Lymphocyte Microcytotoxicity Test Research Articles

Analysis of HLA in Patients with Obstructive Sleep Apnea Syndrome

Background : Obstructive sleep apnea syndrome (OSAS) is believed to have multifactorial causes. The major risk factors for OSAS are obesity, narrowed upper airways, and abnormal cranial-facial structures. A genetic basis for OSAS has been also suggested by reports of families with many members affected. This study analyzed the HLA typing in patients with OSAS to determine the possible role of genetics in OSAS. Methods : Twenty-five Korean patients with OSAS (1 woman and 24 men; age range 30-66 years) were enrolled in this study. A diagnosis of OSAS was made using full-night polysomnography. The control group consisted of 200 healthy Korean people. Serologic typing of the HLA-A and B alleles was performed in all patients using a standard lymphocyte microcytotoxicity test. Analysis of the polymorphic second exons of the HLA-DRB1 gene was performed using a polymerase chain reaction-sequence specific oligonucleotide probe. Results : The allele frequency of HLA-A11 was significantly lower in patients with OSAS compared with the controls (p45) than in the controls (p

First Report of Recombination Between the HLA-DR and HLA-DQ Loci Within a Family

ABSTRACT: Although unusual associations of HLA-DR and HLA-DQ alleles seen in ancestral haplotypes have indicated that recombination between these genes occurred in the past, an actual crossover event between DR and DQ has never been shown within a family. In a study of families with Graves’ disease we have identified an individual from a three generation family who inherited a maternal haplotype that is the result of a recombinational event between the HLA-DR and the HLA-DQ loci on her chromosomes. Family members were typed for HLA class I by the lymphocyte microcytotoxicity test and for HLA class II by polymerase chain reaction (PCR) with sequence-specific primers or with sequence-specific oligonucleotide probes after PCR. Based on linkage disequilibrium it is likely that the recombinant haplotype is present in the proband rather than his brother. This haplotype was subsequently inherited by one of the proband’s sons. The data presented support the conclusion that the recombinant haplotype resulted from a crossover event between the mother’s DRB1 and DQA1 genes. Thus, recombination between the HLA-DR and HLA-DQ genes has been demonstrated within this family; a recombination event not previously described.

HLA antigens in ulcerative colitis and primary sclerosing cholangitis.

The aims of this study were to find out whether the alleles of the HLA class I or II region are associated with susceptibility to ulcerative colitis, and to show whether there is a difference or similarity in HLA associations between primary sclerosing cholangitis and ulcerative colitis. HLA-A, B, C and DR antigens were studied using the standard lymphocyte microcytotoxicity test in 24 Finnish patients with primary sclerosing cholangitis, 77 patients with ulcerative colitis, and 106 controls. HLA-B8 (54%) and DR3 (60%) were associated with primary sclerosing cholangitis. HLA-DR1 (46%) and DR6 (20%) seemed more common in ulcerative colitis than in controls. A positive association with Cw7 was common to both ulcerative colitis (25%) and primary sclerosing cholangitis (33%). Our results indicate that ulcerative colitis is more heterogeneous than primary sclerosing cholangitis in its HLA-DR associations.

Mixed lymphocyte culture studies reveal complexity in the bovine MHC not detected by class I serology

The genetic structure of the bovine major histocompatibility complex (MHC) was investigated using the lymphocyte microcytotoxicity test for class I typing and the mixed lymphocyte culture (MLC) assay for class II typing. Using locally produced alloantisera and antisera from the Third International BoLA Workshop, 14 class I BoLA-A locus alleles were identified in the study population, a single herd of approximately 700 Holstein-Friesian cattle. Eleven of these were alleles recognized in the International Workshop and three were new alleles. An MLC titration assay was employed in conjunction with class I typing to define BoLA haplotypes and identify BoLA complex homozygotes. An embryo transfer family consisting of eight full sibling cattle including one BoLA complex homozygote was produced by half sibling mating. Five other BoLA complex homozygotes were subsequently identified in the herd. Six MLC defined class II haplotypes investigated in detail were designated BoLA-D1, D2, D3, D4, D5 and D7. BoLA-D1 was associated with the class I specificity BoLA-Aw6, D2 with Aw6 and the new class I specificity Ac3, D3 with Aw6 and Aw11, D4 with Aw10, D5 with Aw31 and Aw11, and D7 with Aw20. The discovery of four groups of class I identical-class II disparate haplotypes, and three pairs of class I disparate-class II identical haplotypes indicates the presence of considerable complexity in the BoLA complex that is not detected using class I serology.

Preliminary Report on BoLA Polymorphism in Guernsey Cattle

A total of 95 registered Guernsey cows and heifers sired by 34 bulls were typed for class I antigens encoded by the bovine major histocompatibility complex (BoLA). A panel of alloantisera was used to detect 21 of the 33 internationally recognized BoLA specificities. Fourteen BoLA specificities were detected in the herd using a standard lymphocyte microcytotoxicity test The most frequent BoLA specificity detected was w6; but in 85% of these animals, a subtype of w6, either w17 (13.7%) or UR6.2 (34.7%), could be assigned. The next most common BoLA specificities, by frequency, were w12, w15, UR1, and w21. Specificities not detected were w2, w3, w5, w7, w11, w16, w19, w20, and w24.

The ELY-1 locus controls a di-allelic alloantigenic system on equine lymphocytes.

The ELY-1 locus controls the expression of a polymorphic cell surface antigen of equine lymphocytes which was detected using antibodies generated by alloimmunization with peripheral blood lymphocytes. The ELY-1 antigens were not detected on erythrocytes or platelets by absorption experiments. The two alleles, which have been designated ELY-1.1 and ELY-1.2, are expressed codominantly and appear to constitute a closed system at the population level. In family studies, the ELY-1 antigens segregated as products of an autosomal locus not linked to the major histocompatibility complex (MHC) of the horse. In the complement mediated lymphocyte microcytotoxicity test, antisera to the ELY-1 antigens selectively killed peripheral blood lymphocytes which did not express surface immunoglobulin. The ELY-1 antigens may be useful markers for equine T cells when assayed in this fashion. Three alloantisera were used in immune precipitation of iodinated and solubilized cell surface proteins from peripheral blood lymphocytes. Electrophoresis of the precipitates in sodium dodecyl sulphate (SDS)-polyacrylamide gels demonstrated strong bands in the Mr 180-190K range that were shared in the three different preparations. These results suggest that the ELY-1 allospecificities are expressed on an equine equivalent of the murine T200 molecule.

Antinuclear Antibodies in Psychiatric Patients treated with Psychotropic Drugs

Antinuclear antibodies in sera obtained from 183 psychiatric patients treated with psychotropic drugs were studied.Antinuclear antibodies (ANA) were detected by the indirect immunofluorescent method using rat liver cells as substrate. Farr assay was employed to determined the binding capacity for DNA. The immunoglobulin classes and complement fixing ability (CF) of anti-dsDNA were detected by the indirect immunofluorescent method and the complement fixation method, respectively, using Crithidia luciliae kinetoplasts as the substrate. Anti-ENA was detected by the hemagglutination test (PHA) and double immunodiffusion. Anti-histone was detected by the histone reconstitution of the immunofluorescent test. HLA typing for antigens of the A, B and C was performed by the standard lymphocyte microcytotoxicity test provided by the Terasaki laboratory. Typing for HLA-DR was performed on B lymphocytes isolated on nylon wool column. As controls, 75 normal Japanese individuals were tissue typed with the same antisera.ANA was detected in 37 patients (20.2%). Total doses of carbamazepine in the patients with ANA were significantly larger than in those without ANA. The patients with ANA hada photosensitivity and oral ulceration significantly more than those without ANA. The patients with ANA had a tendency to have an association with HLA-DRw8.Anti-DNA antibodies by Farr assay were detected in 25 patients, but titer of anti-DNA was low. Anti-dsDNA by Crithidia immunofluorescent technique was detected in two patients. However, CF of anti-dsDNA was not detected in any patients. Anti-ENA by PHA technique was detected in 21 patients. The patients with anti-ENA had arthralgia and hypergammaglobulinemia significantly more than those without anti-ENA. Anti-histone antibodies were detected in three patients, who were treated with chlorpromazine.

HLA Antigens in Systemic Lupus Erythematosus

For the purpose of studying immunogenetic factors related to disease susceptibility to systemic lupus erythematosus (SLE), HLA-A, B and DR typing was performed by the standard lymphocyte microcytotoxicity test using 247 typing antisera in 34 Japanese patients with SLE and 51 healthy individuals.The results obtained were as follows: 1) SLE patients with proteinuria had the association with HLA-BW 35 (phenotype frequency 50.0%, Relative Risk=7.5, corrected P<0.10), compared with normal control (11.8%). SLE patients without proteinuria had the association with HLA-BW 55 (phenotype frequency 35.0%, RR=8.6, Pc<0.10), compared with normal control (5.9%).2) Patients with SLE had the strong association with HLA-DRW 9 (phenotype frequency 52.9%, RR=4.6, Pc<0.025), compared with normal control (19.6%).3) All of the peripheral blood lymphocytes obtained from 10 patients with SLE who had anti-RNP antibody reacted with MT 3, which was one of the B-cell alloantisera including three antibodies against HLA-DR 4, DR 7 and DRW 9.4) The linkage disequilibrium between HLA-BW 61 and HLA-DRW 9 was seen in the patients with SLE.

The Immunogenicity of Allograft Knee Joint Transplants

Joint allotransplantation has been employed in the management of osteoarthritis affecting primarily one compartment of the knee. Biological resurfacing and realignment of the knee has been the goal. The immunological results of six patients posttransplant were studied with the lymphocyte microcytotoxicity test. Definite immunogenicity was demonstrated at an early stage. The sera from the recipients posttransplant was noted to abrogate the in vitro cytotoxicity suggesting the presence of blocking factors. The actual clinical significance of this phenomenon was difficult to judge insofar as no evidence of allograft joint rejection was seen.

Studies on Immunogenetics in Broncho-Pulmonary Diseases

Phenotype frequencies and haplotype of HLA antigens in 297 cases with various bronchopulmonary diseases were studied using the method of lymphocyte microcytotoxicity test. The following results were obtained.1. High frequencies of HLA-A2, HLA-A9, HLA-B5, HLA-BW40 were recognized in normal individuals.2. HLA-BW21 in the patients with chronic bronchitis was higher than those in normal individuals (P<0.05). However, there were no differences of frequencies of HLA antigens in chronic pulmonary emphysema, pulmonary tuberculosis, lung cancer, pulmonary sarcoidosis, cystic fibrosis, bronchiectasis, pneumoconiosis, pulmonary alveolar microlithiasis and normal individuals.3. Frequencies of HLA-BW38 in the patient with nasal allergy and frequencies of HLA-BW40 in the patient with bronchial asthma were higher than those in normal individuals (P<0.05). In those various factors, high serum IgE levels and high IgE antibody titers showed to be corelated to HLA-BW40.4. We can not prove asscciated haplotype in atopic families. The other hand, relationship between haplotype HLA-A10, B12 and onset of the disease were suggestted in family study of the patient with hypersensitivity pneumonitis.5. High phenotype frequencies of HLA-B5 were recognized in acute myelocyte leukemia (P<0.05). HLA-B27 was detected in all patients of ankylosing spondylitis.

HL-A histocompatibility antigens in children with coeliac disease (author's transl)

HL-A antigens were determined in 41 unrelated coeliac children and in clinically healthy parents and siblings of 40 of these patients using a lymphocyte microcytotoxicity test. 58.5% of the coeliac patients had phenotype HL-A 8 compared with an HL-A 8 frequency of 16.6% in a control group of 320 unrelated individuals (P less than 0.0005). Excluding five patients not of pure German origin HL-A 8 frequency increases to 63.9%. The increase of HL-A 1 frequency in coeliac patients is attributed to linkage disequilibrium between HL-A 8 and HL-A 1. The haplotype HL-A 1.8 frequency was significantly increased in coeliac children (P less than 0.0001) with frequency elevation also in parents (P approximately 0.025) but not in siblings. Furthermore, an increase in frequency of HL-A 12 and a decreased frequency of HL-A 7 and HL-A 9 was found in coeliac patients. Five clinically healthy siblings had the same HL-A haplotypes as their affected sisters and brothers.

Immunogenicity of virus-induced rat osteosarcoma

The immunogenicity of a virus-induced rat osteosarcoma was studied utilizing the lymphocyte microcytotoxicity test. Intratibial injection of murine sarcoma virus (Moloney) resulted in the development of palpable tumors at the injection site which on histopathological examination appeared to be osteosarcomas. In 73 per cent of animals injected these tumors progressed and metastasized to the lungs. Lymphocytes from these "progressor" animals demonstrated an ability to kill osteosarcoma cells in vitro (as quantitated in the microcytotoxicity test) while serum from these animals abrogated or blocked the cell-mediated cytotoxicity. In the remaining animals the tumors either failed to develop or regressed spontaneously. Lymphocytes from these "regressor" animals also demonstrated cytolytic activity against osteosarcoma cells in vitro, but serum failed to block the lymphocyte-mediated cytolysis. Both regressor and progressor groups demonstrated humoral cytotoxic antibodies to tumor antigen on the basis of the ability of their serum to kill tumor cells in vitro.

Predominance of histocompatibility antigen HL-A8 in patients with gluten-sensitive enteropathy.

HL-A phenotypes were determined in 24 unrelated patients with gluten-sensitive enteropathy (GSE) using a lymphocyte microcytotoxicity test. 21 of the 24 patients had HL-A8 in the second segregant series, a frequency of 0.875. In contrast, the HL-A8 frequency in 200 normal individuals was 0.215 (difference significant at P < 0.002), and in 6 patients with villous atrophy due to tropical sprue or hypogammaglobulinemia the HL-A8 frequency was 0.17 (difference from normal not significant). The HL-A types in the families of three HL-A8 positive patients with GSE indicated that the HL-A8 antigen was inherited as an autosomal dominant. Frequencies of the other HL-A antigens in the GSE group did not differ significantly from that of the normal group. These findings are compatible with the hypothesis that GSE is due to the presence of an abnormal "immune response (Ir) gene," leading to the production of pathogenic antigluten antibody or, alternatively, to the presence of a particular membrane configuration leading to the binding of gluten to epithelial cells with subsequent tissue damage.

The synergistic action of antibodies. A method to increase the sensitivity of the lymphocyte microcytotoxicity test.

Abstract. A method, based on the synergistic action of two antibody systems, is suggested as a lymphocyte cross‐match test. To donor lymphocytes in the serum of the recipient a rabbit anti‐human lymphocyte antibody, in sublytic quantities, is added before the addition of the complement. Results obtained with this test in tissue typing sera are compared to the lymphocyte microcytotoxicity test. It could be shown that the new test is more sensitive than the ordinary microcytotoxicity test.

The Synergistic Action of Antibodies

A method, based on the synergistic action of two antibody systems, is suggested as a lymphocyte cross-match test. To donor lymphocytes in the serum of the recipient a rabbit anti-human lymphocyte antibody, in sublytic quantities, is added before the addition of the complement. Results obtained with this test in tissue typing sera are compared to the lymphocyte microcytotoxicity test. It could be shown that the new test is more sensitive than the ordinary microcytotoxicity test.