Function of IgD on lymphocyte activation and effect of hIgD-Fc-Ig fusion protein on human PBMC proliferation

Wen-sheng CHEN,Qiong HUANG,Yu-jing WU,Heng-shi CHEN,Jin DONG,Wei WEI

(Institute of Clinical Pharmacology,Anhui Medical University,Key Laboratory of Antiinflammatory and Immune Medicine,Ministry of Education,Hefei 230032,China)

T3-6

Function of IgD on lymphocyte activation and effect of hIgD-Fc-Ig fusion protein on human PBMC proliferation

Wen-sheng CHEN,Qiong HUANG,Yu-jing WU,Heng-shi CHEN,Jin DONG,Wei WEI

(Institute of Clinical Pharmacology,Anhui Medical University,Key Laboratory of Antiinflammatory and Immune Medicine,Ministry of Education,Hefei 230032,China)

OBJECTIVEThis study aimed to investigate the influence of IgD on T/B cell activation and construct hIgD-Fc-Ig fusion protein to competitive inhibition IgD binding with IgDR.METHODST/B cells were sorted by magnetic cell sorting.The differences of mIgD and IgD-R level between different T/B cell subtypes were detected by FCM.Serum IgD level was detected by ELISA.Human IgD-Fc-IgG1-Fc sequence was amplified by cross-PCR and then subcloned into PET28a(+)empty vector.After prokaryotic expression through escherichia coli,we obtained the hIgD-Fc-Ig fusion protein by affinity chromatograph.Western blot was used to identify the hIgD-Fc-Ig fusion protein.Human peripheral blood monouclear cells(PBMC)and fibroblast like synoviocytes(FLS)proliferation were detected using a cell counting kit-8(CCK-8).RESULTSThe percentage of CD3+/CD4+,CD3+/IgD+,CD3+/CD4+/IgD+,CD3+/IgD-R+and CD3+/CD4+/IgD-R+cells increased significantly in RA patients comparing to healthy people.IgD can stimulate PBMC proliferation.IgD(1,3,10,30 μg·mL-1)stimulate PBMC proliferation significantly after 24 h.We obtained stable and active hIgD-Fc-Ig fusion protein.The hIgD-Fc-Ig fusion protein showed no effect on PBMC proliferation.But it could downregulate human IgD protein promoting proliferation effects in human PBMC.CONCLUSIONThis result suggests that IgD and IgDR play an important role on T/B cell activation in RA patients and the hIgD-Fc-Ig fusion protein may competitively inhibit IgD′s function and may play an therapeutic role in autoimmune diseases.

immunoglobulin D；fusion protein；hIgD-Fc-Ig；PBMC；proliferation

The projectsupported by National Natural Science Foundation of China(81330081,81202596);Specialized Research Fund for the Doctoral Program of Higher Education(20123420110003);Program for Tackling Key Problems in Science and Technology by Anhui Province(1301042098);and China Postdoctoral Science Foundation(2013M540508)

Wei WEI,Tel:13605510563,E-mail:wwei@ahmu.edu.cn