Postdoctoral research fellow
Thomas E. Starzl Transplantation Institute
Investigation of the potential clinical significance of the expression of NeuGc on pig aortas and corneas
Whayoung Lee1, Cassandra Long 1, Burcin Ekser2, Eric Walters3, Jagdeece Ramsoondar 4, David Ayares4, A. Joseph Tector 2, David K. C. Cooper 1, Hidetaka Hara 1.
1Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsuburgh, PA, United States; 2Department of Surgery, Indiana University School of Medicine, Indianapolis, IN, United States; 3National Swine Resource and Research Center, Columbia, MO, United States; 4Revivicor, Blacksburg, VA, United States
Purpose: The recent introduction of pigs expressing neither galactose-α1,3-galactose (Gal) nor N-glycolylneuraminic acid (NeuGc) has taken xenotransplantation one step closer to the clinic. Lack of NeuGc expression has been shown to be associated with reduced human antibody binding to pig peripheral blood mononuclear cells (PBMCs), but binding to pig aortas or corneas or to cultured cells has not been examined. Our aims were (i) to confirm the lack of NeuGc expression on vascular (aorta) and avascular (cornea) tissues and on cultured cells (aortic endothelial cells [AECs]; corneal endothelial cells [CECs]) of GTKO/NeuGcKO pigs, and (ii) to investigate whether the absence of NeuGc reduced human antibody binding to the tissue and cells.
Methods: Wild-type (WT) pigs (n=2), GTKO pigs from different sources (Revivicor [n=3]; National Swine Resource and Research Center [NSRRC, n=2]), and GTKO/NeuGcKO pigs from different sources (Revivicor [n=8]; Indiana University School of Medicine [n=2]) were used for the study. Human tissue and cultured cells were used as negative controls. Immunofluorescence staining was performed using anti-Gal and anti-NeuGc antibodies, and to determine human IgM and IgG binding to aortic and corneal tissue. Flow cytometric analysis was used to determine Gal and NeuGc expression on cultured AECs and CECs and to measure human IgM/IgG binding to these cells.
Results: Both Gal and NeuGc were detected on WT pig aortas and corneas. Tissues and cells from distinct GTKO pigs lacked Gal expression, but demonstrated different expression levels of NeuGc (NSRRC > Revivicor). There was no significant difference in human antibody binding to the AECs or CECs from these two pigs, regardless of the variable NeuGc expression. Neither human nor GTKO/NeuGcKO pigs expressed Gal or NeuGc. Human IgM and IgG antibody binding to aortas and corneas from all GTKO and GTKO/NeuGcKO pigs was reduced compared to binding to those of WT pigs. Nevertheless, a low level of human antibody binding was detected even to GTKO/NeuGcKO aortas and corneas. Human antibody binding to GTKO/NeuGcKO AECs was significantly less than to GTKO AECs (p<0.05), but there was no significant difference in human antibody binding between GTKO and GTKO/NeuGcKO CECs (possibly due to the relatively low expression of NeuGc on cultured CECs compared to AECs).
Conclusions: (i)The absence of NeuGc on GTKO aortic tissue and AECs is associated with reduced human antibody binding, and possibly will provide better outcome in clinical xenotransplantation using vascularized organs. (ii) In contrast, for clinical corneal xenotransplantation, the absence of NeuGc expression on GTKO/NeuGcKO pig corneas may not prove an advantage over GTKO corneas. (iii) Efforts should be made to identify potential antigens other than Gal and NeuGc on pig aortas and corneas.
08:00 - 09:00
|Progress in Tissue & Cell Xenotransplantation||Heart valves||Room 110|
15:30 - 16:30
|Non-Gal Antibodies||Investigation of the potential clinical significance of the expression of NeuGc on pig aortas and corneas||Room 109|
15:30 - 16:30
|Non-Gal Antibodies||The potential significance of NeuGc in the structural valve deterioration of bioprosthetic heart valves||Room 109|