# Reduction in Xenogeneic Epitopes on Porcine Endothelial Cells by Periodate Oxidation

## TL;DR

This paper explores using periodate oxidation to reduce immune-triggering sugars on pig cells for safer human xenotransplantation.

## Contribution

A chemical method using sodium periodate is proposed to reduce xeno-reactive glycan epitopes on porcine cells.

## Key findings

- Periodate oxidation reduced αGal epitope binding and human antibody reactivity on porcine endothelial cells.
- Cell viability was preserved when using 2 mM NaIO4 at 4°C for 60 minutes.
- Oxidation was effective under both static and flow conditions without affecting cell survival.

## Abstract

Background: Patterns of humoral immune responses represent a major hurdle in terms of pig-to-human xenotransplantation approaches. The best-known xenogeneic glycan antigens present in pigs are the αGal (Galili antigen) and the non-human sialic acid Neu5Gc. As there are further differences between porcine and human cellular surface glycosylation, a much broader range of glycan epitopes with xeno-reactive relevance can be anticipated. Therefore, we set out to chemically modify porcine cellular surface glycans in a global approach by applying sodium periodate (NaIO4) oxidation. Methods: Porcine endothelial cells were exposed to oxidation with 1 to 5 mM NaIO4 for different time periods at 37 °C or 4 °C and under static or dynamic conditions. The impact on cellular survival was determined by applying live/dead assays. Oxidation of αGal-epitopes was assessed by fluorescence microscopy-based quantification of isolectin-B4 (IL-B4) staining. Overall immunogenicity of porcine cells was determined by human serum antibody binding. Results: Treatment of porcine endothelial cells and tissues with NaIO4 led to reduced binding of the αGal-specific IL-B4 and/or human serum antibodies. NaIO4 was revealed to be cytotoxic when performed at elevated temperatures and for a prolonged time. However, by applying 2 mM NaIO4 for 60 min at 4 °C, a high extent of cellular viability and a relevant reduction in detectable αGal epitope were observed. No differences were detected irrespectively on whether the cells were oxidized under static or flow conditions. Conclusions: Glycan epitopes on living cells can be oxidized with NaIO4 while maintaining their viability. Accordingly, this strategy holds promise to prevent immune reactions mediated by preformed anti-glycan antibodies.