Establishment of a mesenchymal stem cells model stably expressing hCD4 and hCCR5
10.3969/j.issn.1671-8348.2017.32.001
- VernacularTitle:表达hCD4和hCCR5基因的骨髓间充质干细胞模型的建立
- Author:
Yajing LI
1
;
Fuyan ZHUGE
;
Juan LIANG
;
Jinyang HE
;
Ning TAN
;
Changchun ZENG
Author Information
1. 广东医科大学附属龙华中心医院
- Keywords:
mesenchymal stem cells;
HIV-1;
cell model;
human CD4 and CCR5;
gene expression
- From:
Chongqing Medicine
2017;46(32):4465-4468,4472
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a kind of mesenchymal stem cells(MSCs) model that could express hCD4 and hCCR5 to study the field of human immunodeficiency virus type 1 (HIV-1).Methods Lentiviral vectors containing the genes of hCD4 and hCCR5 under the transcriptional control of cytomegalovirus promoter were designed.MSCs were transfected by the lentiviral vectors at optimum multiplicity of infection.Transduction efficiencies of hCD4 and hCCR5 in MSCs were analyzed by quantitative real-time polymerase chain reaction(RT-PCR),Western blot,and immunofluorescence staining.Subsequently,the transfected human MSCs were infected with HIV-1,and the expression of HIVRNA in the MSCs was detected by RT-PCR.Results The MSCs model con-taining hCD4 and hCCR5 and supporting normal HIV-1 infection was constructed.qRT-PCR showed that MSCs upon infection with lentiviral vectors were highly expressed in hCD4 and CCR5 mRNA sequences(P＜0.01).Western blot detection showed the positive bands of 55.0 × 103 (hCD4) and 40.6 × 103 (hCCR5).The results of immunofluorescence staining revealed that hCD4 and CCR5 were expressed on the surface of MSCs.Such results were not found in cells infected with empty lentiviral vectors.And the susceptibility of the hCD4/CCR5 transgenic MSCs to the HIV-1 was further indicated by the detection of HIV-1 RNA in the culture supernatants and cell lysates(P＜0.05).Conclusion The MSCs model that could highly express hCD4 and hCCR5 was established to support normal HIV-1 infection,which can be used to investigate the development of new therapies and vaccines against HIV.
