Potent immunosuppression by a bivalent molecule binding to CD200R and TGF-betaR.
BACKGROUND
The novel immunosuppressive molecule, CD200, has been reported to induce immunoregulation after interaction with its receptor(s), CD200R(s), in part at least through augmented induction of regulatory T-cell populations. Independent studies have also described increased expression of indoleamine-2,3-dioxygenase after CD200R triggering, whereas others have provided evidence that TGF-beta is important for the induction or function of many populations of regulatory T cells. We have asked whether a hybrid molecule in which a soluble fusion protein containing CD200, CD200Fc, was linked to TGF-beta through a glycine linker (Gly6) functions as a superior immunosuppressant molecule when compared with CD200Fc or TGF-beta alone, or in combination.


METHODS
The hybrid molecule CD200FcGly6TGF-beta was expressed by transient transfection in CHO cells and purified over a protein A column. Functional activity of this and recombinant CD200Fc or TGF-beta alone were assessed in mixed leukocyte cultures (MLCs) and in skin graft rejection in vivo.


RESULTS
Immunosuppression mediated by CD200FcGly6TGF-beta is dependent on both functional CD200 and TGF-beta moieties, as indicated by inhibition of suppression using anti-CD200 or anti-TGF-beta antibodies. Using as responder cells, using antigen-presenting cell from mice with a deletion of the CD200R gene and responder T cells from mice with siRNA-mediated suppression of expression of the TGF-betaII receptor, we show that suppression follows binding to TGF-betaRII on T cells, and CD200R1 on antigen-presenting cells. Indoleamine-2,3-dioxygenase inhibitors did not attenuate suppression by CD200FcGly6TGF-beta.


CONCLUSIONS
CD200FcGly6TGF-beta is a potent immunosuppressant in vivo and in vitro.
Gorczynski
Reginald M
RM
Department of Surgery, Transplant Research Division, University Health Network, Toronto, ON, M5G1L7, Canada. rgorczynski@uhnres.utoronto.ca
Chen
Zhiqi
Z
Shivagnahnam
Saaiis
S
Taseva
Adriana
A
Wong
Karrie
K
Yu
Kai
K
Khatri
Ismat
I
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D001483:Base Sequence; D016466:CHO Cells; D002455:Cell Division; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers; D007108:Immune Tolerance; D007074:Immunoglobulin G / Q000235:genetics; D007165:Immunosuppression Therapy / Q000379:methods; D016753:Interleukin-10 / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011485:Protein Binding; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D013154:Spleen / Q000166:cytology / Q000276:immunology / Q000502:physiology; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics
D015703:Antigens, CD; C492346:CD200 receptor, mouse; C529001:CD200-FC, mouse; D017931:DNA Primers; D007074:Immunoglobulin G; D008562:Membrane Glycoproteins; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D016753:Interleukin-10; C042739:antigens, CD200