THE HUMAN ANTIPORCINE CELLULAR REPERTOIRE - INVITRO STUDIES OF ACQUIRED AND INNATE CELLULAR RESPONSIVENESS

Discordant xenogeneic transplantation offers a potentially unlimited s ource of donor organs from easily bred, nonendangered, physiologically compatible animals, but has been limited by the inevitable occurrence of hyperacute rejection (HAR). The potential existence of cell-mediat ed discordant graft rejection has remained obscured by HAR, and hence is incompletely understood. To define the cellular elements capable of recognition of and subsequent response against discordant tissue in a clinically applicable species combination, we have studied the in vit ro interaction of human peripheral blood lymphocytes against 3 porcine B lymphoblastoid cell lines and 6 primary porcine endothelial cell po pulations. PBL from all individuals tested (n=10) proliferated in resp onse to culture for 72 hr in xenogeneic mixed lymphocyte culture (XMLC ) with cell lines expressing porcine MHC (SLA) class II antigens, whil e endothelial cultures lacking SLA class II generally failed to evoke a response. The proliferative response to class II-positive cells was attenuated by addition of anti-SLA class II antibody but not by anti-S LA class I antibody. Two endothelial populations expressing class II s timulated an inhibitable proliferative response. The magnitude of the short-term proliferative xenogeneic response was similar to that evoke d by fully mismatched allogeneic human B lymphoblastoid stimulators. A dditionally, extended XMLC was performed with PBL from 3 individuals. All populations responded with continued proliferation when repeatedly stimulated by porcine cells. This was characterized not only by T cel l growth, but by prominent NK cell growth as well. Elucidation of the TCR Vbeta chain usage patterns by semi-quantitative PCR documented sel ection of TCR transcripts from gene family Vbeta 2 in each group, comp lemented by a heterogeneous mixture of other transcripts including Vbe ta 17.1, 20.1, and 6.1. suggesting that direct human TCR binding of po rcine cells occurs, and that it is likely to be an individualistic res ponse complemented by a more homogeneous NK response. A Cr-51 release assay was utilized to demonstrate that unprimed PBL could also lyse po rcine target cells. This cytotoxic response was maintained despite the complete removal of T cells, suggesting that porcine-directed NK cell activity is present prior to the maturation of any T cell response. C ytolysis was also demonstrated in serum-free medium and thus was not m ediated solely by antibody-dependent cellular cytotoxicity. Chinese ha mster ovary cells transfected with the human T cell receptor accessory molecule CD4 were used to study the ability of this molecule to stabi lize the interaction between the human TCR and SLA class II. Strong bi nding was demonstrated in 2 of 3 porcine lines tested, indicating that some but not all porcine class II alleles can be bound by human CD4. These data demonstrate that a human T and NK cell-mediated response di rected against discordant porcine antigens, most notably SLA class II, is physiologically possible given successful abrogation of HAR. The T cell recognition of xenogeneic class II may be facilitated by direct TCR binding with stabilization of this interaction by CD4, but a cytot oxic NK response may precede the maturation of specific T cell recogni tion.