Suppression of human T-cell responses to beta-cells by activation of B7-H4 pathway.
B7-H4, a recently described member of the B7 family of cosignal molecules, is thought to be involved in the regulation of cellular and humoral immune responses through receptors on activated T and B cells. Human islet cells express positive B7-H4 mRNA in RT-PCR assays, but not B7-H4 protein on cell surface in flow cytometric analyses. To investigate the regulatory effects of activation of the B7-H4 pathway on the function of activated T cells of patients with type 1 diabetes (T1D), we have used our in vitro human experimental system, including human beta-cell antigen-specific T-cell clones and human beta-cell lines CM and HP62, as well as primary islet cells. B7-H4.Ig protein was purified from the culture supernatant of 293T cells transfected by a B7-H4.Ig plasmid (pMIgV, containing a human B7-H4 cDNA and a mouse IgG2a Fc cDNA). Our preliminary studies showed that immobilized fusion protein human B7-H4.Ig (coated with 5 microg/ml for 2 h at 37 degrees C), but not control Ig, clearly inhibited the proliferation of activated CD4+ and CD8+ T cells of patients induced by anti-CD3 antibody in CFSE assays. B7-H4.Ig also arrested cell cycle progression of T cells in G0/G1 phase and induced T-cell apoptosis as measured by BrdU-7-AAD flow cytometric analysis. To determine the cytoprotective effects of B7-H4, we developed transfectants of human beta-cell lines CM and HP62 and islet cells transfected with the B7-H4.Ig plasmid, using empty vector transfectants as controls. The results demonstrate that cell-associated B7-H4.Ig expressed on human beta-cells clearly inhibits the cytotoxicity of the T-cell clones to targeted human beta-cells in 51Cr release cytotoxicity assays. Activation of the B7-H4 pathway may represent a novel immunotherapeutic approach to inhibit T-cell responses for the prevention of beta-cell destruction in T1D.
Ou
Dawei
D
Department of Surgery, University of British Columbia, Vancouver, BC, V5Z 1L8, Canada. daweiou@interchange.ubc.ca
Wang
Xiaojie
X
Metzger
Daniel L
DL
Ao
Ziliang
Z
Pozzilli
Paolo
P
James
Roger F L
RF
Chen
Lieping
L
Warnock
Garth L
GL
D000293:Adolescent; D000328:Adult; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D018122:B7-1 Antigen / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D017252:CD3 Complex / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000187:drug effects; D018414:CD8-Positive T-Lymphocytes / Q000166:cytology / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002478:Cells, Cultured; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D005260:Female; D006801:Humans; D007136:Immunoglobulins / Q000378:metabolism / Q000502:physiology; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D007371:Interferon-gamma / Q000378:metabolism; D008297:Male; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D060907:V-Set Domain-Containing T-Cell Activation Inhibitor 1
D000911:Antibodies, Monoclonal; D018122:B7-1 Antigen; D017252:CD3 Complex; D007136:Immunoglobulins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D060907:V-Set Domain-Containing T-Cell Activation Inhibitor 1; C475826:VTCN1 protein, human; D007371:Interferon-gamma