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1.
Breast Cancer Res Treat ; 142(1): 101-7, 2013 Nov.
Article in English | MEDLINE | ID: mdl-24129976

ABSTRACT

Perceptions among women with breast cancer about the relative importance of different potential chemotherapy side effects is not well understood. A survey was performed by women receiving chemotherapy for breast cancer. Grade I/II (mild to moderate) and III/IV (moderate to severe) descriptions of nine common chemotherapy side effects were assigned preference weights using the standard gamble technique. For each hypothetical side effect, patients could choose to stay in the respective side effect state or take a gamble between full health (probability p) or being dead (1 - p). For each side effect, p was varied until the patient was indifferent between these options. The survey also included questions about the importance of survival, slowing cancer growth, and quality of life. This analysis included 69 patients; mean age 54 years (range 35-84), representing all cancer stages. Standard gamble preferences were lowest (i.e., least preferred) for grade III/IV nausea/vomiting (0.621), indicating that patients would, on average, risk a 38 % chance of being dead to avoid having grade III/IV nausea/vomiting for the rest of their lives. The next least preferred side effects were grade III/IV diarrhea (0.677) and grade III/IV sensory neuropathy (0.694). Survival appeared more important than slowing cancer growth and maintaining quality of life across cancer stages. Nevertheless, patients with advanced disease placed less importance on survival (p = 0.09) and higher importance on quality of life (p = 0.05). These standard gamble utilities provide unique insights into chemotherapy toxicities from the patient perspective. Differences in the relative importance of overall survival and quality of life with treatment existed between patients with different stages of disease. These studies should be expanded as the data may also be used to calculate quality-adjusted life expectancy in cost-effectiveness evaluations of breast cancer chemotherapies.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/adverse effects , Breast Neoplasms/complications , Breast Neoplasms/psychology , Drug-Related Side Effects and Adverse Reactions/psychology , Patient Preference , Adult , Aged , Aged, 80 and over , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Cost-Benefit Analysis , Cross-Sectional Studies , Female , Humans , Middle Aged , Neoplasm Staging , Quality of Life , Surveys and Questionnaires
2.
Lupus ; 22(10): 1038-45, 2013 Sep.
Article in English | MEDLINE | ID: mdl-23963432

ABSTRACT

OBJECTIVE: The objective was to develop a conceptual model illustrating the relationships between the physician-patient relationship and patient outcomes, including health status and regimen satisfaction, in systemic lupus erythematosus (SLE). METHODS: This was a cross-sectional survey of a geographically diverse sample of adults with SLE in the United States. Patients completed a Web-based survey that focused on physician interactions, clinical management, and patient outcomes, including patient perception of treatment regimen and health status. All survey variables related to physician interactions and patient perceptions of their health and satisfaction were evaluated for incorporation into a patient-centered model using cluster analysis. Structural equation modeling (SEM) was conducted to assess the inter-relationships observed among the variables to inform the development of a conceptual model of SLE patient-centered care. RESULTS: A total of 302 SLE patients completed the survey. The majority of patients were female (94.3%) with a mean age of 46 years. The cluster analysis resulted in six main factors: 1) physician interactions, 2) current health and hope, 3) satisfaction with treatment, 4) bedside manner, 5) discussion of lupus impacts during physician visits, and 6) steroid treatment. The significant relationships among the factors showed that positive physician interactions, such as including the patient in treatment decisions, were associated with higher satisfaction with treatment regimen and patients feeling that SLE was well controlled, a more favorable perception of current health, and being more hopeful about future health. Among the components of physician interactions, setting goals with patients is particularly important, as this was significantly associated with the patient being more hopeful about future health. Being steroid free was significantly related to higher treatment satisfaction. CONCLUSION: The study findings informed a conceptual model of SLE patient-centered care that may be used to create more targeted education programs in the management of SLE, with the goal to improve patient outcomes.


Subject(s)
Lupus Erythematosus, Systemic/therapy , Physician-Patient Relations , Adult , Cluster Analysis , Cross-Sectional Studies , Female , Health Status , Humans , Male , Middle Aged , Patient Education as Topic , Patient-Centered Care
3.
J Neurosci Res ; 66(3): 497-505, 2001 Nov 01.
Article in English | MEDLINE | ID: mdl-11746368

ABSTRACT

As oligodendrocytes mature they progress through a series of distinct differentiation steps characterized by the expression of specific markers. One such marker, polysialic acid found on the neural cell adhesion molecule (NCAM), is detected by antibodies and is present on progenitor oligodendrocytes, but is not detected to the same extent on mature oligodendrocytes. Two closely related polysialyltransferases, ST8Sia II (STX) and ST8Sia IV (PST) have been cloned previously and shown to synthesize polysialic acid on NCAM and other glycoproteins. To determine whether or not polyalpha2,8sialyltransferases are downregulated during the differentiation of oligodendrocytes, the enzyme activity and expression of ST8Sia II and ST8Sia IV mRNA at two stages of maturation in JS12/1 and JS3/16 oligodendrocytes were examined. Differentiation in both oligodendroglial cell lines was accompanied by more than a 50% reduction in the biosynthesis of polymers of alpha2,8sialic acid when fetuin was used as substrate. Most interestingly, extracts of JS12/1 mature cells synthesized 60% more short oligomers of alpha2,8sialic acid than the progenitor cells, whereas JS3/16 mature cells synthesized barely detectable amounts of the short oligomers. Transcripts for ST8Sia IV mRNA were present in both JS12/1 and JS3/16 and were reduced when the biosynthesis was markedly reduced. In contrast ST8Sia II mRNA was barely detectable in JS3/16 cells and although detectable in JS12/1 cells, there was no clear modulation with maturation. These results were supported by the examination of the brains of rats from embryonic to Day 21 ages. The enzyme activity and mRNA experiments show that polyalpha2,8sialyltransferase itself is down regulated to cause the reduction in sialyl polymers on mature oligodendrocytes. Moreover, ST8Sia IV is responsible for the polysialylation of NCAM in oligodendrocytes.


Subject(s)
Cell Differentiation/physiology , Neural Cell Adhesion Molecules/metabolism , Oligodendroglia/enzymology , Polymers/metabolism , RNA, Messenger/metabolism , Sialic Acids/biosynthesis , Sialyltransferases/genetics , Aging/physiology , Animals , Animals, Newborn , Cells, Cultured , Central Nervous System/cytology , Central Nervous System/enzymology , Central Nervous System/growth & development , Fetus , Gene Expression Regulation, Developmental/physiology , Gene Expression Regulation, Enzymologic/physiology , Oligodendroglia/cytology , Rats , Rats, Sprague-Dawley , Stem Cells/enzymology , Transcription, Genetic/physiology
4.
J Neurosci Res ; 64(4): 371-9, 2001 May 15.
Article in English | MEDLINE | ID: mdl-11340644

ABSTRACT

The myelin-deficient (MD) rat has a point mutation in its proteolipid protein (PLP) gene that causes severe dysmyelination and oligodendrocyte cell death. Using an in vitro model, we have shown that MD oligodendrocytes initially differentiate similarly to wild-type cells, expressing galactocerebroside, 2',3'-cyclic nucleotide 3'-phosphodiesterase, and myelin basic protein. However, at the time when PLP expression would normally begin, the MD oligodendrocytes die via an apoptotic pathway involving caspase activation. The active form of caspase-3 was detected, along with the cleavage products of poly-(ADP-ribose) polymerase (PARP) and spectrin, major targets of caspase-mediated proteolysis. A specific inhibitor of casapse-3, Ac-DEVD-CMK, reduced apoptosis in MD oligodendrocytes, but the rescued cells did not mature fully or express myelin-oligodendrocyte glycoprotein. These results suggest that mutant PLP affects not only cell death but also oligodendrocyte differentiation.


Subject(s)
Apoptosis/physiology , Caspases/metabolism , Myelin Proteolipid Protein/deficiency , Oligodendroglia/metabolism , Animals , Animals, Newborn , Apoptosis/drug effects , Caspase 3 , Caspase Inhibitors , Cell Differentiation/drug effects , Cell Differentiation/physiology , Cells, Cultured , Culture Media/pharmacology , Cysteine Proteinase Inhibitors/pharmacology , Male , Myelin Proteolipid Protein/genetics , Oligodendroglia/drug effects , Point Mutation/genetics , Rats , Rats, Mutant Strains
5.
J Neurosci Res ; 61(4): 376-87, 2000 Aug 15.
Article in English | MEDLINE | ID: mdl-10931524

ABSTRACT

Myelin, a multilamellar membrane structure that facilitates nerve conduction, is synthesized in the central nervous system (CNS) by oligodendrocytes. Gtx, a member of the homeodomain family of transcriptional factors, is a candidate regulator of myelin gene expression, because it is uniquely expressed in myelinating oligodendrocytes in postnatal rodent brain. To analyze the regulatory activity of Gtx, we first identified the optimal Gtx-binding sequence using an in vitro DNA-binding assay. This sequence, (A/T)TTAATGA, contains a TAAT core and is similar, but not identical, to that of other homeodomain protein binding sites. When coexpressed in cultured cells along with a minimal promoter containing five tandem repeats of this optimal Gtx-binding sequence, Gtx demonstrated repressor activity, which was also present when Gtx was tethered to DNA by way of the strong GAL4 DNA-binding domain. Truncations of the GAL4-Gtx fusion identified a portable repressor domain within a relatively proline/alanine-rich region N-terminal to the Gtx homeodomain. Cotransfection of a Gtx expression vector into a variety of cell lines, including oligodendrocytes, along with constructs containing portions of the PLP, MBP, or Gtx promoters fused to a reporter gene, however, did not modulate transcription from any of these promoter constructs. These data support the notion that the oligodendrocyte-specific homeodomain protein Gtx can act as a transcriptional repressor. In addition, they suggest that interaction of Gtx with other, as yet undefined, transcriptional regulators modifies Gtx activity in oligodendrocytes.


Subject(s)
Genes, Regulator/physiology , Homeodomain Proteins/physiology , Oligodendroglia/physiology , Repressor Proteins/physiology , Transcription Factors/physiology , Amino Acid Sequence , Animals , Cell Line , Genetic Vectors/chemistry , Homeodomain Proteins/chemistry , Molecular Sequence Data , Rats , Repressor Proteins/chemistry , Transcription Factors/chemistry , Transfection/genetics
6.
J Neurosci Res ; 60(6): 725-32, 2000 Jun 15.
Article in English | MEDLINE | ID: mdl-10861784

ABSTRACT

Prior reports demonstrated that cells of the oligodendroglial lineage are susceptible to excitotoxic necrosis mediated by alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid glutamate receptors (AMPA-GluR), and also showed that these cells express the high affinity neurotrophin receptors, TrkC and TrkA. We now report that: a) oligodendroglial progenitors (OP) and immature oligodendroglia are more vulnerable to AMPA-GluR-mediated excitotoxicity than are mature oligodendroglia; b) TrkC expression falls substantially during differentiation of cultured OP to mature oligodendroglia, whereas TrkA expression increases markedly; and c) neurotrophin-3, and to a lesser extent, nerve growth factor, protect the oligodendroglial lineage against AMPA-GluR-mediated excitotoxicity.


Subject(s)
Neurotoxins/metabolism , Neurotrophin 3/pharmacology , Oligodendroglia/drug effects , Oligodendroglia/physiology , Receptors, AMPA/physiology , Animals , Calcium/metabolism , Cell Line , Cells, Cultured , Cellular Senescence/physiology , Drug Resistance , Intracellular Membranes/metabolism , Oligodendroglia/cytology , Oligodendroglia/metabolism , Osmolar Concentration , Rats , Rats, Sprague-Dawley , Receptors, Nerve Growth Factor/metabolism , Stem Cells/drug effects , Stem Cells/metabolism , Stem Cells/physiology
7.
J Neurobiol ; 43(1): 1-17, 2000 Apr.
Article in English | MEDLINE | ID: mdl-10756062

ABSTRACT

Oligodendrocyte maturation is regulated by multiple secreted factors present in the brain during critical stages of development. Whereas most of these factors promote oligodendrocyte proliferation and survival, members of the bone morphogenetic protein family (BMPs) recently have been shown to inhibit oligodendrocyte differentiation in vitro. Oligodendrocyte precursors treated with BMPs differentiate to the astrocyte lineage. Given that cells at various stages of the oligodendrocyte lineage have distinct responses to growth factors, we hypothesized that the response to BMP would be stage-specific. Using highly purified, stage-specific cultures, we found that BMP has distinct effects on cultured oligodendrocyte preprogenitors, precursors, and mature oligodendrocytes. Oligodendrocyte preprogenitors (PSA-NCAM+, A2B5-) treated with BMP2 or BMP4 developed a novel astrocyte phenotype characterized by a morphological change and expression of glial fibrillary acidic protein (GFAP) but little glutamine synthetase expression and no labeling with A2B5 antibody. In contrast, treating oligodendrocyte precursors with BMPs resulted in the accumulation of cells with the traditional type 2 astrocyte phenotype (GFAP+, A2B5+). However, many of the cells with an astrocytic morphology did not express GFAP or glutamine synthetase unless thyroid hormone was present in the medium. The addition of fibroblast growth factor along with BMP to either oligodendrocyte preprogenitor or the oligodendrocyte precursor cells inhibited the switch to the astrocyte lineage, whereas platelet-derived growth factor addition had no effect. Treatment of mature oligodendrocytes with BMP elicited no change in morphology or expression of GFAP. These data suggest that as cells progress through the oligodendrocyte lineage, they show developmentally restricted responses to the BMPs.


Subject(s)
Bone Morphogenetic Proteins/metabolism , Bone Morphogenetic Proteins/pharmacology , Cell Lineage/drug effects , Oligodendroglia/drug effects , Oligodendroglia/metabolism , Transforming Growth Factor beta , Animals , Anti-Inflammatory Agents/pharmacology , Antineoplastic Agents/pharmacology , Astrocytes/cytology , Astrocytes/drug effects , Astrocytes/metabolism , Bone Morphogenetic Protein 2 , Bone Morphogenetic Protein 4 , Brain/drug effects , Brain/embryology , Brain/metabolism , Carcinogens/pharmacology , Cell Culture Techniques , Cell Lineage/genetics , Cell Size/drug effects , Cell Size/physiology , Dexamethasone/pharmacology , Fibroblast Growth Factor 2/metabolism , Fibroblast Growth Factor 2/pharmacology , Gangliosides/analysis , Gangliosides/metabolism , Glial Fibrillary Acidic Protein/analysis , Glial Fibrillary Acidic Protein/metabolism , Oligodendroglia/cytology , Phorbol Esters/pharmacology , Platelet-Derived Growth Factor/metabolism , Platelet-Derived Growth Factor/pharmacology , Rats , Rats, Sprague-Dawley , Stem Cells/cytology , Stem Cells/drug effects , Stem Cells/metabolism , Thyroid Hormones/metabolism , Thyroid Hormones/pharmacology , Tretinoin/metabolism , Tretinoin/pharmacology
8.
Mol Pharmacol ; 57(4): 667-78, 2000 Apr.
Article in English | MEDLINE | ID: mdl-10727511

ABSTRACT

The glial glutamate transporter GLT-1 may be the predominant Na(+)-dependent glutamate transporter in forebrain. Expression of GLT-1 correlates with astrocyte maturation in vivo and increases during synaptogenesis. In astrocyte cultures, GLT-1 expression parallels differentiation induced by cAMP analogs or by coculturing with neurons. Molecule(s) secreted by neuronal cultures contribute to this induction of GLT-1, but little is known about the signaling pathways mediating this regulation. In the present study, we determined whether growth factors previously implicated in astrocyte differentiation regulate GLT-1 expression. Of the six growth factors tested, two [epidermal growth factor (EGF) and transforming growth factor-alpha] induced expression of GLT-1 protein in cultured astrocytes. Induction of GLT-1 protein was accompanied by an increase in mRNA and in the V(max) for Na(+)-dependent glutamate transport activity. The effects of dibutyryl-cAMP and EGF were additive but were independently blocked by inhibitors of protein kinase A or protein tyrosine kinases, respectively. The induction of GLT-1 in both EGF- and dibutyryl-cAMP-treated astrocytes was blocked by inhibitors targeting phosphatidylinositol 3-kinase (PI3K) or the nuclear transcription factor-kappaB. Furthermore, transient transfection of astrocyte cultures with a constitutively active PI3K construct was sufficient to induce expression of GLT-1. These data suggest that independent but converging pathways mediate expression of GLT-1. Although an EGF receptor-specific antagonist did not block the effects of neuron-conditioned medium, the induction of GLT-1 by neuron-conditioned medium was completely abolished by inhibition of PI3K or nuclear factor-kappaB. EGF also increased expression of GLT-1 in spinal cord organotypic cultures. Together, these data suggest that activation of specific signaling pathways with EGF-like molecules may provide a novel approach for limiting excitotoxic brain injury.


Subject(s)
ATP-Binding Cassette Transporters/biosynthesis , Astrocytes/metabolism , ErbB Receptors/agonists , NF-kappa B/metabolism , Phosphatidylinositol 3-Kinases/metabolism , ATP-Binding Cassette Transporters/genetics , ATP-Binding Cassette Transporters/metabolism , Amino Acid Transport System X-AG , Animals , Bucladesine/metabolism , Cells, Cultured , Culture Media, Conditioned/metabolism , Epidermal Growth Factor/metabolism , ErbB Receptors/metabolism , RNA, Messenger/metabolism , Rats , Signal Transduction , Sodium/metabolism , Spinal Cord/metabolism , Transfection , Transforming Growth Factor alpha/metabolism , Tritium
9.
Proc Natl Acad Sci U S A ; 96(5): 2439-44, 1999 Mar 02.
Article in English | MEDLINE | ID: mdl-10051661

ABSTRACT

Proper dorsal-ventral patterning in the developing central nervous system requires signals from both the dorsal and ventral portions of the neural tube. Data from multiple studies have demonstrated that bone morphogenetic proteins (BMPs) and Sonic hedgehog protein are secreted factors that regulate dorsal and ventral specification, respectively, within the caudal neural tube. In the developing rostral central nervous system Sonic hedgehog protein also participates in ventral regionalization; however, the roles of BMPs in the developing brain are less clear. We hypothesized that BMPs also play a role in dorsal specification of the vertebrate forebrain. To test our hypothesis we implanted beads soaked in recombinant BMP5 or BMP4 into the neural tube of the chicken forebrain. Experimental embryos showed a loss of the basal telencephalon that resulted in holoprosencephaly (a single cerebral hemisphere), cyclopia (a single midline eye), and loss of ventral midline structures. In situ hybridization using a panel of probes to genes expressed in the dorsal and ventral forebrain revealed the loss of ventral markers with the maintenance of dorsal markers. Furthermore, we found that the loss of the basal telencephalon was the result of excessive cell death and not a change in cell fates. These data provide evidence that BMP signaling participates in dorsal-ventral patterning of the developing brain in vivo, and disturbances in dorsal-ventral signaling result in specific malformations of the forebrain.


Subject(s)
Bone Morphogenetic Proteins/physiology , Eye Abnormalities/etiology , Holoprosencephaly/etiology , Homeodomain Proteins , Prosencephalon/embryology , Trans-Activators , Zebrafish Proteins , Animals , Apoptosis , Body Patterning/drug effects , Bone Morphogenetic Protein 4 , Bone Morphogenetic Protein 5 , Bone Morphogenetic Proteins/toxicity , Chick Embryo , DNA-Binding Proteins/genetics , Drug Implants , Eye Proteins , Gene Expression Regulation, Developmental/drug effects , Hedgehog Proteins , In Situ Nick-End Labeling , PAX2 Transcription Factor , PAX6 Transcription Factor , Paired Box Transcription Factors , Prosencephalon/drug effects , Prosencephalon/pathology , Proteins/genetics , Proto-Oncogene Proteins/genetics , Recombinant Proteins/toxicity , Repressor Proteins , Telencephalon/abnormalities , Telencephalon/embryology , Transcription Factors/genetics , Wnt Proteins , Wnt4 Protein
10.
J Cell Biochem ; 76(2): 270-9, 1999 Dec.
Article in English | MEDLINE | ID: mdl-10618643

ABSTRACT

Oligodendrocyte differentiation is accompanied by dramatic changes in gene expression as well as cell cycle arrest. To determine whether cell cycle arrest is sufficient to induce the changes in cell phenotype associated with differentiation, we inhibited oligodendrocyte precursor proliferation in vitro by overexpressing p27, a cyclin kinase inhibitor, using a recombinant adenovirus. Ectopic expression of p27 efficiently inhibited oligodendrocyte precursor cell division, even in the presence of exogenous mitogens, by blocking the activity of the cyclin-dependent kinase, cdk2. Although the cells had stopped dividing, they did not express galactocerebroside (GalC) or myelin basic protein (MBP), changes associated with oligodendrocyte differentiation, suggesting that they had not differentiated. After removal of exogenous mitogens, however, adenovirus-expressing oligodendrocyte precursors differentiated with a temporal profile similar to that of control, uninfected oligodendrocytes, as indicated by expression of GalC and MBP. We conclude that cell cycle arrest is not sufficient to induce differentiation of dividing oligodendrocyte precursors, and that modulation of additional, as yet unknown, signaling pathways is required for this to occur.


Subject(s)
CDC2-CDC28 Kinases , Cell Cycle Proteins , Cell Cycle/physiology , Cell Differentiation/physiology , Microtubule-Associated Proteins/metabolism , Oligodendroglia/cytology , Oligodendroglia/metabolism , Tumor Suppressor Proteins , Adenoviridae/genetics , Animals , Cells, Cultured , Cyclin-Dependent Kinase 2 , Cyclin-Dependent Kinase 5 , Cyclin-Dependent Kinase Inhibitor p27 , Cyclin-Dependent Kinases/antagonists & inhibitors , Enzyme Inhibitors/metabolism , Gene Expression , Genetic Vectors , Microtubule-Associated Proteins/genetics , Protein Serine-Threonine Kinases/antagonists & inhibitors , Rats , Stem Cells/cytology , Stem Cells/metabolism
11.
J Neurosci Res ; 54(5): 623-34, 1998 Dec 01.
Article in English | MEDLINE | ID: mdl-9843153

ABSTRACT

Mutations in the proteolipid protein gene (PLP/plp), which encodes the major intrinsic membrane protein in central nervous system (CNS) myelin, cause inherited dysmyelination in mammals. One of these mutants, the myelin-deficient (md) rat, has severe dysmyelination that is associated with oligodendrocyte cell death. Using the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling (TUNEL) assay, which labels apoptotic cells, we find that cell death is increased in multiple white matter tracts of md rats. The tracts that myelinate the earliest show the earliest increase in cell death, and cell death persists for at least 22 days, the lifespan of these mutant animals. In all tracts, and at all developmental ages examined, apoptotic cells expressed the markers of mature oligodendrocytes, such as myelin basic protein, myelin-associated glycoprotein, and the Rip antigen, but not chondroitin sulfate proteoglycan, a marker of oligodendrocyte precursors. Mature oligodendrocytes fail to accumulate in md brain because they die before they fully mature.


Subject(s)
Apoptosis , Brain/pathology , Demyelinating Diseases/pathology , Myelin Proteolipid Protein/deficiency , Oligodendroglia/pathology , Age Factors , Animals , Biomarkers , Brain/metabolism , Demyelinating Diseases/genetics , Demyelinating Diseases/metabolism , Female , In Situ Nick-End Labeling , Male , Myelin Proteolipid Protein/physiology , Oligodendroglia/metabolism , Point Mutation , Rats , Rats, Mutant Strains
12.
J Biol Chem ; 273(42): 27741-8, 1998 Oct 16.
Article in English | MEDLINE | ID: mdl-9765312

ABSTRACT

The central nervous system expression of myelin basic protein (MBP) is restricted to oligodendrocytes and is developmentally regulated; these regulatory features are transcriptionally mediated. We have previously shown that the proximal 149 nucleotides of the MBP promoter were both necessary and sufficient to activate the transcription of MBP in cultured oligodendrocytes, but not in other cell types. Sequences within the distal portion of this promoter, which contains a nuclear factor 1 (NF1) binding site, repressed activation of the MBP promoter in Cos-7 cells, but not in oligodendrocytes. We now describe a sequence upstream of and partially overlapping the NF1 site that activates the MBP promoter in oligodendrocytes, but not in Cos-7 cells. A protein complex binds to this site, designated MEBA (myelinating glia-enriched DNA binding activity), and is enriched in nuclear extracts prepared from the brain, oligodendrocytes, and Schwann cells. The amount of MEBA parallels MBP expression and myelinogenesis in the developing brain and parallels new MBP expression as purified oligodendrocytes differentiate. Mutational analyses of binding and function distinguish MEBA, an activator, from NF1, a repressor of MBP transcription, and suggest that MEBA consists of at least two proteins. Because the binding sites of MEBA and NF1 overlap, we suggest that MEBA may either compete with or modify NF1 binding, thereby activating the MBP promoter in oligodendrocytes.


Subject(s)
Myelin Basic Protein/genetics , Oligodendroglia/metabolism , Promoter Regions, Genetic , Animals , Binding Sites/genetics , Cell Differentiation , Cells, Cultured , DNA Footprinting , DNA Mutational Analysis , DNA-Binding Proteins/metabolism , Gene Expression Regulation , Humans , Models, Genetic , Myelin Basic Protein/biosynthesis , NFI Transcription Factors , Oligodendroglia/cytology , Protein Binding , Sequence Deletion , Transcription Factors/metabolism
14.
J Neurosci ; 17(17): 6657-68, 1997 Sep 01.
Article in English | MEDLINE | ID: mdl-9254678

ABSTRACT

We have investigated the patterns of postnatal brain expression and DNA binding of Gtx, a homeodomain transcription factor. Gtx mRNA accumulates in parallel with the RNAs encoding the major structural proteins of myelin, myelin basic protein (MBP), and proteolipid protein (PLP) during postnatal brain development; Gtx mRNA decreases in parallel with MBP and PLP mRNAs in the brains of myelin-deficient rats, which have a point mutation in the PLP gene. Gtx mRNA is expressed in differentiated, postmitotic oligodendrocytes but is not found in oligodendrocyte precursors or astrocytes. These data thus demonstrate that Gtx is expressed uniquely in differentiated oligodendrocytes in postnatal rodent brain and that its expression is regulated in parallel with the major myelin protein mRNAs, encoding MBP and PLP, under a variety of physiologically relevant circumstances. Using a Gtx fusion protein produced in bacteria, we have confirmed that Gtx is a sequence-specific DNA-binding protein, which binds DNA sequences containing a core AT-rich homeodomain binding site. Immunoprecipitation of labeled DNA fragments encoding either the MBP or PLP promoter regions with this fusion protein has identified several Gtx-binding fragments, and we have confirmed these data using an electrophoretic mobility shift assay. In this way we have identified four Gtx binding sites within the first 750 bp of the MBP promoter and four Gtx binding sites within the first 1. 3 kb of the PLP promoter. In addition, inspection of the PLP promoter sequence demonstrates the presence of six additional Gtx binding sites. These data, taken together, strongly suggest that Gtx is important for the function of differentiated oligodendrocytes and may be involved in the regulation of myelin-specific gene expression.


Subject(s)
Homeodomain Proteins/physiology , Myelin Sheath/physiology , Nerve Tissue Proteins/physiology , Oligodendroglia/physiology , Aging/metabolism , Animals , Animals, Newborn/growth & development , Apoproteins/genetics , Binding Sites , Brain/growth & development , Brain/metabolism , Cells, Cultured , DNA-Binding Proteins/metabolism , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Mutation , Myelin Basic Protein/genetics , Myelin Proteolipid Protein/genetics , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Nervous System Malformations , Oligodendroglia/cytology , Oligodendroglia/metabolism , Promoter Regions, Genetic , RNA, Messenger/metabolism , Rats , Thymus Gland/metabolism , Tissue Distribution
15.
J Neurosci Res ; 46(4): 456-64, 1996 Nov 15.
Article in English | MEDLINE | ID: mdl-8950705

ABSTRACT

Remyelination in the CNS following demyelinating disease may be accomplished by surviving mature oligodendrocytes that dedifferentiate, proliferate, migrate, and finally regenerate myelin. We previously reported that basic fibroblast growth factor (bFGF) induces oligodendrocytes in primary mixed glial cultures to dedifferentiate and synthesize DNA (Grinspan et al.: J Neurosci Res 36:672-680, 1993). We now show that this effect is direct and not mediated through the effects of bFGF on other cell types, because we were able to demonstrate similar changes in oligodendrocyte phenotype in enriched oligodendrocyte cultures prepared by immunopanning. The bFGF-induced reversion to the precursor stage of the oligodendroglial lineage can be blocked by agents that inhibit entry to the cell cycle; thus oligodendroglial dedifferentiation is dependent on proliferation. We also report that 2 days of bFGF treatment inhibits oligodendroglial apoptosis. However, when oligodendroglia are prevented from entering the cell cycle in the presence of bFGF, apoptotic cell death is increased. Thus, bFGF induces oligodendroglial dedifferentiation if oligodendroglial DNA synthesis can occur but causes oligodendroglial apoptosis when oligodendroglial DNA synthesis is prevented.


Subject(s)
Fibroblast Growth Factor 2/pharmacology , Oligodendroglia/drug effects , Animals , Aphidicolin/pharmacology , Apoptosis/drug effects , Brain/cytology , Cell Cycle , Cell Differentiation/drug effects , Cell Division/drug effects , Cells, Cultured , DNA Fragmentation , DNA Replication/drug effects , Myelin Sheath/physiology , Nerve Regeneration , Oligodendroglia/cytology , Rats , Thymidine/pharmacology
16.
J Neurosci ; 16(19): 6107-18, 1996 Oct 01.
Article in English | MEDLINE | ID: mdl-8815893

ABSTRACT

Programmed cell death during development resulting from the lack of appropriate survival factors has been demonstrated in both neurons and oligodendrocytes and occurs mostly in the form of apoptosis. We now demonstrate that Schwann cells in the rat sciatic nerve undergo apoptosis during early postnatal development and that the amount of apoptosis is markedly increased by axotomy. The apoptotic Schwann cells express the low-affinity nerve growth factor receptor but not myelin-related proteins, indicating that they are in the premyelinating state. Apoptosis resulting from normal development or from axotomy can be inhibited markedly by exogenous neuregulin. Consistent with this, the neuregulin receptor components erbB2 and erbB3 are expressed and phosphorylated in developing sciatic nerve. These data suggest that Schwann cell number in developing peripheral nerve is regulated by apoptosis through competition for axonally derived neuregulin.


Subject(s)
Apoptosis , Axons/physiology , Glycoproteins/physiology , Receptors, Nerve Growth Factor/metabolism , Schwann Cells/physiology , Sciatic Nerve/physiology , Aging/physiology , Animals , Animals, Newborn/growth & development , Animals, Newborn/physiology , Apoptosis/drug effects , Denervation , ErbB Receptors/metabolism , Glycoproteins/pharmacology , Myelin Sheath/physiology , Neuregulins , Phenotype , Phosphorylation , Proto-Oncogene Proteins/metabolism , Rats , Rats, Sprague-Dawley , Receptor, ErbB-2/metabolism , Receptor, ErbB-3 , Sciatic Nerve/cytology , Sciatic Nerve/growth & development
17.
J Neurosci ; 15(12): 8281-94, 1995 Dec.
Article in English | MEDLINE | ID: mdl-8613761

ABSTRACT

We have examined the expression of a gap junction protein, connexin32 (Cx32), in Schwann cells and oligodendrocytes. In peripheral nerve, Cx32 is found in the paranodal myelin loops and Schmidt-Lanterman incisures of myelinating Schwann cells, and the levels of Cx32 protein and mRNA change in parallel with those of other myelin-related genes during development, Wallerian degeneration, and axonal regeneration. In the central nervous system, Cx32 is found in oligodendrocytes and their processes, but not in compact myelin, and the levels of Cx32 protein and mRNA increase during development in parallel with those of the other myelin genes. Thus, Cx32 is expressed as part of the myelinating phenotype of both Schwann cells and oligodendrocytes, indicating that this gap junction protein plays in important role in the biology of myelin-forming cells.


Subject(s)
Central Nervous System/metabolism , Connexins/metabolism , Myelin Proteins/metabolism , Peripheral Nerves/metabolism , Animals , Axons/physiology , Cell Communication , Colforsin/pharmacology , Connexins/genetics , Myelin Proteins/genetics , Myelin Sheath/physiology , Oligodendroglia/metabolism , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Schwann Cells/metabolism , Schwann Cells/physiology , Gap Junction beta-1 Protein
18.
J Neurocytol ; 24(10): 725-34, 1995 Oct.
Article in English | MEDLINE | ID: mdl-8586993

ABSTRACT

Oligodendroglia synthesize myelin in the CNS. In vitro, oligodendroglia may be identified by the binding of monoclonal antibodies against galactocerebroside, a myelin-specific galactolipid. Oligodendroglial trophic factor is a protein mitogen for cells of the oligodendroglial lineage. When oligodendroglia in cerebral white matter cultures are treated with oligodendroglial trophic factor, galactocerebroside-positive cells undergo mitosis but fail to express the myelin structural proteins, myelin basic protein and proteolipid protein. Oligodendroglia treated with oligodendroglial trophic factor, however, do express 2',3'-cyclic nucleotide 3'-phosphodiesterase and myelin-associated glycoprotein in a manner similar to oligodendroglia treated with platelet-derived growth factor. Oligodendroglial trophic factor, therefore, generates a population of somewhat 'immature' oligodendroglia, which are galactocerebroside, myelin-associated glycoprotein and 2', 3'-cyclic nucleotide 3' phosphodiesterase positive but myelin basic protein and proteolipid protein negative.


Subject(s)
Gene Expression , Mitogens/pharmacology , Myelin Proteins/genetics , Nerve Growth Factors/pharmacology , Neuroglia/drug effects , Neuroglia/metabolism , 2',3'-Cyclic-Nucleotide Phosphodiesterases/genetics , Animals , Antibodies, Monoclonal , Brain/metabolism , Cells, Cultured , Galactosylceramides/analysis , Myelin Basic Protein/genetics , Myelin Proteolipid Protein/genetics , Myelin-Associated Glycoprotein/genetics , Platelet-Derived Growth Factor/pharmacology , RNA, Messenger/analysis , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley
19.
J Neurosci Res ; 41(4): 540-51, 1995 Jul 01.
Article in English | MEDLINE | ID: mdl-7473886

ABSTRACT

Mature oligodendroglia, which synthesize and express lipids and proteins characteristic of myelin, are generated from precursor cells which are formed in germinal matrix, then migrate widely through the neuraxis. We now demonstrate that these precursor cells can be recognized at a very early stage by their surface expression of polysialylated neural cell adhesion molecules (PSA-NCAM), and only later bind anti-ganglioside antibodies that had previously been used to recognize "O-2A" oligodendroglial precursor cells. PSA-NCAM expression by these cells is likely to be of functional significance, since a recent study demonstrated that O-2A cells become immobile when stripped of PSA-NCAM. Platelet-derived growth factor (PDGF) proved to be a survival factor for these PSA-NCAM+cells, and in a defined medium, PDGF was sufficient to ensure maturation of immunopurified PSA-NCAM+cells to oligodendroglia.


Subject(s)
Cell Survival , Oligodendroglia/drug effects , Platelet-Derived Growth Factor/pharmacology , Animals , Cells, Cultured , Immunohistochemistry , In Situ Hybridization , Prosencephalon/metabolism , Rats , Rats, Inbred Strains , Time Factors
20.
J Neurochem ; 64(6): 2572-80, 1995 Jun.
Article in English | MEDLINE | ID: mdl-7760037

ABSTRACT

Pharmacological and molecular biological studies provide evidence for subtypes of sodium-dependent high-affinity glutamate (Glu) transport in the mammalian CNS. At least some of these transporters appear to be selectively expressed in different brain regions or by different cell types. In the present study, the properties of L-[3H]Glu transport were characterized using astrocyte-enriched cultures prepared from cerebellum and cortex. In both brain regions, the kinetic data for sodium-dependent transport were consistent with a single site with Km values of 91 +/- 17 microM in cortical glial cells and 66 +/- 23 microM in cerebellar glial cells. The capacities were 6.1 +/- 1.6 nmol/mg of protein/min in cortical glial cells and 8.4 +/- 0.9 nmol/mg of protein/min in cerebellar glial cells. The potencies of approximately 40 excitatory amino acid analogues for inhibition of sodium-dependent transport into glial cells prepared from cortex and cerebellum were examined, including compounds that are selective inhibitors of transport in synaptosomes prepared from either cerebellum or cortex. Of the analogues tested, 14 inhibited transport activity by > 50% at 1 mM concentrations. Unlike L-[3H]Glu transport in synaptosomes prepared from cerebellum or cortex, there were no large differences between the potencies of compounds for inhibition of transport measured in glial cells prepared from these two brain regions. With the exception of (2S,1'R,2'R)-2-(carboxycyclopropyl)glycine and L-alpha-aminoadipate, all of the compounds examined were approximately 10-200-fold less potent as inhibitors of L-[3H]Glu transport measured in glial cells than as inhibitors of transport measured in synaptosomes prepared from their respective brain regions.(ABSTRACT TRUNCATED AT 250 WORDS)


Subject(s)
Glutamic Acid/metabolism , Neuroglia/metabolism , Sodium/physiology , Animals , Binding, Competitive , Biological Transport/drug effects , Cells, Cultured , Cerebellum/cytology , Cerebellum/metabolism , Cerebral Cortex/cytology , Cerebral Cortex/metabolism , Excitatory Amino Acids/pharmacology , Rats , Rats, Sprague-Dawley , Synaptosomes/metabolism
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