ABSTRACT
Objective To investigate the extraction and purification methods of serum specific endothelial cell antibody of renal transplant recipients with rejection after renal transplantation.Methods Human umbilical vein endothelial cell (HUVEC)was isolated and cultured.The serum samples of the renal transplant recipients with poor renal function after renal transplantation were collected.Specific endothelial cell antibody was screened out with flow cytometry;antibodies against human leukocyte antigen (HLA)and major histocompatibility complex class Ⅰ-related chain A (MICA)were detected by Luminex platform.After the existence of specific endothelial cell antibody in the serum sample was confirmed,specific endothelial cell antibody was absorbed with HUVEC.The cell was washed and then the absorbed antibody was eluted from the cell membrane.Antibody IgG in the eluent was purified and concentrated again with Protein-A /G magnetic beads.Antibody activity in the eluent was detected by flow cytometry and the purified specific endothelial cell antibody (IgG)was identified by SDS-polyacrylamide gel (SDS-PAGE)and Western blot.Results In the serum of 386 renal transplant recipients,the serum samples of 5 renal transplant recipients with serum creatinine (Scr) >400 μmoI /L,negative anti-HLA antibody,negative anti-MICA antibody and median fluorescence intensity (MFI) >16 were selected.Purified specific endothelial cell antibody IgG showed immunoglobulin heavy chain (purity > 95%)by SDS-PAGE gel.Flow cytometry showed that the purified antibody had the feature of rebinding with the surface antigen of vascular endothelial cell.Conclusions The purification method of using human umbilical vein endothelial cell to absorb specific endothelial cell antibody in the serum of renal transplant recipients may obtain good effect.
ABSTRACT
Organ transplantation is the effective method to replace the function of the patient failed organ. But it is very disappoint that recipients have to receive the long-term immunosuppression regimens for prevention of allograft rejection. To induce allograft immune tolerance without immunosuppressant is in great demand. Although several tolerance strategies for organ transplant have been proposed, even some has already been tested in the 1st clinical trial, these strategies haven ' t approached to ideal efficacy. Helminths are remarkably successful parasites to achieve immunological tolerance to host immune response. It is now well established that the parasites′ success is the result of active immunomodulation of their hosts ' immune response. We suggest that injecting B cells from donor spleen and helminth soluble antigens, recipient might become tolerance to donor organ, but not tolerated to other antigens. Research based on this approach has great translated value for future clinic practice.
ABSTRACT
<p><b>OBJECTIVE</b>To review the role of polymorphism of major histocompatibility complex class I-related chain A (MICA) gene and antibodies against MICA antigens in transplant immunology.</p><p><b>DATA SOURCES</b>The data used in this review were mainly from our own results and from the relevant English language literatures published from 1999 to 2010. Some data presented in this review are in press.</p><p><b>STUDY SELECTION</b>Articles regarding MICA gene discovery and pioneering finding of antibodies against MICA antigen and allograft rejection were selected. This review chronicles the development of our understanding of the role that MICA antigens and antibodies may play in organ transplantation.</p><p><b>RESULTS</b>Polymorphic glycoprotein MICA antigens were detected on freshly isolated human umbilical cord endothelial cells, but not on peripheral lymphocytes. Antibodies were found and typing of recipients and donors by sequencing the MICA alleles has established that de novo antibodies produced in kidney transplant recipients are directed at mismatched MICA epitopes and are associated with acute rejection and chronic transplant failure. The specificity of antibodies against the epitopes of MICA antigens were well characterized by donor MICA typing, single antigen array testing with antibody absorption and elution. Acute graft-versus-host disease was observed in stem-cell recipients who were mismatched for MICA.</p><p><b>CONCLUSIONS</b>Immunization against mismatched MICA epitopes encountered in donor organs after transplantation may result in antibodies against MICA alleles. Testing for MICA donor-specific antibodies (DSA) which are associated with early failure of kidney transplants may be helpful for identifying some of the targets of antibodies against antigens other than the human leukocyte antigen (HLA) and for improving transplantation outcome.</p>