Abstract
Our group previously investigated the levels of anti-Gal and anti-nonGal IgM and IgG in a cohort of 75 healthy humans of various backgrounds, and found some significant differences related to factors such as age, gender, ABO blood group, diet, vaccination history, and geographic location during childhood. We have now expanded our cohort (n = 84) to investigate the levels of anti-Neu5Gc and anti-nonGal/nonNeu5Gc antibodies in healthy humans. Anti-nonGal and anti-nonGal/nonNeu5Gc human IgM and IgG binding to pRBCs and pAECs from GTKO/CD46 and GTKO/CD46/Neu5GcKO pigs were measured by flow cytometry. Anti-Gal and anti-Neu5Gc IgM and IgG levels were measured by ELISA. In summary, (i) the great majority (almost 100%) of humans had anti-Neu5Gc IgM and IgG antibodies that bound to pAECs and approximately 50% had anti-Neu5Gc antibodies that bound to pRBCs, (ii) there was significantly less human antibody binding to pig cells that did not express either Gal or Neu5Gc compared with those that did not express Gal alone, (iii) the levels of both IgM and IgG binding to GTKO/CD46/Neu5GcKO pRBCs and pAECs were low, (iv) the level of anti-Neu5Gc IgG was higher in men than women, (v) the level did not change with age or diet, and there was some variability associated with (vi) previous vaccination history and (vii) the geographic region in which the individual spent his or her childhood. Our study confirms that human antibody binding to RBCs and AECs from GTKO/CD46/Neu5GcKO pigs is greatly reduced compared to binding to GTKO/CD46 cells. However, all humans appear to have a low level of antibody that binds to pAECs that is not directed to either Gal or Neu5Gc. Our findings require consideration in planning clinical trials of xenotransplantation.
Highlights
Because of inactivation of the α1,3-galactosyltransferase gene, humans do not express galactose-α1,3-galactose (Gal) [1]
As our group has previously reported [23, 24], both Gal and Nglycolylneuraminic acid (Neu5Gc) were expressed on wildtype pig red blood cells (RBCs) and aortal endothelial cells (AECs), but hCD46 was not (Fig 1)
RBCs and AECs from GTKO/CD46 pigs expressed Neu5Gc, but not Gal; the AECs expressed hCD46, but RBCs did not
Summary
Because of inactivation of the α1,3-galactosyltransferase gene, humans do not express galactose-α1,3-galactose (Gal) [1]. Humans develop natural antibodies directed to this oligosaccharide that is expressed on many wild-type (i.e., genetically-unmodified) pig cells [1,2,3,4]. Binding of primate anti-Gal antibodies to pig Gal epitopes results in hyperacute rejection (defined as graft failure within 24 hours after transplantation) or early graft failure [5,6,7]. The availability of pigs homozygous for α1,3-galactosyltransferase gene-knockout (GTKO) [8] has enabled pig-to-baboon organ transplantation to be carried out in the absence of Gal epitopes [9, 10]. We investigated the influence of age, gender, ABO blood group, diet, history of vaccination, and geographic location during childhood on antibody levels, the major conclusions from which are summarized in S1 Table
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