A comparison of fetal and adult porcine islets with regard to Gal alpha (1,3)Gal expression and the role of human immunoglobulins and complement in islet cell cytotoxicity

Background. It is still debated whether fetal or adult porcine islets shoul d be the preferred choice for future clinical islet xenotransplantation, Ea ch type of islet preparation has advantages and disadvantages compared with the other. Here we present a direct comparison between fetal and adult por cine islets with regard to Gal alpha(1,3)Gal expression, immunoglobulin and complement binding, and cytotoxicity after exposure to fresh human serum, Method. Islet single cell suspensions were prepared from adult and fetal is lets by trypsin digestion, Fluorescein isothiocyanate conjugated Bandeiraea simplicifolia isolectin B-4 (BS-IB4) and affinity-purified chicken anti-Ga l alpha(1,3)Gal antibody was used to detect Gal alpha(1,3)Gal expression. I mmunoglobulin and complement binding to the islet cells and cytotoxicity fo r islet cells was compared after incubation with fresh and heat-inactivated human sera and with an immune serum from a diabetic patient who received a fetal porcine islet transplant, Furthermore, two pools of human AB sera we re depleted of porcine endothelial cell cytotoxic human anti-Gal alpha(1,3) Gal antibodies by absorption and were used to analyze the effect of Gal alp ha(1,3)Gal antibody removal on islet cell cytotoxicity, Results. Fetal islet cells readily bound both BS-IB4 and the chicken anti-G al alpha(1,3)Gal antibody, None of 10 adult porcine islet preparations were stained by BS-IB4. In comparison, IgY anti-Gal Ab binding was detected in two of eight adult islet isolations, whereas the other six preparations sho wed marginal/no binding. After incubation of fetal islet cells with fresh h uman serum, C3c binding was strongly positive and IgM binding variable, wit h occasional binding of IgG and no detectable binding of IgA. Adult islet c ells were also strongly positive for C3c but did not bind detectable amount s of IgM, IgG, or IgA, Immune sera from a patient who had received fetal po rcine islets showed the presence of induced antibodies that bound to fetal islet cells and to porcine peripheral blood lymphocytes, whereas binding to adult islet cells was barely detectable. Fresh human sera showed a high an d similar level of complement-mediated lytic activity for both adult islet cells (78+/-22%) and fetal islet cells (75+/-16%). Cytotoxicity for fetal i slet cells and peripheral blood lymphocytes was significantly reduced when the corresponding sera were depleted of anti-Gal antibodies before use (P=0 .002 and P=0.003, respectively). In contrast, no difference in cytotoxicity for adult islet cells was detected when anti-Gal-depleted human sera were used. Conclusion. Gal alpha(1,3)Gal expression is occasionally detectable on adul t porcine islet cells, but not as readily and at a lower level, compared wi th fetal islet cells. Thus, as porcine fetal islets mature to adult islets, the expression of the Gal alpha(1,3)Gal epitope gradually diminishes. Cons equently, cytotoxic anti-Gal alpha(1,3)Gal antibodies in human serum play a n important role in the lysis of fetal but not adult porcine islet cells.