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1.
Autoimmunity ; 42(3): 183-97, 2009 Mar.
Article in English | MEDLINE | ID: mdl-19301199

ABSTRACT

The MerTK receptor tyrosine kinase is an important negative regulator of dendritic cell function and is required to prevent B cell autoimmunity in vivo. It is not currently known however, if any causal relationship exists between these two aspects of MerTK function. We sought to determine if dendritic cells (DC) from mice lacking MerTK (mertk(- / - ) mice) have characteristics that may aid in the development of B cell autoimmunity. Specifically, we found that mertk(- / - ) mice contain an elevated number of splenic DC, and this population contains an elevated proportion of cells secreting the critical B cell pro-survival factor, B cell activating factor (BAFF). Elevated numbers of BAFF-secreting cells were also detected among mertk(- / - ) bone marrow-derived dendritic cell (BMDC) populations. This was observed in both resting BMDC, and BMDC stimulated with lipopolysaccharide (LPS) or treated with exogenous apoptotic cells. We also found that DC in general have a pro-survival effect on resting B cells in co-culture. However, despite containing more BAFF-secreting cells, mertk(- / - ) BMDC were not superior to C57BL/6 or baff-deficient BMDC at promoting B cell survival. Furthermore, using decoy receptors, we show that DC may promote B cell survival and autoimmunity through a BAFF-and a proliferation-inducing ligand-independent mechanism.


Subject(s)
B-Cell Activating Factor/metabolism , Dendritic Cells/metabolism , Proto-Oncogene Proteins/physiology , Receptor Protein-Tyrosine Kinases/physiology , Age Factors , Animals , Antibodies, Antinuclear/blood , Antibodies, Antinuclear/immunology , Autoimmunity/physiology , B-Cell Activating Factor/blood , B-Cell Activating Factor/pharmacology , B-Lymphocytes/cytology , B-Lymphocytes/physiology , Bone Marrow Cells/cytology , Bone Marrow Cells/drug effects , Cell Count , Cell Differentiation/drug effects , Cell Survival/physiology , Chromatin/immunology , Coculture Techniques , DNA/immunology , Dendritic Cells/cytology , Dendritic Cells/drug effects , Dendritic Cells, Follicular/cytology , Dendritic Cells, Follicular/metabolism , Gene Expression/genetics , Interferon-gamma/pharmacology , Lipopolysaccharides/pharmacology , Mice , Mice, Inbred C57BL , Mice, Knockout , Recombinant Proteins/pharmacology , Spleen/cytology , Spleen/metabolism , Tumor Necrosis Factor Ligand Superfamily Member 13/physiology , c-Mer Tyrosine Kinase
2.
J Neuropathol Exp Neurol ; 60(11): 1062-74, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11706936

ABSTRACT

Twitcher (twi/twi) is a murine model of a human genetic demyelinating disease, globoid cell leukodystrophy (Krabbe disease). The affected mice usually die before reaching age 45 days, having demyelination associated with extensive glial activation. The twi/twi mice that receive wild-type bone marrow transplantation (BMT) survive up to 3 times longer with improved pathology. We hypothesize that immune-related molecules such as cytokines and chemokines are partly responsible for the demyelination in twi/twi, and that the decrease in the expression of such molecules following BMT contributes to clinico-pathological improvement. Cells expressing TNF-alpha, MCP-1, and MIP-1beta were conspicuous in the twi/twi CNS accompanied by infiltration of Ia+ and CD8+/CD3- hematogenous cells. These cells decreased gradually after BMT TNF-alpha mRNA and mRNA of C-C chemokine families, including MCP-1, IP-10, MIP-1alpha, MIP-1beta, and RANTES, were upregulated in the twi/twi CNS but downregulated gradually following BMT. In twi/twi that survived to 20 wk of age, cells expressing TNF-alpha, MCP-1, MIP-1beta, Ia, or CD8 were hardly detected and pathology was clearly improved. These results are consistent with the hypothesis that cytokine expression in glial cells contributes (to some extent) to the pathogenesis of demyelinating lesions in the twi/twi mice.


Subject(s)
Bone Marrow Transplantation/immunology , Cytokines/metabolism , Leukodystrophy, Globoid Cell/immunology , Leukodystrophy, Globoid Cell/therapy , Animals , Astrocytes/chemistry , Astrocytes/pathology , CD3 Complex/analysis , CD8 Antigens/analysis , Chemokine CCL2/analysis , Chemokine CCL2/metabolism , Chemokine CCL3 , Chemokine CCL4 , Cytokines/analysis , Down-Regulation/immunology , Immunohistochemistry , Interleukin-10/analysis , Interleukin-10/metabolism , Leukodystrophy, Globoid Cell/pathology , Macrophage Inflammatory Proteins/analysis , Macrophage Inflammatory Proteins/metabolism , Mice , Mice, Inbred C57BL , Mice, Neurologic Mutants , Mice, Transgenic , Microglia/chemistry , Microglia/pathology , Microscopy, Electron , Nerve Fibers, Myelinated/immunology , Nerve Fibers, Myelinated/ultrastructure , Tumor Necrosis Factor-alpha/analysis , Tumor Necrosis Factor-alpha/metabolism
3.
Nat Neurosci ; 4(11): 1116-22, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11600888

ABSTRACT

Here we used mice lacking tumor necrosis factor-alpha (TNF alpha) and its associated receptors to study a model of demyelination and remyelination in which these events could be carefully controlled using a toxin, cuprizone. Unexpectedly, the lack of TNF alpha led to a significant delay in remyelination as assessed by histology, immunohistochemistry for myelin proteins and electron microscopy coupled with morphometric analysis. Failure of repair correlated with a reduction in the pool of proliferating oligodendrocyte progenitors (bromodeoxyuridine-labeled NG2(+) cells) followed by a reduction in the number of mature oligodendrocytes. Analysis of mice lacking TNF receptor 1 (TNFR1) or TNFR2 indicated that TNFR2, not TNFR1, is critical to oligodendrocyte regeneration. This unexpected reparative role for TNF alpha in the CNS is important for understanding oligodendrocyte regeneration/proliferation, nerve remyelination and the design of new therapeutics for demyelinating diseases.


Subject(s)
Antigens, CD/metabolism , Myelin Sheath/metabolism , Oligodendroglia/physiology , Receptors, Tumor Necrosis Factor/metabolism , Stem Cells/physiology , Tumor Necrosis Factor-alpha/metabolism , Animals , Antigens, CD/genetics , Apoptosis , B-Lymphocytes/metabolism , Brain Chemistry , Corpus Callosum/metabolism , Corpus Callosum/ultrastructure , Cuprizone/administration & dosage , Cuprizone/toxicity , Demyelinating Diseases/chemically induced , Disease Models, Animal , Humans , Immunohistochemistry , In Situ Nick-End Labeling , Macrophages/metabolism , Male , Mice , Mice, Knockout , Microglia/metabolism , Monoamine Oxidase Inhibitors/pharmacology , Myelin Sheath/pathology , Myelin Sheath/ultrastructure , Oligodendroglia/cytology , Oligodendroglia/drug effects , Receptors, Tumor Necrosis Factor/genetics , Receptors, Tumor Necrosis Factor, Type I , Receptors, Tumor Necrosis Factor, Type II , Stem Cells/ultrastructure , Tumor Necrosis Factor-alpha/genetics , Up-Regulation
4.
J Neurosci ; 21(18): 7046-52, 2001 Sep 15.
Article in English | MEDLINE | ID: mdl-11549714

ABSTRACT

Interleukin-1beta (IL-1beta) is a proinflammatory cytokine associated with the pathophysiology of demyelinating disorders such as multiple sclerosis and viral infections of the CNS. However, we demonstrate here that IL-1beta appears to promote remyelination in the adult CNS. In IL-1beta(-/-) mice, acute demyelination progressed similarly to wild-type mice and showed parallel mature oligodendrocyte depletion, microglia-macrophage accumulation, and the appearance of oligodendrocyte precursors. In contrast, IL-1beta(-/-) mice failed to remyelinate properly, and this appeared to correlate with a lack of insulin-like growth factor-1 (IGF-1) production by microglia-macrophages and astrocytes and to a profound delay of precursors to differentiate into mature oligodendrocytes. Thus, IL-1beta may be crucial to the repair of the CNS, presumably through the induction of astrocyte and microglia-macrophage-derived IGF-1.


Subject(s)
Central Nervous System/metabolism , Demyelinating Diseases/physiopathology , Interleukin-1/metabolism , Regeneration/physiology , Animals , Antigens, Differentiation/biosynthesis , Astrocytes/metabolism , Astrocytes/pathology , Cell Count , Central Nervous System/drug effects , Central Nervous System/pathology , Chelating Agents , Corpus Callosum/drug effects , Corpus Callosum/metabolism , Corpus Callosum/pathology , Cuprizone , Demyelinating Diseases/chemically induced , Demyelinating Diseases/pathology , Disease Progression , Glutathione S-Transferase pi , Glutathione Transferase/biosynthesis , Insulin-Like Growth Factor I/biosynthesis , Insulin-Like Growth Factor I/genetics , Interleukin-1/genetics , Interleukin-1/pharmacology , Isoenzymes/biosynthesis , Macrophages/metabolism , Macrophages/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Microglia/metabolism , Microglia/pathology , Myelin Sheath/metabolism , Oligodendroglia/drug effects , Oligodendroglia/metabolism , Oligodendroglia/pathology , RNA, Messenger/metabolism , Regeneration/drug effects , Stem Cells/cytology , Stem Cells/metabolism , Up-Regulation/drug effects
5.
J Immunol ; 167(5): 2964-71, 2001 Sep 01.
Article in English | MEDLINE | ID: mdl-11509646

ABSTRACT

Chemokines are small chemotactic cytokines that modulate leukocyte recruitment and activation during inflammation. Here, we describe the role of macrophage inflammatory protein-1alpha (MIP-1alpha) during cuprizone intoxication, a model where demyelination of the CNS features a large accumulation of microglia/macrophage without T cell involvement or blood-brain barrier disruption. RNase protection assays showed that mRNA for numerous chemokines were up-regulated during cuprizone treatment in wild-type, C57BL/6 mice. RANTES, inflammatory protein-10, and monocyte chemoattractant protein-1 showed greatest expression with initiation of insult at 1-2 wk of treatment, whereas MIP-1alpha and beta increased later at 4-5 wk, coincident with peak demyelination and cellular accumulation. The function of MIP-1alpha during demyelination was tested in vivo by exposing MIP-1alpha knockout mice (MIP-1alpha(-/-)) to cuprizone and comparing pathology to wild-type mice. Demyelination at 3.5 wk of treatment was significantly decreased in MIP-1alpha(-/-) mice ( approximately 36% reduction), a result confirmed by morphology at the electron microscopic level. The delay in demyelination was correlated to apparent decreases in microglia/macrophage and astrocyte accumulation and in TNF-alpha protein levels. It was possible that larger effects of the MIP-1alpha deficiency were being masked by other redundant chemokines. Indeed, RNase protection assays revealed increased expression of several chemokine transcripts in both untreated and cuprizone-treated MIP-1alpha(-/-) mice. Nonetheless, despite this possible compensation, our studies show the importance of MIP-1alpha in demyelination in the CNS and highlight its effect, particularly on cellular recruitment and cytokine regulation.


Subject(s)
Demyelinating Autoimmune Diseases, CNS/etiology , Macrophage Inflammatory Proteins/deficiency , Animals , Astrocytes/pathology , Blood-Brain Barrier/physiology , Chemokine CCL3 , Chemokine CCL4 , Chemokine CCL5/genetics , Chemokines/genetics , Cuprizone/toxicity , Demyelinating Autoimmune Diseases, CNS/immunology , Demyelinating Autoimmune Diseases, CNS/pathology , Macrophage Inflammatory Proteins/genetics , Macrophage Inflammatory Proteins/physiology , Macrophages/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Microglia/pathology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Chemokine/genetics , Tumor Necrosis Factor-alpha/biosynthesis , Up-Regulation/drug effects
6.
Nature ; 411(6834): 207-11, 2001 May 10.
Article in English | MEDLINE | ID: mdl-11346799

ABSTRACT

Apoptosis is fundamental to the development and maintenance of animal tissues and the immune system. Rapid clearance of apoptotic cells by macrophages is important to inhibit inflammation and autoimmune responses against intracellular antigens. Here we report a new function for Mer, a member of the Axl/Mer/Tyro3 receptor tyrosine kinase family. mer(kd) mice with a cytoplasmic truncation of Mer had macrophages deficient in the clearance of apoptotic thymocytes. This was corrected in chimaeric mice reconstituted with bone marrow from wild-type animals. Primary macrophages isolated from mer(kd) mice showed that the phagocytic deficiency was restricted to apoptotic cells and was independent of Fc receptor-mediated phagocytosis or ingestion of other particles. The inability to clear apoptotic cells adequately may be linked to an increased number of nuclear autoantibodies in mer(kd) mice. Thus, the Mer receptor tyrosine kinase seems to be critical for the engulfment and efficient clearance of apoptotic cells. This has implications for inflammation and autoimmune diseases such as systemic lupus erythematosus.


Subject(s)
Apoptosis , Macrophages, Peritoneal/immunology , Phagocytosis , Protein-Tyrosine Kinases/metabolism , Proto-Oncogene Proteins/metabolism , Receptor Protein-Tyrosine Kinases , Thymus Gland/cytology , Animals , Antibodies, Antinuclear/immunology , Apoptosis/drug effects , Bone Marrow Transplantation , Cell Adhesion , Cells, Cultured , Crosses, Genetic , Cytochalasin B/pharmacology , Dexamethasone/pharmacology , Female , Flow Cytometry , Immunohistochemistry , Listeria monocytogenes/immunology , Macrophages, Peritoneal/cytology , Macrophages, Peritoneal/metabolism , Macrophages, Peritoneal/ultrastructure , Male , Mice , Mice, Inbred C57BL , Mice, Mutant Strains , Microscopy, Electron, Scanning , Microspheres , Mutation/genetics , Protein-Tyrosine Kinases/genetics , Proto-Oncogene Proteins/genetics , Radiation Chimera/immunology , Receptors, Fc/immunology , Thymus Gland/drug effects , Thymus Gland/immunology , Thymus Gland/ultrastructure , c-Mer Tyrosine Kinase
7.
J Neurochem ; 77(4): 1067-76, 2001 May.
Article in English | MEDLINE | ID: mdl-11359872

ABSTRACT

We studied markers of myelin content and of the rate of myelination in brains of mice between 8 and 20 weeks of age. During the 12-week time-course, control animals showed slight increases in the content of oligodendroglial-specific cerebroside, as well as cholesterol (enriched in, but not specific to, myelin). In contrast, synthesis of these lipids, as assayed by in vivo incorporation of (3)H(2)O, was substantial, indicating turnover of 0.4% and 0.7% of total brain cerebroside and cholesterol, respectively, each day. We also studied mice exposed to a diet containing 0.2% of the copper chelator, cuprizone. After 6 weeks 20%, and by 12 weeks, over 30% of brain cerebroside was gone. Demyelination was accompanied by down-regulation of mRNA expression for enzymes controlling myelin lipid synthesis (ceramide galactosyl transferase for cerebroside; hydroxymethylglutaryl-CoA reductase for cholesterol), and for myelin basic protein. Synthesis of myelin lipids was also greatly depressed. The 20% cerebroside deficit consequent to 6 weeks of cuprizone exposure was restored 6 weeks after return to a control diet. During remyelination, expression of myelin-related mRNA species, as well as cerebroside and cholesterol synthesis were restored to normal. However, in contrast to the steady state metabolic turnover in the control situation, all the cerebroside and cholesterol made were accumulated. To the extent that accumulating cerebroside is targeted for eventual inclusion in myelin (discussed) the rate of its synthesis is proportional to remyelination. With our assay, in vivo rates of cerebroside synthesis can be determined for a time window of the order of hours. This offers greater temporal resolution and accuracy relative to classical methods assaying accumulation of myelin components at time intervals of several days. We propose this experimental design, and the reproducible cuprizone model, as appropriate for studies of how to promote remyelination.


Subject(s)
Brain/drug effects , Cerebrosides/biosynthesis , Cuprizone/pharmacology , Myelin Basic Protein/metabolism , Myelin Sheath/drug effects , Animals , Biomarkers , Brain/metabolism , Brain/physiology , Brain Stem/drug effects , Brain Stem/metabolism , Cerebellum/drug effects , Cerebellum/metabolism , Cholesterol/biosynthesis , Galactosyltransferases/genetics , Gene Expression Regulation, Enzymologic/drug effects , Hydroxymethylglutaryl CoA Reductases/genetics , Male , Mice , Mice, Inbred C57BL , Myelin Sheath/physiology , N-Acylsphingosine Galactosyltransferase , Time Factors , Transcription, Genetic/drug effects
8.
Neuropathol Appl Neurobiol ; 27(1): 50-8, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11299002

ABSTRACT

Exposure of young adult C57BL/6 mice to cuprizone in the diet initiated profound and synchronous demyelination of the corpus callosum, which was virtually complete by 4 weeks of exposure. Interestingly, even in the face of a continued exposure to cuprizone, there was spontaneous remyelination 2 weeks later. This remyelination preferentially involved smaller calibre axons. There was a suggestion of yet another cycle of demyelination (at 10 weeks) and remyelination (at 12 weeks), but by 16 weeks of exposure, the regenerative capacity was exhausted and the animals were near death. The relapsing-remitting pattern suggests this may be a useful model for certain human demyelinating disorders. In contrast to the above chronic model, the corpus callosum from mice exposed to cuprizone for only 6 weeks continued to remyelinate, with 67% of the axons being myelinated or remyelinated at 10 weeks. Interestingly, a significant reduction in the mean value for axonal diameter was observed during acute demyelination. Upon remyelination, however, the axonal calibre distribution returned to near-normal. In contrast, when mice were maintained on a cuprizone diet for 16 weeks, the mean value for axonal diameter was reduced to 60% of normal. These results provide further evidence that the interactions between oligodendrocytes and axons alter axonal calibre.


Subject(s)
Axons/pathology , Central Nervous System/pathology , Demyelinating Diseases/pathology , Amidines , Animals , Axons/drug effects , Axons/ultrastructure , Cell Size/drug effects , Central Nervous System/drug effects , Chronic Disease , Corpus Callosum/drug effects , Corpus Callosum/pathology , Cuprizone , Demyelinating Diseases/chemically induced , Disease Models, Animal , Fluorescent Dyes , Mice , Mice, Inbred C57BL , Microscopy, Electron , Nerve Fibers, Myelinated/pathology , Nerve Fibers, Myelinated/ultrastructure , Recurrence , Remission, Spontaneous
9.
Brain Pathol ; 11(1): 107-16, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11145196

ABSTRACT

Myelin of the adult CNS is vulnerable to a variety of metabolic, toxic, and autoimmune insults. That remyelination can ensue, following demyelinating insult, has been well demonstrated. Details of the process of remyelination are, however difficult to ascertain since in most experimental models of demyelination/remyelination the severity, localization of lesion site, or time course of the pathophysiology is variable from animal to animal. In contrast, an experimental model in which massive demyelination can be reproducibly induced in large areas of mouse brain is exposure to the copper chelator, cuprizone, in the diet. We review work from several laboratories over the past 3 decades, with emphasis on our own recent studies, which suggest an overall picture of cellular events involved in demyelination/remyelination. When 8 week old C57BL/6 mice are fed 0.2% cuprizone in the diet, mature olidgodendroglia are specifically insulted (cannot fulfill the metabolic demand of support of vast amounts of myelin) and go through apoptosis. This is closely followed by recruitment of microglia and phagoctytosis of myelin. Studies of myelin gene expression, coordinated with morphological studies, indicate that even in the face of continued metabolic challenge, oligodendroglial progenitor cells proliferate and invade demyelinated areas. If the cuprizone challenge is terminated, an almost complete remyelination takes place in a matter of weeks. Communication between different cell types by soluble factors may be inferred. This material is presented in the context of a model compatible with present data -- and which can be tested more rigorously with the cuprizone model. The reproducibility of the model indicates that it may allow for testing of manipulations (e.g. available knockouts or transgenics on the common genetic background, or pharmacological treatments) which may accelerate or repress the process of demyelination and or remyelination.


Subject(s)
Central Nervous System/drug effects , Chelating Agents/pharmacology , Cuprizone/pharmacology , Demyelinating Diseases/physiopathology , Models, Animal , Oligodendroglia/physiology , Animals , Mice , Mice, Inbred C57BL , Microglia , Myelin Sheath
10.
J Neurochem ; 76(1): 77-86, 2001 Jan.
Article in English | MEDLINE | ID: mdl-11145980

ABSTRACT

Myelination, during both normal development and with respect to disorders of myelination, is commonly studied by morphological and/or biochemical techniques that assay as their end-points the extent of myelination. The rate of myelination is potentially a more useful parameter, but it is difficult and time-consuming to establish, requiring a complete developmental study with labor-intensive methodology. We report herein development of methodology to assay the absolute rate of myelination at any desired time during development. This involves intraperitoneal injection of (3)H(2)O to label body water pools, followed by determination of label in the myelin-specific lipid, cerebroside. The absolute amount of cerebroside synthesized can then be calculated from the specific radioactivity of body water and knowledge of the number of hydrogens from water incorporated into cerebroside. During development, the rate of cerebroside synthesis correlated well with the rate of accumulation of the myelin-specific components, myelin basic protein and cerebroside. For purposes of control, we also tested other putative, albeit less quantitative, indices of the rate of myelination. Levels of mRNA for ceramide galactosyltransferase (rate-limiting enzyme in cerebroside synthesis) and for myelin basic protein did not closely correlate with myelination at all times. Cholesterol synthesis closely matched the rate of cholesterol accumulation but did not track well with myelination. Synthesis of fatty acids did not correlate well with accumulation of either fatty acids (phospholipids) or myelin markers. We conclude that measurement of cerebroside synthesis rates provides a good measure of the rate of myelination. This approach may be useful as an additional parameter for examining the effects of environmental or genetic alterations on the rate of myelination.


Subject(s)
Brain Chemistry , Brain/metabolism , Lipid Metabolism , Myelin Sheath/metabolism , Aging/metabolism , Animals , Biomarkers , Body Water/metabolism , Cerebrosides/analysis , Cerebrosides/biosynthesis , Cholesterol/analysis , Cholesterol/metabolism , Fatty Acids/analysis , Fatty Acids/biosynthesis , Female , Galactosyltransferases/genetics , Galactosyltransferases/metabolism , Hydroxymethylglutaryl CoA Reductases/genetics , Hydroxymethylglutaryl CoA Reductases/metabolism , Lipids/analysis , Male , Mice , Mice, Inbred C57BL , Myelin Basic Protein/genetics , Myelin Basic Protein/metabolism , N-Acylsphingosine Galactosyltransferase , RNA, Messenger/analysis , RNA, Messenger/metabolism , Tritium
12.
Mol Cell Neurosci ; 16(4): 338-49, 2000 Oct.
Article in English | MEDLINE | ID: mdl-11085872

ABSTRACT

Evidence suggests that interferon-gamma (IFN-gamma), a proinflammatory cytokine secreted by activated T lymphocytes, contributes a deleterious effect to immune-mediated demyelinating disorders such as multiple sclerosis and experimental autoimmune encephalomyelitis (EAE). Nevertheless, mouse strains that are normally resistant to EAE induction become susceptible when the gene encoding either IFN-gamma or its receptor is mutated, demonstrating that the role that this cytokine plays in demyelinating disorders is complex. We have examined the effect of IFN-gamma in a chemically induced model of CNS demyelination. Mice that receive through their diet the copper chelator cuprizone display extensive demyelination of the corpus callosum. Remarkably, transgenic mice that ectopically express low levels of IFN-gamma in the CNS did not display evidence of demyelination when treated with cuprizone, nor did they shows signs of oligodendroglial death, astrogliosis, or microgliosis, which are typically seen in treated animals. Myelin protein gene expression was, however, dramatically reduced in both the treated control and the transgenic animals, indicating that demyelination is not an obligatory consequence of a large diminution of myelin protein synthesis. Interestingly, the CNS of the IFN-gamma-expressing mice contained elevated levels of insulin-like growth factor I, which has been demonstrated to have a protective effect against the demyelinating action of cuprizone.


Subject(s)
Corpus Callosum/immunology , Demyelinating Diseases/immunology , Interferon-gamma/immunology , Animals , Astrocytes/pathology , Chelating Agents , Corpus Callosum/pathology , Cuprizone , Demyelinating Diseases/chemically induced , Demyelinating Diseases/genetics , Disease Models, Animal , Gene Expression/immunology , Gliosis/chemically induced , Gliosis/genetics , Gliosis/immunology , Insulin-Like Growth Factor I/genetics , Interferon-gamma/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Microglia/pathology , Myelin Proteins/genetics , Myelin Proteins/immunology
13.
Blood ; 96(9): 2973-80, 2000 Nov 01.
Article in English | MEDLINE | ID: mdl-11049973

ABSTRACT

To investigate the mechanism by which macrophage inflammatory protein-1alpha (MIP-1alpha) affects graft-versus-host disease (GVHD), the expression and function of MIP-1alpha in 2 murine models of GVHD were evaluated. In irradiated class I and class II disparate recipients, the expression of messenger RNA (mRNA) and protein for MIP-1alpha was significantly increased in GVHD target organs after transfer of allogeneic lymphocytes compared to syngeneic lymphocytes. When lymphocytes unable to make MIP-1alpha were transferred, there was a decrease in the production of MIP-1alpha in the liver, lung, and spleen of bm1 (B6.C-H2(bm1)/By) and bm12 (B6.C-H2(bm12)/KhEg) recipients compared to the transfer of wild-type splenocytes. At day 6 there was a 4-fold decrease in the number of transferred CD8(+) T cells in the lung and approximately a 2-fold decrease in the number of CD8(+) T cells in the liver and spleen in bm1 recipients after transfer of MIP-1alpha-deficient (MIP-1alpha(-/-)) splenocytes compared to wild-type (MIP-1alpha(+/+)) splenocytes. These differences persisted for 13 days after splenocyte transfer. In contrast, the number of donor CD4(+) T cells found in the liver and lung was significantly increased after the transfer of MIP-1alpha(-/-) compared to wild-type splenocytes in bm12 recipients from day 6 through day 10. Thus, the transfer of allogeneic T cells was associated with the enhanced expression of MIP-1alpha in both a class I and class II mismatch setting. However, the increased expression only led to enhanced recruitment of CD8(+), but not CD4(+), donor T cells. Production of MIP-1alpha by donor T cells is important in the occurrence of GVHD and functions in a tissue-dependent fashion.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Chemokines/genetics , Graft vs Host Disease/immunology , Liver/immunology , Lung/immunology , Lymphocyte Transfusion , Macrophage Inflammatory Proteins/genetics , Spleen/immunology , T-Lymphocytes/immunology , Animals , Cell Line , Chemokine CCL3 , Chemokine CCL4 , Crosses, Genetic , Disease Models, Animal , Green Fluorescent Proteins , Luminescent Proteins/genetics , Macrophage Inflammatory Proteins/deficiency , Macrophage Inflammatory Proteins/physiology , Mice , Mice, Inbred C57BL , Mice, Inbred Strains , Mice, Knockout , Mice, Transgenic , Transcription, Genetic , Transplantation, Homologous , Transplantation, Isogeneic
14.
J Neurosci Res ; 61(3): 251-62, 2000 Aug 01.
Article in English | MEDLINE | ID: mdl-10900072

ABSTRACT

We have documented changes in the oligodendrocyte population during demyelinating insult to the adult CNS. Feeding of cuprizone to adult mice led to apoptotic death of mature oligodendrocytes followed by profound demyelination of the corpus callosum. A regenerative response was initiated even during active demyelination. Oligodendrocyte progenitors have begun to proliferate and then accumulate within the lesion. Many of these cells may have migrated from the sub-ventricular zone and fornix before their accumulation in the demyelinating corpus callosum. The accumulation of differentiating oligodendrocyte progenitors was followed closely by the reappearance of mature oligodendrocytes and remyelination. Interestingly, an increase in IGF-1 mRNA was detected at Week 3 through Week 7, suggesting potential involvement in remyelination. Other factors, however, such as PDGF, NT3, FGF, jagged, and notch remained unchanged. These results suggest that the mature oligodendroglial population depleted by apoptosis is replaced by a newly formed oligodendroglial population derived from progenitors; these accumulate and seem to differentiate during remyelination.


Subject(s)
Apoptosis , Demyelinating Diseases/pathology , Insulin-Like Growth Factor I/biosynthesis , Myelin Sheath/pathology , Oligodendroglia/pathology , Stem Cells/pathology , Animals , Cell Count , Cell Differentiation , Corpus Callosum/metabolism , Corpus Callosum/pathology , Cuprizone , Demyelinating Diseases/chemically induced , Immunohistochemistry , Male , Mice , Mice, Inbred C57BL , Oligodendroglia/metabolism
15.
J Neurosci ; 20(15): 5703-8, 2000 Aug 01.
Article in English | MEDLINE | ID: mdl-10908609

ABSTRACT

Metabolic insult results in apoptosis and depletion of mature oligodendrocytes during demyelination. To examine the role of insulin-like growth factor-1 (IGF-1) during acute demyelination and remyelination in the adult CNS, we exposed transgenic mice that continuously express IGF-1 (IGF-1 tg) to cuprizone intoxication. Demyelination was observed within the corpus callosum in both wild-type and IGF-1 tg mice 3 weeks after exposure to cuprizone. Wild-type mice showed significant apoptotic mature oligodendrocytes and a dramatic loss of these cells within the lesion that resulted in near complete depletion and demyelination by week 5. In contrast, the demyelinated corpus callosum of the IGF-1 tg mice was near full recovery by week 5. This rapid recovery was apparently caused by survival of the mature oligodendrocyte population because apoptosis was negligible, and by week 4, the mature oligodendrocyte population was completely restored. Furthermore, despite demyelination in both wild-type and IGF-1 tg mice, oligodendrocyte progenitors accumulated only in the absence of mature oligodendrocytes and failed to accumulate if the mature oligodendrocytes remained as demonstrated in the IGF-1 tg mice. These results suggest that IGF-1 may be important in preventing the depletion of mature oligodendrocytes in vivo and thus facilitates an early recovery from demyelination.


Subject(s)
Apoptosis/physiology , Demyelinating Diseases/physiopathology , Insulin-Like Growth Factor I/metabolism , Oligodendroglia/cytology , Acute Disease , Animals , Brain Chemistry/drug effects , Brain Chemistry/physiology , Cell Survival/physiology , Chelating Agents/poisoning , Corpus Callosum/pathology , Corpus Callosum/physiopathology , Cuprizone/poisoning , Demyelinating Diseases/chemically induced , Demyelinating Diseases/genetics , Female , Gene Expression/physiology , Insulin-Like Growth Factor I/genetics , Macrophages/cytology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Microglia/cytology , Recovery of Function/physiology , Stem Cells/cytology
16.
Am J Pathol ; 156(6): 1849-54, 2000 Jun.
Article in English | MEDLINE | ID: mdl-10854208

ABSTRACT

The twitcher mouse is a murine model of globoid cell leukodystropy, a genetic demyelinating disease caused by a mutation of the galactosylceramidase gene. Demyelination of the central nervous system commences around 20 postnatal days. Using GFP-transgenic mice as donors, the distribution of hematogenous cells after bone marrow transplantation was investigated in the twitcher mice. Bone marrow transplantation was carried out at 8 postnatal days. In twitcher chimeric mice examined before 30 postnatal days, numerous GFP(+) cells were detected in spleen and peripheral nerve but only a few were detected in the liver, lung, and spinal white matter. In contrast, at 35 to 40 postnatal days when demyelination is evident, many GFP(+) cells with ameboid form were detected in the white matter of the spinal cord, brainstem, and cerebrum. Approximately half of these GFP(+) cells were co-labeled with Mac-1. In twitcher chimeric mice examined after 100 postnatal days, the majority of GFP/Mac-1 double-positive cells displayed the morphological features of ramified microglia with fine delicate processes and was distributed diffusely in both gray and white matter. These results suggest that a significant number of donor hematogenous cells are able to infiltrate into the brain parenchyma, repositioning themselves into areas previously occupied by microglia, and to ameliorate lethality.


Subject(s)
Blood Cells/transplantation , Bone Marrow Transplantation , Indicators and Reagents , Luminescent Proteins/metabolism , Postoperative Care , Tissue Donors , Animals , Blood Cells/metabolism , Central Nervous System/metabolism , Green Fluorescent Proteins , Luminescent Proteins/blood , Luminescent Proteins/genetics , Mice , Mice, Inbred C57BL , Mice, Neurologic Mutants , Mice, Transgenic , Peripheral Nerves/metabolism , Reference Values , Tissue Distribution , Viscera/metabolism
17.
Nature ; 398(6729): 723-8, 1999 Apr 22.
Article in English | MEDLINE | ID: mdl-10227296

ABSTRACT

We have generated and analysed null mutations in the mouse genes encoding three structurally related receptors with tyrosine kinase activity: Tyro 3, Axl, and Mer. Mice lacking any single receptor, or any combination of two receptors, are viable and fertile, but male animals that lack all three receptors produce no mature sperm, owing to the progressive death of differentiating germ cells. This degenerative phenotype appears to result from a failure of the tropic support that is normally provided by Sertoli cells of the seminiferous tubules, whose function depends on testosterone and additional factors produced by Leydig cells. Tyro 3, Axl and Mer are all normally expressed by Sertoli cells during postnatal development, whereas their ligands, Gas6 and protein S, are produced by Leydig cells before sexual maturity, and by both Leydig and Sertoli cells thereafter. Here we show that the concerted activation of Tyro 3, Axl and Mer in Sertoli cells is critical to the role that these cells play as nurturers of developing germ cells. Additional observations indicate that these receptors may also be essential for the tropic maintenance of diverse cell types in the mature nervous, immune and reproductive systems.


Subject(s)
Intercellular Signaling Peptides and Proteins , Proto-Oncogene Proteins , Receptor Protein-Tyrosine Kinases , Receptor Protein-Tyrosine Kinases/physiology , Sertoli Cells/physiology , Spermatogenesis/physiology , Animals , Gene Expression , Leydig Cells/physiology , Ligands , Male , Mice , Mice, Mutant Strains , Neural Cell Adhesion Molecules/genetics , Oncogene Proteins/genetics , Phenotype , Protein S , Proteins , Receptor Protein-Tyrosine Kinases/genetics , Spermatogenesis/genetics , c-Mer Tyrosine Kinase , Axl Receptor Tyrosine Kinase
18.
Brain Res ; 825(1-2): 59-67, 1999 Apr 17.
Article in English | MEDLINE | ID: mdl-10216173

ABSTRACT

Microglia in primary afferent projection territories are activated and proliferate after peripheral nerve injury. However, it is not known whether stimulation of peripheral nerves by noxious stimuli applied to their receptive fields activates microglial cells in the spinal cord. This study was designed to investigate the response of microglia in the lumbar spinal cord and in the brainstem to a tonic noxious stimulus. Thirty-two male Sprague-Dawley rats received subcutaneous injections of 5% formalin (50 microliter) into the plantar surface of the right hind paw, and 24 rats were injected with 50 microliter saline as a control. The lumbar spinal cord and brainstem were evaluated for immunoreactivity (IR) to complement receptor C3bi (monoclonal antibody OX-42) and major histocompatibility complex class II (monoclonal antibody OX-6) on postinjection hours 0, 2, 4 and 8 and days 1, 3, 7, 14 and 28. A qualitative and quantitative increase of OX-42-IR microglial cells were observed in the medial portion of the dorsal horn and in the gracile nucleus of the brainstem on the side ipsilateral to the formalin injection, starting on days 1-3 and peaking on day 7 postinjection. OX-6-positive cells were scattered both in gray and white matter, but no difference was detected between the two sides of the spinal cord or between formalin-injected and control animals. This is the first study that reports that subcutaneous injection of formalin into the rat's hind paw induces microglial activation in the spinal cord as well as in the brainstem. Although we have not determined whether these responses result from nociceptor activity, peripheral inflammation, or degeneration of primary afferents and/or central neurons, this method provides a simple, effective and stable animal model that will permit the future study of the mechanisms that contribute to microglial activation and its pathophysiological consequences.


Subject(s)
Microglia/physiology , Nociceptors/physiology , Pain/physiopathology , Animals , Antibodies, Monoclonal/pharmacology , Brain Stem/chemistry , Brain Stem/cytology , Complement C3b/analysis , Complement C3b/immunology , Disinfectants , Formaldehyde , Hindlimb , Histocompatibility Antigens Class II/immunology , Male , Pain/chemically induced , Rats , Rats, Sprague-Dawley , Spinal Cord/chemistry , Spinal Cord/cytology
19.
J Immunol ; 162(6): 3498-503, 1999 Mar 15.
Article in English | MEDLINE | ID: mdl-10092806

ABSTRACT

The regulation of monocyte function and the inhibition of TNF-alpha production during bacterial sepsis are critical in attenuating adverse host responses to endotoxemia. To study the function of a novel receptor tyrosine kinase, mer, that is expressed in monocytes, we generated mice (merkd) that lack the signaling tyrosine kinase domain. Upon LPS challenge, merkd animals died of endotoxic shock (15/17, 88.2%), whereas control wild-type mice survived (1/15, 6.7% died). Susceptible merkd mice exhibited edema, leukocyte infiltration, and signs of endotoxic shock that correlated with higher levels of TNF-alpha found in the serum of merkd mice as compared with wild-type control animals. Death due to LPS-induced endotoxic shock in merkd mice was blocked by administration of anti-TNF-alpha Ab, suggesting that overproduction of this cytokine was principally responsible for the heightened suseptibility. The increase in TNF-alpha production appeared to be the result of a substantial increase in the LPS-dependent activation of NF-kappa B nuclear translocation resulting in greater TNF-alpha production by macrophages from merkd mice. Thus, Mer receptor tyrosine kinase signaling participates in a novel inhibitory pathway in macrophages important for regulating TNF-alpha secretion and attenuating endotoxic shock.


Subject(s)
Lipopolysaccharides/toxicity , Neural Cell Adhesion Molecules/physiology , Proto-Oncogene Proteins , Receptor Protein-Tyrosine Kinases/physiology , Shock, Septic/prevention & control , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Tumor Necrosis Factor-alpha/biosynthesis , Animals , Crosses, Genetic , Drug Resistance/genetics , Macrophages/metabolism , Mice , Mice, Inbred DBA , Mice, Knockout , Neural Cell Adhesion Molecules/genetics , Neural Cell Adhesion Molecules/metabolism , Receptor Protein-Tyrosine Kinases/genetics , Receptor Protein-Tyrosine Kinases/metabolism , Sepsis/immunology , Sepsis/mortality , Sepsis/prevention & control , Shock, Septic/etiology , Shock, Septic/genetics , Tumor Necrosis Factor-alpha/metabolism , c-Mer Tyrosine Kinase
20.
Mol Cell Neurosci ; 12(4-5): 220-7, 1998 Nov.
Article in English | MEDLINE | ID: mdl-9828087

ABSTRACT

When C57BL/6J mice, 8 weeks of age, received 0.2% Cuprizone in their diet, extensive demyelination in corpus callosum was detectable after 3 weeks, and there was massive demyelination by 4 weeks. As expected, the accumulation of phagocytically active microglia/macrophages correlated closely with demyelination. When Cuprizone was removed from the diet, remyelination was soon initiated; after 6 weeks of recovery, myelin levels were near-normal and phagocytic cells were no longer prominent. Steady-state levels of mRNA for myelin-associated glycoprotein, myelin basic protein, and ceramide galactosyltransferase were already profoundly depressed after 1 week of Cuprizone exposure and were only 10-20% of control values after 2 weeks. Unexpectedly, upregulation of mRNA for these myelin genes did not correlate with initiation of remyelination but rather with accumulation of microglia/macrophages. After 6 weeks of exposure to Cuprizone, mRNA levels were at control levels or higher-in the face of massive demyelination. This suggests that in addition to effecting myelin removal, microglia/macrophages may simultaneously push surviving oligodendroglia or their progenitors toward myelination.


Subject(s)
Brain/metabolism , Cuprizone/toxicity , Demyelinating Diseases/metabolism , Demyelinating Diseases/pathology , Gene Expression Regulation/drug effects , Myelin Proteins/genetics , Myelin Sheath/physiology , Animals , Brain/drug effects , Brain/pathology , Corpus Callosum/drug effects , Corpus Callosum/pathology , Cuprizone/administration & dosage , Demyelinating Diseases/chemically induced , Diet , Galactosyltransferases/genetics , Macrophages/physiology , Mice , Mice, Inbred C57BL , Microglia/physiology , Myelin Basic Protein/genetics , Myelin Sheath/pathology , Myelin Sheath/ultrastructure , Myelin-Associated Glycoprotein/genetics , N-Acylsphingosine Galactosyltransferase , Phagocytosis , RNA, Messenger/genetics , Transcription, Genetic
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