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J A Frelinger

Publications and source records attributed to J A Frelinger.

At least 109 records · Page 6Linked to original sources
PubMed1985

Polymorphic class II sequences linked to the rat major histocompatibility complex (RT1) homologous to human DR and DQ sequences.

Until recently, the analysis of Class II genes linked to the rat major histocompatibility complex, RT1, has been confined to serologic and electrophoretic analysis of their gene products. To obtain a more definitive estimate of the number and relative polymorphism of RT1 Class II sequences, we performed Southern blot analysis of rat genomic DNA employing human cDNA probes specific for Class II heavy and light chain genes. Southern blots of EcoRI and BamHI digests of genomic DNA from ten inbred strains, expressing eight RT1 haplotypes, were hybridized with the human DQ beta or DR beta cDNA that are homologous to Class II light chain sequences. Four to eight bands were observed to hybridize with the light chain cDNA: band sizes ranged from 2.5 to 28 kb. Restriction fragment patterns were polymorphic; the only identical patterns observed were those associated with RT1 haplotypes with identical RT1.B regions. The number and size of bands hybridizing with DQ beta and DR beta suggested a minimum of four light chain sequences in each haplotype. Southern blots of BamHI and EcoRI digests of genomic DNA from the same strains were hybridized with a DR alpha cDNA that is homologous to Class II heavy chain sequences. All RT1 haplotypes expressed either a 10.0-kb or 13.0-kb band when digested with BamHI, and either a 17-kb or 3.7-kb band when digested with EcoRI. Considerably less polymorphism was detected with the DR alpha probe; this observation is consistent with previously reported limited protein polymorphism of the rat equivalent of the I-E alpha subunit. The size and number of bands hybridizing with the DR alpha probe suggests a minimum of two heavy chain sequences. These observations suggest that the RT1 complex includes more Class II sequences than have been observed in serologic and electrophoretic analyses of Class II gene products. Furthermore, the level of polymorphism of RT1 Class II sequences appears to be comparable with mouse and human Class II sequences.

Animals↗
PubMed1984

Distinct epitopes are recognized by cytolytic T lymphocyte clones on the same class I molecule: direct demonstration using DNA-transfected targets and long-term cytolytic T cell clones.

By producing long-term, stable, cytolytic T lymphocyte clones and utilizing targets expressing only a single gene product derived from the stimulator mouse strain, we have been able to directly demonstrate that T cells recognize distinct epitopes expressed on a single H-2 molecule. These multiple determinants are distinguishable by inhibition patterns with monoclonal antibodies (mAb). When two T cell clones, P-2.14 and P-2.17, are tested on an L cell transfected with the Dp gene (lambda 12a), the T cells kill the transfected targets as well as blasts derived from Dp mouse strains. mAb 7-16.10 inhibits recognition and killing of Dp targets by both P-2.14 and P-2.17. This mAb recognizes a private specificity H-2.m22. Interestingly mAb 11-20.3 which also recognizes the H-2.m22 specificity inhibits clone P-2.14 but not P-2.17. The mAb 7-16.10, however, competes with 11-20.3 for binding to the surface of L cells expressing the Dp gene. Thus the two T cells must recognize an overlapping specificity. Other mAb which bind to the H-2Dp molecule are unable to inhibit either of these two cytolytic T cell clones. Paradoxically, any of the mAb when tested individually are sufficient to inhibit the polyclonal response derived from in vitro mixed lymphocyte culture. Therefore, by using targets expressing only a single H-2 molecule derived by DNA-mediated gene transfer and cytolytic T cell clones we have been able to directly demonstrate the presence of multiple epitopes on a single molecule and define their inhibition with mAb directed to that same molecule.

Animals↗
PubMed1984

Cloning and identification of the H-2Dp gene.

We have cloned six different class I genes from a B10.P sperm library. After cotransfection with the herpes simplex tk gene, one L-cell line was found to react with six H-2Dp-specific monoclonal antibodies. The cell line L12a did not react with Kp-specific monoclonal antibodies. This identification was confirmed by mapping a 2.5 kb Bam H1 restriction fragment present in the lambda 12a DNA clone to the D-TL region of H-2p. Only a single 8.8 kb Bam H1 fragment can be assigned to Kp by restriction fragment length polymorphism, while many others map to the D-TL interval. A restriction map of lambda 12a is presented.

Animals↗
PubMed1984

Functional characterization of the H-2Dp gene product.

A lambda clone containing the Dp gene was used to transform L cells. The Dp product expressed was identified by two-dimensional gel electrophoresis and flow cytometry. The Dp product expressed by the L cells was recognized by Dp-specific but not Kp-specific killer T cells. This killing was inhibited by monoclonal antibodies specific for Dp but not Kp or Kk antigens. Similarly, lymphocytic choriomeningitis virus (LCMV) killer T cells from B10.P mice were able to kill LCMV-infected L12a cells, but not LCMV-infected Ltk+. Again only Dp monoclonal antibodies could inhibit this killing.

Animals↗
PubMed1984

Production and characterization of T cell clones specific for mouse hepatitis virus, strain JHM: in vivo and in vitro analysis.

Mouse hepatitis virus, strain JHM, is a coronavirus capable of inducing both acute and chronic central nervous system disease. To characterize the cell-mediated immune response directed toward JHMV antigens, we have generated and analyzed the properties of four T cell clones reactive to JHMV antigens. All four clones are of the Ly-1+2- phenotype and show an antigen-specific, I-Ab-restricted proliferative response that is dependent on antigen-presenting cells (APC). The fine specificity of I-A restriction was revealed by their differential recognition of JHMV antigen in the context of mutant I-A molecules present on B6.C-H-2bm12 APC. Two clones were unable to recognize JHMV antigen presented on bm12 APC, whereas a third clone showed an equivalent response with both bm12 and wild type B6 APC. In the analysis of the viral specificity, three of the clones showed distinct cross-reactions with MHV-2 and very little if any cross-reaction with MHV-A59. In addition, three of three clones tested responded well to sucrose gradient-purified, UV-inactivated JHMV, indicating that their specificity is directed toward virus structural rather than nonstructural proteins. Finally, two of the clones were analyzed for their ability to induce a DTH reaction when injected together with antigen into the footpads of various mouse strains. Consistent with the in vitro data, both clones produced a marked DTH response only in those strains carrying the I-Ab genotype. These data demonstrate, both in vivo and in vitro, that at least one component of the cell-mediated immune response to JHMV is characterized by an Ly-1+2-, I-A-restricted T cell specific for JHMV structural proteins.

Animals↗
PubMed1983

Brain Ia antigens have a bone marrow origin.

Our results, using radiation-induced bone marrow chimeras, demonstrate that the Ia antigen found in the brains of such animals is produced by cells having precursors in the bone marrow. These cells are not immediately blood borne since no IgM is detected in these brains. This rules out the obvious possibility of B-lymphocyte contamination as the source of Ia in the brain cell preparations. It thus appears that the central nervous system, like many other nonlymphoid organs, has a source of Ia-positive cells that are derived from bone marrow precursors.

Animals↗
PubMed1983

Antigenic relationships of murine coronaviruses: analysis using monoclonal antibodies to JHM (MHV-4) virus.

Monoclonal antibodies were produced to JHMV-DL, a neurotropic member of the mouse hepatitis virus (MHV) or murine coronavirus group. Of 23 antibodies isolated, 10 were specific for the major envelope glycoprotein, gp180/90, 10 for the nucleocapsid protein, pp60, and 3 for the minor envelope glycoprotein, gp25. Eleven different MHV isolates were used in antibody binding assays to study antigenic relationships among the viruses. Each MHV isolate tested had a unique pattern of antibody binding, indicating that each is a distinct strain. Conservation of JHMV-DL antigenic determinants varied among the three proteins, with pp60 showing intermediate conservation, gp180/90 little conservation, and gp25 marked conservation in the different MHV strains. Monoclonal antibodies to pp60 proved most useful in delineating antigenic relationships among MHV strains. These antigenic groups correlated with pathogenic types, indicating that pp60 may be one of the gene products which mediates the distinct disease patterns manifested by different murine coronaviruses.

Animals↗
PubMed1983

Evidence for extensive polymorphism of class I genes in the rat major histocompatibility complex (RT1).

The major histocompatibility complex of the rat (RT1) has been poorly characterized with respect to the number, linkage, and polymorphism of class I genes. To estimate the number of class I RT1 genes and the relative extent of their polymorphism, we performed Southern blot analysis with liver DNA from rat strains expressing eight RT1 haplotypes. After digestion with EcoRI and BamHI, the DNA was separated on agarose gels, blotted onto nitrocellulose, and hybridized with mouse H-2 cDNA probes, pH-2III and pH-2IIa. Ten to 20 EcoRI and 13 to 20 BamHI bands hybridized with pH-2III and pH-2IIa; restriction fragment length patterns were observed to be highly polymorphic. The restriction fragments associated with different RT1 haplotypes differed by 17-70%; this range is similar to the differences observed between mouse H-2 haplotypes. The same restriction fragment pattern was observed in DNA from three different rat strains sharing the same RT1 allele, confirming that the patterns were RT1-associated. Further, the RT1l and RT1lvl haplotypes, which differ at a single previously identified RT1-linked locus, were associated with EcoRI restriction pattern differences of 39-50%, confirming the supposition that RT1 class I genes identified by previous serological and T-cell-mediated assays have identified only a minority of the actual number of RT1-linked class I genes. In summary, the results reported in this communication demonstrate that the RT1 complex encompasses a large family of highly polymorphic class I genes similar to the H-2 and HL-A complexes of mouse and man.

Alleles↗
PubMed1983

Low-density mononuclear cells. Potent stimulators of the human MLR.

We have applied purification strategies similar to those used to purify murine dendritic cells to human peripheral blood in an attempt to enrich for stimulators of the human mixed lymphocyte reaction (MLR). Equilibrium density centrifugation of peripheral blood mononuclear cells yields a population of low density cells that are potent stimulators of a human MLR. The stimulator cells are Dr+, nonlymphocytic, and weakly adherent. Strongly adherent monocytes, also present in the low-density cell population, do not stimulate a human MLR. This contrasts with other human MLR studies that ascribe stimulatory activity to adherent monocytes, and it indicates functional and morphological heterogeneity among monocytes.

Cell Separation↗
PubMed1982

Acute autoimmune encephalomyelitis in mice. II. Susceptibility is controlled by the combination of H-2 and histamine sensitization genes.

The expression of acute experimental autoimmune encephalomyelitis (EAE) in mice is controlled by several dominant genes, H-2 and histamine sensitization genes. SJL/J and SWR/J, which are H-2s and H-2q, respectively, are susceptible to EAE and sensitive to Bordetella pertussis histamine-sensitizing factor (HSF), which produces a vasoactive amine hypersensitivity. Other H-2s or H-2q strains such as A.SW, B10.Q and several others do not develop acute EAE and are not sensitive to B. pertussis HSF. One strain tested, DDD (KsIsD?) is HSF sensitive but does not develop EAE (presumably because it lacks the appropriate responder H-2 haplotype). However, F1 hybrids between B10.S and DDD are sensitive to HSF and develop EAE. The induction and effector phases of acute EAE are apparently controlled by the combination of H-2 and HSF genes. A combination of the correct H-2 hapotype and histamine sensitivity is required for the development of acute EAE.

Acute Disease↗
PubMed1982

Identification of a BALB/c H-2Ld gene by DNA-mediated gene transfer.

Gene transfer and immunoselection were used in the identification of a BALB/c genomic clone containing an H-2Ld gene (clone 27.5). Transformation of thymidine kinase-negative C3H mouse L cells with the cloned 27.5 DNA together with the herpes simplex virus tk gene produced transformants expressing Ld molecules detected by radioimmune assay with monoclonal hybridoma antibodies to Ld antigens. The foreign Ld gene products expressed by cloned mouse L cell transformants were shown to be virtually indistinguishable from BALB/c spleen Ld molecules by two-dimensional electrophoretic analysis of H-2Ld immunoprecipitates. These results indicate that the genomic clone 27.5 contains a functional BALB/c H-2Ld gene and demonstrate the usefulness of this approach for identifying the gene products encoded by cloned genes which are members of a multigene family. Furthermore, the ability to place cell-surface recognition molecules on the surfaces of foreign cells provides a powerful opportunity for functional analyses of these molecules.

Animals↗
PubMed1982

Specific recognition of the product of a transferred major histocompatibility complex gene by cytotoxic T lymphocytes.

Mouse L cells transfected with a genomic clone containing the H-2Ld gene (8-5 cells) were shown to function as targets for H-2Ld-specific cytotoxic T lymphocytes (CTL). The CTL-mediated lysis of 8-5 cells was shown to be H-2Ld specific by the use of (i) CTL with restricted reactivity, (ii) unlabeled target inhibiton, and (iii) monoclonal antibody inhibiton. We also demonstrated that 8-5 cells could function as targets for antibody-plus-complement-mediated cell lysis. Specificity was confirmed by using H-2Ld-specific monoclonal antibodies. These experiments demonstrate that the gene products of a major histocompatibility complex genomic clone can be functionally expressed in a foreign cell and can mediate immunologically specific cellular interactions.

Animals↗