Substantial evidence has been accumulated suggesting that T cells in patients with epithelial ovarian carcinoma (EOC) exhibit an antigen-driven immune response directed against the tumor cells. In ...the context of human leukocyte antigen (HLA), this suggests its possible involvement in the disease. Therefore, we examined the distribution of the HLA-DRB1*, -DQA1*, and -DQB1* alleles in 47 patients with EOC and 67 healthy Caucasian women. The frequency of D
70 and E
71 polymorphic residues of the DRB1 alleles was significantly reduced in EOC patients versus controls (
pD
70E
71 = 0.009), suggesting a protective role against the disease. The DQα residues R
52 and Y
11R
55 were increased in the patients (
p = 0.008 and 0.012, respectively). Because residues 11 and 55 participate in the formation of pocket 1, they may be functionally important amino acid positions that influence disease susceptibility. The frequency of the DQα susceptibility epitope (R
52Y
11R
55) among the DRβD
70E
71-positive EOC patients was increased when compared with DRβD
70E
71-positive controls (EOC, 100%; control, 52%;
p = 0.028). Among individuals without the DQα susceptibility epitope, the distribution of DRβD
70E
71-positive cases was significantly different between EOC patients and controls (EOC, 0%; control, 60%;
p = 0.039). Therefore, it appears that the presence of DQα susceptibility elements overrides the protective effect of the DRβD
70E
71 epitope and suggests an interactive relationship between DRβ and DQα epitopes that may be of importance for disease susceptibility. Because positions DRβ 70,71 and DQα 52 have been implicated in immunologic diseases, it is likely that besides being critical for T-cell recognition, they may also play a role in T-cell development and acquisition of the T-cell repertoire.
cDNAs coding for the HLA class II DR and DQ alpha and beta chains of the diabetogenic haplotypes DR3 and DR4 were introduced into a mammalian expression vector and transfected into L-cell mouse ...fibroblasts to produce cells expressing individual human class II molecules. Stable L transfectants were generated expressing each of the DR or DQ isotypes of the cis-encoded alpha and beta chains of the DR3 or DR4 haplotypes, as well as the trans-encoded alpha and beta chains of the DQ molecules of the two haplotypes. However, isotype mismatched combinations (DR alpha/DQ beta or DQ alpha/DR beta) did not result in any stable transfectants. The stable DQ L-cell transfectants obtained, along with homozygous B-cell lines expressing the DQ2 and DQ8 specificities, were tested against a large panel of twentyone anti-HLA class II monoclonal antibodies (mAbs). Their unusual reactivity patterns are described including the failure of most "pan-DQ" mAbs to react with all DQ expressing L-cell transfectants. Interestingly, some mAbs react with certain alpha beta heterodimers expressed on B-LCL but fail to recognize the same heterodimers expressed on the transfectants. This is suggestive of minor structural modifications that class II molecules undergo depending on the cells they are expressed on.
Four patients with a history of multiple blood transfusions who awaited renal transplantation were tested for human immunodeficiency virus (HIV) infection and found to be positive on enzyme ...immunoassay (EIA) and negative on Western blot. None of these patients had any clinical evidence of HIV infection. Absorption of these patients' sera with B-lymphoblastoid cell lines (B-LCL) positive for the serologic specificities DR3, DR4 (Dw4, Dw10, Dw14), and DR5 resulted in EIAs that were negative for HIV. Treatment of the B-LCL with an anti-DR monoclonal antibody (L243) interfered with the absorption of the serum sample by B-LCL. This indicates that the initial false-positive EIA results may be due to HLA antibodies. Furthermore, it was shown that these HLA antibodies are not limited in specificity to the HLA type of the host cell used in the preparation of the EIA reagents, but can consist of other DR specificities.
Functional and structural heterogeneity of HLA class I molecules was sought among five donors serologically identical for A2, A3, B7, and Bw44. Functional differences were identified by cell-mediated ...lympholysis (CML) after allogeneic mixed lymphocyte reaction among the five donors. Structural differences were characterized by high resolution two-dimensional electrophoretic maps of the class I HLA proteins synthesized by peripheral blood lymphocytes of these donors. Cells from three donors showed no CML-defined differences from one another; their HLA protein maps were identical. The cells of one donor recognized an A3-associated target antigen on the cells of all the other donors; her HLA map revealed a unique protein with altered isoelectric point. Another donor's cells differed by two CML-detected antigens: one was identified as a variant of Bw44 ("44.2") and the other was associated with Cw4. This donor's two-dimensional HLA map showed two novel charged proteins. By using these data, a two-dimensional map locating HLA-A2, -A3, -A3', -B7, -Bw44.1, -Bw44.2, and -Cw4 was prepared. Because each of three CML-detected antigens was correlated with a protein of distinctive charge, our results and the available published data raise the possibility that amino acid substitution producing charge variation may be a particularly important mechanism in the generation of CML-detectable HLA diversity.
Phorbol-12-myristate-13-acetate (PMA) is a potent tumor promoter. However, the mechanism of its effect is still unknown. In the present study we use two dimensional gel electrophoresis to show that ...the PMA effect on platelets is associated with enhanced phosphorylation of a series of polypeptides at 44K which migrate close to but distinct from a previously reported 47K protein. We identified these proteins as the class I molecules of the human histocompatibility antigens (HLA A,B). We further demonstrate that the PMA effect is also associated with a dramatic phosphorylation of HLA antigens in HL-60 leukemic cells and in human lymphocytes, showing that an increase in phosphorylation of HLA antigens is intimately related to the signal of PMA in various cellular systems. Immunoprecipitation of HLA proteins resulted in coprecipitation of phosphorylated myosin light chain (20K). HLA antigens are transmembrane proteins which interact with cytoskeletal elements, probably via their intracellular region, which has been previously shown to be phosphorylated. It is suggested that phosphorylation of HLA membrane proteins may represent an important mechanism in the effects induced by PMA.
Fifteen DR4-bearing haplotypes from twelve patients with insulin-dependent diabetes mellitus (IDDM) were analyzed serologically, cellularly, and biochemically. The HLA-Dw composition of these ...DR4-positive haplotypes was Dw4 (46%), Dw14 (22%), and Dw10 (33%). The biochemical analysis by two-dimensional electrophoresis (2D-PAGE) of the DR beta chains showed that each Dw specificity is characterized by a specific DR4 beta chain that appears to be identical in normal and diabetic individuals. Analysis of DQ beta chains in the DR4-bearing haplotypes revealed that certain Dw specificities such as Dw4 are characterized by the presence of either the DQw7 (formerly DQw3.1) or DQw8 (formerly DQw3.2) alleles, which generate the Dw4.1 or Dw4.2 subtypes, respectively. Others such as Dw14 and Dw10 are characterized by the presence of the DQw8 allele. In our sample of 12 patients the Dw4.2 (Dw4, DR4 beta I-4 DQw8) and Dw10 (Dw10, DR4 beta I-1, DQw8) subtypes were predominant. It is concluded that individual DR beta and DQ beta gene products from the DR4-bearing haplotype of IDDM patients are identical to those of normal control subjects and that Dw14 as well as Dw10 are involved in disease susceptibility. We suggest that disease susceptibility may be influenced by more than one locus within the HLA-D region.
We examined different antisera and varying serum dilutions for their effects on the sensitivity of immunofixation electrophoresis (IFE) for detecting and characterizing serum IgG paraproteins. ...Different antisera reacted unequally with a single paraprotein and conversely, a single antiserum reacted unequally with several different paraproteins. The dilution of serum was extremely important; the adoption of a single dilution for all sera could result in failure to detect low-concentration or poorly immunoreactive paraproteins. While the application of a standardized amount of paraprotein is recommended, it must be emphasized that no single antiserum can be relied on to provide unequivocal detection of all paraproteins of that particular isotype.
Current immunologic theory defines a role for HLA-A, -B, -C proteins in cells presenting foreign antigens to cytotoxic T lymphocytes. No clear function, however, is assigned to the prominently ...represented HLA molecules on the T cells themselves. To investigate this question, the synthesis and turnover of HLA-A, -B, -C molecules in human peripheral lymphocytes was studied. Newly synthesized HLA-A, -B, -C and beta 2-microglobulin molecules were identified among total lymphocyte proteins by two-dimensional gel electrophoresis and specific immunoselection. In resting T lymphocytes, the polymorphic set of HLA-A, -B, -C proteins was among the most prominently synthesized and rapidly turned over of total cell proteins. Its half-life was 6 to 7 hr, which is considerably shorter than that of total cell protein. HLA and beta 2M were the major proteins of the plasma membrane undergoing active synthesis and turnover in resting T lymphocytes. After growth stimulation, the rate of HLA synthesis increased to a lesser degree than total protein synthesis. Survival of newly synthesized HLA was increased, however, indicating reduction in HLA turnover. The result was the accumulation of HLA in the plasma membrane. It is suggested that continuous degradation and replacement reflects the biochemical nature of the participation of HLA molecules in immunologic surveillance by quiescent lymphocytes. Cessation of HLA turnover, with accumulation of surface HLA molecules, is a biochemical adjustment related to T cell activation.
