RESUMO
BACKGROUND AND OBJECTIVES: Patients undergoing induction/reinduction chemotherapy for haematologic malignancies (HM) are at risk for invasive fungal infections (IFIs). In 2015, Duke University Hospital (DUH) implemented a new standardised fungal prophylaxis protocol for adult patients undergoing induction chemotherapy for acute lymphocytic leukaemia, acute myelocytic leukaemia and myelodysplastic syndrome. This study assessed the impact of protocol implementation on (1) use of antifungal prophylaxis, throughout the at-risk period and (2) patient outcomes such as IFI and mortality. METHODS: Retrospective, observational study of adult HM patients admitted to DUH for induction/reinduction chemotherapy pre- (7/1/2013-12/31/2014) and post- (1/1/2015-10/31/2016) implementation of standardised antifungal prophylaxis protocol (which recommended posaconazole as the first-line agent). Patients were followed for up to 100 days after initiation of induction chemotherapy to evaluate use of antifungal prophylaxis and patient outcomes. RESULTS: 218 patients with haematologic malignancies were included (90 pre, 128 post). Use of antifungal prophylaxis increased from 81.1% (pre) to 97.7% (post) (p < .0001). Overall, 71% received posaconazole as initial antifungal prophylaxis (64.4% pre, 75.7% post). Approximately one-fourth of patients (25.6%, pre vs 26.6%, post) developed an IFI (proven/probable or possible using modified EORTC definitions) (p = .868); 100-day mortality remained stable (18.9% pre vs 18.8% post, respectively, p = .979). Lack of antifungal prophylaxis and older age (≥60 years) were associated with higher risk of IFI. CONCLUSION: Implementation of a standardised protocol with posaconazole as the primary agent was associated with increased use of antifungal prophylaxis among patients undergoing induction/reinduction chemotherapy for haematologic malignancies in our hospital. Lack of antifungal prophylaxis was an independent predictor of IFIs, underscoring the importance of prophylaxis in this at-risk population.
Assuntos
Neoplasias Hematológicas , Leucemia Mieloide Aguda , Adulto , Antifúngicos/uso terapêutico , Neoplasias Hematológicas/complicações , Neoplasias Hematológicas/tratamento farmacológico , Humanos , Leucemia Mieloide Aguda/complicações , Leucemia Mieloide Aguda/tratamento farmacológico , Estudos Retrospectivos , Triazóis/uso terapêuticoRESUMO
Trichodysplasia spinulosa (TS) is a rare skin condition caused by trichodysplasia spinulosa-associated polyomavirus (TSPyV). It affects immunosuppressed patients, and <50 cases have been reported. The majority of these cases are seen in solid organ transplant recipients. TS often poses a diagnostic and therapeutic challenge because of its rarity and resemblance with other skin conditions. Several forms of treatment are usually tried prior to establishing a definitive diagnosis. Oral valganciclovir and topical cidofovir have been found to give the best results and hence are the most commonly used agents once the diagnosis is established. Here, we present two cases with a review of literature of TS in solid organ transplant recipients, focusing on time to develop the condition post-transplant, immunosuppression regimen used, and treatment initiated both before and after a definitive diagnosis.
Assuntos
Doenças do Cabelo , Humanos , Hospedeiro Imunocomprometido , Terapia de Imunossupressão , Polyomavirus , Infecções por PolyomavirusRESUMO
Treatment options for drug-resistant cytomegalovirus (CMV) are limited. Letermovir is a novel antiviral recently approved for CMV prophylaxis following hematopoietic cell transplantation, but its efficacy in other settings is unknown. We recently used letermovir for salvage treatment in four solid organ transplant recipients with ganciclovir-resistant CMV retinitis. All patients improved clinically without known adverse drug events. However, three patients failed to maintain virologic suppression, including two patients who developed genotypically confirmed resistance to letermovir while on therapy.
Assuntos
Acetatos/uso terapêutico , Antivirais/uso terapêutico , Retinite por Citomegalovirus/tratamento farmacológico , Citomegalovirus/efeitos dos fármacos , Quinazolinas/uso terapêutico , Citomegalovirus/genética , Farmacorresistência Viral , Ganciclovir/farmacologia , Humanos , Transplante de Órgãos/efeitos adversos , Terapia de Salvação , TransplantadosRESUMO
African tick bite fever is the most commonly encountered travel-associated rickettsiosis, occurring in as many as 5% of travelers returning from rural subequatorial Africa. This case report illustrates that rifampin represents an effective alternative to doxycycline for treatment of African tick bite fever in some selective situations.
Assuntos
Antibacterianos , Rifampina , Rickettsiose do Grupo da Febre Maculosa , Antibacterianos/administração & dosagem , Antibacterianos/uso terapêutico , Doxiciclina/efeitos adversos , Humanos , Masculino , Pessoa de Meia-Idade , Rickettsia , Rifampina/administração & dosagem , Rifampina/uso terapêutico , Rickettsiose do Grupo da Febre Maculosa/diagnóstico , Rickettsiose do Grupo da Febre Maculosa/tratamento farmacológicoRESUMO
This article evaluates the use of a "canthal cutdown" technique in orbital compartment syndrome in a cadaveric model. Twelve cadaver orbits were used to simulate orbital compartment syndrome using a blood analog solution. Two pressure probes, in different orbital locations, were used to monitor orbital pressure. Pressure was monitored during successive procedures: canthotomy, cantholysis, and canthal cutdown. Orbits were then re-injected with solution, simulating an active orbital hemorrhage, and pressure measurements were recorded over a 10-minute duration. No statistically significant difference was found between the two orbital pressure monitoring devices at each measurement point (p = 0.99). Significant pressure reductions, for both probes, were observed after canthal cutdown compared to initial measurement after injection of 20 mL blood analog (p < 0.001 and p = 0.005). When comparing the orbital pressure following canthotomy and inferior cantholysis versus canthal cutdown, the cutdown procedure provided an additional 74% in orbital pressure reduction (p =0.01). After re-injection of 10 mL of solution and 10 minutes of egress, pressure returned to baseline (probe 1: baseline 7 mm Hg vs. post-cutdown at 10 minutes 7 mm Hg; p = 0.83; and probe 2: 5 mm Hg vs. 5 mm Hg; p = 0.83). The canthal cutdown technique provides further reduction in orbital pressure versus canthotomy and cantholysis alone. The technique may be effective for treatment of static orbital compartment syndrome and temporizing treatment of compartment syndrome from active orbital hemorrhages.
Assuntos
Síndromes Compartimentais/cirurgia , Descompressão Cirúrgica/métodos , Pálpebras/cirurgia , Procedimentos Cirúrgicos Oftalmológicos , Doenças Orbitárias/cirurgia , Cadáver , Síndromes Compartimentais/etiologia , Humanos , Pressão Intraocular , Doenças Orbitárias/etiologia , Hemorragia Retrobulbar/complicações , Hemorragia Retrobulbar/cirurgiaAssuntos
Acanthamoeba , Encéfalo/patologia , Confusão/parasitologia , Cefaleia/parasitologia , Encefalite Infecciosa/parasitologia , Idoso , Encéfalo/diagnóstico por imagem , Evolução Fatal , Transplante de Coração , Humanos , Encefalite Infecciosa/diagnóstico , Imageamento por Ressonância Magnética , Masculino , TransplantadosRESUMO
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.
Assuntos
Tecido Adiposo Marrom/fisiopatologia , Regulação da Temperatura Corporal/genética , Proteínas de Choque Térmico/metabolismo , Doença de Huntington/etiologia , Fatores de Transcrição/metabolismo , Animais , Temperatura Corporal/genética , Células Cultivadas , Modelos Animais de Doenças , Proteínas de Choque Térmico/genética , Doença de Huntington/genética , Doença de Huntington/metabolismo , Camundongos , Camundongos Transgênicos , Mitocôndrias/fisiologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Transdução de Sinais/genética , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
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.
Assuntos
Apoptose , Proteínas Morfogenéticas Ósseas/metabolismo , Neoplasias Encefálicas/metabolismo , Meduloblastoma/metabolismo , Comunicação Parácrina , Retinoides/farmacologia , Fator de Crescimento Transformador beta , Animais , Proteína Morfogenética Óssea 2 , Receptores de Proteínas Morfogenéticas Ósseas , Neoplasias Encefálicas/patologia , Feminino , Perfilação da Expressão Gênica , Humanos , Meduloblastoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Análise de Sequência com Séries de Oligonucleotídeos , Receptores de Fatores de Crescimento/genética , Receptores de Fatores de Crescimento/metabolismo , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
Huntington disease (HD) is fatal in humans within 15-20 years of symptomatic disease. Although late stage HD has been studied extensively, protein expression changes that occur at the early stages of disease and during disease progression have not been reported. In this study, we used a large two-dimensional gel/mass spectrometry-based proteomics approach to investigate HD-induced protein expression alterations and their kinetics at very early stages and during the course of disease. The murine HD model R6/2 was investigated at 2, 4, 6, 8, and 12 weeks of age, corresponding to absence of disease and early, intermediate, and late stage HD. Unexpectedly the most HD stage-specific protein changes (71-100%) as well as a drastic alteration (almost 6% of the proteome) in protein expression occurred already as early as 2 weeks of age. Early changes included mainly the up-regulation of proteins involved in glycolysis/gluconeogenesis and the down-regulation of the actin cytoskeleton. This suggests a period of highly variable protein expression that precedes the onset of HD phenotypes. Although an up-regulation of glycolysis/gluconeogenesis-related protein alterations remained dominant during HD progression, late stage alterations at 12 weeks showed an up-regulation of proteins involved in proteasomal function. The early changes in HD coincide with a peak in protein alteration during normal mouse development at 2 weeks of age that may be responsible for these massive changes. Protein and mRNA data sets showed a large overlap on the level of affected pathways but not single proteins/mRNAs. Our observations suggest that HD is characterized by a highly dynamic disease pathology not represented by linear protein concentration alterations over the course of disease.
Assuntos
Doença de Huntington/metabolismo , Doença de Huntington/patologia , Proteínas/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Desenvolvimento Embrionário , Metabolismo Energético , Feminino , Regulação da Expressão Gênica , Doença de Huntington/genética , Cinética , Masculino , Redes e Vias Metabólicas , Camundongos , Fenótipo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de Tempo , Extratos de TecidosRESUMO
We integrated five sets of proteomics data profiling the constituents of cerebrospinal fluid (CSF) derived from Huntington disease (HD)-affected and -unaffected individuals with genomics data profiling various human and mouse tissues, including the human HD brain. Based on an integrated analysis, we found that brain-specific proteins are 1.8 times more likely to be observed in CSF than in plasma, that brain-specific proteins tend to decrease in HD CSF compared with unaffected CSF, and that 81% of brain-specific proteins have quantitative changes concordant with transcriptional changes identified in different regions of HD brain. The proteins found to increase in HD CSF tend to be liver-associated. These protein changes are consistent with neurodegeneration, microgliosis, and astrocytosis known to occur in HD. We also discuss concordance between laboratories and find that ratios of individual proteins can vary greatly, but the overall trends with respect to brain or liver specificity were consistent. Concordance is highest between the two laboratories observing the largest numbers of proteins.
Assuntos
Encéfalo/metabolismo , Proteínas do Líquido Cefalorraquidiano/metabolismo , Doença de Huntington/líquido cefalorraquidiano , Animais , Proteínas do Líquido Cefalorraquidiano/genética , Perfilação da Expressão Gênica , Humanos , Laboratórios , Camundongos , Especificidade de Órgãos , ProteômicaAssuntos
Síndromes Compartimentais/diagnóstico , Técnicas de Diagnóstico Oftalmológico/instrumentação , Órbita , Doenças Orbitárias/diagnóstico , Pressão , Adulto , Exoftalmia/diagnóstico por imagem , Traumatismos Oculares/etiologia , Traumatismos Oculares/cirurgia , Humanos , Pressão Intraocular , Masculino , Manometria/instrumentação , Fraturas Orbitárias/etiologia , Fraturas Orbitárias/cirurgia , Tomografia Computadorizada por Raios X , Ferimentos por Arma de Fogo/etiologia , Ferimentos por Arma de Fogo/cirurgiaRESUMO
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.
Assuntos
Encéfalo/metabolismo , Evolução Molecular , Expressão Gênica , Adulto , Idoso , Animais , Sequência Conservada , Feminino , Perfilação da Expressão Gênica , Variação Genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Especificidade de Órgãos , Distribuição TecidualRESUMO
Huntington's disease (HD) is a fatal neurodegenerative condition caused by expansion of the polyglutamine tract in the huntingtin (Htt) protein. Neuronal toxicity in HD is thought to be, at least in part, a consequence of protein interactions involving mutant Htt. We therefore hypothesized that genetic modifiers of HD neurodegeneration should be enriched among Htt protein interactors. To test this idea, we identified a comprehensive set of Htt interactors using two complementary approaches: high-throughput yeast two-hybrid screening and affinity pull down followed by mass spectrometry. This effort led to the identification of 234 high-confidence Htt-associated proteins, 104 of which were found with the yeast method and 130 with the pull downs. We then tested an arbitrary set of 60 genes encoding interacting proteins for their ability to behave as genetic modifiers of neurodegeneration in a Drosophila model of HD. This high-content validation assay showed that 27 of 60 orthologs tested were high-confidence genetic modifiers, as modification was observed with more than one allele. The 45% hit rate for genetic modifiers seen among the interactors is an order of magnitude higher than the 1%-4% typically observed in unbiased genetic screens. Genetic modifiers were similarly represented among proteins discovered using yeast two-hybrid and pull-down/mass spectrometry methods, supporting the notion that these complementary technologies are equally useful in identifying biologically relevant proteins. Interacting proteins confirmed as modifiers of the neurodegeneration phenotype represent a diverse array of biological functions, including synaptic transmission, cytoskeletal organization, signal transduction, and transcription. Among the modifiers were 17 loss-of-function suppressors of neurodegeneration, which can be considered potential targets for therapeutic intervention. Finally, we show that seven interacting proteins from among 11 tested were able to co-immunoprecipitate with full-length Htt from mouse brain. These studies demonstrate that high-throughput screening for protein interactions combined with genetic validation in a model organism is a powerful approach for identifying novel candidate modifiers of polyglutamine toxicity.
Assuntos
Degeneração Neural/genética , Degeneração Neural/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/metabolismo , Animais , Drosophila melanogaster/efeitos dos fármacos , Humanos , Proteína Huntingtina , Imunoprecipitação , Camundongos , Modelos Neurológicos , Peptídeos/toxicidade , Ligação Proteica , Mapeamento de Interação de Proteínas , Reprodutibilidade dos TestesRESUMO
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.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Doenças do Sistema Nervoso Central/tratamento farmacológico , Sistemas de Liberação de Medicamentos , Peptídeos/farmacologia , Animais , Antígenos CD/química , Antígenos CD/efeitos dos fármacos , Antígenos CD/genética , Antígenos CD/farmacologia , Sistema Nervoso Central/efeitos dos fármacos , Cistina/química , Cistina/genética , Humanos , Inflamação/tratamento farmacológico , Inflamação/patologia , Camundongos , Neuropeptídeos/química , Neuropeptídeos/farmacologia , Neurotensina/química , Neurotensina/farmacologia , Peptídeos/química , Ligação Proteica/efeitos dos fármacos , Receptores da Transferrina/química , Receptores da Transferrina/efeitos dos fármacos , Receptores da Transferrina/genéticaRESUMO
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.
Assuntos
Artrite Experimental , Corticosteroides , Animais , Artrite Experimental/tratamento farmacológico , Cartilagem , Humanos , Peptídeos , Ratos , EsteroidesRESUMO
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.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/deficiência , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Perfilação da Expressão Gênica/métodos , Doença de Huntington/metabolismo , Degeneração Neural/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/genética , Corpo Estriado/patologia , Humanos , Doença de Huntington/genética , Doença de Huntington/patologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Degeneração Neural/genética , Degeneração Neural/patologia , Ratos , Ratos Endogâmicos LewRESUMO
Small sensors are useful for in vivo measurements and probing small spaces. In this paper, we compare two methods of fabrication of small, cylindrical carbon-fiber microelectrodes: flame-etching and electrochemical etching. With both methods, microelectrodes can be fabricated with tip diameters of 1 to 3 microm. Electrodes were tested with fast-scan cyclic voltammetry. Flame etching resulted in electrodes that have larger S/N ratios and higher currents per unit area for 1 microM dopamine than normal carbon-fiber microelectrodes or electrochemically etched electrodes. Therefore, the increased sensitivity is not just a property of size. The flame-etched surfaces had nanometer-scale surface features that were not observed on the other electrodes and exhibited increased sensitivity for other electroactive compounds found in the brain, including ascorbic acid, DOPAC, and serotonin. Faster kinetics and a faster response to a step change in dopamine were also observed, when the applied waveform was -0.4 to 1.0 V and back at 400 V/s. The sensitivity of the flame-etched electrodes was enhanced by overoxidizing the surface. The flame-etched electrodes were used to detect dopamine release in anesthetized rats after a single stimulation pulse. The small flame-etched electrodes will facilitate measurements of low concentrations in discrete brain regions or small organisms.
Assuntos
Carbono , Microeletrodos , Análise de Injeção de Fluxo , Cinética , Microscopia Eletrônica de Varredura , Neurotransmissores/análise , Oxirredução , Sensibilidade e EspecificidadeRESUMO
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.
Assuntos
Bancos de Espécimes Biológicos , Neoplasias Encefálicas/patologia , Imuno-Histoquímica , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Animais , Linhagem Celular Tumoral , Criança , Pré-Escolar , Metilação de DNA/genética , Feminino , Genômica , Humanos , Masculino , Camundongos , Mutação , PediatriaRESUMO
The expansion of a polyglutamine tract in the ubiquitously expressed huntingtin protein causes Huntington's disease (HD), a dominantly inherited neurodegenerative disease. We show that the activity of the cholesterol biosynthetic pathway is altered in HD. In particular, the transcription of key genes of the cholesterol biosynthetic pathway is severely affected in vivo in brain tissue from HD mice and in human postmortem striatal and cortical tissue; this molecular dysfunction is biologically relevant because cholesterol biosynthesis is reduced in cultured human HD cells, and total cholesterol mass is significantly decreased in the CNS of HD mice and in brain-derived ST14A cells in which the expression of mutant huntingtin has been turned on. The transcription of the genes of the cholesterol biosynthetic pathway is regulated via the activity of sterol regulatory element-binding proteins (SREBPs), and we found an approximately 50% reduction in the amount of the active nuclear form of SREBP in HD cells and mouse brain tissue. As a consequence, mutant huntingtin reduces the transactivation of an SRE-luciferase construct even under conditions of SREBP overexpression or in the presence of an exogenous N-terminal active form of SREBP. Finally, the addition of exogenous cholesterol to striatal neurons expressing mutant huntingtin prevents their death in a dose-dependent manner. We conclude that the cholesterol biosynthetic pathway is impaired in HD cells, mice, and human subjects, and that the search for HD therapies should also consider cholesterol levels as both a potential target and disease biomarker.