Down-regulation of CXCR4 expression in MT4 cells by a recombinant vector expressing antisense RNA to CXCR4 and its potential anti-HIV-1 effect.
CXC-chemokine receptor (CXCR4) is one principle co-receptor for the entry of T cell line (T)-tropic HIV-1 virus into a cell. In order to find more efficacious therapeutic possibilities for people with an HIV-1 infection, we explored the inhibitory effects of antisense RNA on CXCR4 expression in MT4 cells. First, we used to RT-PCR to obtain DNA fragments from healthy adult peripheral blood mononuclear cells; these fragments targeted the initiation region of CXCR4 mRNA translation. We then constructed a recombinant retroviral vector, pLXSN-X4a (containing antisense RNA to CXCR4). After packaging by PA317 cells, the pseudovirion of the recombinant vector had formed and succeeded in transfecting MT4 cells (a kind of T-tropic HIV-1 susceptibility cell line). The PCR and RT-PCR results showed that the recombinant vector had integrated into the genome of MT4 cells and had been transcribed. The expression of CXCR4 on the surface of MT4 cells transfected with antisense RNA was reduced by 30%, compared with those cells transfected with blank vector or untransfected cells. No change in the DNA synthesis rates or in cell proliferation was found in any of the transfected cells. After a challenge with HIV-1 SF33, the cells transfected with antisense RNA vector (pLXSN-X4a) produced reduced p24 levels compared with the cells transfected with blank vector (pLXSN) or untransfected cells. These results indicated that these CXCR4-antisense expressing cells could resist T-tropic HIV-1 infection and could retain normal biological functions. These studies provide useful data for further experiments in this area.
Xing
Hui Chun
HC
Department of Infectious Diseases, Peking University First Hospital, Beijing 100034, PR China.
Xu
Xiao Yuan
XY
Liu
Zhen
Z
Wang
Qin Huan
QH
Yu
Min
M
Si
Chong Wen
CW
D002461:Cell Line, Transformed; D015536:Down-Regulation; D005434:Flow Cytometry; D005822:Genetic Vectors; D016655:HIV Core Protein p24 / Q000096:biosynthesis / Q000235:genetics; D015658:HIV Infections / Q000188:drug therapy*; D015497:HIV-1 / Q000187:drug effects*; D006801:Humans; D016372:RNA, Antisense / Q000235:genetics / Q000494:pharmacology*; D019718:Receptors, CXCR4 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D016655:HIV Core Protein p24; D016372:RNA, Antisense; D019718:Receptors, CXCR4