RESUMEN

BACKGROUND AND PURPOSE: The transient receptor potential (TRP) ion channel TRPM3 functions as a noxious heat sensor, plays a key role in acute pain sensation and inflammatory hyperalgesia in rodents. Despite its potential as a novel analgesic drug target, little is known about the expression, function and modulation in the humans. EXPERIMENTAL APPROACH: We studied TRPM3 in freshly isolated human dorsal root ganglion (hDRG) neurons and human stem cell-derived sensory (hSCDS) neurons. Expression was analysed at the mRNA level using RT-qPCR. Channel function was assessed using Fura-2-based calcium imaging and whole-cell patch-clamp recordings. KEY RESULTS: TRPM3 was detected at the mRNA level in both hDRG and hSCDS neurons. The TRPM3 agonists pregnenolone sulphate (PS) and CIM0216 evoked robust intracellular Ca2+ responses in 52% of hDRG and 58% of hSCDS neurons. Whole-cell patch-clamp recordings in hSCDS neurons revealed pregnenolone sulphate (PS)- and CIM0216-evoked currents exhibiting the characteristic current-voltage relation of TRPM3. PS-induced calcium responses in hSCDS neurons were reversed in a dose-dependent manner by the flavonoid isosakuranetin and by antiseizure drug primidone. Finally, the µ-opioid receptor agonist DAMGO and the GABAB receptor agonist baclofen inhibited PS-evoked TRPM3 responses in a subset of hSCDS neurons. CONCLUSION AND IMPLICATIONS: These results provide the first direct evidence of functional expression of the pain receptor TRPM3 in human sensory neurons, largely mirroring the channel's properties observed in mouse sensory neurons. hSCDS neurons represent a valuable and readily accessible in vitro model to study TRPM3 regulation and pharmacology in a relevant human cellular context.

Asunto(s)

Canales Catiónicos TRPM , Animales , Ganglios Espinales , Humanos , Hiperalgesia , Ratones , Técnicas de Placa-Clamp , Células Receptoras Sensoriales

RESUMEN

Successful pregnancy requires the establishment of a complex dialogue between the implanting embryo and the endometrium. Knowledge regarding molecular candidates involved in this early communication process is inadequate due to limited access to primary human endometrial epithelial cells (EEC). Since pseudo-pregnancy in rodents can be induced by mechanical scratching of an appropriately primed uterus, this study aimed to investigate the expression of mechanosensitive ion channels in EEC. Poking of EEC provoked a robust calcium influx and induced an increase in current densities, which could be blocked by an inhibitor of mechanosensitive ion channels. Interestingly, RNA expression studies showed high expression of PIEZO1 in EEC of mouse and human. Additional analysis provided further evidence for the functional expression of PIEZO1 since stimulation with Yoda1, a chemical agonist of PIEZO1, induced increases in intracellular calcium concentrations and current densities in EEC. Moreover, the ion channel profile of human endometrial organoids (EMO) was validated as a representative model for endometrial epithelial cells. Mechanical and chemical stimulation of EMO induced strong calcium responses supporting the hypothesis of mechanosensitive ion channel expression in endometrial epithelial cells. In conclusion, EEC and EMO functionally express the mechanosensitive PIEZO1 channel that could act as a potential target for the development of novel treatments to further improve successful implantation processes.

Asunto(s)

Endometrio/metabolismo , Canales Iónicos/metabolismo , Organoides/metabolismo , Animales , Endometrio/citología , Células Epiteliales/metabolismo , Femenino , Humanos , Ratones

RESUMEN

The cation channel TRPM8 plays a central role in the somatosensory system, as a key sensor of innocuously cold temperatures and cooling agents. Although increased functional expression of TRPM8 has been implicated in various forms of pathological cold hypersensitivity, little is known about the cellular and molecular mechanisms that determine TRPM8 abundance at the plasma membrane. Here we demonstrate constitutive transport of TRPM8 towards the plasma membrane in atypical, non-acidic transport vesicles that contain lysosomal-associated membrane protein 1 (LAMP1), and provide evidence that vesicle-associated membrane protein 7 (VAMP7) mediates fusion of these vesicles with the plasma membrane. In line herewith, VAMP7-deficient mice exhibit reduced functional expression of TRPM8 in sensory neurons and concomitant deficits in cold avoidance and icilin-induced cold hypersensitivity. Our results uncover a cellular pathway that controls functional plasma membrane incorporation of a temperature-sensitive TRP channel, and thus regulates thermosensitivity in vivo.

Asunto(s)

Membrana Celular/metabolismo , Frío , Hiperestesia/genética , Proteínas R-SNARE/genética , Células Receptoras Sensoriales/metabolismo , Canales Catiónicos TRPM/metabolismo , Vesículas Transportadoras/metabolismo , Animales , Calcio/metabolismo , Femenino , Ganglios Espinales/metabolismo , Células HEK293 , Humanos , Hiperestesia/inducido químicamente , Hiperestesia/metabolismo , Proteínas de Membrana de los Lisosomas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microscopía Fluorescente , Técnicas de Placa-Clamp , Pirimidinonas/toxicidad , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ganglio del Trigémino/metabolismo

RESUMEN

PURPOSE: To examine the reliability, validity, and sensitivity to change of the 20-item version and the Rasch-refined 15-item version of the Upper Extremity Functional Index (UEFI-20 and UEFI-15, respectively) and to determine the impact of arm dominance on the positive minimal clinically important difference (pMCID). METHODS: Adults with upper-extremity (UE) dysfunction completed the UEFI-20, Upper Extremity Functional Scale (UEFS), Pain Limitation Scale, and Pain Intensity Scale at their initial physiotherapy assessment (Time 1); 24-48 hours later (Time 2); and 3 weeks into treatment or at discharge, whichever came first (Time 3). Demographics, including working status, were obtained at Time 1. Global ratings of change (GRC) were provided by the treating physiotherapist and patient at Time 3. The UEFI-15 was calculated from relevant items in the UEFI-20. The intra-class correlation coefficient (ICC) and minimal detectable change (MDC) quantified test-retest reliability (Time 1-Time 2). Cross-sectional convergent validity was determined by the association (Pearson's r) between Time 1 measures of function and pain. Known-groups validity was evaluated with a one-way ANOVA across three levels of working status. Longitudinal validity was determined by the association (Pearson's r) between function and pain change scores (Time 1-Time 3). Receiver operating characteristic (ROC) curves estimated the pMCID using Time 1-Time 3 change scores and average patient/therapist GRC. RESULTS: Reliability for the UEFI-20 and UEFI-15 was the same (ICC=0.94 for both measures). MDC values were 9.4/80 for the UEFI-20 and 8.8/100 for the UEFI-15. Cross-sectional, known-groups, and longitudinal validity were confirmed for both UEFI measures. pMCID values were 8/80 for the UEFI-20 and 6.7/100 for the UEFI-15; pMCID was higher for people whose non-dominant arm was affected. CONCLUSIONS: Both UEFI measures show acceptable reliability and validity. Arm dominance affects pMCID. The UEFI-15 is recommended because it measures only one dimension: UE function.

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Objectif : Étudier la fiabilité, la validité et la sensibilité au changement des versions à 20 questions et à 15 questions raffinées par Rasch de l'Indice fonctionnel des membres supérieurs (IFMS-20 et IFMS-15, respectivement) et déterminer l'effet du bras dominant sur la différence minimale positive cliniquement importante (pDMCI). Méthodes : Les adultes ayant une dysfonction des membres supérieurs (MS) ont répondu au questionnaire IFMS-20, aux questionnaires de l'Échelle fonctionnelle des membres supérieurs (EFMS), de l'Échelle de limitation de la douleur et de l'Échelle de l'intensité de la douleur au cours de leurs premières évaluations en physiothérapie (moment 1); de 24 à 48 heures plus tard (moment 2) et 3 semaines après le début du traitement ou le congé, selon l'échéance la plus rapprochée (moment 3). On a réuni des données démographiques, y compris sur leur état de travailleur, au cours du moment 1. Le physiothérapeute traitant et le patient ont fourni des évolutions globales du changement (EGC) au moment 3. On a calculé le résultat du questionnaire IFMS-15 à partir de questions pertinentes contenues dans la version IFMS-20. Le coefficient de corrélation intracatégorie (CCI) et le changement détectable minimal (CDM) ont quantifié la fiabilité de test­retest (moment 1­moment 2). La validité convergente transversale a été déterminée par le lien (r de Pearson) entre les mesures de fonction et de douleur prises au moment 1. On a évalué la validité des groupes connus au moyen d'une analyse bidirectionnelle des écarts (ANOVA) entre trois niveaux d'état de fonctionnement. La validité longitudinale a été déterminée en fonction du lien (r de Pearson) entre les scores de changement de la fonction et de la douleur (moment 1­moment 3). Des courbes des caractéristiques opérationnelles du récepteur (COR) ont estimé la pDMCI à partir des scores de changement entre le moment 1 et le moment 3 et les EGC moyennes patient/thérapeute. Résultats : La fiabilité des questionnaires IFMS-20 et IFMS-15 a été la même (CCI=0,94 pour les deux mesures). Les valeurs du CDM se sont établies à 9,4/80 pour le questionnaire IFMS-20 et à 8,8/100 pour le questionnaire IFMS-15. La validité transversale, de groupes connus et longitudinale a été confirmée pour les deux mesures de l'IFMS. Les valeurs de la pDMCI s'établissaient à 8/80 pour le questionnaire IFMS-20 et à 6,7/100 pour le questionnaire IFMS-15; la pDMCI était plus élevée chez les personnes dont le bras non dominant était atteint. Conclusions : Les deux mesures de l'IFMS montrent une fiabilité et une validité acceptables. Le bras dominant a un effet sur la pDMCI. On recommande le questionnaire IFMS-15 parce qu'il mesure une dimension seulement: la fonction des membres supérieurs.

RESUMEN

CONTEXT: Adolescents with differentiated thyroid cancer (DTC) require lifelong monitoring with a high possibility of reoperation or radioactive iodine. Although adult DTC survivors have similar or slightly worse quality of life (QOL), this has not been evaluated in the pediatric population. OBJECTIVE: Our objective was to compare QOL and anxiety in adolescents with DTC to patients with acquired autoimmune hypothyroidism. DESIGN, SETTING, AND PATIENTS: In this cross-sectional pilot study, three validated questionnaires were administered to 16 adolescents with DTC and 16 controls for assessment of QOL and anxiety levels. These included teen and parent PedsQL, Multidimensional Anxiety Scale for Children, and Coddington Life Events Scales for Adolescents. The contribution of age, time since diagnosis, and biochemical variables were compared with the outcome measures. RESULTS: There were 16 DTC patients (seven males); 13 had papillary carcinoma, one had follicular carcinoma, and two had mixed type. At diagnosis, five DTC patients had lymph node involvement and two had lung metastases, although at time of assessment, only one DTC patient had lymph node involvement. DTC patients were older than control subjects (P=0.004) and had lower TSH levels than control subjects at time of assessment (P=0.013). QOL and anxiety levels did not differ between DTC patients compared with control subjects and with previously reported scores in a healthy cohort. QOL and anxiety level parameters were not influenced by age, time since diagnosis, or free T4 levels measured at the time of assessment. CONCLUSIONS: Adolescents with DTC have similar QOL and anxiety levels compared with autoimmune hypothyroidism patients and with a healthy normative population.

Asunto(s)

Adenocarcinoma Folicular/psicología , Trastornos de Ansiedad/diagnóstico , Trastornos de Ansiedad/psicología , Carcinoma Papilar/psicología , Calidad de Vida , Neoplasias de la Tiroides/psicología , Adenocarcinoma Folicular/secundario , Adolescente , Carcinoma Papilar/secundario , Estudios de Casos y Controles , Diferenciación Celular , Niño , Estudios Transversales , Femenino , Enfermedad de Hashimoto/psicología , Humanos , Neoplasias Pulmonares/psicología , Neoplasias Pulmonares/secundario , Metástasis Linfática , Masculino , Proyectos Piloto , Psicología del Adolescente , Psicología Infantil , Encuestas y Cuestionarios/normas , Neoplasias de la Tiroides/patología , Tiroiditis Autoinmune

RESUMEN

Transient receptor potential melastatin-3 (TRPM3) is a broadly expressed Ca(2+)-permeable nonselective cation channel. Previous work has demonstrated robust activation of TRPM3 by the neuroactive steroid pregnenolone sulfate (PS), but its in vivo gating mechanisms and functions remained poorly understood. Here, we provide evidence that TRPM3 functions as a chemo- and thermosensor in the somatosensory system. TRPM3 is molecularly and functionally expressed in a large subset of small-diameter sensory neurons from dorsal root and trigeminal ganglia, and mediates the aversive and nocifensive behavioral responses to PS. Moreover, we demonstrate that TRPM3 is steeply activated by heating and underlies heat sensitivity in a subset of sensory neurons. TRPM3-deficient mice exhibited clear deficits in their avoidance responses to noxious heat and in the development of inflammatory heat hyperalgesia. These experiments reveal an unanticipated role for TRPM3 as a thermosensitive nociceptor channel implicated in the detection of noxious heat.

Asunto(s)

Calor/efectos adversos , Hiperalgesia/metabolismo , Umbral del Dolor/fisiología , Células Receptoras Sensoriales/metabolismo , Canales Catiónicos TRPM/metabolismo , Acrilamidas/uso terapéutico , Animales , Conducta Animal/efectos de los fármacos , Glucemia/efectos de los fármacos , Glucemia/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Calcio/metabolismo , Bloqueadores de los Canales de Calcio/farmacología , Capsaicina/farmacología , Línea Celular Transformada , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Adyuvante de Freund/efectos adversos , Ganglios Espinales/citología , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/efectos de la radiación , Humanos , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/genética , Inflamación/inducido químicamente , Inflamación/genética , Inflamación/metabolismo , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Planta de la Mostaza , Nifedipino/farmacología , Dimensión del Dolor/efectos de los fármacos , Dimensión del Dolor/métodos , Umbral del Dolor/efectos de los fármacos , Técnicas de Placa-Clamp , Aceites de Plantas/farmacología , Pregnenolona/efectos adversos , Células Receptoras Sensoriales/efectos de los fármacos , Canal Catiónico TRPA1 , Canales Catiónicos TRPM/antagonistas & inhibidores , Canales Catiónicos TRPM/deficiencia , Canales Catiónicos TRPM/genética , Telemetría/métodos , Factores de Tiempo , Transfección/métodos , Canales de Potencial de Receptor Transitorio/deficiencia , Canales de Potencial de Receptor Transitorio/genética , Ganglio del Trigémino/citología

RESUMEN

Topical application of nicotine, as used in nicotine replacement therapies, causes irritation of the mucosa and skin. This reaction has been attributed to activation of nicotinic acetylcholine receptors (nAChRs) in chemosensory neurons. In contrast with this view, we found that the chemosensory cation channel transient receptor potential A1 (TRPA1) is crucially involved in nicotine-induced irritation. We found that micromolar concentrations of nicotine activated heterologously expressed mouse and human TRPA1. Nicotine acted in a membrane-delimited manner, stabilizing the open state(s) and destabilizing the closed state(s) of the channel. In the presence of the general nAChR blocker hexamethonium, nociceptive neurons showed nicotine-induced responses that were strongly reduced in TRPA1-deficient mice. Finally, TRPA1 mediated the mouse airway constriction reflex to nasal instillation of nicotine. The identification of TRPA1 as a nicotine target suggests that existing models of nicotine-induced irritation should be revised and may facilitate the development of smoking cessation therapies with less adverse effects.

Asunto(s)

Nicotina/farmacología , Agonistas Nicotínicos/farmacología , Células Receptoras Sensoriales/efectos de los fármacos , Células Receptoras Sensoriales/metabolismo , Resistencia de las Vías Respiratorias/efectos de los fármacos , Resistencia de las Vías Respiratorias/genética , Animales , Antipruriginosos/farmacología , Biofisica , Células CHO , Calcio , Canales de Calcio/genética , Células Cultivadas , Cricetinae , Cricetulus , Estimulación Eléctrica , Líquido Extracelular/efectos de los fármacos , Líquido Extracelular/metabolismo , Humanos , Mecamilamina/farmacología , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Mentol/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Planta de la Mostaza , Proteínas del Tejido Nervioso/genética , Antagonistas Nicotínicos/farmacología , Técnicas de Placa-Clamp/métodos , Aceites de Plantas/farmacología , Pletismografía Total/métodos , Células Receptoras Sensoriales/citología , Canal Catiónico TRPA1 , Factores de Tiempo , Transfección/métodos , Canales de Potencial de Receptor Transitorio/genética , Ganglio del Trigémino/citología