Acquired haemolytic anaemia due to isohaemagglutinins of donor origin following ABO-minor-incompatible kidney transplantation.

Of 1041 renal allograft transplantations performed in our centre, 142 (13.6%) were carried out with ABO-minor-incompatible kidneys. Anti-recipient ABO antibodies were found in two of 34 patients treated with cyclosporin. These two cases of severe but self-limited haemolysis due to anti-A1 and anti-B, respectively, are reported in detail. Among 108 azathioprine-treated patients no evidence of the disorder was found. The condition should be suspected in any recipient with an unexpected reduction in haematrocrit or other signs of haemolysis after ABO-minor-incompatible organ transplantation.

Lymphocytotoxic cross-matching performed on spleen cells: immunomagnetic technique versus current KN (Kissmeyer-Nielsen) technique.

This paper compares the immunomagnetic (IM) and the KN lymphocytotoxic cross-matching techniques. Only sera from patients with panel reactive antibodies and spleen cells from cadaveric donors were used. A panel study involving 60 combinations revealed 11 (18%) discrepancies. Four T-cell cross-matches were KN negative while IM positive. Inversely, six T-cell cross-matches were IM negative while KN positive (two) or doubtfully positive (four). Regarding HLA class II antibodies, one combination was found IM positive but KN negative. Out of 106 cadaveric renal allograft transplantations performed during 1987 with assistance from our laboratory, IM cross-matching was retrospectively performed on 33 KN T-cell cross-match negative donor-recipient pairs: two combinations were found T-cell cross-match positive. The corresponding allografts were not lost owing to hyperacute rejection involving performed antibodies. We recommend the IM technique for HLA typing, but more experience needs to be gained before we can recommend the technique for routine cross-matching prior to transplantation.

A standard microcytotoxicity technique for quantitative analysis of lymphocyte subsets. A comparison with indirect immunofluorescence, evaluated by microscopy or flow cytometry.

A standard complement-dependent microcytotoxicity (CDC) technique was used for quantitative analysis of T-lymphocyte subsets in human peripheral blood and the results compared to those obtained by indirect immunofluorescence microscopy and flow cytometry. The monoclonal antibodies OKT3, OKT4 and OKT8 were used in the CDC method for detection of total-T cells, T-helper and T-suppressor cells respectively. The CDC technique provided reproducible results (CV, 3-7%) correlating well with both immunofluorescence techniques. This observation was valid both for healthy persons (n = 21) and for patients (n = 10) with immunological disorders. The correct antibody dilution, correction for background and the use of eosin staining are considered critical for the usefulness of this technique. The method has several advantages: it is widely used for histocompatibility testing, only simple equipment is necessary, and the amount of monoclonal antibody required per test is small.

Immunogenetic composition of 94 consecutive Danish families investigated with respect to bone marrow transplantation.

The immunogenetic composition of 94 patients needing bone marrow transplantation and their core families primarily investigated to select family bone marrow donors have been further analysed to test for association between disease and HLA-region markers. From this material it is shown, that in the primary immunogenetic analysis of the family, inclusion of mixed lymphocyte culture analysis increases donor possibilities by approximately 14% when a reciprocal negative MLC response and phenotypic HLA-DR compatibility are accepted as criteria for transplantation. Further, the results indicate, that no association between HLA and leukemia seems to exist.

Plaque-forming cells in man. III. Generation of plaque-forming cells in allogeneic in vitro cultures of HLA-D/DR-incompatible B and T lymphocytes.

We have investigated the ability of allogeneic, irradiated T lymphocytes to induce proliferation and immunoglobulin (Ig) secretion in untreated peripheral blood B lymphocytes. Non-mitogen-activated co-cultures of isolated T and B lymphocytes from selected, full-house HLA-A,B and D/DR antigen-phenotyped donors were reconstituted in a ratio of 4:1. Proliferation was assessed on day 5-6 of culture by the 3H-thymidine incorporation technique, and the Ig secretion was monitored on day 6 with a protein A plaque-forming cell (PFC) assay. B lymphocytes were able to differentiate into PFC, and the number of plaques was significantly higher in cultures of cells with two HLA-D/DR antigen incompatibilities than in those sharing one antigen. In cultures of peripheral blood lymphocytes with no HLA-D/DR antigen difference, only a few PFC developed. HLA-A and B antigens had no influence on the response. Further, monocytes were not an absolute requirement for allogeneic activation of B cells. Sonicated T cells and culture supernatants from allogeneic T- and B-cells cultures were not able to induce PFC formation in B lymphocytes. Our results indicate that the PFC response obtained in non-mitogen-activated cultures of allogeneic T and B lymphocytes is dependent on HLA-D/DR disparity or on genes encoded in the HLA-D/DR region.

Cell-mediated PPD-specific cytotoxicity against human monocyte targets: evidence for restriction by class II HLA antigens.

Human Purified Protein Derivative of tuberculin- (PPD-) specific cytotoxic cells have been detected in cultures of peripheral blood mononuclear cells stimulated for 6 days with PPD. These cytotoxic cells are demonstrated by their ability to lyse PPD-pulsed autologous monocyte target cells, but not unpulsed targets. In a series of checkerboard experiments each involving 3-5 randomly combined donors, effector cells from 35 donors have been tested in autologous and 130 allogeneic combinations. Analysis of results from the pooled allogeneic combinations reveals that HLA-B - and even more pronounced HLA-DR - antigen sharing correlates positively to high lysis. No effect of HLA-A antigen sharing is found. A more detailed analysis shows that the effect of HLA-B sharing may be fully accounted for by HLA-B-DR linkage disequilibrium. The results thus indicate that cell-mediated PPD specific cytotoxicity is HLA-restricted. Further, the correlation to HLA-DR sharing indicates that the restriction element in this system in all probability is a class II antigen.

The impact of HLA-DR antigen matching on the survival of cadaveric renal allografts. A prospective one-center analysis.

The impact of HLA-DR antigen matching on the survival of cadaveric renal allografts was assessed in 158 consecutive transplants performed in our unit since early 1978. In 41 donor-recipient pairs with two shared HLA-DR antigens, the actuarial graft survival rate at 6 months was 73% as compared with 51% in 76 transplants with one HLA-DR antigen shared and 32% in 41 transplants with zero shared HLA-DR antigens. This finding is highly significant (P for heterogeneity [PH] = 0.0005 and P for trend, [PT] = 0.0001). Our data clearly indicate that HLA-DR antigen sharing is more beneficial than merely avoiding HLA-DR incompatibility. But, the frequent antigen HLA-DRw6 was not taken into account in this study due to difficulties in its identification. We found no evidence that the observed beneficial effect of HLA-DR matching could be explained by interaction of other prognostic factors, such as sex, age, previous transplantation, diabetes mellitus or pretransplant blood transfusion. Patients who did not receive blood transfusion prior to transplantation had a significantly lower graft survival rate than those who did (13% vs. 56% at 6 months). HLA-DR matching was found to have a powerful effect on graft survival even among pretransplant blood transfused recipients (PH = 0.002, PT = 0.0006). We conclude that selection of recipients for transplantation should attempt to achieve HLA-DR identical combinations.

Planned immunization of human volunteers with HLA-DR incompatible lymphocytes I. Serological aspects.

Aiming at the production of anti HLA--DR test sera, eight healthy human volunteers were immunized by repeated intradermal injections of lymphocytes which were selected to be incompatible for one HLA--DR antigen, and matched as well as possible for HLA--A,--B,--C antigens. One out of 3 recipients immunized exclusively against HLA--DR produced lymphocytotoxic HLA-DR antibodies. The remaining 5 recipients were immunized against 1 or more HLA--A,--B,--C antigens in addition to one HLA--DR antigen. After 3 immunizations, 3 of these reacted with strong HLA--A or --B antibody production; however, only one showed a parallel anti HLA-DR antibody response detectable by complement dependent lymphocytotoxicity. Testing of the recipient sera in the antibody dependent cell-mediated cytotoxicity (ADCC) assay revealed that 6 of the 8 recipients did react early to the immunizations with HLA specific antibody production. However, in spite of repeated booster injections it was not possible to obtain more than the above-mentioned 2 sera with HLA--DR antibodies strong enough to react in the lymphocytotoxicity microtechnique.

Human B-blast specific target determinants in CML: a family study.

Human B blast specific target determinants, selectively identified on PWM stimulated purified B lymphoblasts by in vitro generated CTLs, have previously been studied in the population and showed association to and inclusion of HLA-DR geneproducts. This report indicates that B blast target determinants are products of genes which in a codominant mendelain way segregate with the HLA haplotypes in 4 selected families. Furthermore tests of families with HLA-B/D, DR and HLA-D, DR/GLO recombinations show that human B blast specific target determinants are coded from loci (locus) in the HLA-D region, between HLA-B and GLO.

Familial progressive myoclonic epilepsy. A clinical, genetical, biochemical and patho-anatomical study of a family with a 6-year follow-up.

Six siblings, including 4 cases of myoclonic epilepsy, their parents and 2 grandmothers were subjected to systematic investigation, and the patients were followed-up. The genetic studies revealed in the mother's family a patient with Lafora bodies demonstrated at autopsy. No chromosome abnormalities were found nor any linkage to the HLA system. The affected family members were characterized biochemically by an increased excretion of total glycosaminoglycans and/or an abnormal electrophoretic pattern of urinary glycosaminoglycans with an increased proportion of low-sulfated glycosaminoglycans. In the healthy family members this pattern of electrophoresis could also be demonstrated in the father and the paternal grandmother. Based on the biochemical results and the genetic studies it is suggested that the family members with progressive familial myoclonic epilepsy present a combination of at least 2 hereditary defects. The course of the disease has been relatively benign and treatment with sodium valproate, baclofen and clonazepam has shown quite satisfying results. In consequence of the biochemical findings combined treatment with A and E vitamins has been initiated.

Hereditary angioneurotic edema and HLA types in two Danish families.

HLA types were determined in 19 patients and 9 healthy members of 2 Danish families with hereditary angioneurotic edema. The study revealed no connections between hereditary angioneurotic edema and the HLA system.

HLA-DR genes and antigens in the Danish population. A study of 500 unrelated Danes.

Five hundred unrelated Danes, including 202 healthy individuals, 35 cadaveric kidney donors, and 263 patients in terminal uraemia were typed for the HLA-DR antigens DR1-w8. The HLA-DR gene frequencies of healthy Danes are in good agreement with frequencies estimated during The Eighth International Histocompatibility Workshop of the Scandinavian population, but differ to some extent from other groups of European Caucasians. This indicates geographical variation in the distribution of DR genes within Europe. Very close associations were found between the HLA-DR antigens and their corresponding HLA-D specificities, except for Dw4 and Dw6, which were included in DR4 and DRw6, respectively. Linkage disequilibrium was calculated between alleles of two loci (HLA-A, -DR; -B, -DR and -C, -DR alleles was assessed for multiple loci by using haplotype data from 32 HLA-A, -B, -C, -D, -DR, and Bf typed Danish families. The haplotype data showed that HLA-DR4 is strongly associated with HLA-B15 only in haplotypes together with HLA-Dw4. This indicates genetic heterogeneity of HLA-DR4 which, however, does not appear serologically in this study.

Immune thrombocytopenia due to antazoline (Antistina).

Antazoline (Antistina)-induced thrombocytopenic purpura occurring on three occasions in a 21-year-old woman is reported. After withdrawal of the drug ther recovered promptly. In vitro investigation demonstrated the presence of an antibody in serum, which in association with antazoline caused complement fixation when added to test platelets. Also platelet agglutinins could be detected in the patient's serum when antazoline was added. The reactions were drug specific and they could still be demonstrated 9 months after the last exposure to the drug.