Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 21
Filtrar
Más filtros

Banco de datos
Tipo del documento
Intervalo de año de publicación
1.
Cell Metab ; 4(5): 349-62, 2006 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-17055784

RESUMEN

Huntington's disease (HD) is a fatal, dominantly inherited disorder caused by polyglutamine repeat expansion in the huntingtin (htt) gene. Here, we observe that HD mice develop hypothermia associated with impaired activation of brown adipose tissue (BAT). Although sympathetic stimulation of PPARgamma coactivator 1alpha (PGC-1alpha) was intact in BAT of HD mice, uncoupling protein 1 (UCP-1) induction was blunted. In cultured cells, expression of mutant htt suppressed UCP-1 promoter activity; this was reversed by PGC-1alpha expression. HD mice showed reduced food intake and increased energy expenditure, with dysfunctional BAT mitochondria. PGC-1alpha is a known regulator of mitochondrial function; here, we document reduced expression of PGC-1alpha target genes in HD patient and mouse striatum. Mitochondria of HD mouse brain show reduced oxygen consumption rates. Finally, HD striatal neurons expressing exogenous PGC-1alpha were resistant to 3-nitropropionic acid treatment. Altered PGC-1alpha function may thus link transcription dysregulation and mitochondrial dysfunction in HD.


Asunto(s)
Tejido Adiposo Pardo/fisiopatología , Regulación de la Temperatura Corporal/genética , Proteínas de Choque Térmico/metabolismo , Enfermedad de Huntington/etiología , Factores de Transcripción/metabolismo , Animales , Temperatura Corporal/genética , Células Cultivadas , Modelos Animales de Enfermedad , Proteínas de Choque Térmico/genética , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Ratones , Ratones Transgénicos , Mitocondrias/fisiología , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Transducción de Señal/genética , Factores de Transcripción/genética , Transcripción Genética
2.
Nat Med ; 9(8): 1033-8, 2003 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-12872164

RESUMEN

The mechanisms of retinoid activity in tumors remain largely unknown. Here we establish that retinoids cause extensive apoptosis of medulloblastoma cells. In a xenograft model, retinoids largely abrogated tumor growth. Using receptor-specific retinoid agonists, we defined a subset of mRNAs that were induced by all active retinoids in retinoid-sensitive cell lines. We also identified bone morphogenetic protein-2 (BMP-2) as a candidate mediator of retinoid activity. BMP-2 protein induced medulloblastoma cell apoptosis, whereas the BMP-2 antagonist noggin blocked both retinoid and BMP-2-induced apoptosis. BMP-2 also induced p38 mitogen-activated protein kinase (MAPK), which is necessary for BMP-2- and retinoid-induced apoptosis. Retinoid-resistant medulloblastoma cells underwent apoptosis when treated with BMP-2 or when cultured with retinoid-sensitive medulloblastoma cells. Retinoid-induced expression of BMP-2 is thus necessary and sufficient for apoptosis of retinoid-responsive cells, and expression of BMP-2 by retinoid-sensitive cells is sufficient to induce apoptosis in surrounding retinoid-resistant cells.


Asunto(s)
Apoptosis , Proteínas Morfogenéticas Óseas/metabolismo , Neoplasias Encefálicas/metabolismo , Meduloblastoma/metabolismo , Comunicación Paracrina , Retinoides/farmacología , Factor de Crecimiento Transformador beta , Animales , Proteína Morfogenética Ósea 2 , Receptores de Proteínas Morfogenéticas Óseas , Neoplasias Encefálicas/patología , Femenino , Perfilación de la Expresión Génica , Humanos , Meduloblastoma/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Trasplante de Neoplasias , Análisis de Secuencia por Matrices de Oligonucleótidos , Receptores de Factores de Crecimiento/genética , Receptores de Factores de Crecimiento/metabolismo , Trasplante Heterólogo , Células Tumorales Cultivadas
3.
PLoS Genet ; 3(4): e59, 2007 Apr 20.
Artículo en Inglés | MEDLINE | ID: mdl-17447843

RESUMEN

Many neurodegenerative diseases have a hallmark regional and cellular pathology. Gene expression analysis of healthy tissues may provide clues to the differences that distinguish resistant and sensitive tissues and cell types. Comparative analysis of gene expression in healthy mouse and human brain provides a framework to explore the ability of mice to model diseases of the human brain. It may also aid in understanding brain evolution and the basis for higher order cognitive abilities. Here we compare gene expression profiles of human motor cortex, caudate nucleus, and cerebellum to one another and identify genes that are more highly expressed in one region relative to another. We separately perform identical analysis on corresponding brain regions from mice. Within each species, we find that the different brain regions have distinctly different expression profiles. Contrasting between the two species shows that regionally enriched genes in one species are generally regionally enriched genes in the other species. Thus, even when considering thousands of genes, the expression ratios in two regions from one species are significantly correlated with expression ratios in the other species. Finally, genes whose expression is higher in one area of the brain relative to the other areas, in other words genes with patterned expression, tend to have greater conservation of nucleotide sequence than more widely expressed genes. Together these observations suggest that region-specific genes have been conserved in the mammalian brain at both the sequence and gene expression levels. Given the general similarity between patterns of gene expression in healthy human and mouse brains, we believe it is reasonable to expect a high degree of concordance between microarray phenotypes of human neurodegenerative diseases and their mouse models. Finally, these data on very divergent species provide context for studies in more closely related species that address questions such as the origins of cognitive differences.


Asunto(s)
Encéfalo/metabolismo , Evolución Molecular , Expresión Génica , Adulto , Anciano , Animales , Secuencia Conservada , Femenino , Perfilación de la Expresión Génica , Variación Genética , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Especificidad de Órganos , Distribución Tisular
4.
Sci Transl Med ; 12(533)2020 03 04.
Artículo en Inglés | MEDLINE | ID: mdl-32132215

RESUMEN

On-target, off-tissue toxicity limits the systemic use of drugs that would otherwise reduce symptoms or reverse the damage of arthritic diseases, leaving millions of patients in pain and with limited physical mobility. We identified cystine-dense peptides (CDPs) that rapidly accumulate in cartilage of the knees, ankles, hips, shoulders, and intervertebral discs after systemic administration. These CDPs could be used to concentrate arthritis drugs in joints. A cartilage-accumulating peptide, CDP-11R, reached peak concentration in cartilage within 30 min after administration and remained detectable for more than 4 days. Structural analysis of the peptides by crystallography revealed that the distribution of positive charge may be a distinguishing feature of joint-accumulating CDPs. In addition, quantitative whole-body autoradiography showed that the disulfide-bonded tertiary structure is critical for cartilage accumulation and retention. CDP-11R distributed to joints while carrying a fluorophore imaging agent or one of two different steroid payloads, dexamethasone (dex) and triamcinolone acetonide (TAA). Of the two payloads, the dex conjugate did not advance because the free drug released into circulation was sufficient to cause on-target toxicity. In contrast, the CDP-11R-TAA conjugate alleviated joint inflammation in the rat collagen-induced model of rheumatoid arthritis while avoiding toxicities that occurred with nontargeted steroid treatment at the same molar dose. This conjugate shows promise for clinical development and establishes proof of concept for multijoint targeting of disease-modifying therapeutic payloads.


Asunto(s)
Artritis Experimental , Corticoesteroides , Animales , Artritis Experimental/tratamiento farmacológico , Cartílago , Humanos , Péptidos , Ratas , Esteroides
5.
J Mol Biol ; 432(14): 3989-4009, 2020 06 26.
Artículo en Inglés | MEDLINE | ID: mdl-32304700

RESUMEN

The impenetrability of the blood-brain barrier (BBB) to most conventional drugs impedes the treatment of central nervous system (CNS) disorders. Interventions for diseases like brain cancer, neurodegeneration, or age-associated inflammatory processes require varied approaches to CNS drug delivery. Cystine-dense peptides (CDPs) have drawn recent interest as drugs or drug-delivery vehicles. Found throughout the phylogenetic tree, often in drug-like roles, their size, stability, and protein interaction capabilities make CDPs an attractive mid-size biologic scaffold to complement conventional antibody-based drugs. Here, we describe the identification, maturation, characterization, and utilization of a CDP that binds to the transferrin receptor (TfR), a native receptor and BBB transporter for the iron chaperone transferrin. We developed variants with varying binding affinities (KD as low as 216 pM), co-crystallized it with the receptor, and confirmed murine cross-reactivity. It accumulates in the mouse CNS at ~25% of blood levels (CNS blood content is only ~1%-6%) and delivers neurotensin, an otherwise non-BBB-penetrant neuropeptide, at levels capable of modulating CREB signaling in the mouse brain. Our work highlights the utility of CDPs as a diverse, easy-to-screen scaffold family worthy of inclusion in modern drug discovery strategies, demonstrated by the discovery of a candidate CNS drug delivery vehicle ready for further optimization and preclinical development.


Asunto(s)
Barrera Hematoencefálica/efectos de los fármacos , Enfermedades del Sistema Nervioso Central/tratamiento farmacológico , Sistemas de Liberación de Medicamentos , Péptidos/farmacología , Animales , Antígenos CD/química , Antígenos CD/efectos de los fármacos , Antígenos CD/genética , Antígenos CD/farmacología , Sistema Nervioso Central/efectos de los fármacos , Cistina/química , Cistina/genética , Humanos , Inflamación/tratamiento farmacológico , Inflamación/patología , Ratones , Neuropéptidos/química , Neuropéptidos/farmacología , Neurotensina/química , Neurotensina/farmacología , Péptidos/química , Unión Proteica/efectos de los fármacos , Receptores de Transferrina/química , Receptores de Transferrina/efectos de los fármacos , Receptores de Transferrina/genética
6.
J Neurosci ; 27(43): 11758-68, 2007 Oct 24.
Artículo en Inglés | MEDLINE | ID: mdl-17959817

RESUMEN

Many pathways have been proposed as contributing to Huntington's disease (HD) pathogenesis, but generally the in vivo effects of their perturbation have not been compared with reference data from human patients. Here we examine how accurately mechanistically motivated and genetic HD models recapitulate the striatal gene expression phenotype of human HD. The representative genetic model was the R6/2 transgenic mouse, which expresses a fragment of the huntingtin protein containing a long CAG repeat. Pathogenic mechanisms examined include mitochondrial dysfunction; profiled in 3-nitropropionic acid-treated rats, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice, and PGC-1alpha knock-out mice; and depletion of brain-derived neurotrophic factor (BDNF) using heterozygous and forebrain-specific BDNF-knock-out mice (BDNF(HET), Emx-BDNF(KO)). Based on striatal gene expression, we find the BDNF models, both heterozygous and homozygous knock-outs, to be more like human HD than the other HD models. This implicates reduced trophic support as a major pathway contributing to striatal degeneration in HD. Because the majority of striatal BDNF is synthesized by cortical neurons, the data also imply that cortical dysfunction contributes to HD's hallmark effects on the basal ganglia. Finally, the results suggest that striatal lesions caused by mitochondrial toxins may arise via pathways different from those that drive neurodegeneration in HD. Based on these findings, we present a testable model of HD pathogenesis that, unlike most models, begins to account for regional specificity in human HD and the absence of such specificity in genetic mouse models of HD.


Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/deficiencia , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica/métodos , Enfermedad de Huntington/metabolismo , Degeneración Nerviosa/metabolismo , Animales , Factor Neurotrófico Derivado del Encéfalo/genética , Cuerpo Estriado/patología , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/patología , Ratones , Ratones Noqueados , Ratones Transgénicos , Degeneración Nerviosa/genética , Degeneración Nerviosa/patología , Ratas , Ratas Endogámicas Lew
7.
Nat Med ; 24(11): 1752-1761, 2018 11.
Artículo en Inglés | MEDLINE | ID: mdl-30349086

RESUMEN

Brain tumors are the leading cause of cancer-related death in children. Genomic studies have provided insights into molecular subgroups and oncogenic drivers of pediatric brain tumors that may lead to novel therapeutic strategies. To evaluate new treatments, better preclinical models adequately reflecting the biological heterogeneity are needed. Through the Children's Oncology Group ACNS02B3 study, we have generated and comprehensively characterized 30 patient-derived orthotopic xenograft models and seven cell lines representing 14 molecular subgroups of pediatric brain tumors. Patient-derived orthotopic xenograft models were found to be representative of the human tumors they were derived from in terms of histology, immunohistochemistry, gene expression, DNA methylation, copy number, and mutational profiles. In vivo drug sensitivity of targeted therapeutics was associated with distinct molecular tumor subgroups and specific genetic alterations. These models and their molecular characterization provide an unprecedented resource for the cancer community to study key oncogenic drivers and to evaluate novel treatment strategies.


Asunto(s)
Bancos de Muestras Biológicas , Neoplasias Encefálicas/patología , Inmunohistoquímica , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Animales , Línea Celular Tumoral , Niño , Preescolar , Metilación de ADN/genética , Femenino , Genómica , Humanos , Masculino , Ratones , Mutación , Pediatría
8.
BMC Bioinformatics ; 7: 211, 2006 Apr 19.
Artículo en Inglés | MEDLINE | ID: mdl-16623940

RESUMEN

BACKGROUND: Gene expression microarray experiments are expensive to conduct and guidelines for acceptable quality control at intermediate steps before and after the samples are hybridised to chips are vague. We conducted an experiment hybridising RNA from human brain to 117 U133A Affymetrix GeneChips and used these data to explore the relationship between 4 pre-chip variables and 22 post-chip outcomes and quality control measures. RESULTS: We found that the pre-chip variables were significantly correlated with each other but that this correlation was strongest between measures of RNA quality and cRNA yield. Post-mortem interval was negatively correlated with these variables. Four principal components, reflecting array outliers, array adjustment, hybridisation noise and RNA integrity, explain about 75% of the total post-chip measure variability. Two significant canonical correlations existed between the pre-chip and post-chip variables, derived from MAS 5.0, dChip and the Bioconductor packages affy and affyPLM. The strongest (CANCOR 0.838, p < 0.0001) correlated RNA integrity and yield with post chip quality control (QC) measures indexing 3'/5' RNA ratios, bias or scaling of the chip and scaling of the variability of the signal across the chip. Post-mortem interval was relatively unimportant. We also found that the RNA integrity number (RIN) could be moderately well predicted by post-chip measures B_ACTIN35, GAPDH35 and SF. CONCLUSION: We have found that the post-chip variables having the strongest association with quantities measurable before hybridisation are those reflecting RNA integrity. Other aspects of quality, such as noise measures (reflecting the execution of the assay) or measures reflecting data quality (outlier status and array adjustment variables) are not well predicted by the variables we were able to determine ahead of time. There could be other variables measurable pre-hybridisation which may be better associated with expression data quality measures. Uncovering such connections could create savings on costly microarray experiments by eliminating poor samples before hybridisation.


Asunto(s)
Artefactos , Perfilación de la Expresión Génica/instrumentación , Modelos Genéticos , Análisis de Secuencia por Matrices de Oligonucleótidos/instrumentación , Garantía de la Calidad de Atención de Salud/métodos , Simulación por Computador , Interpretación Estadística de Datos , Diseño de Equipo , Análisis de Falla de Equipo , Modelos Estadísticos , Control de Calidad , Reproducibilidad de los Resultados , Sensibilidad y Especificidad
9.
Sci Transl Med ; 7(284): 284ra58, 2015 Apr 22.
Artículo en Inglés | MEDLINE | ID: mdl-25904742

RESUMEN

A fundamental problem in cancer drug development is that antitumor efficacy in preclinical cancer models does not translate faithfully to patient outcomes. Much of early cancer drug discovery is performed under in vitro conditions in cell-based models that poorly represent actual malignancies. To address this inconsistency, we have developed a technology platform called CIVO, which enables simultaneous assessment of up to eight drugs or drug combinations within a single solid tumor in vivo. The platform is currently designed for use in animal models of cancer and patients with superficial tumors but can be modified for investigation of deeper-seated malignancies. In xenograft lymphoma models, CIVO microinjection of well-characterized anticancer agents (vincristine, doxorubicin, mafosfamide, and prednisolone) induced spatially defined cellular changes around sites of drug exposure, specific to the known mechanisms of action of each drug. The observed localized responses predicted responses to systemically delivered drugs in animals. In pair-matched lymphoma models, CIVO correctly demonstrated tumor resistance to doxorubicin and vincristine and an unexpected enhanced sensitivity to mafosfamide in multidrug-resistant lymphomas compared with chemotherapy-naïve lymphomas. A CIVO-enabled in vivo screen of 97 approved oncology agents revealed a novel mTOR (mammalian target of rapamycin) pathway inhibitor that exhibits significantly increased tumor-killing activity in the drug-resistant setting compared with chemotherapy-naïve tumors. Finally, feasibility studies to assess the use of CIVO in human and canine patients demonstrated that microinjection of drugs is toxicity-sparing while inducing robust, easily tracked, drug-specific responses in autochthonous tumors, setting the stage for further application of this technology in clinical trials.


Asunto(s)
Antineoplásicos/química , Ensayos de Selección de Medicamentos Antitumorales/métodos , Linfoma/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Animales , Biomarcadores , Línea Celular Tumoral , Ciclofosfamida/análogos & derivados , Ciclofosfamida/química , Perros , Doxorrubicina/química , Sistemas de Liberación de Medicamentos , Resistencia a Antineoplásicos/efectos de los fármacos , Humanos , Ratones , Ratones Desnudos , Ratones SCID , Trasplante de Neoplasias , Prednisolona/química , Serina-Treonina Quinasas TOR/metabolismo , Vincristina/química
10.
Neuroreport ; 20(12): 1098-103, 2009 Aug 05.
Artículo en Inglés | MEDLINE | ID: mdl-19590393

RESUMEN

Neuroinflammation is a prominent feature of many neurodegenerative diseases, however, little is known about neuroinflammation in Huntington's disease. We used quantitative real time-PCR to compare the expression level of neuroinflammation-associated mediators in the striatum, cortex, and cerebellum from post-mortem Huntington's disease patient samples with controls. We found increased expression of several key inflammatory mediators, including CCL2 and IL-10, specifically in the striatum of Huntington's disease patients, the main area affected by this pathology. Remarkably, we also found upregulation of IL-6, IL-8, and MMP9, in the cortex and notably the cerebellum, a brain area commonly thought to be spared by Huntington's disease. Our data suggest that neuroinflammation is a prominent feature associated with Huntington's disease and may constitute a novel target for therapeutic intervention.


Asunto(s)
Cerebelo/inmunología , Corteza Cerebral/inmunología , Cuerpo Estriado/inmunología , Enfermedad de Huntington/inmunología , Adulto , Anciano , Cerebelo/metabolismo , Corteza Cerebral/metabolismo , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Cuerpo Estriado/metabolismo , Femenino , Expresión Génica , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Interleucina-10/genética , Interleucina-10/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Interleucina-8/genética , Interleucina-8/metabolismo , Masculino , Metaloproteinasa 9 de la Matriz/genética , Metaloproteinasa 9 de la Matriz/metabolismo , Persona de Mediana Edad , ARN Mensajero/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo
11.
Hum Mol Genet ; 16(15): 1845-61, 2007 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-17519223

RESUMEN

To test the hypotheses that mutant huntingtin protein length and wild-type huntingtin dosage have important effects on disease-related transcriptional dysfunction, we compared the changes in mRNA in seven genetic mouse models of Huntington's disease (HD) and postmortem human HD caudate. Transgenic models expressing short N-terminal fragments of mutant huntingtin (R6/1 and R6/2 mice) exhibited the most rapid effects on gene expression, consistent with previous studies. Although changes in the brains of knock-in and full-length transgenic models of HD took longer to appear, 15- and 22-month CHL2(Q150/Q150), 18-month Hdh(Q92/Q92) and 2-year-old YAC128 animals also exhibited significant HD-like mRNA signatures. Whereas it was expected that the expression of full-length huntingtin transprotein might result in unique gene expression changes compared with those caused by the expression of an N-terminal huntingtin fragment, no discernable differences between full-length and fragment models were detected. In addition, very high correlations between the signatures of mice expressing normal levels of wild-type huntingtin and mice in which the wild-type protein is absent suggest a limited effect of the wild-type protein to change basal gene expression or to influence the qualitative disease-related effect of mutant huntingtin. The combined analysis of mouse and human HD transcriptomes provides important temporal and mechanistic insights into the process by which mutant huntingtin kills striatal neurons. In addition, the discovery that several available lines of HD mice faithfully recapitulate the gene expression signature of the human disorder provides a novel aspect of validation with respect to their use in preclinical therapeutic trials.


Asunto(s)
Cuerpo Estriado/metabolismo , Expresión Génica , Enfermedad de Huntington/genética , Mutación , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/genética , Animales , Encéfalo/metabolismo , Encéfalo/patología , Modelos Animales de Enfermedad , Dosificación de Gen , Humanos , Proteína Huntingtina , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Ratones , Ratones Mutantes , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Proteínas Nucleares/metabolismo , Fenotipo , ARN Mensajero/metabolismo
12.
Hum Mol Genet ; 15(6): 965-77, 2006 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-16467349

RESUMEN

Huntington's disease (HD) pathology is well understood at a histological level but a comprehensive molecular analysis of the effect of the disease in the human brain has not previously been available. To elucidate the molecular phenotype of HD on a genome-wide scale, we compared mRNA profiles from 44 human HD brains with those from 36 unaffected controls using microarray analysis. Four brain regions were analyzed: caudate nucleus, cerebellum, prefrontal association cortex [Brodmann's area 9 (BA9)] and motor cortex [Brodmann's area 4 (BA4)]. The greatest number and magnitude of differentially expressed mRNAs were detected in the caudate nucleus, followed by motor cortex, then cerebellum. Thus, the molecular phenotype of HD generally parallels established neuropathology. Surprisingly, no mRNA changes were detected in prefrontal association cortex, thereby revealing subtleties of pathology not previously disclosed by histological methods. To establish that the observed changes were not simply the result of cell loss, we examined mRNA levels in laser-capture microdissected neurons from Grade 1 HD caudate compared to control. These analyses confirmed changes in expression seen in tissue homogenates; we thus conclude that mRNA changes are not attributable to cell loss alone. These data from bona fide HD brains comprise an important reference for hypotheses related to HD and other neurodegenerative diseases.


Asunto(s)
Encéfalo/metabolismo , Perfilación de la Expresión Génica , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Adulto , Anciano , Axones/metabolismo , Encéfalo/patología , Muerte Celular/genética , Femenino , Humanos , Enfermedad de Huntington/patología , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero/biosíntesis , Transducción de Señal/genética
13.
Stat Med ; 24(15): 2281-98, 2005 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-15889452

RESUMEN

Which significance test is carried out when the number of repeats is small in microarray experiments can dramatically influence the results. When in two sample comparisons both conditions have fewer than, say, five repeats traditional test statistics require extreme results, before a gene is considered statistically significant differentially expressed after a multiple comparisons correction. In the literature many approaches to circumvent this problem have been proposed. Some of these proposals use (empirical) Bayes arguments to moderate the variance estimates for individual genes. Other proposals try to stabilize these variance estimate by combining groups of genes or similar experiments. In this paper we compare several of these approaches, both on data sets where both experimental conditions are the same, and thus few statistically significant differentially expressed genes should be identified, and on experiments where both conditions do differ. This allows us to identify which approaches are most powerful without identifying many false positives. We conclude that after balancing the numbers of false positives and true positives an empirical Bayes approach and an approach which combines experiments perform best. Standard t-tests are inferior and offer almost no power when the sample size is small.


Asunto(s)
Teorema de Bayes , Interpretación Estadística de Datos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Animales , Simulación por Computador , Dermatoglifia del ADN/métodos , Reacciones Falso Positivas , Humanos , Enfermedad de Huntington/genética , Ratones
14.
Hum Mol Genet ; 14(13): 1863-76, 2005 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-15888475

RESUMEN

Huntington's disease (HD) is an incurable and fatal neurodegenerative disorder. Improvements in the objective measurement of HD will lead to more efficient clinical trials and earlier therapeutic intervention. We hypothesized that abnormalities seen in the R6/2 mouse, a greatly accelerated HD model, might highlight subtle phenotypes in other mouse models and human HD. In this paper, we identify common gene expression changes in skeletal muscle from R6/2 mice, Hdh(CAG(150)) homozygous knock-in mice and HD patients. This HD-triggered gene expression phenotype is consistent with the beginnings of a transition from fast-twitch to slow-twitch muscle fiber types. Metabolic adaptations similar to those induced by diabetes or fasting are also present but neither metabolic disorder can explain the full phenotype of HD muscle. The HD-induced gene expression changes reflect disease progression. This raises the possibility that muscle gene expression may be used as an objective biomarker to complement clinical HD-rating systems. Furthermore, an understanding of the molecular basis of muscle dysfunction in HD should provide insight into mechanisms involved in neuronal abnormalities and neurodegeneration.


Asunto(s)
Regulación de la Expresión Génica , Enfermedad de Huntington/metabolismo , Músculo Esquelético/metabolismo , Animales , Biomarcadores , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Humanos , Enfermedad de Huntington/genética , Enfermedad de Huntington/patología , Ratones , Ratones Noqueados , Fibras Musculares de Contracción Rápida/metabolismo , Fibras Musculares de Contracción Rápida/patología , Fibras Musculares de Contracción Lenta/metabolismo , Fibras Musculares de Contracción Lenta/patología , Músculo Esquelético/patología , Análisis de Secuencia por Matrices de Oligonucleótidos
15.
Hum Mol Genet ; 14(20): 3065-78, 2005 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-16183657

RESUMEN

In postmortem Huntington's disease brains, mutant htt is present in both nuclear and cytoplasmic compartments. To dissect the impact of nuclear and extranuclear mutant htt on the initiation and progression of disease, we generated a series of transgenic mouse lines in which nuclear localization or nuclear export signal sequences have been placed N-terminal to the htt exon 1 protein carrying 144 glutamines. Our data indicate that the exon 1 mutant protein is present in the nucleus as part of an oligomeric or aggregation complex. Increasing the concentration of the mutant transprotein in the nucleus is sufficient for and dramatically accelerates the onset and progression of behavioral phenotypes. Furthermore, nuclear exon 1 mutant protein is sufficient to induce cytoplasmic neurodegeneration and transcriptional dysregulation. However, our data suggest that cytoplasmic mutant exon 1 htt, if present, contributes to disease progression.


Asunto(s)
Núcleo Celular/metabolismo , Modelos Animales de Enfermedad , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Péptidos/metabolismo , Animales , Encéfalo/metabolismo , Encéfalo/patología , Encéfalo/ultraestructura , Citoplasma/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Proteína Huntingtina , Sustancias Macromoleculares , Ratones , Ratones Transgénicos , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Señales de Localización Nuclear , Proteínas Nucleares/química , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fenotipo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Prueba de Desempeño de Rotación con Aceleración Constante , Transcripción Genética , Transgenes/genética
16.
Hum Mol Genet ; 11(19): 2207-21, 2002 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-12217949

RESUMEN

We present a simple method to assign approximate P-values to gene expression changes detected with Affymetrix oligonucleotide arrays and software. The method pools data for groups of genes and a small number of like-to-like comparisons in order to estimate the significance of changes observed for single genes in comparisons of experimental interest. Statistical significance levels are based on the observed variability in the fractional majority of probe pairs that indicate increasing or decreasing differential expression in comparisons of technical replicates. From this reference distribution or error model, we compute the expected frequency for fractional majorities in comparisons for N > or = 2. These computed distributions are the source of P-value estimates for changes seen in the experimental comparisons. The method is intended to complement the Affymetrix software and to rationalize gene selection for experimental designs involving limited replication.


Asunto(s)
Perfilación de la Expresión Génica , Análisis de Secuencia por Matrices de Oligonucleótidos , Animales , Interpretación Estadística de Datos , Modelos Animales de Enfermedad , Ratones
17.
Hum Mol Genet ; 11(19): 2223-32, 2002 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-12217950

RESUMEN

A randomization procedure to evaluate the significance level and the false-discovery rate in complex microarray experiments is proposed. A related graph can be used to compare different test statistics that can be used to analyze the same experiment. This graph is closely related to receiver operator characteristic (ROC) curves. The proposed method is applied to a subset of the data from a cell-line experiment related to Huntington's disease. A small simulation study compares the effectiveness of the proposed procedure with the significance analysis of microarrays (SAM) procedure.


Asunto(s)
Interpretación Estadística de Datos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Simulación por Computador , Enfermedad de Huntington/genética , Modelos Genéticos , Curva ROC , Distribución Aleatoria , Análisis de Regresión
18.
Hum Mol Genet ; 11(17): 1967-76, 2002 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-12165558

RESUMEN

Kennedy's disease is a degenerative disease of motor neurons in which the causative mutation is expansion of a CAG/polyglutamine tract near the 5' end of the androgen receptor gene. The mutant protein misfolds, aggregates, and interacts abnormally with other proteins, leading to a novel, toxic gain of function and an alteration of normal function. We used a cell culture model to explore the mechanisms underlying the alterations in androgen receptor function conferred by the mutation. Here we show that cells expressing the wild-type androgen receptor with 24 CAG repeats respond to ligand by showing trophic effects including prolonged survival in low serum, whereas cells expressing the mutant receptor with 65 CAG repeats do not show a robust trophic response. This partial loss of function correlates with decreased levels of the mutant protein due to its preferential degradation by the ubiquitin-proteasome pathway. Expression analysis using oligonucleotide arrays confirms that the mutant receptor has undergone a partial loss of function, and fails to regulate a subset of genes whose expression is normally affected by ligand activation of the wild-type receptor. The mutant receptor has also undergone several functionally important post-translational modifications in the absence of ligand that the wild-type receptor undergoes in the presence of ligand, including acetylation and phosphorylation. These modifications correlate with a ligand-independent gain of function exhibited by the mutant receptor in expression analysis. Our findings suggest that polyglutamine expansion alters androgen receptor function by promoting its degradation and by modifying its activity as a transcription factor.


Asunto(s)
Regulación de la Expresión Génica/fisiología , Neuronas Motoras/metabolismo , Atrofia Muscular Espinal/metabolismo , Péptidos/metabolismo , Receptores Androgénicos/metabolismo , Animales , Supervivencia Celular , Células Cultivadas , Perfilación de la Expresión Génica , Humanos , Ratones , Modelos Biológicos , Neuronas Motoras/patología , Atrofia Muscular Espinal/patología , Mutagénesis Sitio-Dirigida , Análisis de Secuencia por Matrices de Oligonucleótidos , Péptido Hidrolasas/metabolismo , Péptidos/genética , Receptores Androgénicos/genética , Congéneres de la Testosterona/farmacología , Transcripción Genética , Expansión de Repetición de Trinucleótido/genética , Ubiquitina/metabolismo
19.
J Cell Biochem ; 90(5): 1068-78, 2003 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-14624465

RESUMEN

Large-scale gene expression measurements with oligonucleotide microarrays have contributed tremendously to biological research. However, to distinguish between relevant expression changes and falsely identified positives, the source and magnitude of errors must be understood. Here, we report a source of biological variability in microarray experiments with stably transfected cell lines. Mouse embryonic fibroblast (MEF/3T3) and rat schwannoma (RT4) cell lines were generated to provide regulatable schwannomin expression. The expression levels of 29 samples from five different mouse embryonic fibroblast clonal cell lines and 18 samples from 3 RT4 cell lines were monitored with oligonucleotide microarrays. Using hierarchical clustering, we determined that the changes in gene expression induced by schwannomin overexpression were subtle when compared with those detected as a consequence of clonal selection during generation of the cell lines. The hierarchical clustering implies that significant alterations of gene expression were introduced during the transfection and selection processes. A total of 28 genes were identified by Kruskal-Wallis rank test that showed significant variation between clonal lines. Most of them were related to cytoskeletal function and signaling pathways. Based on these analyses, we recommend that replications of experiments with several selected cell lines are necessary to assess biological effects of induced gene expression.


Asunto(s)
Fibroblastos/metabolismo , Perfilación de la Expresión Génica , Heterogeneidad Genética , Neurilemoma/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos/estadística & datos numéricos , Proyectos de Investigación/tendencias , Animales , Línea Celular , Análisis por Conglomerados , Citoesqueleto/metabolismo , Regulación de la Expresión Génica , Ratones , Ratas , Reproducibilidad de los Resultados , Transducción de Señal , Transfección
20.
Hum Mol Genet ; 11(17): 1911-26, 2002 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-12165554

RESUMEN

Previous analyses of gene expression in a mouse model of Huntington's disease (R6/2) indicated that an N-terminal fragment of mutant huntingtin causes downregulation of striatal signaling genes and particularly those normally induced by cAMP and retinoic acid. The present study expands the regional and temporal scope of this previous work by assessing whether similar changes occur in other brain regions affected in Huntington's disease and other polyglutamine diseases and by discerning whether gene expression changes precede the appearance of disease signs. Oligonucleotide microarrays were employed to survey the expression of approximately 11,000 mRNAs in the cerebral cortex, cerebellum and striatum of symptomatic R6/2 mice. The number and nature of gene expression changes were similar among these three regions, influenced as expected by regional differences in baseline gene expression. Time-course studies revealed that mRNA changes could only reliably be detected after 4 weeks of age, coincident with development of early pathologic and behavioral changes in these animals. In addition, we discovered that skeletal muscle is also a target of polyglutamine-related perturbations in gene expression, showing changes in mRNAs that are dysregulated in brain and also muscle-specific mRNAs. The complete dataset is available at www.neumetrix.info.


Asunto(s)
Encéfalo/metabolismo , Enfermedad de Huntington/genética , Músculo Esquelético/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas Nucleares/genética , Péptidos/genética , Animales , Secuencia de Bases , Northern Blotting , Western Blotting , Encéfalo/patología , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica , Humanos , Proteína Huntingtina , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Técnicas para Inmunoenzimas , Masculino , Ratones , Ratones Transgénicos , Datos de Secuencia Molecular , Músculo Esquelético/patología , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Péptidos/metabolismo , ARN Mensajero/metabolismo
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA