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制备了工程化靶向融合蛋白XE-TNFαm2。其中,XE为HIV/SIV辅助受体CXCR4的第二胞外域。TNFαm2是经突变改型的TNFα,其毒副作用已降低18倍,己用于临床治疗恶性肿瘤。所用的整合有HIV的标准细胞株J-Lat Tat-GFP(H2/9855),为美国NIH艾滋病试剂中心所赠送。其中,一个经缺失突变后的HIV被整合在Jurkat细胞的染色体上,成为5′LTR-Tat-GFP-3′LTR。不同剂量的XE-TNFαm2加于一定量的JurkatH2/9855细胞后,流式细胞仪检测结果表明,荧光蛋白的表达量随着处理时间的延续而增加,并有XE-TNFαm2剂量的依赖关系。这一结果表明,XE-TNFαm2可强力激活潜伏于细胞染色体中的HIV,使之重新繁殖起来。鉴于己有的研究表明,XE-TNFαm2可杀灭受HIV/SIV感染的细胞。据此,当重新繁殖的HIV开始出芽时,其gp120必然出现在宿主细胞表面,且此gp120必然被XE-TNFαm2中的XE所结合,并其TNFαm2的杀伤信号将转导进入细胞。这样,这些宿主细胞将被杀灭。细胞的死亡导致未成熟HIV繁殖的中止。最后,在重新繁殖且成熟起来的HIV导致细胞破碎并释放出细胞之前,细胞内尚无感染力的未成熟HIV将同死亡的宿主细胞一起被清除。
The engineered targeting fusion protein XE-TNFαm2 was prepared. Among them, XE is the second extracellular domain of HIV / SIV co-receptor CXCR4. TNFαm2 is a mutated TNFα that has been 18-fold less toxic and has been used clinically to treat malignant tumors. The HIV-containing standard cell line J-Lat Tat-GFP (H2 / 9855) was used as a gift from the NIH AIDS Reagent Center. Among them, a deletion of the mutated HIV was integrated in the chromosome of Jurkat cells, become 5’LTR-Tat-GFP-3’LTR. After different doses of XE-TNFαm2 were added to a certain amount of JurkatH2 / 9855 cells, the results of flow cytometry showed that the expression of fluorescent protein increased with the prolongation of treatment time and the dose dependence of XE-TNFαm2. This result suggests that XE-TNFαm2 potently activates HIV that is latent in the chromosomes of cells and reproduces them. In view of previous studies, it has been shown that XE-TNFαm2 can kill cells infected with HIV / SIV. Accordingly, when repopulated HIV begins to sprout, its gp120 must appear on the surface of host cells, and this gp120 must be bound by XE in XE-TNFa2 and its TNFa2 killing signal will be transduced into the cell. In this way, these host cells will be killed. Cell death leads to termination of immature HIV multiplication. Finally, immature HIV cells that are uninfected in the cell will be eliminated along with the dead host cells before repopulating and maturing HIV causes the cells to break up and release the cells.