L.P. de Waal

Pretransplantation Blood Transfusion Revisited

BACKGROUND Blood transfusion before organ transplantation has a beneficial effect on allograft survival; the mechanism of this effect has remained a mystery. In murine models, the presence of common histocompatibility antigens in the blood donor and the recipient favors the induction of allograft tolerance. METHODS To investigate the effect of HLA compatibility between blood donor and recipient on the induction of allograft tolerance, we determined the relative frequency of cytotoxic T-lymphocyte precursors specific for donor cells before and at several times after blood transfusion in 23 patients awaiting a first renal transplant. We correlated the results with the presence of shared HLA antigens. RESULTS T-cell nonresponsiveness against donor cells developed after blood transfusion in 10 of the 23 patients. Tolerance developed only if the blood donor and the recipient had one HLA haplotype or at least one HLA-B and one HLA-DR antigen in common (as was observed in 9 of these 10 patients). Tolerance developed relatively late after blood transfusion (one to two months) and was long-lasting. No decline in the T-cell response against donor alloantigens was observed in any of the 13 patients who received transfusions without having HLA-antigen compatibility with the donor. CONCLUSIONS Blood transfusion in which there is a common HLA haplotype or shared HLA-B and HLA-DR antigens induces tolerance to donor antigens. This finding may lead to the development of new strategies with which to induce tolerance for transplantation after blood transfusion. Perhaps transplant donors will be selected not only by HLA-antigen matching, but also on the basis of acceptable HLA-antigen mismatches associated with T-cell non-responsiveness induced by selected blood transfusion.

IgG antibodies against an HLA antigen are associated with activated cytotoxic T cells against this antigen, IgM are not

Pre-existing alloantibodies against the mismatched HLA-A and HLA-B antigens of the donor, when present in current sera, are believed to be detrimental to kidney graft survival. When these antibodies are present only in historical sera, their immunoglobulin class has reported to be important with respect to the expected graft survival, IgG antibodies being associated with poor graft prognosis and IgM with reasonable graft survival. In the present study, we have tested whether the immunoglobulin class of anti-HLA antibodies is reflected in the activation state of CTLs directed against these HLA antigens as measured by their in vitro resistance or sensitivity to CsA. The results indicate that the presence of IgG anti-HLA antibodies is associated with the presence of activated CTLs (CsA resistant), whereas in the case of IgM antibodies, mainly naive CTLs (CsA sensitive) are found. This observation may explain the different prognoses of historical positive crossmatches due to IgG versus IgM alloantibodies.

HLA typing in congenital toxoplasmosis.

HLA-A, HLA-B, HLA-C, and HLA-D typing was performed in 47 mothers of patients suffering from ocular toxoplasmosis to investigate whether an immunogenetic predisposition exists for developing congenital toxoplasmosis in their offspring. No significant association between any HLA antigen was observed in the mothers of patients with ocular toxoplasmosis, although a total absence of the HLA-B51 antigen was found in this group. HLA-A, HLA-B, and HLA-C typing was also performed in their children (52 patients with ocular toxoplasmosis), to investigate a possible relation between the severity of ocular toxoplasmosis and an eventual immunogenetic factor. In the patients with ocular toxoplasmosis an increased frequency of the HLA-Bw62 antigen was observed in correlation with severe ocular involvement.

Familial differences in endotoxin-induced TNF release in whole blood and peripheral blood mononuclear cells in vitro; relationship to TNF gene polymorphism

Healthy volunteers show large interindividual differences in endotoxin-induced TNF release in vitro and certain HLA class II types can be related to phenotypic TNF resporise. To investigate the possibility of a genetic basis for endotoxin responsiveness, we tested TNF release in whole blood and PBMNC after stimulation by endotoxin in 47 relatives of 7 healthy volunteers. All volunteers were HLA-typed and the TNF-gene associated Nco1 and DNA microsatellite polymorphisms were determined. A significant difference in TNF release by PBMNC and the Nco1 genotype could be established, showing a lower response of TNFB*2 homozygotes than TNFB*1 homozygotes (165 vs 265, 413 vs 703 and 462 vs 832 pg/106 PBMNC for 1, 10 and 100 ng/ml of endotoxin respectively; P < 0.05). The highest endotoxin-induced TNF release was observed in TNFB*1/TNFB*2 heterozygotes (340, 911 and 1,149 pg/10 6 PBMNC respectively; P < 0.05 compared to TNFB*1 homozygotes and P < 0.0005 when compared to TNFB*2 homozygotes). TNFa and TNFb microsatellite typing revealed extensive polymorphism, showing a significantly lower TNF release in whole blood in individuals with TNFa2, -a6 and -a10 alleles than in individuals with TNFa4 and -a11 microsatellite haplotypes.

High-affinity cytotoxic T lymphocytes after non-HLA-sharing blood transfusion--the other side of the coin.

Previously, we have shown that pretransplantation blood transfusion modulates the T cell repertoire to a great extent. Patients receiving a BT from a donor sharing one HLA haplotype with the patient (HLA-sharing BT) develop CTL nonresponsiveness against cells of the BT donor and show a selective decrease in the usage of T cell receptor V beta families. The present study has focused on the analysis of the T cell repertoire in patients receiving an HLA mismatched (non-HLA-sharing) BT. CTL precursor frequencies were measured against single class I-mismatched antigens in split-well analysis. In addition, blocking studies of CTL-target cell interaction were performed with anti-CD8 monoclonal antibodies. The results demonstrate that non-HLA-sharing BT immunizes and induces the generation of CD8 independent, high-affinity CTL against immunogenic class I-mismatched antigens. Such HLA class I antigens might become nonacceptable mismatches in subsequent organ transplantation.

Comparison Of T Cell Responses In Patients With A Long-term Surviving Renal Allograft Versus A Long-term Surviving Liver Allograft: It's a Different World

The aim of the present study was to analyze whether acquired transplantation tolerance had developed in patients with a long-term surviving renal or liver allograft. Analysis of antidonor cytotoxic T cell precursor frequencies was performed in 31 renal allograft recipients and 9 liver allograft recipients with good graft function 2 years after transplantation. The results demonstrated that, before transplantation, normal antidonor T cell responses were generated in both groups of patients. Two years after transplantation, donor-specific CTL nonresponsiveness had developed in a minority of the renal transplant recipients. In contrast, 8 out of 9 liver transplant recipients showed donor-specific mixed lymphocyte culture and CTL nonresponsiveness. These findings indicate that development of donor-specific T cell nonresponsiveness is not a common event after kidney transplantation, whereas liver transplantation seems to induce, at least in vitro, a state of donor-specific T cell nonresponsiveness.

Frequency analysis of HLA-specific cytotoxic T lymphocyte precursors in humans.

Frequencies of HLA class 1-specific cytotoxic T lymphocyte precursors (CTLp) from 33 responders were determined in 115 responder/stimulator combinations. In each combination there was a single HLA-A or HLA-B antigen mismatch. A wide range of CTLp frequency (CTLpf) values was found for most A and B locus antigens. Some A locus antigens appeared less immunogenic than other A locus antigens. The effect of additional C locus differences was negligible. The relationship between responder and stimulator HLA antigens is of minor importance because HLA-specific CTLpf against crossreactive (CREG) and subtype antigens were not significantly lower than CTLpf against non-CREG antigens. The CTLpf did not correlate with Bw4 or Bw6 mismatches. The existence of a broad range of values for HLA class I-specific CTLpf is of general interest. We have arbitrarily subdivided the CTLpf values into high, medium, low, and very low. In about 20% of the combinations the HLA-specific CTLpf were low or not even detectable in our assay. In contrast, HLA-specific CTLpf in combinations with multiple HLA antigen differences were regularly high. Our results confirm the high values of allospecific CTLpf in general but simultaneously point to unexpected variations. Frequency analysis of HLA-specific CTLp may be considered a new parameter in clinical transplantation for the selection of appropriate donor/recipient pairs.

Haematopoietic stem cells can induce specific skin graft acceptance across full MHC barriers.

Previously, we and others have demonstrated in several animal models that the establishment of stable haematopoietic chimerism through allogeneic bone marrow transfusion provides an effective means for the development of specific transplantation tolerance. However, a major limitation to the clinical application of allogeneic bone marrow transfusion in immunosuppressed recipients for induction of tolerance to solid grafts, is the risk of graft-versus-host disease (GVHD). Therefore, it is important to identify the cell population needed for the induction of mixed chimerism and tolerance. Haematopoietic stem cells have the capacity of self-renewal and multilineage differentiation, and have been shown to reduce the risk of GVHD. We studied transfusion of two rich sources of stem cells, namely allogeneic fetal liver cells and a subset of purified bone marrow-derived progenitor cells (c-kit+) into anti-T cell monoclonal antibody-treated, low-dose irradiated recipient mice. Our data revealed that stable multilineage mixed chimerism and permanent donor-specific tolerance for skin, even when transplanted directly following conditioning, can be successfully achieved in this way, with no signs of GVHD.

5' regulatory nucleotide sequence of an HLA-A*0101null allele.

Abstract We have previously demonstrated an HLA-A*0101null allele segregating in a family with the HLA-B8, -Cw7, -DR3, -DR52, -DQ2 haplotype. In the present study the regulatory elements with known transcription enhancement activity of the silenced HLA-A*0101 allele were analyzed. In the enhancer B element, a T was substituted for a C at position  – 106, whereas no other alterations were found in the adjacent 5′ section of the HLA-A*0101null allele. This substitution was not seen in the enhancer B elements of the corresponding genes involved in normal HLA-A*0101 membrane expression. Comparison of enhancer B element sequences of classical functional major histocompatibility complex (MHC) class I alleles demonstrated a high degree of conservation. In contrast, many MHC class I pseudogenes showed mutation in their enhancer B boxes. These results may indicate that the single mutation detected in the enhancer B element plays a pivotal role in the abolishment of membrane expression of the HLA-A*0101null allele.

A new public antigen shared by all HLA-A locus products except HLA-A23, -A24, -A32, and -A25 is probably influenced by the amino acid residue at position 79 in the α1 domain

One of the most striking features of HLA gene products is their high degree of polymorphism. The most effort was aimed at the definition of private HLA antigens, and as a result, almost a hundred HLA alleles received official recognition by the WHO Nomenclature Committee. However, thus far little attention has been focused on the definition and the biological relevance of HLA antigenic determinants, shared among various HLA products. Public HLA specificities have been defined by pregnancy sera and by monoclonal antibodies (mAb) (Konoeda et al. 1986, Parham et al. 1986, Spear et al. 1985). An important reason for defining public HLA specificities is the recognition that public specificities represent polymorphic epitopes that can function as alloantigens. This would have important implications for the study of HLA and disease associations and for transplantation studies (Konoeda et al. 1986). Two of the public HLA determinants first described were the Bw4 and Bw6 specificities (Van Rood 1962). Originally, they were identified by pregnancy sera as products of a diallelic system in genetic linkage with the HLA-B-locus-encoded alloantigens. It was shown that these public determinants were spatially distinct from the private determinants of the H L A B locus alleles (Parham 1983). More recently, it became evident that the expression of Bw4 and Bw6 epitopes is not strictly limited to HLA-B locus antigens. The Bw4 epitope appears to be present on HLA-A23, HLA-A24, and HLA-A32 molecules as well, and HLA-Cw3 carries a determinant related to Bw6 (Muller et al. 1982, Kostyu et al. 1980). In the present study, we describe a new public HLAspecificity recognized by a mouse lymphocytotoxic mAb. This mAb, 3Gl l , is of the IgM isotype and recognizes a determinant shared by all HLA-A locus products with the exception of the Bw4-associated HLA-A antigens.