María José Castro

Preformed antibodies detected by cytotoxic assay or multibead array decrease liver allograft survival: Role of human leukocyte antigen compatibility

The significance of human leukocyte antigen (HLA) compatibility and preformed antibodies in liver transplantation remains unclear. The objectives of this study were to evaluate, in a single‐center cohort comprising 896 liver transplants, whether the degree of donor‐recipient compatibility and preformed antibodies modified graft survival. Univariate Kaplan‐Meier analysis demonstrated that donor‐recipient HLA compatibility had a marginal impact on allograft survival. As for compatibility at individual antigen loci, 2 mismatches at HLA‐A conferred a survival advantage in retransplanted allografts (P = 0.011). HLA‐B and HLA‐DR loci did not play a significant role in outcome in any pathology. The concordance of results on preformed antibodies detected by complement‐dependent cytotoxicity (CDC) and a multiple bead assay (Luminex® xMAP) showed a strong correlation between both techniques (P < 0.0001). Both CDC‐detected and Luminex‐detected antibodies were associated with shorter graft survival within the first year post‐transplant (P = 0.01 and P = 0.016, respectively). Positive CDC T crossmatches and Luminex‐detected HLA class II antibodies played a significant role in decreasing graft survival (P = 0.043 and P = 0.0019 at 1 year, respectively, and P = 0.005 and P = 0.038 at 5 years, respectively). A correlation was also observed between the presence of preformed Luminex‐detected class II or Luminex I and II antibodies and allograft rejection (P = 0.001 and P = 0.042, respectively). In conclusion, although HLA typing is not a prerequisite for transplantation, screening of HLA antibodies with Luminex techniques and CDC crossmatch may be useful in the detection of at‐risk patients that could benefit from increased surveillance and tailored therapy following transplantation. Liver Transpl, 2008.

Evolution of MHC-G in primates: a different kind of molecule for each group of species

When MHC-G molecules in primates (New World and Old World monkeys, Anthropoids and humans) were compared phylogenetically, very different evolutionary patterns within each species were found; their molecules did not have a straight forward and linear development throughout the postulated evolutionary pathway of primates. The earlier New World monkeys (South America) had relatively more alleles and the polymorphism was placed in the T-cell receptor (TcR), NK receptors and antigen binding sites; MHC-G probably works as a classical class I presenting molecule in these monkeys. MHC-G intron 2 from New World monkeys does not show the typical 23 bp deletion found in all other more recent primate species. Thus, it is possible that MHC-G molecules in New World monkeys belong to a different lineage than the MHC from higher primates. Another early lineage, Eurasian Old World monkeys, shows stop codons at exon 3: MHC-G proteins lacking the alpha2 domain may functionally suffice or otherwise reading-through stop-codon translational mechanisms may exist, as shown for other genes. Orangutans show lower (but significant) polymorphism than New World monkeys at NK, TcR and antigen binding regions; gorilla and chimpanzee show very low polymorphism. Humans only show three different HLA-G proteins with changes not affecting NK, TcR or antigen binding sites. It is observed that the more exposed the mother to allogeneic fetuses (polygamy), the less polymorphic HLA-G is observed within a given species. The data are concordant with the postulated immune inhibitory function for MHC-G in Old World monkeys, anthropoids and humans both at placental and inflammatory level.

Detection of HLA-E and -G DNA alleles for population and disease studies

Abstract. HLA-E and -G genes show a restricted polymorphism encoding for molecules whose variability is limited at the peptide binding site. Fourteen alleles that give rise to only three productive proteins for HLA-G (*0101, *0103 and *0104) and five alleles with three different proteins for HLA-E (*0101, *0102 and *0103) have been described. Expression of these molecules is low and found in many tissues for HLA-E; HLA-G protein is expressed in extravillous trophoblast cells and thymic epithelium. Molecular studies have shown how HLA-G and HLA-E bind to natural killer (NK) cells immunoglobulin and lectin-type inhibitory receptors. HLA-E may act as a sentinel of the cell; if classical class I and HLA-G are being expressed, HLA-E molecules may reach the cell surface and inhibit the lysis by NK cells. Most findings are consistent with the hypothesis that HLA-E and -G proteins may be tolerogenic molecules at either the T-cell receptor (TcR) (inflammation, graft rejection) or NK level, switching off cells which usually attack foreign (including foetus) or self (autoimmune) antigens. A low HLA-E and -G polymorphism is observed in humans, and their allele frequencies are mostly homogeneous in the populations tested so far. Many studies to detect these alleles are now being performed in isolated populations and also in pregnancy-associated pathologies. In the present paper, standard and detailed techniques to detect HLA-E and -G DNA polymorphism are reported and discussed.

Association of early kidney allograft failure with preformed IgA antibodies to β2-glycoprotein I.

In the current immunosuppressive therapy era, vessel thrombosis is the most common cause of early graft loss after renal transplantation. The prevalence of IgA anti-β2-glycoprotein I antibodies (IgA-aB2GPI-ab) in patients on dialysis is elevated (>30%), and these antibodies correlate with mortality and cardiovascular morbidity. To evaluate the effect of IgA-aB2GPI-ab in patients with transplants, we followed all patients transplanted from 2000 to 2002 in the Hospital 12 de Octubre prospectively for 10 years. Presence of IgA-aB2GPI-ab in pretransplant serum was examined retrospectively. Of 269 patients, 89 patients were positive for IgA-aB2GPI-ab (33%; group 1), and the remaining patients were negative (67%; group 2). Graft loss at 6 months post-transplant was significantly higher in group 1 (10 of 89 versus 3 of 180 patients in group 2; P=0.002). The most frequent cause of graft loss was thrombosis of the vessels, which was observed only in group 1 (8 of 10 versus 0 of 3 patients in group 2; P=0.04). Multivariate analysis showed that the presence of IgA-aB2GPI-ab was an independent risk factor for early graft loss (P=0.04) and delayed graft function (P=0.04). There were no significant differences regarding patient survival between the two groups. Graft survival was similar in both groups after 6 months. In conclusion, patients with pretransplant IgA-aB2GPI-ab have a high risk of early graft loss caused by thrombosis and a high risk of delayed graft function. Therefore, pretransplant IgA-aB2GPI-ab may have a detrimental effect on early clinical outcomes after renal transplantation.

Renal Transplantation Dramatically Reduces IgA Anti-beta-2-glycoprotein I Antibodies in Patients with Endstage Renal Disease

IgA anti-beta-2-glycoprotein I (aB2GPI) antibodies have been related to vascular pathology in the general population and mainly in hemodialyzed patients (prevalence 33%) in whom an elevated incidence of thrombosis and mortality is found. In this paper we have studied the presence of IgA aB2GPI antibodies at pretransplant and their evolution after transplantation with a cross-sectional-based follow-up study of a cohort of 288 endstage renal disease (ESRD) patients treated with kidney transplantation. Pretransplant IgA aB2GPI levels were elevated 31.7 ± 4.2 U/mL without differences in age or type of dialysis. Patients with different etiologies of ESRD showed higher levels of IgA aB2GPI than blood donors, except the groups of non-IgA glomerular disease and systemic erythematosus lupus, whose nonsignificant differences were observed. IgA aB2GPI antibodies dropped immediately after transplantation (10.7 ± 1.0 U/mL, P < 0.0001), coinciding with a high degree of immunosuppression, and remained significantly lower than that observed in pretransplant status. Prevalence of patients with elevated antibodies was also less in transplanted patients (8.9% versus 30.4%, P < 0.0001). Among, positivity for IgA aB2GPI was higher than in patients who had received their first transplant that those were retransplanted. This finding could have important clinical implications and can suggest new therapeutic strategies in patients with IgA aB2GPI antibodies.

TRANSCRIPTION AND WEAK EXPRESSION OF HLA-DRB6 : A GENE WITH ANOMALIES IN EXON 1 AND OTHER REGIONS

Abstract HLA-DRB6 is one of the human major histocompatibility complex (MHC) genes present in DR1, DR2, and DR10 haplotypes (approximately 26% of individuals). It shows several anomalies in human and non-human primates, including exon 2 stop codons (non-randomly grouped between codons 74 and 94) and a promoter region, and an exon 1 coming from an inserted retrovirus. It has been shown that not only chimpanzee but also human Mhc-DRB6 lack the usual 3’ untranslated (UT) polyadenylation signal, and in the present work it was found that the human DRB6 gene coming from an HLA-DR2 haplotype is effectively transcribed after transfection in mouse L cells, and that HLA-DRB6 molecules may be expressed on the cell surface. DRB6 transcription level is remarkably lower in human than in chimpanzee. Moreover, their exons 1 (both taken from the 3’LTR region of a mammary tumor retrovirus) are also different; this shows that these viral insertions may be an important mechanism for different evolutionary changes in orthologous genes of different species. The pathways by which DRB6 molecules may be expressed on the membrane are unclear but other examples of truncated protein expression have also been described, even within the human major histocompatibility complex (i. e., in HLA-G). Finally, the presence of mature HLA-DRB6 mRNA molecules supports the notion that splicing may take place even in the absence of a canonical 3’UT polyadenylation signal.

The Presence of Pretransplant Antiphospholipid Antibodies IgA Anti-β-2-Glycoprotein I as a Predictor of Graft Thrombosis After Renal Transplantation.

Background Vessel thrombosis is a severe complication after renal transplantation. Antibodies anti-&bgr;-2 glycoprotein-I of IgA isotype (IgA-aB2GP1) have been linked to thrombotic events and mortality in hemodialysis patients. Methods All kidney transplanted patients from 2000 to 2011 (n = 1375) in our hospital were followed up for 2 years, evaluating 3 time periods. Results At transplantation, 401 patients were positive for IgA-aB2GPI (29.2%, group 1), and the remaining patients were negative (group 2). Graft loss at 6 months posttransplantation was higher in group 1 (18% vs 7.2%; P < 0.001). The most frequent cause of early graft loss was vessel thrombosis, especially in group 1 (12.2% vs 2.6% of patients; P < 0.001). In fact, vessel thrombosis was the most important cause of graft loss in the 3 time periods, irrespective of demographic changes and introduction of transplantation with asystolic donors. Notably, IgA-aB2GP1 was an independent risk factor for graft thrombosis (odds ratio, 5.047; P < 0.001). Furthermore, the presence of IgA-aB2GP1 was associated with early graft loss and delayed graft function. Mortality at 24 months was also higher in group 1. Conclusions In conclusion, pretransplant IgA-aB2GP1 was the main risk factor for graft thrombosis and early graft loss. Further research should be made on whether anticoagulation in antibody-positive patients could ameliorate this catastrophic complication.

The evolution of the MHC-G gene does not support a functional role for the complete protein.

Summary: The study of the MHC‐G gene evolution during nearly 40 million years does not support a role for the full molecule. The MHC‐G‐like proteins of New World monkeys are probably classical presenting molecules. Old World Cercopithecinae monkeys do not have a full MHC‐G molecule and human individuals homozygous for the HLA‐G null allele are healthy and do not show birth pathologies.

Human leukocyte antigen haplotypes and HFE mutations in Spanish hereditary hemochromatosis and sporadic porphyria cutanea tarda

Background and Aims:  It has been postulated that the HFE C282Y mutation (linked to human leukocyte antigen [HLA]‐A3‐B7 haplotype) is not only responsible for hereditary hemochromatosis; HLA class I alleles would also contribute to the disease pathogenesis. In addition, H63D mutation linked to HLA‐A29‐B44 would also be pathogenetic, particularly in the Mediterranean Basin and throughout the world. However, sporadic porphyria cutanea tarda (s‐PCT) has also been linked to these HFE mutations. In the present work, we have studied HFE mutations and HLA genes to test these hypotheses.

An evolutionary overview of the MHC-G polymorphism: clues to the unknown function(s)

The functions of the major histocompatibility complex-G (MHC-G) molecule are still unknown. The idea that this molecule may be involved in preventing the rejection of fetuses is suggested by only indirect evidence. In the present paper, we review the structure, in vitro function and tissue distribution of MHC-G genes and proteins in different primate species. From available data, we conclude the following. First, the nomenclature of MHC-G alleles needs to be revised. Rhesus monkey A/G gene cannot be orthologous to MHC-G because of a lack of structural similarity. Cotton-top tamarin G molecules (which are also structurally similar to E molecules) cannot be orthologous to classical class I molecules. Second, selective pressure to maintain a low degree of polymorphism appears to operate only at the peptide-binding region (PBR) of the MHC-G molecule. Thus, this observation contradicts the idea that the MHC-G leader peptide is the only functional part of the molecule, and suggests that G proteins may have an antigen presenting function to clonotypic T cell receptors besides the ability to interact with NK receptors. Third, MHC-G may also have a function in the thymus, because it is expressed in the thymic epithelium. MHC-G may be important for creating an appropriate T-cell repertoire. Fourth, the presence of HLA-G proteins in tumours and the specific absence of soluble HLA-G mRNA isoforms in the tissues taken from patients with autoimmune diseases suggest that MHC-G may have a role in the immune response and inflammation control.