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J Magram

Publications and source records attributed to J Magram.

At least 37 records · Page 2Linked to original sources
PubMed1996

IL-12 antagonism induces T helper 2 responses, yet exacerbates cardiac allograft rejection. Evidence against a dominant protective role for T helper 2 cytokines in alloimmunity.

IL-12 promotes Th1 development and inhibits the generation of Th2 by inducing IFN-gamma production. In several experimental models, Th2 are preferentially induced in the absence of IL-12. It was proposed that the preferential induction of Th2 by IL-12 antagonism would inhibit Th1-driven rejection responses, thereby promoting allograft acceptance. To test this possibility, mouse cardiac allograft recipients were treated with either anti-IL-12 Abs, or with the IL-12 receptor antagonist p40 homodimer. Unmodified rejection is characterized by a Th1-dominated response, with Th2 cytokines being absent or only weakly expressed within the allograft. Though both forms of IL-12 antagonism induced Th2 cytokine expression within the allograft, these treatments surprisingly exacerbated graft rejection relative to control animals. Interestingly, IL-12 antagonism did not inhibit IFN-gamma gene expression or in vivo sensitization of IFN-gamma-producing cells. Similar observations were made when IL-12 p40 knockout mice were used as allograft donors and recipients, verifying that IL-12 was not required for Th1 development. Further, IL-12 antagonism was associated with strong expression of p40 and weak expression of p35 within the graft. Neither p35 nor p40 mRNAs were detectable in control allografts. These data indicate that while IL-12 antagonism does induce Th2 cytokine expression within cardiac allografts, Th2 cytokines do not play a dominant protective role in the rejection process. Further, the Th2-inducing activity of IL-12 antagonism is not related to decreased IFN-gamma production, but may reflect altered regulation of IL-12 itself.

Animals↗
PubMed1996

HLA-DR4-IE chimeric class II transgenic, murine class II-deficient mice are susceptible to experimental allergic encephalomyelitis.

To investigate the development of HLA-DR-associated autoimmune diseases, we generated transgenic (Tg) mice with HLA-DRA-IE alpha and HLA-DRB1*0401-IE beta chimeric genes. The transgene-encoded proteins consisted of antigen-binding domains from HLA-DRA and HLA-DRB1*0401 molecules and the remaining domains from the IE(d)-alpha and IE(d)-beta chains. The chimeric molecules showed the same antigen-binding specificity as HLA-DRB1*0401 molecules, and were functional in presenting antigens to T cells. The Tg mice were backcrossed to MHC class II-deficient (IA beta-, IE alpha-) mice to eliminate any effect of endogenous MHC class II genes on the development of autoimmune diseases. As expected, IA alpha beta or IE alpha beta molecules were not expressed in Tg mice. Moreover, cell-surface expression of endogenous IE beta associated with HLA-DRA-IE alpha was not detectable in several Tg mouse lines by flow cytometric analysis. The HLA-DRA-IE alpha/HLA-DRB1*0401-IE beta molecules rescued the development of CD4+ T cells in MHC class II-deficient mice, but T cells expressing V beta 5, V beta 11, and V beta 12 were specifically deleted. Tg mice were immunized with peptides, myelin basic protein (MBP) 87-106 and proteolipid protein (PLP) 175-192, that are considered to be immunodominant epitopes in HLA-DR4 individuals. PLP175-192 provoked a strong proliferative response of lymph node T cells from Tg mice, and caused inflammatory lesions in white matter of the CNS and symptoms of experimental allergic encephalomyelitis (EAE). Immunization with MBP87-106 elicited a very weak proliferative T cell response and caused mild EAE. Non-Tg mice immunized with either PLP175-192 or MBP87-106 did not develop EAE. These results demonstrated that a human MHC class II binding site alone can confer susceptibility to an experimentally induced murine autoimmune disease.

Amino Acid Sequence↗
PubMed1996

Genetically resistant mice lacking interleukin-12 are susceptible to infection with Leishmania major and mount a polarized Th2 cell response.

Mice with homologous disruption of the gene coding for either the p35 subunit or the p40 subunit of interleukin-12 (IL-12) and derived from a strain genetically resistant to infection with Leishmania major have been used to study further the role of this cytokine in resistance to infection and the differentiation of functional CD4+ T cell subsets in vivo. Wild-type 129/Sv/Ev mice are resistant to infection with L. major showing only small lesions which resolve spontaneously within a few weeks and develop a type 1 CD4+ T cell response. In contrast, mice lacking bioactive IL-12 (IL-12p35-/- and IL-12p40-/-) developed large, progressing lesions. Whereas resistant mice were able to mount a delayed-type hypersensitivity (DTH) response to Leishmania antigen, susceptible BALB/c mice as well as IL-12-deficient 129/Sv/Ev mice did not show any DTH reaction. To characterize the functional phenotype of CD4+ T cells triggered in infected wild-type mice and IL-12-deficient mice, the expression of mRNA for interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) in purified CD4+ lymph node cells was analyzed. Wild-type 129/Sv/Ev mice showed high levels of mRNA for IFN-gamma and low levels of mRNA for IL-4 which is indicative of a Th1 response. In contrast, IL-12- deficient mice and susceptible BALB/c mice developed a strong Th2 response with high levels of IL-4 mRNA and low levels of IFN-gamma mRNA in CD4+ T cells. Similarly, lymph node cells from infected wild-type 129 mice produced predominantly IFN-gamma in response to stimulation with Leishmania antigen in vitro whereas lymph node cells from IL-12-deficient mice and susceptible BALB/c mice produced preferentially IL-4. Taken together, these results confirm in vivo the importance of IL-12 in induction of Th1 responses and protective immunity against L. major.

Animals↗
PubMed1996

Reduced incidence and severity of collagen-induced arthritis in interleukin-12-deficient mice.

Collagen-induced arthritis (CIA) is an animal model for rheumatoid arthritis. The disease is elicited by immunization of genetically susceptible DBA/1 mice with type II collagen, resulting in a debilitating arthritis characterized by inflammation and involvement of multiple joints. We investigated the role of endogenous interleukin (IL)-12 in the pathogenesis of this disease by undertaking an analysis of IL-12-deficient mice on the DBA/1 genetic background after immunization with type II collagen. Both the incidence and severity of disease were significantly reduced in mice unable to produce biologically active IL-12. Concomitant decreases were observed in serum levels of pathogenic, collagen-specific IgG2a antibodies and collagen-induced secretion of interferon-gamma by immune splenocytes in vitro, consistent with an impaired T helper-1 response. There were, however, a few animals which developed severe disease in a single paw in spite of this highly diminished Th1 response. Taken together, these results demonstrate an important role for IL-12 in the pathogenesis of CIA, although it is not absolutely required for disease development.

Animals↗
PubMed1996

Interleukin-12 an integral cytokine in the immune response.

Interleukin 12 (IL-12) is a heterodimeric cytokine that is produced primarily by antigen-presenting cells and plays a primary role in the induction of cell-mediated immunity. This function is promoted by the IL-12 induced production of interferon-gamma (IFN-gamma) from both resting and activated NK and T cells, by the proliferative activity of IL-12 on activated NK and T cells, by enhancing the cytotoxic activity of NK cells, and by supporting cytotoxic T lymphocyte generation. IL-12 and IL-12-induced IFN-gamma promote the development of naive T cells into Th1 cells and the proliferation and IFN-gamma secretion by differentiated Th1 cells in response to antigen. IL-12 has been found to exhibit many of these activities in vivo, as well as in vitro, and thus IL-12 plays an important role in both innate resistance and antigen-specific adaptive immunity to intracellular bacterial, fungal, and protozoan pathogens. Due to its effects on T cells, recombinant IL-12 has been shown to have therapeutic activity in a variety of mouse tumor and infectious disease models and is being evaluated in clinical trials in human cancer patients. IL-12 also appears to play a role in the genesis of some forms of immunopathology, including endotoxin-induced shock and some autoimmune diseases associated with aberrant Th1 activity. Therefore, IL-12 antagonists may also have therapeutic potential in the treatment of auto immune disorders.

Animals↗
PubMed1996

IL-12-deficient mice are defective in IFN gamma production and type 1 cytokine responses.

IL-12 is a cytokine that can exert regulatory effects on T and NK cells and promote Th1 responses. To delineate further the physiologic role of IL-12 in immunity, mice deficient for this cytokine were generated. IL-12-deficient mice were impaired but not completely lacking in the ability to produce IFN gamma following endotoxin administration and to mount a Th1 response in vivo, as measured by antigen-induced IFN gamma secretion by immune lymph node cells in vitro. In contrast, secretion of IL-4 was enhanced, while proliferation and secretion of IL-2 and IL-10 were normal following antigen stimulation. DTH responses were significantly reduced in IL-12-deficient mice, but no defect in allogeneic CTL responses was observed. These results indicate that IL-12 plays an essential role in regulating IFN gamma production and in facilitating normal DTH responses. However, other phenomena associated with Th1 responses and cell-mediated immunity, i.e., IL-2 secretion and CTL generation, were not compromised in the absence of IL-12.

Animals↗
PubMed1995

Regulation of interleukin 12 p40 expression through an NF-kappa B half-site.

Interleukin 12 (IL-12) is an inducible cytokine composed of 35- and 40-kDa subunits that is critical for promoting T helper type 1 development and cell-mediated immunity against pathogens. The 40-kDa subunit, expressed by activated macrophages and B cells, is induced by several pathogens in vivo and in vitro and is augmented or inhibited by gamma interferon (IFN-gamma) or IL-10, respectively. Control of IL-12 p40 expression is therefore important for understanding resistance and susceptibility to a variety of pathogens, including Leishmania major and perhaps human immunodeficiency virus. In this report, we provide the first characterization of IL-12 p40 gene regulation in macrophages. We localize inducible activity of the promoter to the sequence -122GGGGAATTTTA-132 not previously recognized to bind Rel family transcription factors. We demonstrate binding of this sequence to NF-kappa B (p50/p65 and p50/c-Rel) complexes in macrophages activated by several p40-inducing pathogens and provide functional data to support a role for NF-kappa B family members in IL-12 p40 activation. Finally, we find that IFN-gamma treatment of cells enhances this binding interaction, thus potentially providing a mechanism for IFN-gamma augmentation of IL-12 production by macrophages.

Animals↗
PubMed1995

Vascular endothelial cell lineage-specific promoter in transgenic mice.

Vascular endothelial cells play essential roles in the function and development of the cardiovascular system. However, due to the lack of lineage-specific markers suitable for molecular and biochemical analyses, very little is known about the molecular mechanisms that regulate endothelial cell differentiation. We report the first vascular endothelial cell lineage-specific (including angioblastic precursor cells) 1.2 kb promoter in transgenic mice. Moreover, deletion analysis of this promoter region in transgenic embryos revealed multiple elements that are required for the maximum endothelial cell lineage-specific expression. This is a powerful molecular tool that will enable us to identify factors and cellular signals essential for the establishment of vascular endothelial cell lineage. It will also allow us to deliver genes specifically into this cell type in vivo to test specifically molecules that have been implicated in cardiovascular development. Furthermore, we have established embryonic stem (ES) cells from the blastocysts of the transgenic mouse that carry the 1.2 kb promoter-LacZ reporter transgene. These ES cells were able to differentiate in vitro to form cystic embryoid bodies (CEB) that contain endothelial cells determined by PECAM immunohistochemistry. However, these in vitro differentiated endothelial cells did not express the LacZ reporter gene. This indicates the lack of factors and/or cellular interactions which are required to induce the expression of the reporter gene mediated by this 1.2 kb promoter in this in vitro differentiation system. Thus this system will allow us to screen for the putative inducers that exist in vivo but not in vitro. These putative inducers are presumably important for in vivo differentiation of vascular endothelial cells.

Animals↗
PubMed1991

Dominant male sterility in mice caused by insertion of a transgene.

While examining a series of transgenic mouse lines carrying the HCK protooncogene, we encountered one line in which males hemizygous for the transgene were sterile. The sterile males mated normally but failed to impregnate females. Light and electron microscopy revealed that spermatogenesis proceeds normally until nuclear condensation, which occurs but gives rise to a variety of abnormally shaped nuclei. Expression of the transgene was not detectable. Thus, the insertion itself probably caused the abnormal phenotype by disrupting a gene (or genes) important in spermatogenesis. The mutation is genetically dominant, causing an abnormal phenotype even though the sterile mice carry an ostensibly normal counterpart of the disrupted locus. The mutant phenotype is completely penetrant only in some genetic backgrounds, suggesting a modifying influence from a second locus. Junctions between the inserted transgene and adjoining cellular DNA were cloned, allowing us to confirm the heterozygous nature of the genetic disruption and to detect and associated deletion. We have designated the mutation Lvs (lacking vigorous sperm) and presume that it may define a previously undescribed locus important in spermatogenesis.

Animals↗
PubMed1989

Beta-globin enhancers target expression of a heterologous gene to erythroid tissues of transgenic mice.

To examine the role of human beta-globin enhancers in tissue-specific and developmental regulation, a hybrid beta-globin-simian virus 40 gene was analyzed in transgenic mice. A beta-globin DNA fragment containing two previously defined enhancers stimulated transcription from the simian virus 40 promoter in a tissue- and stage-specific pattern similar to that of the normal beta-globin gene. These results help to define the functions of beta-globin regulatory elements and suggest an approach for targeted expression of heterologous genes in erythroid cells in vivo.

Aging↗
PubMed1987

Upstream G gamma-globin and downstream beta-globin sequences required for stage-specific expression in transgenic mice.

The human G gamma-globin and beta-globin genes are expressed in erythroid cells at different stages of human development, and previous studies have shown that the two cloned genes are also expressed in a differential stage-specific manner in transgenic mice. The G gamma-globin gene is expressed only in murine embryonic erythroid cells, while the beta-globin gene is active only at the fetal and adult stages. In this study, we analyzed transgenic mice carrying a series of hybrid genes in which different upstream, intragenic, or downstream sequences were contributed by the beta-globin or G gamma-globin gene. We found that hybrid 5'G gamma/3'beta globin genes containing G gamma-globin sequences upstream from the initiation codon were expressed in embryonic erythroid cells at levels similar to those of an intact G gamma-globin transgene. In contrast, beta-globin upstream sequences were insufficient for expression of 5'beta/3'G gamma hybrid globin genes or a beta-globin-metallothionein fusion gene in adult erythroid cells. However, beta-globin downstream sequences, including 212 base pairs of exon III and 1,900 base pairs of 3'-flanking DNA, were able to activate a 5'G gamma/3'beta hybrid globin gene in fetal and adult erythroid cells. These experiments suggest that positive regulatory elements upstream from the G gamma-globin and downstream from the beta-globin gene are involved in the differential expression of the two genes during development.

Animals↗
PubMed1986

Correction of murine beta-thalassemia by gene transfer into the germ line.

A murine beta-thalassemia was corrected by the transfer of cloned beta-globin genes into the mouse germ line. The cloned mouse beta maj-globin gene or the cloned human beta-globin gene was introduced into mice deficient in beta-globin synthesis because of a deletion of the beta maj-globin gene. Both introduced genes produced functional beta-globin chains, leading to a reduction in one case, and elimination in another case, of the anemia and associated abnormalities of the red blood cells.

Animals↗