The development of transgenic pigs has revolutionized the field of Xenotransplantation (XTx) improving the survival of pigs to baboons XTx.
Although recent pig to human XTx has been performed, there are few opportunities in the clinical setting to analyze the immune response humans develop after exposure to pig organs or tissues.
A better understanding of the pig to human immune response, will be a step toward the clinical application of XTx.
Aim: Given the importance of Xenoantibodies (XAb) in initiating xenograft rejection, we have analyzed in patients exposed to pig hepatocytes following Bioartificial Liver (BAL) treatment: 
1) the long-term XAb response to pig endothelial cells (PEC) from wild-type (wt) and transgenic pigs lacking the Gal epitope (GalT-KO)
2) the cytotoxicity of patients' plasma to wt and GalT-KO PEC
Material & Methods: The BAL was used as a support treatment in patients with acute liver failure (ALF) awaiting liver transplantation (LTx) and consists of patients’ plasma perfused through a hollow-fiber cartridge containing porcine hepatocytes. 
Six long-term follow-up ALF patients treated with BAL were included in this study. Each patient received 1 to 6 BAL treatments; 3 patients subsequently received a LTx, the other patients recovered without the need of LTx. 
Patients' plasma collected before (day 0) and at different time-points post-BAL treatment (up to 1 year) was analyzed by ELISA for IgG and IgM XAb binding to PEC from wt and GalT-KO pigs, and to bovine thyroglobulin. The cytotoxicity of patients' plasma was analyzed by FACS using  wt and GalT-KO PEC. 
Results: Anti-pig IgG and IgM XAb to wt, GalT-KO PEC, and thyroglobulin were present in patients' plasma before pig exposure (day 0), with high XAb variability among patients. Repetitive BAL treatments were associated with a decreased IgG/IgM XAb, indicating that XAb were absorbed during BAL treatments. A marked increase of XAb was observed by 1-2 weeks post-BAL treatments followed by a gradual decrease over time; the increase being more pronounced for anti-Gal XAb. 
The cytotoxicity assay showed that patient's plasma on day 0 has a degree of cytotoxicity to wt PEC and minimal cytotoxicity to GalT-KO. By 1-2 weeks post-BAL a sharp increase of cytotoxicity to wt was observed, but minimal cytotoxicity to GalT-KO-PEC.
The high anti-pig XAb did not increase the incidence of allograft rejection in patients who had a subsequent LTx.
Conclusions
- Patients' plasma contains both IgG and IgM XAb that bind to Gal and non-Gal epitopes on wt and GalT-KO PEC. - An immediate XAb absorption was observed during repetitive BAL treatments, followed by an increase IgG and IgM XAb titers by 1-2 weeks post-BAL. 
- Human XAb produced in response to pig hepatocytes exposure are cytotoxic to wt PEC; however low cytotoxicity was observed toward GalT-KO PEC.
- The 3 patients who received a subsequent LTx after BAL treatments, did not have allograft rejection despite the high anti-pig XAb titers.