Immunological effects of serum antibodies in heart valve patients
2Department of Chemistry, University of California
3Complex Carbohydrates Research Center, The University of Georgia
4Genos, Glycoscience Research Laboratory
Animal-derived bioprosthetic heart valves (BHVs) are used to replace diseased heart valves, however, their susceptibility to structural valve deterioration through calcification limits their efficiency. BHVs contain the immunogenic glycans N-glycolylneuraminic acid (Neu5Gc) and galactose-α1,3-galactose (αGal). Previous studies revealed that BHVs recipients induce IgG against these foreign glycans within one-month post-implantation, and these antibodies (Abs) promote BHV calcification. Other studies suggest that the glycosylation of circulating serum Abs is altered in inflammatory conditions. We hypothesize that IgM isotype could also contribute to BHV calcification, and that glycosylation of serum Abs is altered in BHV recipients, reflects inflammatory responses leading to BHV calcification. We investigated IgM immune response in a large cohort of BHV recipients compared to mechanical heart valve or coronary artery bypass graft, before and after implantation. ELISA analysis showed that anti-Neu5Gc IgM or anti-Gal IgM levels did not significantly differ between BHV patients and control patients. To further examine glycosylation of serum Abs, the methodologies for Ab glycosylation analysis had been established, initially on a small set of patients` samples. Glycosylation of total serum and affinity-purified anti-Neu5Gc Abs were examined by printed microarrays using multiple glycan-binding lectins, and total IgG-Fc N-glycosylation examined by HILIC-UHPLC and by mass spectrometry IgG glycoproteomics. This initial glycosylation analysis revealed consistent glycosylation patterns by all these technologies, particularly detecting N-glycans with core fucose as the most abundant, as well as N-glycans with terminal sialic acid, galactose and mannose, yet each technology provides unique information that together build up the complete information on Ab glycosylation. These technologies will further be used to investigate the glycosylation of total serum samples, purified Ig isotypes, and affinity-purified anti-Neu5Gc Abs in BHV-recipients that have confirmed early signs of calcification compared to control patients over time. Altogether, these data suggest that anti-Neu5Gc IgM and anti-Gal IgM Abs do not significantly contribute to BHV calcification, in contrast to the role of IgG Abs. Further investigation of Ab glycosylation could potentially be used to monitor progression of BHV disease. In particular, unique Ab-glycosylation patterns could serve as potential markers for inflammation and early calcification in BHV-recipients compared to the control patients.