Experimental study on induction of allo-hyporesponsiveness by ICOS-Ig in vitro and in vivo ZHANG
10.3760/cma.j.issn.0254-1785.2009.07.001
- VernacularTitle:可诱导共刺激分子与人IgG Fc融合蛋白诱导同种免疫低应答的体内外实验
- Author:
Peng ZHANG
;
Qin QIN
;
Zhen-meng WANG
;
Qian SHEN
;
- Publication Type:Journal Article
- Keywords:
Antigens,CD28;
Immunoglobulin G;
Recombinant fusion proteins;
Immunosuppression;
Lymphocyte culture teat,mixed
- From:
Chinese Journal of Organ Transplantation
2009;30(7):389-393
- CountryChina
- Language:Chinese
-
Abstract:
Objective To express human inducible eostimulator (ICOS) extracellular region and IgG Fc fusion protein, and analyze their function in allogenie lymphocyte proliferation in vitro and in vivo. Methods Human ICOS extraeellular region and IgG Fc fragment were cloned into a soluble expression vector. ICOS-Ig fusion protein was expressed and purified in CHO cells. To monitor primary MLR, Balb/c spleen T cells were isolated as responder cells, and irradiated C57BL/6 spleen cells as stimulator cells. 50 μg/ml ICOS-Ig or IgG was added to primary MLR cultures. The cells responsive rates were detected by 3 H-TdR methods. ELISA tested supernatants for eytokines (IL-2,IL-4, IL-10 and IFN-γ). T cells of each group in primary MLR were cultured as responder cells for secondary MLR, and irradiated C57BL/6 (donor) or C3H (third party) spleen cells as stimulator cells. Similar indexes were detected in secondary MLR. Then vital dye CFSE was used to study alloreactive T cell proliferation in vivo. CFSE-labeled C57BL/6 spleen cells were transferred to irradiated Balb/c mice. Mice were then intraperitoneally injected with 0. 2 mg IgG, ICOS-Ig or CsA each day.At the 3rd day after transfection, the spleen cells of the mice were harvested to detect CD4+ CFSE+ and CD8+ CFSE+ by FACS. Results In primary MLR, ICOS-Ig inhibited allogenic T-cell proliferation with inhibition rate being (58 ± 8)% in 50 μg/ml, and increased IFN-γ secretion. In secondary MLR, ICOS-Ig specifically inhibited the proliferation of donor spleen cells with inhibition rate being (42±8)%, and in ICOS-Ig group the levels of IL-4 and IL-10 were lower and the level of IFN-γ higher than in IgG group. However, ICOS-Ig didn't inhibit the proliferation of third-party spleen cells. In the CFSE dye assay, CFSE intensity of CD4+ and CD8+ T cells in ICOS-Ig and CsA groups was stronger than that in control group (P < 0. 05), while CFSE intensity in combined treatment group were even stronger than that in ICOS-Ig and CsA groups (P<0. 05). Conclusion ICOS-Ig could inhibit allo-reactive T cell proliferation in vitro and in vivo, and induce donor-specific T cell hyporesponsiveness specifically.
