Mac-1-negative B-1b phenotype of natural antibody-producing cells, including those responding to Galα1,3Gal epitopes in α1,3-galactosyltransferase-deficient mice

Human natural Abs against Gal alpha1-3Gal beta1-4GlcNAc (Gal) epitopes are a major barrier to xenotransplantation, Studies in this report, which use combined multiparameter flow cytometric sorting and enzyme-linked immunospot assay, demonstrate that anti-Gal IgM-producing cells are found exclusively in a small B cell subpopulation (i.e., CD21(-/low) IgM(high) B220(low) CD5(-) Mac-1(-)493(-) cells) in the spleens of alpha1,3-galactosyltransferase-deficient mice. All IgM-producing cells were detected in a similar splenic subpopulation of alpha1,3-galactosyltransferase-deficient and wild-type mice. A higher frequency of B cells with anti-Gal surface IgM receptors was observed in the peritoneal cavity than in the spleen, but these did not actively secrete Abs, and showed phenotypic properties of B-lb cells (CD21(-/low) IgM(high) CD5(-) CD43(+) Mac-1(+)). However, these became Mac-1(-) and developed anti-Gal Ab-producing activity after in vitro culture with LPS, The splenic B cells with anti-Gal receptors consisted of both Mac-1(+) B-1b cells and Mac-1(-) B-1b-like cells. The latter comprised most anti-Gal IgM-producing cells, Our studies indicate that anti-Gal natural IgM Abs are produced by a Bib-like, Mac-1(-) splenic B cell population and not by plasma cells or B-la cells, They are consistent with a model whereby B-lb cells lose Mac-1 expression upon Ag exposure and that these, rather than plasma cells, become the major IgM Ab-producing cell population.