Biological activity of pig islet-cell reactive IgG antibodies in xenotransplanted diabetic patients.
BACKGROUND
The IgG antibody response in type I diabetic patients, transplanted with fetal pig islet-like cell-clusters, was investigated using purified immunoglobulin G (IgG) fractions from sera collected 7 to 9 yr after transplantation. From our earlier studies, we knew that the immunological specificities of xenoreactive IgG1 and IgG2 antibodies are different, and that IgG1 antibodies, in contrast to the IgG2 population, are mainly directed against non-Galalpha1,3Gal epitopes. In this study our goal was to establish whether xenoreactive IgG1 and IgG2 antibodies react with pig islet cells and, if so, to identify the target cell type, the biological function as well as the specificity of such antibodies for islet cell antigens. Sera from xenotransplanted patients were compared with those of patients with diabetes, selected for high titres of islet-cell specific autoantibodies.


METHODS
IgG antibody fractions from patient sera were purified on a protein G column. Surface expression of target antigens was studied using flow cytometry as well as immunofluorescence microscopy. The biological function of islet-cell reactive sera was tested using antibody dependent cellular cytotoxicity with both xenogeneic adult pig islet cells and allogeneic human islet cells as targets. Antibody specificity was assessed using 2D Western blots with both fetal and adult pig islet as well as human islet cell antigenic preparations.


RESULTS
Some of the diabetic patients, who have been transplanted with xenogeneic fetal pig islet cells, continue to produce xenospecific IgG1 and IgG2 antibodies for 7 to 9 yr post-transplantation. A separate analysis of IgG1 and IgG2 antibodies showed that IgG1 antibodies react with pig islet beta cells, whereas IgG2 antibodies mainly react with non-endocrine pig cells. Such antibodies are xenospecific, as they were found to mediate antibody-dependent cellular cytotoxicity of adult pig, but not human islet target cells. The reverse was true for antibodies from non-transplanted diabetic patients with high titres of autoantibodies against beta cells. Fluorescence analysis as well as 2D gel Western blots revealed that the reactivity was variable between patient samples, indicating that target antigens for non-Galalpha1,3Gal-specific antibodies are heterogeneous.


CONCLUSIONS
Thus, xenotransplantation of diabetic patients induces islet-beta cell reactive xenospecific IgG1 antibodies, which are biologically active and can mediate antibody-dependent cellular cytotoxicity of pig islet cells.
Division of Clinical Immunology, Department of Laboratory Medicine, Karolinska Institutet, Huddinge, Sweden. ellinor.lindeborg@labmed.ki.se
Ellinor
E
Lindeborg
Makiko
M
Kumagai-Braesch
Annika
A
Tibell
Birger
B
Christensson
Erna
E
Möller
D000818:Animals; D000910:Antibodies, Heterophile / Q000097:blood*; D001323:Autoantibodies / Q000097:blood; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology* / Q000601:surgery*; D006085:Graft Survival; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood*; D016381:Islets of Langerhans Transplantation / Q000276:immunology* / Q000473:pathology; D013552:Swine; D013997:Time Factors; D014183:Transplantation, Heterologous / Q000276:immunology* / Q000401:mortality / Q000473:pathology
D000910:Antibodies, Heterophile; D001323:Autoantibodies; D007074:Immunoglobulin G; C024211:islet cell antibody