Protective effect of human CD40-Ig fusion protein in a murine model of acute graft-versus-host disease
10.3760/j.issn:0366-6999.2001.07.004
- VernacularTitle:人CD40-Ig融合蛋白对小鼠GVHD的保护作用研究
- Author:
HeZhong LIU
1
;
Ning MAO
;
Chunmei HOU
;
Xiusen LI
;
Beifen SHEN
;
Pei-Hsien TANG
Author Information
1. Academy of Military Medical Sciences
- Keywords:
CD40-Ig;
fusion protein;
graft-versus-host disease
- From:
Chinese Medical Journal
2001;114(7):685-689
- CountryChina
- Language:Chinese
-
Abstract:
Abstract:Objective To investigate the protective effects of blocking CD40/CD40L interactions with human CD40-Ig fusion protein in a murine graft-versus-host disease model.Methods Human CD40 gene extracellular region was inserted into plasmid pIG1, which contains genomic human IgG1 Fc gene. A transient vector containing CD40-Fc fusion gene was transfected into COS-7 cells. The CD40-Ig fusion protein was detected through enzyme-linked immunosorbent assay (ELISA). A constitutive vector was also generated by ligating the CD40-Fc fusion gene into pcDNA3.1 and transfecting it into CHO cells. CD40-Ig was purified by protein A affinity chromatography. SDS-PAGE, Western blot and ligand binding assay were used to identify the qualities of CD40-Ig. Murine acute graft-versus-host disease (GVHD) was induced by intravenous injection of C57BL/6J (H-2b) spleen cells into sub-lethally irradiated BALB/c (H-2d) mice. Protective effects against murine graft-versus-host disease by in vivo administration of CD40-Ig were evaluated.Results Mammalian expression vectors pIG/40Ig and p3.1/40Ig were constructed as described above. Chimeric proteins were expressed in COS-7 and CHO cell culture supernatant and confirmed by ELISA and Western blot. SDS-PAGE showed that fusion proteins had a disulfide-bonded dimeric structure and existed as homodimer. Purified CD40-Ig could bind to CD40L. In vivo administration of CD40-Ig could prevent the development of GVHD and significantly prolong the mean survival time of mice with graft-versus-host disease.Conclusions These results demonstrate that CD40/CD40L interactions play an important role in the pathogenesis of graft-versus-host disease and suggest clinical potential for CD40-Ig in the prevention and treatment of human graft-versus-host disease.
