RESUMEN
SignificanceThe modulation of growth hormone secretagogue receptor-1a (GHSR1a) signaling is a promising strategy for treating brain conditions of metabolism, aging, and addiction. GHSR1a activation results in pleiotropic physiological outcomes through distinct and pharmacologically separable G protein- and ß-arrestin (ßarr)-dependent signaling pathways. Thus, pathway-selective modulation can enable improved pharmacotherapeutics that can promote therapeutic efficacy while mitigating side effects. Here, we describe the discovery of a brain-penetrant small molecule, N8279 (NCATS-SM8864), that biases GHSR1a conformations toward Gαq activation and reduces aberrant dopaminergic behavior in mice. N8279 represents a promising chemical scaffold to advance the development of better treatments for GHSR1a-related brain disorders involving the pathological dysregulation of dopamine.
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Encéfalo/metabolismo , Dopamina/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/metabolismo , Receptores de Ghrelina/metabolismo , Animales , Dopamina/genética , Subunidades alfa de la Proteína de Unión al GTP Gq-G11/genética , Masculino , Ratones , Ratones Noqueados , Receptores de Ghrelina/genéticaRESUMEN
OBJECTIVE: Ectopic calcification is an important contributor to chronic diseases, such as osteoarthritis. Currently, no effective therapies exist to counteract calcification. We developed peptides derived from the calcium binding domain of human Alpha-2-HS-Glycoprotein (AHSG/Fetuin A) to counteract calcification. METHODS: A library of seven 30 amino acid (AA) long peptides, spanning the 118 AA Cystatin 1 domain of AHSG, were synthesized and evaluated in an in vitro calcium phosphate precipitation assay. The best performing peptide was modified (cyclic, retro-inverso and combinations thereof) and evaluated in cellular calcification models and the rat Medial Collateral Ligament Transection + Medial Meniscal Tear (MCLT + MMT) osteoarthritis model. RESULTS: A cyclic peptide spanning AA 1-30 of mature AHSG showed clear inhibition of calcium phosphate precipitation in the nM-pM range that far exceeded the biological activity of the linear peptide variant or bovine Fetuin. Biochemical and electron microscopy analyses of calcium phosphate particles revealed a similar, but distinct, mode of action in comparison with bFetuin. A cyclic-inverso variant of the AHSG 1-30 peptide inhibited calcification of human articular chondrocytes, vascular smooth muscle cells and during osteogenic differentiation of bone marrow derived stromal cells. Lastly, we evaluated the effect of intra-articular injection of the cyclic-inverso AHSG 1-30 peptide in a rat osteoarthritis model. A significant improvement was found in histopathological osteoarthritis score and animal mobility. Serum levels of IFNγ were found to be lower in AHSG 1-30 peptide treated animals. CONCLUSIONS: The cyclic-inverso AHSG 1-30 peptide directly inhibits the calcification process and holds the potential for future application in osteoarthritis.
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Calcinosis , Osteoartritis , Humanos , Animales , Bovinos , Ratas , alfa-2-Glicoproteína-HS/metabolismo , Péptidos Cíclicos/farmacología , Péptidos Cíclicos/uso terapéutico , Péptidos Cíclicos/metabolismo , Osteogénesis , Osteoartritis/tratamiento farmacológicoRESUMEN
OBJECTIVE: Since the joint microenvironment and tissue homeostasis are highly dependent on synovial fluid, we aimed to compare the essential chondrocyte signaling signatures of non-osteoarthritic vs end-stage osteoarthritic knee synovial fluid. Moreover, we determined the phenotypic consequence of the distinct signaling patterns on articular chondrocytes. METHODS: Protein profiling of synovial fluid was performed using antibody arrays. Chondrocyte signaling and phenotypic changes induced by non-osteoarthritic and osteoarthritic synovial fluid were analyzed using a phospho-kinase array, luciferase-based transcription factor activity assays, and RT-qPCR. The origin of osteoarthritic synovial fluid signaling was evaluated by comparing the signaling responses of conditioned media from cartilage, synovium, infrapatellar fat pad and meniscus. Osteoarthritic synovial fluid induced pathway-phenotype relationships were evaluated using pharmacological inhibitors. RESULTS: Compared to non-osteoarthritic synovial fluid, osteoarthritic synovial fluid was enriched in cytokines, chemokines and growth factors that provoked differential MAPK, AKT, NFκB and cell cycle signaling in chondrocytes. Functional pathway analysis confirmed increased activity of these signaling events upon osteoarthritic synovial fluid stimulation. Tissue secretomes of osteoarthritic cartilage, synovium, infrapatellar fat pad and meniscus activated several inflammatory signaling routes. Furthermore, the distinct pathway signatures of osteoarthritic synovial fluid led to accelerated chondrocyte dedifferentiation via MAPK/ERK signaling, increased chondrocyte fibrosis through MAPK/JNK and PI3K/AKT activation, an elevated inflammatory response mediated by cPKC/NFκB, production of extracellular matrix-degrading enzymes by MAPK/p38 and PI3K/AKT routes, and enabling of chondrocyte proliferation. CONCLUSION: This study provides the first mechanistic comparison between non-osteoarthritic and osteoarthritic synovial fluid, highlighting MAPKs, cPKC/NFκB and PI3K/AKT as crucial OA-associated intracellular signaling routes.
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Cartílago Articular , Condrocitos , Condrocitos/metabolismo , Líquido Sinovial/metabolismo , Cartílago Articular/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Células Cultivadas , FenotipoRESUMEN
OBJECTIVE: Basic Calcium Phosphate (BCP) crystals play an active role in the progression of osteoarthritis (OA). However, the cellular consequences remain largely unknown. Therefore, we characterized for the first time the changes in the protein secretome of human OA articular chondrocytes as a result of BCP stimulation using two unbiased proteomic analysis methods. METHOD: Isolated human OA articular chondrocytes were stimulated with BCP crystals and examined by Quantitative Reverse Transcription PCR (RT-qPCR) and enzyme-linked immune sorbent assay (ELISA) after twenty-four and forty-eight hours. Forty-eight hours conditioned media were analyzed by label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) and an antibody array. The activity of BCP dependent Transforming Growth Factor Beta (TGF-ß) signaling was analyzed by RT-qPCR and luciferase reporter assays. The molecular consequences regarding BCP-dependent TGF-ß signaling on BCP-dependent Interleukin 6 (IL-6) were investigated using specific pathway inhibitors. RESULTS: Synthesized BCP crystals induced IL-6 expression and secretion upon stimulation of human articular chondrocytes. Concomitant induction of catabolic gene expression was observed. Analysis of conditioned media revealed a complex and diverse response with a large number of proteins involved in TGF-ß signaling, both in activation of latent TGF-ß and TGF-ß superfamily members, which were increased compared to non-stimulated OA chondrocytes. Activity of this BCP driven TGF-ß signaling was confirmed by increased activity of expression of TGF-ß target genes and luciferase reporters. Inhibition of BCP driven TGF-ß signaling resulted in decreased IL-6 expression and secretion with a moderate effect on catabolic gene expression. CONCLUSION: BCP crystal stimulation resulted in a complex and diverse chondrocyte protein secretome response. An important role for BCP-dependent TGF-ß signaling was identified in development of a pro-inflammatory environment.
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Condrocitos , Secretoma , Transducción de Señal , Factor de Crecimiento Transformador beta , Humanos , Fosfatos de Calcio/farmacología , Condrocitos/metabolismo , Cromatografía Liquida , Medios de Cultivo Condicionados , Interleucina-6/metabolismo , Osteoartritis/metabolismo , Proteómica , Espectrometría de Masas en Tándem , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
OBJECTIVE: Osteoarthritis-related cartilage extracellular matrix remodeling is dependent on changes in chondrocyte protein expression. Yet, the role of ribosomes in chondrocyte translation regulation is unknown. In this exploratory study, we investigated ribosomal RNA (rRNA) epitranscriptomic-based ribosome heterogeneity in human articular chondrocytes and its relevance for osteoarthritis. METHODS: Sequencing-based rRNA 2'-O-methylation profiling analysis (RiboMethSeq) was performed on non-OA primary human articular chondrocytes (n = 5) exposed for 14 days to osteoarthritic synovial fluid (14 donors, pooled, 20% v/v). The SW1353 SNORD71 KO cell pool was generated using LentiCRISPRv2/Cas9. The mode of translation initiation and fidelity were determined by dual-luciferase reporters. The cellular proteome was analyzed by LC-MS/MS and collagen type I protein expression was evaluated by immunoblotting. Loading of COL1A1 mRNA into polysomes was determined by sucrose gradient ultracentrifugation and fractionation. RESULTS: We discovered that osteoarthritic synovial fluid instigates site-specific changes in the rRNA 2'-O-me profile of primary human articular chondrocytes. We identified five sites with differential 2'-O-me levels. The 2'-O-me status of 5.8S-U14 (one of identified differential 2'-O-me sites; decreased by 7.7%, 95% CI [0.9-14.5%]) was targeted by depleting the level of its guide snoRNA SNORD71 (50% decrease, 95% CI [33-64%]). This resulted in an altered ribosome translation modus (e.g., CrPV IRES, FC 3, 95% CI [2.2-4.1]) and promoted translation of COL1A1 mRNA which led to increased levels of COL1A1 protein (FC 1.7, 95% CI [1.3-2.0]). CONCLUSIONS: Our data identify a novel concept suggesting that articular chondrocytes employ rRNA epitranscriptomic mechanisms in osteoarthritis development.
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Cartílago Articular , Osteoartritis , Humanos , ARN Ribosómico/metabolismo , Condrocitos/metabolismo , Proteoma , Cromatografía Liquida , Espectrometría de Masas en Tándem , Osteoartritis/metabolismo , Cartílago Articular/metabolismo , ARN Mensajero/metabolismo , Células CultivadasRESUMEN
Chronic neuropathic pain is currently a major health issue in U.S. complicated by the lack of non-opioid analgesic alternatives. Our investigations led to the discovery of major signaling pathways involved in the transition of acute to chronic neuropathic pain and the identification of several targets for therapeutic intervention. Our translational approach has facilitated the advancement of novel medicines for chronic neuropathic pain that are in advanced clinical development and clinical trials.
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Dolor Crónico , Neuralgia , Analgésicos Opioides/uso terapéutico , Dolor Crónico/tratamiento farmacológico , Humanos , Neuralgia/tratamiento farmacológicoRESUMEN
STUDY OBJECTIVE: As clinicians look to nonnarcotic analgesics in the emergency department (ED), it is essential to understand the effectiveness and adverse effects of nonopioid medications in comparison with existing opioid treatments. Studies of intravenous acetaminophen for acute pain in the ED demonstrate mixed results and suffer from small sample sizes and methodological limitations. This study compares intravenous hydromorphone with intravenous acetaminophen in adult ED patients presenting with acute pain. METHODS: This was a prospective, randomized, clinical trial comparing 1 g intravenous acetaminophen with 1 mg intravenous hydromorphone for treatment of adults with severe, acute pain in the ED. The primary outcome was between-group difference in change in numeric rating scale from baseline to 60 minutes postadministration of study medication. Secondary outcomes included the difference in proportion of patients in each group who declined additional analgesia at 60 minutes, received additional medication before 60 minutes, and developed nausea, vomiting, or pruritus. RESULTS: Of 220 subjects randomized, 103 patients in each arm had sufficient data for analysis. At 60 minutes, the mean decrease in numeric rating scale pain score was 5.3 in the hydromorphone arm and 3.3 in the acetaminophen arm, a difference of 2.0 (95% confidence interval [CI] 1.2 to 2.7) favoring hydromorphone. A greater proportion of patients in the hydromorphone arm also declined additional analgesia at 60 minutes (65% versus 44%; difference 21%; (95% CI 8% to 35%). There was no difference in the proportion of patients receiving rescue analgesia before 60 minutes. Significantly more subjects in the hydromorphone group developed nausea (19% versus 3%; difference 16%; 95% CI 4% to 28%) and vomiting (14% versus 3%; difference 11%; 95% CI 0% to 23%). CONCLUSION: Although both 1 mg intravenous hydromorphone and 1 g intravenous acetaminophen provided clinically meaningful reductions in pain scores, treatment with hydromorphone provided both clinically and statistically greater analgesia than acetaminophen, at the cost of a higher incidence of nausea and vomiting.
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Acetaminofén/administración & dosificación , Dolor Agudo/tratamiento farmacológico , Analgésicos no Narcóticos/administración & dosificación , Analgésicos Opioides/administración & dosificación , Servicio de Urgencia en Hospital , Hidromorfona/administración & dosificación , Administración Intravenosa , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Dimensión del Dolor , Estudios Prospectivos , Resultado del TratamientoRESUMEN
Background: Temocillin is an old 'revived' antibiotic that may play an important role in the treatment of febrile urinary tract infection (UTI). Data regarding its activity against current Enterobacteriaceae isolates as well as the performance of routine susceptibility testing methods are, however, scarce. Objectives: To determine the MICs of temocillin for Enterobacteriaceae strains reflecting the current epidemiology and to analyse the accuracy of three commercial methods. Methods: Enterobacteriaceae isolates causing community-acquired UTI were prospectively collected from September 2015 to January 2017 in two French centres. Temocillin MIC was determined by agar dilution (AD) as the reference method and then compared with: (i) susceptibility testing by disc diffusion; (ii) MIC determination by Etest; and (iii) MIC estimation by the Vitek 2 automated system. Results: A total of 762 Enterobacteriaceae were analysed comprising 658 (86.4%) Escherichia coli and 37 (4.9%) ESBL-producing isolates. Susceptibility rate assessed by AD was 99.6% according to the 8 mg/L clinical breakpoint and was significantly lower against the ESBL-producing isolates than the non-ESBL-producing isolates (94.6% versus 99.9%, P < 0.01). The MIC50 and MIC90 for the total set were 3 and 6 mg/L, respectively. According to the 8 mg/L clinical breakpoint, the major error rate was <1% for disc diffusion and Etest, and significantly higher for Vitek 2 (4.3%, P < 0.01), but still low. No very major error was noticed. Conclusions: Temocillin showed a high level of activity against Enterobacteriaceae from community-acquired UTI and good to excellent reliability of routine methods for susceptibility testing in such a setting.
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Antibacterianos/farmacología , Infecciones Comunitarias Adquiridas/orina , Infecciones por Enterobacteriaceae/orina , Enterobacteriaceae/efectos de los fármacos , Penicilinas/farmacología , Infecciones Comunitarias Adquiridas/tratamiento farmacológico , Infecciones Comunitarias Adquiridas/microbiología , Pruebas Antimicrobianas de Difusión por Disco , Enterobacteriaceae/aislamiento & purificación , Infecciones por Enterobacteriaceae/tratamiento farmacológico , Infecciones por Enterobacteriaceae/epidemiología , Francia/epidemiología , Hospitales de Enseñanza , Humanos , Pruebas de Sensibilidad Microbiana , Estudios Prospectivos , Reproducibilidad de los ResultadosRESUMEN
OBJECTIVE: The Hoffa's fat pad (HFP) is an intra-articular adipose tissue which is situated under and behind the patella. It contains immune cells next to adipocytes and secretes inflammatory factors during osteoarthritis (OA). In this study, we compared the release profile of prostanoids, which are involved in inflammation, of HFP from OA patients vs patients with a focal cartilage defect (CD) without evidence for OA on MRI and investigated the prostanoid modulatory anti-inflammatory action of celecoxib on HFP. DESIGN: Prostanoid release was analyzed in conditioned medium of HFP explant cultures from 17 osteoarthritic patients and 12 CD patients, in the presence or absence of celecoxib. Furthermore, gene expression of COX enzymes and expression of genes indicative of a pro-inflammatory or anti-inflammatory phenotype of HFP was analyzed. RESULTS: Prostanoid release by HFP from knee OA patients clustered in two subgroups with high and low prostanoid producers. HFP from high prostanoid producers released higher amounts of PGE2, PGF2α and PGD2 compared to HFP from CD patients. PGE2 release by OA HFP was positively associated with expression of genes known to be expressed by M1 macrophages, indicating a role for macrophages. Celecoxib modulated prostanoid release by HFP, and also modulated the inflammation ratio towards a more favorable anti-inflammatory M2 phenotype, most effectively in patients with higher prostanoid release profiles. CONCLUSION: In knee OA patients with inflamed HFP's, celecoxib may exert positive effects in the knee joint via decreasing the release of prostanoids produced by the HFP and by favorably modulating the anti-inflammatory marker expression in HFP.
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Tejido Adiposo/metabolismo , Celecoxib/farmacología , Inflamación/metabolismo , Imagen por Resonancia Magnética/métodos , Osteoartritis de la Rodilla/terapia , Prostaglandinas/metabolismo , Tejido Adiposo/patología , Adulto , Anciano , Antiinflamatorios no Esteroideos/farmacología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Osteoartritis de la Rodilla/diagnóstico , Osteoartritis de la Rodilla/metabolismoRESUMEN
BACKGROUND: Spontaneous intracerebral hemorrhage (ICH) leaves most survivors dependent at follow-up. The importance of promoting M2-like microglial responses is increasingly recognized as a key element to ameliorate brain injury following ICH. The osmotherapeutic agents, mannitol and hypertonic saline (HTS), which are routinely used to reduce intracranial pressure, have been shown to reduce neuroinflammation in experimental ischemic and traumatic brain injury, but anti-inflammatory effects of osmotherapies have not been investigated in ICH. METHODS: We studied the effects of iso-osmotic mannitol and HTS in rat models of ICH utilizing high-dose and moderate-dose collagenase injections into the basal ganglia, associated with high and low mortality, respectively. We studied the effects of osmotherapies, first given 5 h after ICH induction, and then administered every 12 h thereafter (4 doses total). Immunohistochemistry was used to quantify microglial activation and polarization. RESULTS: Compared to controls, mannitol and HTS increased plasma osmolarity 1 h after infusion (301 ± 1.5, 315 ± 4.2 and 310 ± 2.0 mOsm/kg, respectively), reduced mortality at 48 h (82, 36 and 53%, respectively), and reduced hemispheric swelling at 48 h (32, 21, and 17%, respectively). In both perihematomal and contralateral tissues, mannitol and HTS reduced activation of microglia/macrophages (abundance and morphology of Iba1 + cells), and in perihematomal tissues, they reduced markers of the microglia/macrophage M1-like phenotype (nuclear p65, TNF, and NOS2), increased markers of the microglia/macrophage M2-like phenotype (arginase, YM1, and pSTAT3), and reduced infiltration of CD45 + cells. CONCLUSIONS: Repeated dosing of osmotherapeutics at regular intervals may be a useful adjunct to reduce neuroinflammation following ICH.
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Edema Encefálico/tratamiento farmacológico , Hemorragia Cerebral/tratamiento farmacológico , Diuréticos Osmóticos/farmacología , Inflamación/tratamiento farmacológico , Macrófagos/efectos de los fármacos , Manitol/farmacología , Microglía/efectos de los fármacos , Solución Salina Hipertónica/farmacología , Animales , Edema Encefálico/etiología , Hemorragia Cerebral/complicaciones , Modelos Animales de Enfermedad , Diuréticos Osmóticos/administración & dosificación , Humanos , Inflamación/etiología , Inflamación/metabolismo , Masculino , Ratas , Ratas Wistar , Solución Salina Hipertónica/administración & dosificaciónRESUMEN
Many studies have reported on the effects of cyclooxygenase-2 (COX-2) inhibition on osteogenesis. However, far less is known about the effects of COX-2 inhibition on chondrogenic differentiation. Previous studies conducted by our group show that COX-2 inhibition influences in vitro chondrogenic differentiation. Importantly, this might have consequences on endochondral ossification processes occurring in vivo, such as bone fracture healing, growth plate development and ectopic generation of cartilage. The goal of our study was to investigate, in vivo, the effect of COX-2 inhibition by celecoxib on the cartilaginous phase of three different endochondral ossification scenarios. 10 mg/kg/day celecoxib or placebo were orally administered for 25 d to skeletally-immature New Zealand White rabbits (n = 6 per group). Endochondral ossification during fracture healing of a non-critical size defect in the ulna, femoral growth plate and ectopically-induced cartilaginous tissue were examined by radiography, micro-computed tomography (µ-CT), histology and gene expression analysis. Celecoxib treatment resulted in delayed bone fracture healing, alterations in growth plate development and progression of mineralisation. In addition, chondrogenic differentiation of ectopically-induced cartilaginous tissue was severely impaired by celecoxib. In conclusion, we found that celecoxib impaired the chondrogenic phase of endochondral ossification.
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Celecoxib/farmacología , Condrogénesis/efectos de los fármacos , Inhibidores de la Ciclooxigenasa 2/farmacología , Osteogénesis/efectos de los fármacos , Administración Oral , Animales , Huesos/diagnóstico por imagen , Huesos/efectos de los fármacos , Huesos/metabolismo , Cartílago/diagnóstico por imagen , Cartílago/efectos de los fármacos , Cartílago/metabolismo , Celecoxib/administración & dosificación , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Condrocitos/efectos de los fármacos , Condrocitos/metabolismo , Condrogénesis/fisiología , Inhibidores de la Ciclooxigenasa 2/administración & dosificación , Dinoprostona/metabolismo , Curación de Fractura/efectos de los fármacos , Expresión Génica/efectos de los fármacos , Osteogénesis/fisiología , Conejos , Microtomografía por Rayos XRESUMEN
Disruptions in circadian rhythms and dopaminergic activity are involved in the pathophysiology of bipolar disorder, though their interaction remains unclear. Moreover, a lack of animal models that display spontaneous cycling between mood states has hindered our mechanistic understanding of mood switching. Here, we find that mice with a mutation in the circadian Clock gene (ClockΔ19) exhibit rapid mood-cycling, with a profound manic-like phenotype emerging during the day following a period of euthymia at night. Mood-cycling coincides with abnormal daytime spikes in ventral tegmental area (VTA) dopaminergic activity, tyrosine hydroxylase (TH) levels and dopamine synthesis. To determine the significance of daytime increases in VTA dopamine activity to manic behaviors, we developed a novel optogenetic stimulation paradigm that produces a sustained increase in dopamine neuronal activity and find that this induces a manic-like behavioral state. Time-dependent dampening of TH activity during the day reverses manic-related behaviors in ClockΔ19 mice. Finally, we show that CLOCK acts as a negative regulator of TH transcription, revealing a novel molecular mechanism underlying cyclic changes in mood-related behavior. Taken together, these studies have identified a mechanistic connection between circadian gene disruption and the precipitation of manic episodes in bipolar disorder.
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Potenciales de Acción/genética , Afecto/fisiología , Proteínas CLOCK/genética , Ritmo Circadiano/genética , Neuronas Dopaminérgicas/fisiología , Mutación/genética , Potenciales de Acción/efectos de los fármacos , Adaptación Ocular/efectos de los fármacos , Adaptación Ocular/genética , Animales , Línea Celular Transformada , Dopaminérgicos/farmacología , Neuronas Dopaminérgicas/efectos de los fármacos , Preferencias Alimentarias/efectos de los fármacos , Preferencias Alimentarias/fisiología , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Ratas , Natación , Factores de Tiempo , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismo , Área Tegmental Ventral/citologíaRESUMEN
OBJECTIVES: Studies of public health reporting have only examined multiple episodes of the same communicable disease within an individual. We aimed to characterize Montreal residents with multiple reportable disease episodes from 1990 to 2012, while accounting for all types of reportable diseases. STUDY DESIGN: Retrospective cohort study. METHODS: We performed an exploratory analysis using descriptive statistics, contingency tables, and logistic regression. RESULTS: There were 157,839 individuals with at least one disease report and a total of 179,455 disease reports. The 9.8% of subjects with more than one episode accounted for 20.7% of all reported episodes. Among subjects with four or fewer episodes, 54.0% were women, while 74.3% of subjects with five or more episodes were men. Subjects with multiple episodes were more likely to be reported for sexually transmitted infections than were persons with a single episode [difference of proportions: 10.4% (95% CI: 10.0%-10.9%)] and to reside in the neighbourhood encompassing Montreal's gay village. CONCLUSIONS: Individuals with multiple communicable disease reports place a large burden on public health officials. These results may help guide investigation and prevention efforts to reduce the number of excess episodes.
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Enfermedades Transmisibles/epidemiología , Notificación Obligatoria , Adolescente , Adulto , Distribución por Edad , Anciano , Anciano de 80 o más Años , Canadá/epidemiología , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Distribución por Sexo , Enfermedades de Transmisión Sexual/epidemiología , Adulto JovenRESUMEN
The dopamine transporter is a key protein responsible for regulating dopamine homeostasis. Its function is to transport dopamine from the extracellular space into the presynaptic neuron. Studies have suggested that accumulation of dopamine in the cytosol can trigger oxidative stress and neurotoxicity. Previously, ectopic expression of the dopamine transporter was shown to cause damage in non-dopaminergic neurons due to their inability to handle cytosolic dopamine. However, it is unknown whether increasing dopamine transporter activity will be detrimental to dopamine neurons that are inherently capable of storing and degrading dopamine. To address this issue, we characterized transgenic mice that over-express the dopamine transporter selectively in dopamine neurons. We report that dopamine transporter over-expressing (DAT-tg) mice display spontaneous loss of midbrain dopamine neurons that is accompanied by increases in oxidative stress markers, 5-S-cysteinyl-dopamine and 5-S-cysteinyl-DOPAC. In addition, metabolite-to-dopamine ratios are increased and VMAT2 protein expression is decreased in the striatum of these animals. Furthermore, DAT-tg mice also show fine motor deficits on challenging beam traversal that are reversed with l-DOPA treatment. Collectively, our findings demonstrate that even in neurons that routinely handle dopamine, increased uptake of this neurotransmitter through the dopamine transporter results in oxidative damage, neuronal loss and l-DOPA reversible motor deficits. In addition, DAT over-expressing animals are highly sensitive to MPTP-induced neurotoxicity. The effects of increased dopamine uptake in these transgenic mice could shed light on the unique vulnerability of dopamine neurons in Parkinson's disease.
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Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Neuronas Dopaminérgicas/fisiología , Mesencéfalo/fisiopatología , Trastornos del Movimiento/fisiopatología , Estrés Oxidativo/fisiología , Ácido 3,4-Dihidroxifenilacético/metabolismo , Animales , Antidiscinéticos/farmacología , Muerte Celular/fisiología , Citosol/efectos de los fármacos , Citosol/metabolismo , Dopamina/metabolismo , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/genética , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/patología , Levodopa/farmacología , Mesencéfalo/efectos de los fármacos , Mesencéfalo/patología , Ratones Endogámicos C57BL , Ratones Transgénicos , Destreza Motora/efectos de los fármacos , Destreza Motora/fisiología , Trastornos del Movimiento/tratamiento farmacológico , Trastornos del Movimiento/patología , Trastornos Parkinsonianos/fisiopatología , Proteínas de Transporte Vesicular de Monoaminas/metabolismoRESUMEN
OBJECTIVES: Patient reported outcomes (PROs) are relevant in rheumatology. Variable accessibility and validity of commonly used PROs are obstacles to homogeneity in evidence synthesis. The objective of this project was to provide a comprehensive library of "validated PROs". METHODS: A launch meeting with rheumatologists, PROs methodological experts, and patients, was held to define the library's aims and scope, and basic requirements. To feed the library we performed systematic reviews on selected diseases and domains. Relevant information on PROs was collected using standardised data collection forms based on the COSMIN checklist. RESULTS: The EULAR Outcomes Measures Library (OML), whose aims are to provide and to advise on PROs on a user-friendly manner albeit based on scientific grounds, has been launched and made accessible to all. PROs currently included cover any domain and, are generic or specifically target to the following diseases: rheumatoid arthritis, osteoarthritis, spondyloarthritis, low back pain, systemic lupus erythematosus, gout, osteoporosis, juvenile idiopathic arthritis, and fibromyalgia. Up to 236 instruments (106 generic and 130 specific) have been identified, evaluated, and included. The systematic review for SLE, which yielded 10 specific instruments, is presented here as an example. The OML website includes, for each PRO, information on the construct being measured and the extent of validation, recommendations for use, and available versions; it also contains a glossary on common validation terms. CONCLUSIONS: The OML is an in progress library led by rheumatologists, related professionals and patients, that will help to better understand and apply PROs in rheumatic and musculoskeletal diseases.
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Lupus Eritematoso Sistémico , Evaluación de Procesos y Resultados en Atención de Salud/normas , Manejo de Atención al Paciente , Práctica Clínica Basada en la Evidencia , Humanos , Desarrollo de la Colección de Bibliotecas , Lupus Eritematoso Sistémico/diagnóstico , Lupus Eritematoso Sistémico/terapia , Manejo de Atención al Paciente/métodos , Manejo de Atención al Paciente/normas , Reproducibilidad de los ResultadosRESUMEN
CONTEXT: Subarachnoid hemorrhage (SAH) has a high fatality rate and many suffer from delayed neurological deficits. Biomarkers may aid in the identification of high-risk patients, guide treatment/management and improve outcome. OBJECTIVE: The aim of this review was to summarize biomarkers of SAH associated with outcome. METHODS: An electronic database query was completed, including an additional review of reference lists to include all potential human studies. RESULTS: A total of 298 articles were identified; 112 were reviewed; 55 studies were included. CONCLUSION: This review details biomarkers of SAH that correlate with outcome. It provides the basis for research investigating their possible translation into the management of SAH patients.
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Hemorragia Subaracnoidea/sangre , Animales , Biomarcadores/sangre , Humanos , Pronóstico , Hemorragia Subaracnoidea/diagnóstico , Hemorragia Subaracnoidea/terapia , Resultado del TratamientoRESUMEN
Impaired transmission through glutamatergic circuits has been postulated to play a role in the underlying pathophysiology of schizophrenia. Furthermore, inhibition of the N-methyl-d-aspartate (NMDA) subtype of ionotropic glutamate receptors (NMDAR) induces a syndrome that recapitulates many of the symptoms observed in patients with schizophrenia. Selective activation of metabotropic glutamate receptor subtype 5 (mGlu5) may provide a novel therapeutic approach for treatment of symptoms associated with schizophrenia through facilitation of transmission through central glutamatergic circuits. Here, we describe the characterization of two novel N-aryl piperazine mGlu5 positive allosteric modulators (PAMs): 2-(4-(2-(benzyloxy)acetyl)piperazin-1-yl)benzonitrile (VU0364289) and 1-(4-(2,4-difluorophenyl)piperazin-1-yl)-2-((4-fluorobenzyl)oxy)ethanone (DPFE). VU0364289 and DPFE induced robust leftward shifts in the glutamate concentration-response curves for Ca(2+) mobilization and extracellular signal-regulated kinases 1 and 2 phosphorylation. Both PAMs displayed micromolar affinity for the common mGlu5 allosteric binding site and high selectivity for mGlu5. VU0364289 and DPFE possessed suitable pharmacokinetic properties for dosing in vivo and produced robust dose-related effects in reversing amphetamine-induced hyperlocomotion, a preclinical model predictive of antipsychotic-like activity. In addition, DPFE enhanced acquisition of contextual fear conditioning in rats and reversed behavioral deficits in a mouse model of NMDAR hypofunction. In contrast, DPFE had no effect on reversing apomorphine-induced disruptions of prepulse inhibition of the acoustic startle reflex. These mGlu5 PAMs also increased monoamine levels in the prefrontal cortex, enhanced performance in a hippocampal-mediated memory task, and elicited changes in electroencephalogram dynamics commensurate with procognitive effects. Collectively, these data support and extend the role for the development of novel mGlu5 PAMs for the treatment of psychosis and cognitive deficits observed in individuals with schizophrenia.
Asunto(s)
Antipsicóticos/farmacología , Hipercinesia/tratamiento farmacológico , Memoria a Corto Plazo/efectos de los fármacos , Nootrópicos/farmacología , Piperazinas/farmacología , Receptor del Glutamato Metabotropico 5/agonistas , Receptores de N-Metil-D-Aspartato/metabolismo , Regulación Alostérica , Animales , Antipsicóticos/química , Antipsicóticos/farmacocinética , Antipsicóticos/uso terapéutico , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Células HEK293 , Humanos , Hipercinesia/metabolismo , Hipercinesia/psicología , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Ratones , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Nootrópicos/química , Nootrópicos/farmacocinética , Nootrópicos/uso terapéutico , Piperazinas/química , Piperazinas/farmacocinética , Piperazinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Receptor del Glutamato Metabotropico 5/genética , Receptores de N-Metil-D-Aspartato/genética , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/metabolismo , TransfecciónRESUMEN
OBJECTIVE: Bone morphogenic protein (BMP)-2 and BMP-7 are clinically approved and their recombinant proteins are used for bone tissue regenerative purposes and widely evaluated for cartilage regeneration. Previous comparison of the in vitro chondrogenic characteristics of BMP-2 vs BMP-7 did not address hypertrophic differentiation and characterizing their chondrogenic properties with a focus in on chondrocyte hypertrophy was topic of investigation in this study. DESIGN: Equimolar concentrations of BMP-2 or BMP-7 were added to chondrogenic differentiating ATDC5, human bone marrow stem cells or rabbit periosteal explants. Expression of Col2a1, Sox9, Acan, Col10a1, Runx2, ALP, Mmp13, Mef2c and Bapx1/Nkx3.2 was determined by reverse transcription-quantitative PCR (RT-qPCR) and immunoblotting. Glycosaminoglycan content, cell proliferation capacity and ALP activity were analysed by colourimetric analyses. Expression of Bapx1/Nkx3.2 and Sox9 was targeted by transfection of target specific siRNA duplexes. RESULTS: BMP-2 dose-dependently increased chondrocyte hypertrophy during chondrogenic differentiation of progenitor cells, whereas BMP-7 acted hypertrophy-suppressive and chondro-promotive. Both BMPs did not influence cell proliferation, but they did increase total glycosaminoglycan content. In a candidate approach Bapx1/Nkx3.2 was found to be involved in the BMP-7 mediated suppression of chondrocyte hypertrophy in ATDC5 cells. CONCLUSIONS: BMP-2 and BMP-7 display opposing actions on the chondrogenic outcome of differentiating progenitor cells: BMP-2 acts a specific inducer of chondrocyte hypertrophy, while BMP-7 appears to increase or maintain chondrogenic potential and prevent chondrocyte hypertrophy. Our results pave the way for an application-dependent differential use of BMP-2 or BMP-7.
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Proteína Morfogenética Ósea 2/farmacología , Proteína Morfogenética Ósea 7/farmacología , Condrocitos/efectos de los fármacos , Células Madre/efectos de los fármacos , Animales , Células de la Médula Ósea/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Aumento de la Célula/efectos de los fármacos , Células Cultivadas , Condrocitos/citología , Condrocitos/patología , Condrogénesis/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Proteínas de Homeodominio/fisiología , Humanos , Hipertrofia , Conejos , Células Madre/citología , Factores de Transcripción/fisiologíaRESUMEN
Probably the foremost hypothesis of depression is the 5-hydroxytryptamine (5-HT, serotonin) deficiency hypothesis. Accordingly, anomalies in putative 5-HT biomarkers have repeatedly been reported in depression patients. However, whether such anomalies in fact reflect deficient central 5-HT neurotransmission remains unresolved. We employed a naturalistic model of 5-HT deficiency, the tryptophan hydroxylase 2 (Tph2) R439H knockin mouse, to address this question. We report that Tph2 knockin mice have reduced basal and stimulated levels of extracellular 5-HT (5-HT(Ext)). Interestingly, cerebrospinal fluid (CSF) 5-hydroxyindoleacetic acid (5-HIAA) and fenfluramine-induced plasma prolactin levels are markedly diminished in the Tph2 knockin mice. These data seemingly confirm that low CSF 5-HIAA and fenfluramine-induced plasma prolactin reflects chronic, endogenous central nervous system (CNS) 5-HT deficiency. Moreover, 5-HT(1A) receptor agonist-induced hypothermia is blunted and frontal cortex 5-HT(2A) receptors are increased in the Tph2 knockin mice. These data likewise parallel core findings in depression, but are usually attributed to anomalies in the respective receptors rather than resulting from CNS 5-HT deficiency. Further, 5-HT(2A) receptor function is enhanced in the Tph2 knockin mice. In contrast, 5-HT(1A) receptor levels and G-protein coupling is normal in Tph2 knockin mice, indicating that the blunted hypothermic response relates directly to the low 5-HT(Ext). Thus, we show that not only low CSF 5-HIAA and a blunted fenfluramine-induced prolactin response, but also blunted 5-HT(1A) agonist-induced hypothermia and increased 5-HT(2A) receptor levels are bona fide biomarkers of chronic, endogenous 5-HT deficiency. Potentially, some of these biomarkers could identify patients likely to have 5-HT deficiency. This could have clinical research utility or even guide pharmacotherapy.
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Depresión/sangre , Ácido Hidroxiindolacético/líquido cefalorraquídeo , Receptor de Serotonina 5-HT2A/metabolismo , Neuronas Serotoninérgicas/fisiología , Serotonina/deficiencia , Transmisión Sináptica/fisiología , Triptófano Hidroxilasa/fisiología , Animales , Conducta Animal/efectos de los fármacos , Conducta Animal/fisiología , Biomarcadores/sangre , Biomarcadores/líquido cefalorraquídeo , Biomarcadores/metabolismo , Corticosterona/sangre , Depresión/líquido cefalorraquídeo , Depresión/genética , Modelos Animales de Enfermedad , Líquido Extracelular/metabolismo , Femenino , Fenfluramina/farmacología , Lóbulo Frontal/metabolismo , Técnicas de Sustitución del Gen/métodos , Técnicas de Sustitución del Gen/psicología , Hipocampo/metabolismo , Hipotermia/inducido químicamente , Hipotermia/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Prolactina/sangre , Receptor de Serotonina 5-HT1A/genética , Receptor de Serotonina 5-HT1A/metabolismo , Receptor de Serotonina 5-HT2A/genética , Neuronas Serotoninérgicas/efectos de los fármacos , Neuronas Serotoninérgicas/enzimología , Serotonina/metabolismo , Agonistas del Receptor de Serotonina 5-HT1/farmacología , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/genética , Triptófano Hidroxilasa/genéticaRESUMEN
Congenital myoclonus is a widespread neurologic disorder characterized by hyperexcitability, muscular spasticity and myoclonus associated with marked reduction in neural glycine binding sites. The recessive mouse mutation spastic (spa) is a prototype of inherited myoclonus. Here we show that defects in the gene encoding the beta-subunit of the glycine receptor (Glrb) underlie spa: Glrb maps to the same region of mouse chromosome 3 as spa, and Glrb mRNA is markedly reduced throughout brains of spa mice, most likely as a result of an insertional mutation of a 7.1 kilobase LINE-1 element within intron 6 of Glrb. These results provide evidence that Glrb is necessary for postsynaptic expression of glycine receptor complexes, and suggest Glrb as a candidate gene for inherited myoclonus in other species.