac239感染,但可被SIVmac239感染的CD4+T淋巴细胞产生的细胞因子所捕获,抑制其细胞DNA合成,从而可导致该细胞的功能发生障碍和影响机体体液免疫作用。

Objective：To explore the inhibitive factors which CD4^＋ T lymphocyte secrets after infection with SIVmac239, and whether these factors can inhibit the B lymphocyte proliferation activity and its inhibition mechanism so as to supply some helpful clue for pathogenology research of AIDS. Mehtods：Proliferation activity of MM133 cells treated by the supematant of C8166 cells infected SIVmac239 was tested by MTT assay. The inhibition of DNA synthesis of Supernatant of C8166 cells infected by SIVmac239 on B cells was estimated by 3H thymidine uptake assay, whether or not the apoptosis happened when the DNA synthesis inhibiton was judged by DNA fragmentation detection. Cyclin D1 expression was examined through western blot assay. Re- sults ：SIVmac239 stimulated CD4^＋ T lymphocyte to secret some inhibitive factors for the macques B lymphocyte. These factors could inhibit the DNA synthesis of B cells through 3H thymidine uptake assay,the electrophoresis result failed to show DNA ladder, and there was no Cyelin D1 bands on western blot assay for the concentration of 2% and 12.5%supernantant of C8166 infected by SIVmac239. Conclusions：CD4^＋ T lymphocyte infected by SIVmac239 can produce some inhibitive factors in the macques B lymphocyte,these factors also can inhibit the DNA synthesis of B cells, but the factors did not induce the apoptosis of B cells. The major inhibition mech- anism of the factors in the B cells is to arrest the cells in G1 phase duing cell cycling. A new phenomenon has been found in this experiment,and B cells can not be infected by SlVmac239, but they will be arrested in G1 phase by some factors produced by CD4^＋T Lymphocyte infected with SIVmac239. Thus the function of B lymphocytes and the humoral immunity may be limited.