A non-infectious and quantitative cell-based bioassay for screening HIV entry inhibitors targeting HIV envelope proteins.
- Author:
Min-min LI
1
;
Cheng-lai XIA
;
Qin-chao MAO
;
Shi-bo JIANG
;
Shu-wen LIU
Author Information
- Publication Type:Journal Article
- MeSH: Biological Assay; Cell Fusion; Cell Line; Coculture Techniques; Drug Evaluation, Preclinical; methods; HIV Envelope Protein gp120; metabolism; HIV Envelope Protein gp41; metabolism; HIV Fusion Inhibitors; chemistry; pharmacology; Humans; beta-Galactosidase; metabolism
- From: Journal of Southern Medical University 2010;30(5):941-944
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo develop an objective bioassay for quantitative detection of HIV-induced cell-cell fusion for screening HIV entry inhibitors.
METHODSHL2/3 cells expressing HIV envelope proteins gp120/gp41, Tat, and other HIV proteins were co-cultured with HeLa-CD4-LTR-beta-gal cells expressing CD4 receptor and HIV LTR triggered reporter gene beta-galactosidase. The enzyme activities of beta-galactosidase were detected by a chromogenic substrate, chlorophenol red-beta-galactopyranoside (CPRG). Specific HIV entry inhibitors were used to validate the established detecting method.
RESULTSNo syncytium was formed by mixing HL2/3 and HeLa-CD4-LTR-beta-gal cells. However, the membrane could be fused and the Tat expressed by HL2/3 cells could bind to HIV LTR on HeLa-CD4-LTR-beta-gal cells and trigger the expression of beta-galactosidase. CPRG allowed quantitative and sensitive detection of the activity of beta-galactosidase. Further studies showed that HIV entry inhibitors could inhibit the activity of beta-galactosidase in a dose-dependent manner.
CONCLUSIONWe have developed a simple, cheap, objective and quantitative non-infectious cell-cell fusion bioassay that can be used to screen for anti-HIV agents targeting the virus entry from natural and synthetic compound libraries.
