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Biomedical subjects

F E Ward

Publications and source records attributed to F E Ward.

At least 37 records · Page 2Linked to original sources
PubMed1991

Class I-induced resistance to natural killing: identification of nonpermissive residues in HLA-A2.

Structural characteristics of major histocompatibility complex class I antigens associated with natural killer (NK)-resistance phenomena were examined. Previous research has shown that transfection of class I genomic DNA clones into class I-deficient, NK-sensitive target cell lines results in transfectants exhibiting class I+, NK-resistant phenotypes. In contrast to the HLA-A3, -B7, -B27, and -Bw58 class I molecules, the HLA-A2 class I molecules were shown not to protect target cells from NK activity. Here we show that this nonprotective phenotype maps to the alpha 1 domain of the HLA-A2 molecule by examining the NK-protective capacity of the natural interdomain recombinant HLA-Aw69 molecule. HLA-Aw69, which consists of an alpha 1 domain exhibiting homology with HLA-Aw68, and alpha 2/alpha 3/transmembrane-cytoplasmic domains, exhibiting homologies with HLA-A2, mimics HLA-Aw68 and provides HLA-A,B null target cell (C1R) transfectants with increased resistance to NK. Further, the inability of transfected HLA-A2 to confer protection against NK activity can be completely attributed to the expression of a "nonpermissive" residue at position 74 in the alpha 1 domain. Site-directed mutation of the His-74 residue in HLA-A2 to the Asp-74 (HLA-A3, -Aw68, -Aw69, -B7) residue generates a mutant that provides C1R cell line transfectants an NK-resistant phenotype. As His-74 blocks access to a side pocket in the HLA-A2 antigen-binding cleft, these results support the critical involvement of residues within the peptide-binding groove of class I molecules in determining the NK susceptibility phenotype of class I+ target cells.

Alleles↗
PubMed1990

Dermatitis herpetiformis in two American blacks: HLA type and clinical characteristics.

Dermatitis herpetiformis is a rare blistering skin disease characterized in part by granular IgA deposits at the dermoepidermal junction, an associated gluten-sensitive enteropathy, and a strong association with the human histocompatibility leukocyte antigen (HLA)-A1 (74% of patients with dermatitis herpetiformis), -B8 (88%), -DR3 (95%), and -DQw2 (100%). Dermatitis herpetiformis is rarely seen in American blacks and some investigators have postulated that this finding may be due to the decreased frequency of HLA-A1 and -B8 in American blacks compared with Caucasians (American blacks: HLA-A1 = 15.3%, HLA-B8 = 10.7%; Caucasian: HLA-A1 = 26.4%, HLA-B8 = 18.3%). This report describes two American blacks with dermatitis herpetiformis and reports the results of HLA typing of these subjects for HLA-A, -B, -C, -DR, and -DQ antigens. HLA typing revealed that neither patient expressed HLA-A1 or -B8; however, both patients did express the class II antigens most frequently seen in dermatitis herpetiformis, HLA-DR3 and -DQw2. Comparison of HLA class II antigen frequency in normal American blacks and Caucasians reveals a similar frequency of HLA-DR3 and -DQw2 (American blacks: HLA-DR3 = 27.6%, HLA-DQw2 = 40.9%; Caucasian: HLA-DR3 = 22.6%, HLA-DQw2 = 32.9%). These data confirm the importance of the HLA class II region in the pathogenesis of dermatitis herpetiformis. In addition, these data suggest that the rare occurrence of dermatitis herpetiformis in American blacks is not due to the decreased frequency of the HLA class I antigens -A1 and/or -B8 but rather is related to differences in the HLA class II region not detected by routine HLA typing.

Biopsy↗
PubMed1990

An HLA class II region restriction fragment length polymorphism (RFLP) in patients with dermatitis herpetiformis: association with HLA-DP phenotype.

Dermatitis herpetiformis (DH) is characterized in part by an associated gluten-sensitive enteropathy (GSE), and a strong association with the HLA antigens HLA-A1, -B8, -DR3, and -DQw2, essentially identical to that seen in patients with isolated GSE (celiac disease). A 4.0-kb RsaI RFLP has been identified using a DQ beta-chain cDNA and localized to the HLA-DP beta-chain region. This RFLP has been found more frequently in patients with isolated GSE than in normal HLA matched controls. We have analyzed genomic DNA from 24 patients with DH and 15 HLA-matched controls to determine if this 4.0-kb RsaI RFLP was present in patients with DH. Twenty-one of 24 (87%) of patients with DH were found to have this RFLP as compared to 7 of 10 (70%) HLA-DR3, -DQw2 matched control subjects (p = 0.23). Thus, the 4.0-kb RsaI RFLP detected in patients with isolated GSE is also present in patients with DH; however, its frequency in DH patients does not differ significantly from that of HLA matched controls. Family studies of patients with DH revealed that although the 4.0-kb RsaI RFLP segregated with the HLA-A1, -B8, -DR3, -DQw2 haplotype in one family, it did not segregate with this disease-associated haplotype in two other families. In both patient and control populations, this RFLP was associated with HLA-DPw1 or -DPw3 phenotypes; 25 of 26 (96%) HLA-DPw1 or -DPw3 subjects were found to have this RFLP compared to only 1 of 6 (17%) who did not express HLA-DPw1 or -DPw3 (pc = 0.0009). These population and family data suggest that this 4.0-kb RsaI RFLP is primarily associated with the HLA-DPw1, -DPw3 phenotype, rather than the clinical manifestations of DH. These data further document that the strongest association of DH with HLA antigens remains with HLA-DQw2 and HLA-DR3 antigens.

Cell Line↗
PubMed1989

HLA genotypes and variant alleles in Sudanese families of Arab-Negroid tribal origin.

The indigenous population of the Blue Nile Province, Sudan, is an Arab-Negroid admixture, although some isolates of west African origin (Fallata) have begun to intermix with the indigenous population. Consanguineous marriages are common in these Muslim families. Members of 22 nuclear families were typed for HLA class I and II antigens using complement-dependent microcytotoxicity with Ninth International Workshop (9th WS) and local reagents. Considerable polymorphism was observed at each locus with a majority of the World Health Organization (WHO)-recognized alleles represented in the parental sample, albeit at low frequency. Seven parental haplotypes carried A locus alleles which were not identical to WHO-recognized specificities. All appeared to be Aw19-related specificities. Five B locus serologic variants were observed; all appeared distinct from WHO-recognized specificities. In one family we observed a new DR-DQ association; DR4 segregated with DQw2, rather than with DQw3. As has been observed for several other genetic systems in these tribes, considerable polymorphism was found for all class I and class II gene products in spite of a high level of consanguinity. Tribal admixture and/or a selective advantage in protecting the population against disease may account for this unexpectedly high level of heterozygosity.

Black People↗
PubMed1989

Epitopes within the HLA-B5,B35 cross-reacting group.

The HLA-B5,B35 cross-reacting group is a large and serologically complex antigen family which includes the World Health Organization-recognized specificities HLA-B5,B51,Bw52,B35,Bw53,B18,Bw70,Bw71, and Bw72. In addition, several variants of antigens in this cross-reacting group have been described in the past but have not yet gained official recognition. A genetic basis for the complexity and the protein and molecular bases of this highly cross-reactive and polymorphic cross-reacting group have yet to be established. The potential contributions of shared amino acid sequences, the occurrence of multiple epitopes on a single HLA-B molecule, and the presence of new HLA-C antigens have been difficult to resolve. To address this issue, we have carefully examined the serologic reactions of more than 900 allo- and monoclonal antibodies (Tenth International Workshop, Third Asia-Oceanic Workshop, and local reagents) versus lymphocytes from 92 individuals of diverse ethnic origin (North American Caucasians, North American blacks, Amerindians, Middle Eastern Caucasians), 84 of whom were informative for the HLA-B5,B35 cross-reacting group and related antigens. Our results demonstrate that the HLA-B5,B35 gene products share different combinations of distinct epitopes. We have constructed a model for the evolution of this cross-reacting group by assigning polarity to distinct diversification steps utilizing principles of maximum parsimony.

Antibodies, Monoclonal↗
PubMed1989

A high proportion of ADA point mutations associated with a specific alanine-to-valine substitution.

In 15%-20% of children with severe combined immunodeficiency (SCID), the underlying defect is adenosine deaminase (ADA) deficiency. The overall goal of our research has been to identify the precise molecular defects in patients with ADA-deficient SCID. In this study, we focused on a patient whom we found to have normal sized ADA mRNA by Northern analysis and an intact ADA structural gene by Southern analysis. By cloning and sequencing this patient's ADA cDNA, we found a C-to-T point mutation in exon 11. This resulted in the amino acid substitution of a valine for an alanine at position 329 of the ADA protein. Sequence analysis revealed that this mutation created a new BalI restriction site. Using Southern analyses, we were able to directly screen individuals to determine the frequency of this mutation. By combining data on eight families followed at our institution with data on five other families reported in the literature, we established that five of 13 patients (seven of 22 alleles) with known or suspected point mutations have this defect. This mutation was found to be associated with three different ADA haplotypes. This argues against a founder effect and suggests that the mutation is very old. In summary, a conservative amino acid substitution is found in a high proportion of patients with ADA deficiency; this can easily be detected by Southern analysis.

Adenosine Deaminase↗
PubMed1988

HLA-A and B polymorphisms predate the divergence of humans and chimpanzees.

Major histocompatibility complex (MHC) glycoproteins bind processed fragments of proteins and present them to the receptors of T lymphocytes. The extraordinary polymorphism of class I MHC molecules in man (HLA-A, B and C) and mouse (H-2 K, D and L) poses many questions concerning their diversification and evolution. Comparison of allelic sequences within a species suggests diversity is generated by the assortment of point mutations into varied combinations by mechanisms of recombination and gene conversion. We have now compared class I MHC alleles in two closely related species: humans (Homo sapiens) and chimpanzees (Pan troglodytes). Chimpanzee homologues of HLA-A, HLA-B and a non-classical gene have been identified. No features distinguishing human and chimpanzee alleles could be found. Individual HLA-A or B alleles are more closely related to individual chimpanzee alleles than to other HLA-A or B alleles. These results show that a considerable proportion of contemporary HLA-A and B polymorphism existed before divergence of the chimpanzee and human lines. The stability of the polymorphism indicates that hyper-mutational mechanisms are not necessary to account for HLA-A, B and C diversity.

Alleles↗
PubMed1988

1,25-Dihydroxyvitamin D3 decreases expression of HLA class II molecules in a melanoma cell line.

An HLA class II-positive melanoma cell line, DU-Mel 17, was treated with three compounds known to induce differentiation in various cell lines. Neither retinoic acid nor dibutyryl cAMP altered levels of DR alpha mRNA, but 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) significantly decreased the level of DR alpha mRNA 48 h after treatment. Optimal effect of the hormone on DR alpha mRNA was reached by 72 h. DU-Mel 17 cells were responsive to 1,25(OH)2D3 in a dose-dependent manner, and a reduction in DR alpha mRNA was seen at concentrations as low as 5 x 10(-13) M. The action of 1,25(OH)2D3 on DR alpha mRNA levels was dependent on protein synthesis as evidenced by inhibition of its effect upon addition of cycloheximide. Both DR and DQ protein levels on the surface of DU-Mel 17 were beginning to decline by 72 h after 1,25(OH)2D3 treatment, and by 96 h these proteins were decreased by 65%. 1,25(OH)2D3 was not capable of altering expression of class II molecules on three different class II-positive B lymphoblastoid cell lines, although one of these lines was shown to express the receptor for 1,25(OH)2D3. These findings are important because 1) there is no known physiologic regulator that actively down-regulates class II molecules that are present in and/or on cells, 2) levels of mRNA derived from a very limited number of genes are known to be altered by 1,25(OH)2D3, and 3) they support the contention that 1,25(OH)2D3 may alter the differentiation state of these cells and the activity of the normal counterpart of these cells in an immune response.

Calcitriol↗
PubMed1988

Conversion of a melanoma cell line from an HLA class II positive to negative phenotype after treatment with anti-mycoplasma drugs.

An HLA class II-positive melanoma cell line, DU-Mel 17, expressing both DR and DQ was shown to be contaminated with Mycoplasma hyorhinis. After treatment of the cell line with anti-mycoplasma drugs (a tetracycline derivative and a pleuromutilin), both DR and DQ expression on the cell surface, as well as DR alpha mRNA, were greatly decreased and the cells were rendered mycoplasma-free. Interestingly, drug-treated cells became resistant to geneticin, an unrelated drug. To test whether drug treatment or elimination of mycoplasma caused the abolition of class II expression, contaminated DU-Mel 17 cells were inoculated into peritoneal cavities of nude mice and later removed upon growth of a solid tumor. The cells were shown to be mycoplasma-free, and they expressed DR alpha mRNA, as well as DR and DQ proteins on their surfaces. Although it appears that the drugs were responsible for abating class II molecule expression in DU-Mel 17 cells, the identical drug-treatment regimen on a class II-positive, mycoplasma-free B cell line did not alter DR alpha mRNA or class II protein levels. Based on restriction fragment lengths, the DR alpha gene appeared to be grossly intact after drug treatment, and alterations in methylation patterns were not observed. These studies indicate that anti-mycoplasma treatment of DU-Mel 17 cells resulted in selection of a class II-negative, drug-resistant clone(s). Such treatment may result in a phenotype that is divergent from the parental cell line in one or more characteristics.

Anti-Bacterial Agents↗
PubMed1988

Accelerated development of immunity following transplantation of maternal marrow stem cells into infants with severe combined immunodeficiency and transplacentally acquired lymphoid chimerism.

Transplacentally acquired lymphoid chimerism was detected in two infants with severe combined immunodeficiency (SCID) by two-colour cytofluorographic studies. These cells had no demonstrable function in studies in vitro. Following T cell-depleted maternal bone marrow stem cell transplantation, evidence of T cell function was detected 20 and 50 days later, and transient B cell function was detected 50 days later. These immune functions appeared much sooner than the 90-120 days usually required for T cell function and the 2-2.5 years for B cell function to develop after haplo-identical stem cell transplants into SCID infants without transplacental engraftment. The presence of maternal lymphoid chimerism did not interfere with haplo-identical marrow cell engraftment, even though no pre-transplant immunosuppression was given. This observation suggests that the transplanted maternal marrow stem cell in some way conferred reactivity on the engrafted but apparently non-functional mature T cells that had entered the fetal circulation transplacentally.

B-Lymphocytes↗
PubMed1987

Identification of a deletion in the adenosine deaminase gene in a child with severe combined immunodeficiency.

A patient with adenosine deaminase-deficient severe combined immunodeficiency is described whose defect is secondary to deletion of a portion of the ADA structural gene. In Southern analyses, DNA from this patient does not hybridize to a genomic probe that includes the 3' end of exon 1. This implies that both his parents are heterozygous for deletions of exon 1 sequences. Consistent with this finding, the patient has no detectable adenosine deaminase mRNA by Northern analysis. This is the first report of a deletion mutation as the cause of adenosine deaminase deficiency.

Adenosine Deaminase↗
PubMed1987

Differential expression of the HLA-DR genes in various melanoma cell lines treated with interferon-gamma: methylation of the HLA-DR alpha gene in these lines is not correlated with its expression.

Melanoma cell lines treated with or without interferon-gamma were tested for the presence of DR alpha mRNA and protein. The six lines examined fell into three general categories: two that expressed high levels of DR alpha mRNA and protein before and after interferon-gamma treatment, one that expressed very low levels before treatment with interferon-gamma, but was induced to express high levels after interferon-gamma treatment, and three that expressed very low levels before treatment, and were only slightly inducible after treatment with interferon-gamma. The presence of DR-alpha protein on the melanoma cell surface was always positively correlated with the presence of DR alpha mRNA in the cells. Furthermore, in the cell line that was interferon-gamma-inducible, the time at which DR alpha mRNA and protein appeared and the doses of interferon-gamma needed to induce this appearance were directly correlated. Methylation patterns of the DR alpha gene in these cell lines were also studied in order to determine whether the degree of DR alpha gene methylation among the lines correlated with expression of the gene. Digestion of DNA with the restriction enzyme MspI, which recognizes the sequence 5'CCGG3' and 5'CmCGG3', led to the appearance of a 3.1 kb band from all lines tested. Hpa II digestion, which recognizes 5'CCGG3', but not 5'CmCGG3', led to the appearance of 3.1, 4.4, and 6.7 kb bands in all lines tested except for DUMEL 8, which showed only the 3.1 kb band. Interestingly, DUMEL 8 expressed very low levels of DR alpha mRNA and protein before and after interferon-gamma treatment. We conclude that interferon-gamma has a regulatory effect on DR alpha genes of various melanoma cell lines to varying degrees. This may reflect an effect of interferon-gamma on certain subpopulations of melanocytes in vivo. Our data also indicate that partial methylation of the DR alpha gene does not inhibit its expression. Furthermore, interferon-gamma does not appear to induce expression of the DR gene by altering methylation patterns within the region recognized by our probe.

Cell Line↗
PubMed1986

Development of immunity in human severe primary T cell deficiency following haploidentical bone marrow stem cell transplantation.

Recent advances in the prevention of graft-vs-host disease (GVHD) have allowed the use of haploidentical bone marrow cells for correction of lethal genetic defects of the immune system. Sequential analyses of blood lymphocyte phenotypes and functions were done before and after transplantation of haploidentical marrow stem cells into 17 infants with severe primary T cell deficiencies. The marrow was depleted of post-thymic T cells and most other mature marrow cells by soy lectin agglutination and sheep erythrocyte rosetting. The studies were performed to define the time course and extent of appearance of immune function, and to identify factors leading to resistance to engraftment. No pretransplant immunosuppression was used. T cell function was detected between 34 and 287 days after transplantation, but a sharp rise usually occurred between 84 and 115 days, and normal function was reached between 113 and 210 days. Fifteen of the patients are alive from 6 to 41 mo post-transplantation, 12 have improved or have normal T lymphocyte function, and nine have proven T cell chimerism. Increased immunoglobulins of several isotypes have been noted in 11 patients and specific antibodies in seven patients, although B cell chimerism has been detected in only one patient. B cell function required 2 to 2.5 yr for normalization. No GVHD occurred in 14 patients, and the other three had only transient mild skin rashes. Two patients died of viral infections. Failure to engraft was correlated with some pre-transplant lymphocyte responses to mitogens and allogeneic cells (three cases), but not with the presence of pre-transplant natural killer cell function (five cases) nor with the presence of purine salvage pathway enzyme deficiencies (four cases). The latter, however, was associated with poor lymphoid function in two patients. These studies indicate that the thymic microenvironment of most infants with severe combined immunodeficiency disease is capable of differentiating donor stem cells to mature and functioning T lymphocytes which can cooperate with apparently normal host B cells for antibody production.

Adenosine Deaminase↗
PubMed1986

Further studies on the epitopes of HLA-B7 defined by murine monoclonal antibodies.

Monoclonal antibodies reactive with polymorphic epitopes of HLA-B7 were analyzed by direct and indirect cytotoxicity assays on established panels of HLA typed lymphocytes. This permitted further refinement of their specificity and the identification of various novel reactions. The topographic relationship of polymorphic epitopes on the surface of the B7 molecule was assessed with various serological assays using cell surface B7 or papain solubilized B7 as the antigenic target. These studies focused on monoclonal antibodies recognizing B27 and B7. The results, in combination with those of previously published studies, are used to provide a current assessment of the epitope map of HLA-B7 as defined with mouse monoclonal antibodies. This is compared to the results obtained with alloantisera.

Animals↗
PubMed1985

Successful immune reconstitution in severe combined immunodeficiency despite Epstein-Barr virus and cytomegalovirus infections.

Cytomegalovirus (CMV) and Epstein-Barr virus (EBV), frequently found in the acquired immune deficiency syndrome (AIDS), have been suspected of contributing to the latter immunodeficiency. The ability of normal HLA-identical sibling bone marrow to reconstitute an 8-month-old infant with severe combined immunodeficiency infected with these two viral agents is of interest. After presentation with severe mucocutaneous candidiasis, cavitary pulmonary disease, nodular cutaneous lesions, and hepatic abscesses containing acid-fast organisms, immunologic studies revealed lymphopenia, 1-3% T cells, and no lymphocyte responses to mitogens. Prior to transplantation, the infant's blood B lymphocytes grew spontaneously in culture, suggesting they were infected with EBV. Indeed, an appropriate antibody response to EBV was detected at 2 months post-transplantation. At 3 weeks postgrafting, neutropenia and cholestatic jaundice developed without other signs of graft versus host disease. Liver biopsy demonstrated CMV but no EBV by DNA hybridization. There was evidence of T- and B-cell function by 2 weeks postgrafting, including vigorous in vivo and in vitro responses to candida. Although the blood lymphocyte T4:T8 ratio was inverted at 2 weeks, it reverted to normal by 6 weeks post-transplantation. All clinical disease resolved by 8 months and karotyping revealed all T and B lymphocytes to be XX. Thus, despite infections with both CMV and EBV, complete immunologic reconstitution was achieved in this, the most severe of all genetically determined immunodeficiency conditions, arguing against these viruses having a major role in the failure of bone marrow transplantation in AIDS.

Bone Marrow Transplantation↗