IMMUNE RESPONSES OF HLA-HIGHLY-SENSITIZED AND NONSENSITIZED PATIENTS TO GENETICALLY-ENGINEERED PIG CELLS
We investigated in vitro whether HLA-highly-sensitized patients with end-stage renal disease (ESRD) will be disadvantaged immunologically after a genetically-engineered pig kidney transplant.Methods
Blood was drawn from patients with a cPRA 99-100% (Gp1, n=10) or cPRA 0% (Gp2, n=12), and from healthy volunteers (Gp3, n=10). Serum IgM and IgG binding was measured (i) to Gal and Neu5Gc glycans by ELISA, and (ii) to pRBC, pAEC, and pPBMC from GTKO/CD46 and GTKO/CD46/CMAHKO pigs by flow cytometry. (iii) T and B cell phenotypes were determined by flow cytometry, and (iv) proliferation of T and B cells CFSE-MLR.Results
(i) By ELISA, there was no difference in IgM or IgG binding to Gal or Neu5Gc between Gps1 and 2, but binding was significantly reduced in both groups compared to Gp3. (ii) IgM and IgG binding in Gps1 and 2 was also significantly lower to GTKO/CD46 pig cells than in healthy controls, but there were no differences between the 3 groups in binding to GTKO/CD46/CMAHKO cells. (iii and iv) Gp1 patients had more memory T cells than Gp2, but there was no difference in T or B cell proliferation when stimulated by any pig cells. The proliferative responses in all 3 groups were weakest when stimulated by GTKO/CD46/CMAHKO pPBMC.Conclusions
(i) ESRD was associated with low anti-pig antibody levels. (ii) Xenoreactivity decreased with increased genetic engineering of pig cells. (iii) High cPRA status had no significant effect on antibody binding or T and B cell response.