Evidence for HL-A-linked genes in "juvenile" diabetes mellitus.

HL-A typing of 150 patients who had developed diabetes mellitus by the age of 30 years showed a significant association with HL-A 8 and W 15. The HL-A genotypes were determined in 17 families in which two or more siblings had this type of diabetes. The zygotic assortment of HL-A haplotypes was found to be significantly disturbed from the expected random pattern, with a reduction in the number of siblings showing no identical haplotypes and an appreciable increase in the number with both haplotypes identical. This appears to be most consistent with the presence of a gene or genes pre-disposing to this type of diabetes at a locus closely linked to the HL-A chromosomal loci. This locus appears to have a fundamental role in the susceptibility to juvenile diabetes.     

Cross-reactivity between human and murine lymphocyte antigens: III. Reactivity of H-2 allo- and xenoantisera with human lymphoid cells

H-2 alloantisera and antimouse lymphocyte xenoantisera react with 14%–100% of human lymphocytes from a panel of at least 80 unrelated people. Population and family studies did not reveal HL-A specificity of such lymphocytotoxic antibodies but indicated that the antibodies are directed against polymorphic antigenic determinants inherited in association with HL-A antigens. H-2 allo- and xenoantisera absorbed with human lymphoid cells and a panel of platelets bearing all the known HL-A specificities were still cytolytic when tested against murine lymphocytes, suggesting that only a small proportion of the heterogeneous population of H-2 antibodies react with human lymphocytes. On the other hand, HL-A alloantisera could be absorbed by lymphocytes from certain murine strains. These results suggest that the crossreactivity between human and murine lymphocytes is caused by antigens common to several HL-A (or H-2) types or by antigens linked to HL-A but not identical with them.     

HL-A frequencies in Down's syndrome.

HL-A antigen frequencies were examined in 76 Down's syndrome individuals and 733 normal Caucasians. 10 antigens of the first locus and 15 antigens of the second locus were defined, using a microlymphocytotoxicity technique. No significant differences were observed between the normal and Down's syndrome samples, in contrast to a previous report (Boxer and Yokoyama, 1972) of decreased HL-A antigen frequencies in Down's syndrome individuals. Our results therefore suggest that there is no relationship between trisomy 21-associated immune aberrations and altered HL-A antigen frequencies.     

Linkage group HL-A-MLC-BF (properdin factor B). The site of the Bf locus at the immunogenetic linkage group on chromosome 6.

Genetic linkage between the HL-A and Bf loci could be confirmed in 43 families with 168 offspring. In 4 families, 5 recombinants out of 82 informative meiotic divisions were observed (r = 6.1%). The localisation of the Bf marker system was studied in 3 families with crossovers between HL-A and MLC. From these data the following map order of human chromosome 6 can be proposed: HL-A (1st locus) -- HL-A (2nd locus)--MLC-Bf---PGM(3). The fact that important components of the classical and alternate pathway of complement activation are governed by genes closely linked with HL-A and MLC loci leads to the proposition to include the Bf system into the Major Histocompatibility Complex in man.     

The identification of HL-A antigens on fresh and cultured human endothelial cells.

Human endothelial cells were obtained from the umbilical cord veins of 16 newborns by methods previously described and tested for HL-A antigens by a microcytotoxicity method. HL-A antigens were present on all endothelial cell lines tested. When the HL-A phenotypes of fresh endothelial cells and autologous fetal lymphocytes were compared, a concordance of 70% was observed. When the HL-A phenotypes of maternal lymphocytes and fresh endothelial cells were compared, a maternal contribution to the endothelial cell phenotype was evident in 72% of the possible commmon antigens. Some HL-A antigens were deleted from 11 of 16 endothelial cell lines that were re-typed after 2 weeks in tissue culture. The majority (90%) of deleted antigens were from the second HL-A locus. When three lines of endothelials cells were again re-typed after 6 weeks in culture, no further changes in antigenicity were noted. These findings: a) demonstrate that HL-A antigens are present on human endothelium and suggest that endothelial cells are actively involved in establishing the immunogenicity of a graft, and b) demonstrate that the HL-A antigens on human endothelial cells may be modulated by in vitro culture.     

Maternal HL-A Antibodies and Fetal Sex

In a study of 960 pregnancies a significantly higher male to female birth ratio was found among primigravidae who developed HL-A antibodies, which was highest where these were monospecific. In the whole group male births predominated in women with antibodies to HL-A 1 and 11 (first locus) and HL-A 5, 12, and 13 and TYT (second locus).     

Purification of papain-solubilized histocompatibility antigens from a cultured human lymphoblastoid line, RPMI 4265.

HL-A antigens having specificities HL-A2, HL-A7, HL-A12 have been solubilized by papain treatment of membrane preparations from the cultured human lymphoblastoid cell line RPMI 4265 and purified about 80-fold by chromatography on carboxymethylcellulose, Sephadex G-150, and diethylaminoethylcellulose columns. Separation of HL-A2 from a mixture of HL-A7 and HL-A12 was achieved on the final DEAE-cellulose column. The yield was about 1 mg of protein of each antigen preparation per 100 g of packed, frozen cells. On sodium dodecyl sulfate gel electrophoresis both preparations showed two polypeptide bands. The smaller subunit of 12,000 daltons is common to all HL-A preparations and has been shown to be identical with beta2-microglobulin. The larger subunit is a glycopeptide and in the HL-A7, 12 preparation was resolved into a duplex of 34,000 and 37,000 daltons. The HL-A2 preparation showed a single band at 34,000 daltons. On isoelectric focusing under nondenaturing conditions, the preparation showed multiple bands, all of which contained both subunits and retained antigenic activity. On isoelectric focusing in the presence of 6 M urea a single band for beta2-microglobulin and multiple bands for the larger subunit were seen. This charge heterogeneity of the larger subunit has been shown to be due to variable amounts of sialic acid. When HL-A antigen preparations were subjected to Sephadex G-100 chromatography in the presence of 3 M KCl, no separation of the two subunits was observed.     

C2 deficiency in man. genetic relationship to a mixed lymphocyte reaction determinant (7a*)

Three unrelated individuals with, respectively, lupus erythematosus, polyarteritis, and membranoproliferative glomerulonephritis and totally deficient in the second component of complement are demonstrated to be mutually poorly reactive in mixed lymphocyte culture and homozygous for the mixed lymphocyte reaction determinant (MLR-S or LD) short 7a (7a^*). The gene controlling the elaboration of C2 in man is shown to be separate from, and probably to map outside of, the second locus ofHL-A and theMLR-S locus. Genetic linkage disequilibrium is strongly suggested between HL-A 10, W18, 7a^*, and C2 deficiency.     

HL-A and Kidney Transplantation

AT present the role of HL-A antigens in the rejection of organ transplants is uncertain; we therefore wish to report the results of renal transplantations, under known HL-A compatibility conditions, performed in France since 1959. The 179 cases already published² have now increased to 416 cases³ including 84 sibling (SS), parent-to-child (PC) and 253 unrelated (UR) allografts. Only three cases were excluded from the analysis, which was similar to that previously used². We calculated the probability of incompatibility for the non-determined HL-A antigens, assuming the presence of four HL-A antigens of similar strength in each individual. Differences between antigens having serological similarities (cross-reactions) were considered in a first analysis as incompatibilities and in a second analysis as identities.     

Secondary response of in vitro-primed human lymphocytes to allogeneic cells

In vitro-primed human lymphocytes proliferate in a secondary mixed lymphocyte reaction (MLR) under the control of MLR-S specificities. HL-A antigens are unable to induce a secondary Proliferation. In familial haploidentical combinations, the secondary proliferation is specific for the priming MLR-S specificity, i.e., as early as 24 to 48 hours after the re-stimulation, a clearcut response is observed toward the sensitizing MLR-S specificity. The secondary response is reflected in acceleration of the reaction rather than in the peak of (³H) TdR uptake. However, when either haploidentical familial primed responding cells or unrelated cells primed toward MLR-S homozygous cells were used, no early typing response was observed against unrelated cells. The level of (³H) TdR incorporation toward cells which possessed and those which did not possess the priming specificity was identical until day 3–4. Noneless, the peak response toward cells possessing the priming MLR-S specificity occurs regularly 24 to 48 hours prior to the peak response toward the cells negative for the priming specificity (day 3–4 as opposed to day 5). Technical improvements are therefore needed before such a technique will provide a clearcut MLR-S typing methodology.     

Value of Prospective Tissue Typing in Kidney Transplantation Between HL-A Identical Siblings

The predictability of leucocyte typing in kidney transplantation was assessed by an analysis of 37 kidney transplants from sibling donors. Recipients who were identical for the HL-A antigens with their donors gave highly predictable results. In comparison with those siblings who were incompatible or compatible but not identical their grafts functioned earlier, they required less immunosuppression, and had never had any rejections. They also appeared to have less postoperative morbidity. These results indicate that less immunosuppression than is current in many transplant centres could well be used with benefit in HL-A identical sibling transplants. This could reduce the risk of infection and possibly minimize the adverse effects of steroids on wound healing in these patients.     

Probable genetic linkage between a locus for human urinary pepsinogen and the HL-A loci.

The genetic basis of familial variation in the relative intensities of human urinary pepsinogen isozymes is not completely clear from family studies. An investigation of the linkage relationships of pepsinogen isozyme 5, considering only segregation for the presence or absence of Pg 5, yields a peak lod score of 4.1 at theta = .1 for linkage with HL-A1 or HL-A2. Added to data from segregation interpreted according to a scheme proposed for the inheritance of intensity differences in Pg 5, the peak lod score becomes 3.0 at theta = .2. Data derived from the segregation of pepsinogen isozyme 4, possibly determined by an allele to that controlling the presence or absence of Pg 5, further reduces the total lod score at theta = .2 to 2.9. The results indicate probable linkage between a locus for urinary pepsinogen and the HL-A loci, but are insufficient to permit any conclusion concerning possible heterogeneity in the linkage relationships of Pg 4 and Pg 5 to HL-A.     

HL-A population studies in 445 individuals from Hamburg,respecting 8 LA and 15 Four-antigens including U 18 (W 16) and CM (W 18)

Summary By means of 8 LA (HL-A1, HL-A2, HL-A3, HL-A9, HL-A10, HL-A11, Ba^*, Li) and 15 Four-antisera specificities (HL-A5, HL-A7, HL-A8, HL-A12, HL-A13, R^*, BB, MaKi, LND, U 18, MaPi, CM, ET, AA, FJH) 92% of the LA and 92% of the Four-alleles were identified in 445 unrelated people from Northern Germany (Hamburg). The allele frequencies of the more recently described antigens CM and U 18 are 1.3 and 2.6%, respectively. Both alleles are significantly associated with HL-A10.

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Zusammenfassung Bei Verwendung von 8 LA-(HL-A1, HL-A2, HL-A3, HL-A9, HL-A10, HL-A13, Ba^*, Li) und 15 Four-Antiserenspezifitäten (HL-A5, HL-A7, HL-A8, HL-A12, HL-A13, R^*, BB, MaKi, LND, U 18, MaPi, CM, ET, AA, FJH) konnten an 445 nichtverwandten Personen aus dem norddeutschen Raum 92% aller LA- und 92% aller Four-Allele identifiziert werden. Die Allelhäufigkeiten der erst seit kurzer Zeit beschriebenen Allele CM und U 18 betragen 1,3 bzw. 2,6%. Beide Allele zeigen eine signifikant hohe gametische Assoziation zu HL-A10.

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Supported by the Deutsche Forschungsgemeinschaft.     

Secondary response of in vitro primed human lymphocytes to allogeneic cells

The secondary response of human lymphocytes primed in vitro with allogeneic lymphocytes is reported. Accelerated proliferation is observed ' both against the specific priming cell and against unrelated third party cells, but the intensity of proliferation against the specific cell is usually, but not always, higher than that against third party cells. To clarify the respective roles ofHL-A andMLR-S in the development of this secondary proliferative response, three kinds of cells were used from which MLR-S activity was supposed to have been abolished while serologically-defined HL-A antigens were present: (a) heattreated cells, (b) UV-treated cells, and (c) a recombinant betweenHL-A andMLR-S. Heat treated cells were unsatisfactory for this study, but UV-treated and recombinant cells showed thatMLR-S was sufficient and necessary both for priming and for eliciting a secondary proliferative response. No role could be found forHL-A or for a secondMLR-S locus positioned between the first and secondHL-A loci.     

The zygosity in the human HL-A transplantation system

Summary If by serological methods of tissue typing one antigen only is detected in one or both of the loci LA and 4 of the histocompatibility system HL-A, the question arises whether it is homozygous or heterozygous. The probability thereof is calculated. It depends on the frequency of the known and unknown genes in the HL-A-system. If e.g. the HL-A type of an examinee is 1,3,8 this subject could be homozygous 1,3,8,8 or heterozygous (with an unknown antigen X) 1,3,8,X. The frequencies of homozygous and heterozygous individuals are calculated in all possible combinations of the LA and 4 loci. The results justify always for both the LA-and 4 locus the assumption of heterozygosity, with the one exception that the gene HL-A 2 is under consideration. There the calculations result in a value of 49% for homozygosity (2,2) and 51% for heterozygosity (2,X). Neither alternative is in this case essentially more probable.For transplantations generally heterozygosity can be assumed — with the exception of HL-A 2 — and therefore the minor grade of histocompatibility.National Blood Group Reference Laboratory — WHO (Director: Prof. Dr. P. Speiser)     

Characterization of a serum inhibitor of MLC reactions. III. Specificity: HL-A-related antibodies.

A mixed leukocyte culture (MLC)-inhibitory serum from a healthy multipara, JH, has been characterized with regard to the specificity of its inhibitory antibody. When added directly to MLC, JH serum will inhibit most combinations. However, when lymphocytes intended as responder or stimulator cells in MLC were preincubated with this serum, the specificity narrowed considerably. Four groups of lymphocyte donors were recognized, depending upon whether their lymphocytes were inhibited as responders, as stimulators, as both, or as neither. Absorptions of inhibitory activity, followed by assay of the absorbed sera in pretested MLC combinations, yielded reliable data for determining which donors' cells shared pertinent antigens. An association of MLC inhibition by JH serum with the HL-A types of the involved lymphocytes was observed and these relationships are summarized in Table 4. The three HL-A specificities identified, W19, W29, and 12, correspond with the HL-A typing of the husband of the serum donor. Various cell types absorbed relevant inhibitory activity (against responder and stimulator functions) in the following order of efficiency: LCL cells, B lymphocytes, T lymphocytes, fibroblasts, and erythrocytes. When the above three HL-A specificities were removed by absorption, the serum was no longer inhibitory in any combinations. Whether the inhibitory activity of JH serum is directly related to anti-HL-A antibodies or to antibodies against closely related MLR determinants will depend to a large extent upon the degree of linkage disequilibria found between W19, W29, and 12 antigens and the MLR locus.     

HL-A genotype of patients with acute lymphoblastic leukaemia

Summary 10 patients with acute lymphoblastic leukaemia were tissue-typed for 21 HL-A specificities. Of these, genotypes of 9 pateints were determined by family analyses. Haplotype HL-A1,8 occurred in 5 out of 18 instances. On phenotype basis, a slight increase was observed in the incidence of antigens HL-A1 and HL-A8. No loss of HL-A specificities could be detected on lymphocytes through family analyses.     

Alpha 1 antitrypsin deficiency: a study of the relationship between the Pi system and genetic markers.

Twenty-four members (4 generations) of a family with alpha 1 antitrypsin deficiency were studied in an attempt to determine the chromosomal location of the Pi system locus. Three alpha 1 antitrypsin alleles (PiM, PiI, and PiZ) and five phenotypes (MM, MZ, MI, IZ, and ZZ) were detected in family members. The quinacrine fluorescent banding technique was successfully utilized to reveal eight polymorphic chromosomal markers in family members. Eight red cell antigens and HL-A antigens were identified for each family member. No linkage between the Pi system and chromosomal markers, four polymorphic red cell antigens, and HL-A antigens was detected. On the basis of this family study, the Pi locus as defined by alpha 1 antitrypsin deficiency does not appear to be on chromosomes 2, 3, 13, 14, 21, or 22 within measurable distance of the markers used.     

PGM<Subscript>3</Subscript>: HL-A is Another Linkage in Man

EARLIER work has suggested the existence of a linkage between the histocompatibility region, HL-A and the phosphogluco-mutase locus, PGM₃. We have now produced more convincing evidence after investigating a further set of families.     

The human mixed-lymphocyte culture (MLC) interaction after in vivo allo-immunization. II. HL-A specificity of early proliferative response

Human volunteers were immunized with a single intradermal dose of allogeneic buffycoat cells. When donor and recipient differed for four HL-A antigens belonging to the first and second series, an increased MLC proliferation early in the cultures was observed when responding lymphocytes from the immunized individual were confronted with stimulating cells from his donor. These findings were not observed when the incompatibility between donor and recipient involved only one second series HL-A antigen. The specificity of the altered response was studied by confronting responding lymphocytes from the immunized individual with different third-party stimulating cells. Most of these combinations revealed an early increased proliferation compared to control combinations with responding lymphocytes from nonsensitized individuals. However, the early increased responsiveness was significantly more pronounced in combinations where the stimulating cells shared two or more “private” HL-A determinants with the immunizing donor. It is concluded that a major part of the early increased responsiveness is due to proliferation of lymphocytes with receptors for HL-A (SD) determinants.