Characterization of the novel HLA-B*18:79 allele.

Characterization of the novel HLA B*18:79 allele is described.

Characterization of a novel allele, HLA-B*15:228.

B*15:228 differs from B*15:01:01:01 at three nucleotides in exon 4.

Characterization of a novel allele, HLA-A*03:132.

A*03:132 differs from A*03:01:01:01 at nucleotide 853 (codon 261) in exon 4.

Characterization of four novel HLA alleles, including two in the same haplotype.

Characterization of the novel HLA alleles A*02:330, A*11:108, B*40:175, and B*40:176 is described.

Characterization of three novel HLA alleles with silent nucleotide substitutions.

Characterization of the novel HLA alleles B*35:11:03, B*42:01:03, and B*51:01:35 is described.

Identification of the HLA-Cw*0774 and DQB1*060105 alleles.

Cw*0774 differs from Cw*070201 by one nucleotide within the coding sequence of exons 2-4. DQB1*060105 differs from DQB1*060101 by one nucleotide within the coding sequence of exons 2-3.

A novel HLA-B*44 allele, B*4466, discovered in a potential donor for a hematopoietic stem cell transplant.

This report describes the discovery and characterization of the human leukocyte antigen-B*4466 allele.

Discovery of a novel HLA-Cw*08 allele, Cw*0817.

This report describes the discovery and characterization of the HLA-Cw*0817 allele.

Ectogalactosyltransferase studies in fibroblasts and concanavalin A-stimulated lymphocytes.

In this communication, we have demonstrated that hydrolysis of the nucleotide sugar can cause errors in the detection of an ectoglycosyltransferase. Spleen cell suspensions can incorporate radioactivity when incubated with labeled UDP-galactose, but all the activity is due to decomposition of the nucleotide sugar and uptake of the free sugar. The fibroblast cell lines can incroporate carbohydrate directly from UDP-galactose. Several criteria are presented with can be used to demonstrate that a nucleotide sugar is the direct carbohydrate donor.

Stimulation of glycosaminoglycan synthesis in cultured human dermal fibroblasts by interleukin 1. Induction of hyaluronic acid synthesis by natural and recombinant interleukin 1s and synthetic interleukin 1 beta peptide 163-171.

Hyaluronic acid (HA) is believed to play a critical role in wound healing and in morphogenesis. Factors controlling the production of HA by fibroblasts in normal and pathological states are not completely understood. In this report we have observed that natural human interleukin (IL-1)1 beta and human recombinant (hrIL)-1 alpha and beta are potent stimulators of HA production by fibroblasts in vitro. Hyaluronic acid is the major species of glycosaminoglycan (GAG) stimulated by IL-1 in fibroblasts. PGE2 does not appear to be involved directly in this IL-1 effect on fibroblasts, but stimulation of HA production by IL-1 is dependent on protein synthesis. The synthetic human IL-1 beta peptide 163-171 (Val-Gln-Gly-Glu-Glu-Ser-Asn-Asp-Lys), which has been previously shown to stimulate thymocyte proliferation but not fibroblast PGE2 production, is also able to stimulate fibroblast HA production. The synthesis and secretion of IL-1 by mononuclear phagocytes at sites of inflammation and immune reactions in vivo could potentially serve as a signal for fibroblasts to synthesize HA, which in turn could serve to facilitate and modulate reparative and immune processes by virtue of its ability to alter cell-cell, cell matrix, and cell-membrane receptor interactions.

Transforming growth factor-beta inhibits interferon-gamma-induced HLA-DR expression by cultured human fibroblasts.

This study shows the induction of HLA-DR (DR) in fibroblasts by IFN-gamma and investigates the molecular mechanisms involved in the further DR down-regulation by TGF-beta 1. Kinetics of DR induction on human dermal fibroblasts by IFN-gamma showed that 1 hr of exposure was required to induce detectable levels of DR, and maximal DR expression was achieved only after 2 days of exposure to IFN-gamma. TGF-beta 1 inhibited DR induction by IFN-gamma, although complete inhibition never could be achieved, even with high concentrations of TGF-beta 1 and low concentrations of IFN-gamma. Inhibition was not accounted for by reduction in cell numbers, as TGF-beta 1 stimulated growth of the fibroblasts. Inhibition of DR induction was seen only if TGF-beta 1 was added during the first 24 hr of IFN-gamma treatment. TGF-beta 1 inhibited equally well if the cells were pretreated for as little as 1 hr and then washed before addition of IFN-gamma. TGF-beta 1 did not cause an overall suppression of protein synthesis. Northern blot analysis revealed that TGF-beta 1 greatly reduced the steady-state level of DR beta mRNA induced by IFN-gamma at 24 hr, and then DRP transcripts became undetectable at later stages. It is concluded that early intracellular signals must build up to stimulate maximum DR synthesis, which, later on, are inactivated or degraded by the action of TGF-beta 1. We suggest that these mechanisms regulating DR gene transcription involve the action of genes coding for specific IFN-gamma-inducible transcriptional factors that are turned on and off in an expeditious manner.

A novel HLA-B*51 allele, HLA-B*5149.

Discovery of the novel HLA-B*5149 allele in a North American Caucasian individual is described. It differs from B*510101 by one nucleotide within the coding sequence of exons 1-6. A substitution at nucleotide position 488 in exon 3 changes alanine to glycine in amino acid position 139.

A novel HLA-C locus allele with codon 54 deletion, Cw*0346.

This report describes the discovery and characterization of the novel Cw*0346 allele.

A novel HLA-Cw*05 allele, Cw*0517.

This report describes the discovery and characterization of the HLA-Cw*0517 allele.

A novel DRB1*04 allele.

Discovery of the novel DRB1*0464 allele is described. This allele contains a nucleotide substitution in codon 13 that changes the amino acid histidine coded for in all other DRB1*04 alleles to tyrosine. This allele was found in a parent and one child of a North American Caucasian family with the haplotype: A*03, B*07, DRB1*0464, DRB4*0103, DQB1*0301.

Antigen recognition by T cells. II. Intravenous administration of native or denatured ovalbumin results in tolerance to both forms of the antigen.

In the accompanying report, suppressor T cells were demonstrated that did not recognize cross-reactivity between native and denatured ovalbumin (N-OVA and D-OVA). Here we show that the T cell tolerance induced by i.v. injections of antigen does detect cross-reactivity between N-OVA and D-OVA. Mice that had been immunized with either N-OVA or D-OVA in adjuvant could be rendered profoundly unresponsive if either N-OVA or D-OVA, but not an unrelated protein, were injected i.v. Cross-tolerance was observed in assays of antigen-induced T cell proliferation and helper T cell activity. Tolerance was distinguished from suppressor T cell activity by three criteria: 1) specificity for N-OVA and D-OVA, 2) sensitivity to abrogation by cyclophosphamide, 3) duration of effectiveness. These results confirm observations made by others that suggest that tolerance is mediated by an additional mechanism(s) other than suppressor T cells. Based on a hypothesis that cross-reactivity between native and denatured antigen is related to macrophage processing of antigen, these data also suggest a critical role for processed antigen in the induction of tolerance when antigen is administered i.v.

T-cell-independent stimulation of immunoglobulin secretion in resting human B lymphocytes by the mannose-specific adhesin of Escherichia coli type 1 fimbriae.

Purified Escherichia coli type 1 fimbriae have been shown previously to stimulate T-cell-independent proliferation of human B lymphocytes. The response is mediated by the mannose-specific, lectin-like adhesin protein FimH. Here we show that type 1 fimbriae also stimulate immunoglobulin (Ig) secretion by B cells. The response was maximal at three days of culture and consisted predominantly of the IgM isotype. It was independent of serum components, T lymphocytes, monocytes, and natural killer cells. Highly purified resting B cells were induced to proliferate and secrete Ig in response to the fimbriae. The role of FimH in the response was shown by the failure of FimH- type 1 fimbriae to stimulate and by inhibition of the response with alpha-methyl mannoside. In light of the fact that carbohydrate-binding adhesins have been found on a wide variety of microorganisms, these studies suggest the possibility that responses of other cell types to other microbial adhesins will be discovered.

An IL 2-secreting T cell hybridoma that responds to a self class I histocompatibility antigen in the H-2D region.

A self-reactive T cell hybridoma that secretes IL-2 in response to H-2d haplotype cells resulted from a fusion of BALB/cBy lymph node cells with the AKR thymoma BW5147. The lymph node cells used had been enriched for cells reactive to (TG)-A--L, but neither this antigen nor fetal calf serum were required for stimulation of the hybridoma designated 3DT52.5. The gene product responsible for stimulation mapped to the H-2D region. Allogeneic cells of the b, f, k, q, and s haplotypes failed to stimulate. Not all H-2d haplotype cells were effective stimulators of 3DT52.5. Peritoneal cells and splenic B cells were much more stimulatory than splenic T cells. Most tumor cell lines of H-2d derivation and of B cell or macrophage/monocyte lineage were stimulatory, whereas H-2d T cell lines were not. The capacity to stimulate 3DT52.5 did not correlate with the ability to stimulate I region-restricted hybridomas, or with the ability to be induced to stimulate such hybridomas. Stimulatory cell lines did not apparently produce a soluble factor required for stimulation, and negative cell lines were not inhibitory. The monoclonal antibody 27-11-13, which reacts with H-2D of the b, d, and q haplotypes, inhibited stimulation of 3DT52.5 but did not inhibit stimulation of the sibling hybridoma 3DT18.11, which responds to (TG)-A--L plus I-Ad. Conversely, the monoclonal anti-I-Ad antibody MK-D6 inhibited stimulation of 3DT18.11 but not 3DT52.5. Although it is clear that 3DT52.5 recognizes a class I antigen coded for in the H-2D region, the precise molecular nature of the antigen is unknown. The structure of the antigen receptor on this hybridoma may prove to be of interest when it can be compared with receptors found on T cell hybridomas restricted by class II histocompatibility antigens.

Fragmentation of Escherichia coli type 1 fimbriae exposes cryptic D-mannose-binding sites.

Cells of the gram-negative bacterium Escherichia coli are able to attach to various host cells by means of a mannose-specific adhesin associated with type 1 fimbriae. Here we show that fragmentation of type 1 fimbriae by freezing and thawing results in increased mannose-binding activity as demonstrated by increased hemagglutination, increased stimulation of human lymphocyte proliferation, and increased binding of the mannose-containing enzyme horseradish peroxidase. Increased activity in all three assays was mannose sensitive and was not exhibited by FimH- mutant type 1 fimbriae lacking the adhesin. Scatchard analysis of the data from peroxidase binding assays showed that unfrozen and frozen fimbriae contain binding sites displaying two classes of affinity. Frozen and thawed fimbriae expressed an increase in the number of high-affinity binding sites. These results show that fragmentation of the fimbrial structure exposes cryptic mannose-binding activity associated with type 1 fimbriae, presumably that of internally located adhesin molecules. Our data support earlier observations that adhesin moieties of type 1 fimbriae are located both at the tips and at intervals along the length of the fimbriae. In addition, our data suggest that only the adhesin moieties that are located at the fimbrial tips are functional in binding mannose. Adhesins located along the length of the fimbriae have their mannose-binding activity buried within the fimbrial structure and hence are not functional. We propose an updated model for the structure of type 1 fimbriae that is in agreement with the above observations.

Mitogenic stimulation of human B lymphocytes by the mannose-specific adhesin on Escherichia coli type 1 fimbriae.

Escherichia coli type 1 fimbriae contain in association with the major structural protein a lectin-like adhesin moiety that mediates attachment of E. coli to mannose-containing receptors on the surface of host cells. We have investigated the lymphocyte mitogenic activity of this mannose-specific adhesin by comparing the ability of purified wild type type 1 fimbriae containing the adhesin and mutant type 1 fimbriae lacking the adhesin to stimulate proliferation in human lymphocytes. Both fimbriae stimulated a peak of proliferation at 8 days whereas only the wild type fimbriae stimulated an additional peak of proliferation occurring at 3 days. Proliferation at 3 days but not at 8 days could be blocked by the addition of alpha-methyl-D-mannoside. Neonatal lymphocytes from umbilical cord blood responded to both wild type and mutant fimbriae in a fashion similar to adult cells. Stimulation of separated T and non-T cell populations indicated that the proliferation seen at 3 days was solely due to non-T cells whereas the 8-day response was due to T cell proliferation. The addition of gamma-irradiated T cells did not appear to enhance the 3-day response of the non-T cells. However, the 8-day response by T cells was dependent on the presence of gamma-irradiated non-T cells. In cultures of unseparated cells, wild type fimbriae stimulated more than 75% of the B cells to enter the S and G2 phase at 3 days whereas at 8 days cycling T cells were present in both wild type and mutant fimbriae-stimulated cultures. Taken together, our observations suggest that the adhesin molecule stimulates a polyclonal mitogenic response in B cells that peaks at 3 days, and other structural components of the fimbriae are responsible for evoking an 8-day (probably immune) response in T cells.