Detection of Hanganutziu-Deicher antigens in O-glycans from pig heart tissues by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
BACKGROUND
In the α1,3-galactosyltransferase knockout (α-GalT KO) pig era, identification of the non-Gal epitopes is necessary for successful pig-to-human xenotransplantation. Recently, we successfully detected α-Gal epitopes as well as Hanganutziu-Deicher (H-D) antigens from the N-glycans in the pig heart tissues, which have been considered as promising non-Gal antigens. However, the profiling of O-glycan from pig heart tissues had not been performed owing to the difficulty of O-glycan preparation.


METHODS
In this study, we established the simple and sensitive method to profile O-glycans from pig heart aortic valve, aortic wall, pulmonary valve, pulmonary wall, and cardiac muscle tissues. To liberate O-glycans from the pig heart tissues, we used non-reductive β-elimination reagent and subsequently purified the glycans. After permethylation, the glycans were qualitatively analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS).


RESULTS
The comprehensive O-glycan analysis method was successfully validated using model glycoproteins such as bovine serum fetuin (BSF) and bovine submaxillary gland mucin (BSM) glycoproteins, and their O-glycan profiles were in accordance with the data of previous studies. Next, we applied the method for O-glycan release and characterization to analysis of various pig heart tissues. As a result, total 39, 33, 24, 36, and 25 of O-glycans were detected from aortic valve, aortic wall, pulmonary valve, pulmonary wall, and cardiac muscle, respectively. Furthermore, four in aortic valve, one in aortic wall, one in pulmonary valve, one in pulmonary wall, and one in cardiac muscle were particularly determined as terminally N-glycolylneuraminic acid-linked O-glycans, which is considered to be the H-D antigens.


CONCLUSIONS
Here, we initially described the O-glycan structures of various pig heart tissues, and additionally, the existence of H-D antigen type O-glycans was firstly identified. These results will be fundamental information for overcoming the xenoantigenic carbohydrate-related immunological rejection in pig-to-human heart tissue xenotransplantation.
['Jeong|Hee-Jin|HJ|', 'Adhya|Mausumi|M|', 'Park|Hae-Min|HM|', 'Kim|Yun-Gon|YG|', 'Kim|Byung-Gee|BG|']
D000818:Animals D015478:Antigens, Heterophile D002240:Carbohydrate Sequence D002417:Cattle D060748:Fetuins D064593:Heterografts D006801:Humans D008969:Molecular Sequence Data D015394:Molecular Structure D009077:Mucins D009206:Myocardium D011134:Polysaccharides D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization D034421:Sus scrofa D014181:Transplantation Immunology
Hanganutziu-Deicher antigen O-glycans heart matrix-assisted laser desorption/ionization time-of-flight mass spectrometry xenotransplantation