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Membrane receptors regulate numerous intracellular functions. However, the molecular underpinnings remain poorly understood because most receptors initiate multiple signaling pathways through distinct interaction interfaces that are structurally uncharacterized. We present an integrated computational and experimental approach to model and rationally engineer membrane receptor-intracellular protein systems signaling with novel pathway selectivity. We targeted the dopamine D2 receptor (D2), a G-protein-coupled receptor (GPCR), which primarily signals through Gi, but triggers also the Gq and beta-arrestin pathways. Using this approach, we designed orthogonal D2-Gi complexes, which coupled with high specificity and triggered exclusively the Gi-dependent signaling pathway. We also engineered an orthogonal chimeric D2-Gs/i complex that rewired D2 signaling from a Gi-mediated inhibitory into a Gs-dependent activating pathway. Reinterpreting the evolutionary history of GPCRs in light of the designed proteins, we uncovered an unforeseen hierarchical code of GPCR-G-protein coupling selectivity determinants. The results demonstrate that membrane receptor-cytosolic protein systems can be rationally engineered to regulate mammalian cellular functions. The method should prove useful for creating orthogonal molecular switches that redirect signals at the cell surface for cell-engineering applications.
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Simulação por Computador , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/química , Complexos Multiproteicos/química , Engenharia de Proteínas , Receptores de Dopamina D2/química , Transdução de Sinais , Linhagem Celular , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Humanos , Complexos Multiproteicos/genética , Receptores de Dopamina D2/genéticaRESUMO
BACKGROUND: Obstetric anal sphincter injury is the most frequent cause of fecal incontinence (FI) in young women. However, the relationship between the extent of anal sphincter defects and the severity of long-term FI (at least 1 year after delivery) has been poorly studied. The aim of the present study was to determine if, in the long term, the extent of anal sphincter defects graded at anal endosonography was linked with the severity of FI. METHODS: A retrospective study was conducted on women with a history of vaginal delivery, who presented with FI and had three-dimensional anorectal high-resolution manometry and endoanal ultrasound in our center from January 2015 to 2016. The detailed clinical history of each patient was obtained from the institutional database. The severity of FI was assessed with the Jorge and Wexner continence scale. RESULTS: There were 250 women with a mean age of 60 ± 14 years. Seventy-six (30.4%) had an isolated defect of the internal anal sphincter, 21 (8.4%) had an isolated defect of the external anal sphincter, and 150 (60%) had both internal and external sphincter defects. The extent of IAS and EAS defects was proportionally correlated with the decrease in mean resting anal pressure (p < 0.01) and the decrease in mean squeeze pressure (p = 0.013) measured by 3DHRAM. No significant correlation was found between the extent and location of the defect (IAS, EAS or both) on endoanal ultrasound and the severity of FI. Menopause was the only independent factor significantly associated with the severity of FI. CONCLUSIONS: In our study, no significant correlation was observed between the extent of the anal sphincter defect and the severity of FI. Menopause was the only identified and independent risk factor for FI. These data confirm that, in the long-term, FI is often multifactorial.
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Canal Anal , Incontinência Fecal , Idoso , Canal Anal/diagnóstico por imagem , Canal Anal/patologia , Parto Obstétrico/efeitos adversos , Endossonografia , Incontinência Fecal/complicações , Incontinência Fecal/diagnóstico por imagem , Incontinência Fecal/etiologia , Feminino , Humanos , Manometria , Pessoa de Meia-Idade , Gravidez , Estudos Retrospectivos , UltrassonografiaRESUMO
BACKGROUND: Outside clinical trials, data on systemic reactions (SRs) due to allergen immunotherapy (AIT) are scarce. METHODS: A prospective, longitudinal, web-based survey of 'real-life' respiratory allergen immunotherapy (AIT) clinical practice was conducted in France, Germany and Spain. SRs were recorded and coded according to the Medical Dictionary for Regulatory Activities (MedDRA) and risk factors associated with SRs were identified. RESULTS: A total of 4316 patients (corresponding to 4363 ongoing courses of AIT) were included. A total of 109 SRs were recorded, and 90 patients (2.1%) presented at least one SR. Most of the SRs occurred in subcutaneous allergen immunotherapy (SCIT) (89%, n = 97). The most frequently reported symptoms were urticaria, rhinitis, dyspnoea and cough. Respiratory symptoms appeared before skin symptoms. Most SRs occurred during the up-dosing phase (75.8%) and were mild in severity (71.6%). Intramuscular adrenaline was administered in 17 SRs, but only 65% of these were subsequently classified as anaphylaxis. Independent risk factors for SRs during SCIT were as follows: the use of natural extracts (odds ratio, OR) [95% confidence interval (CI)] = 2.74 [1.61-4.87], P = 0.001), the absence of symptomatic allergy medications (1.707 [1.008-2.892], P = 0.047), asthma diagnosis (1.74 [1.05-2.88], P = 0.03), sensitization to animal dander (1.93 [1.21-3.09], P = 0.006) or pollen (1.16 [1.03-1.30], P = 0.012) and cluster regimens (vs rush) (4.18 [1.21-14.37], P = 0.023). A previous episode of anaphylaxis increased the risk for anaphylaxis in SCIT (OR [95% CI] = 17.35 [1.91-157.28], P = 0.01). CONCLUSION: AIT for respiratory allergy is safe, with a low number of SRs observed in real-life clinical practice. A personalized analysis of risk factors could be used to minimize SRs.
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Dessensibilização Imunológica/efeitos adversos , Hipersensibilidade/epidemiologia , Vigilância da População , Adolescente , Adulto , Alérgenos/administração & dosagem , Alérgenos/imunologia , Dessensibilização Imunológica/métodos , Europa (Continente)/epidemiologia , Feminino , Humanos , Hipersensibilidade/diagnóstico , Hipersensibilidade/imunologia , Hipersensibilidade/terapia , Imunização , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Estudos Longitudinais , Masculino , Razão de Chances , Estudos Prospectivos , Fatores de Risco , Testes Cutâneos , Inquéritos e Questionários , Avaliação de Sintomas , Adulto JovemRESUMO
We report the generation of free space terahertz (THz) pulses with energy up to 8.3 ± 0.2 µJ from an encapsulated interdigitated ZnSe Large Aperture Photo-Conductive Antenna (LAPCA). An aperture of 12.2 cm2 is illuminated using a 400 nm pump laser with multi-mJ energies at 10 Hz repetition rate. The calculated THz peak electric field is 331 ± 4 kV/cm with a spectrum characterized by a median frequency of 0.28 THz. Given its relatively low frequency, this THz field will accelerate charged particles efficiently having very large ponderomotive energy of 15 ± 1 eV for electrons in vacuum. The scaling of the emission is studied with respect to the dimensions of the antenna, and it is observed that the capacitance of the LAPCA leads to a severe decrease in and distortion of the biasing voltage pulse, fundamentally limiting the maximum applied bias field and consequently the maximum energy of the radiated THz pulses. In order to demonstrate the advantages of this source in the strong field regime, an open-aperture Z-scan experiment was performed on n-doped InGaAs, which showed significant absorption bleaching.
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AIM: Anorectal manometry is the most common test used to explore anorectal disorders. The recent three-dimensional high-resolution anorectal manometry (3D-HRAM) technique appears to be able to provide new topographic information. Our objective was to develop an automated analysis of 3D-HRAM images to diagnose anal sphincter defects and compare the results with those of endoanal ultrasonography (EUS), which is considered to be the gold standard. METHOD: All patients being tested in our department for faecal incontinence or dyschezia by 3D-HRAM and EUS were eligible for the study. 3D-HRAM was used to record resting and squeeze pressure, reflecting internal and external anal sphincter function, respectively. A software platform was designed to automatically analyse the 3D-HRAM images and calculate a diagnostic score for any anal sphincter defect compared with EUS. RESULTS: A total of 206 (91% female) patients of mean age of 54 years were included in the study. A sphincter defect was diagnosed by EUS in 130 (63%). The diagnostic scores from the 3D-HRAM automated analysis for an internal anal sphincter defect showed a sensitivity of 65% and a specificity of 65%. For an external anal sphincter defect, the sensitivity was 43% and the specificity 87%. CONCLUSION: Our study developed a method based on 3D-HRAM to automatically diagnose sphincter defects, allowing a systematic and comprehensive analysis of the test recordings. Compared with EUS, the 3D-HRAM image analysis procedure revealed poor sensitivity and specificity.
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Canal Anal/diagnóstico por imagem , Automação/instrumentação , Incontinência Fecal/diagnóstico , Imageamento Tridimensional , Manometria/métodos , Adulto , Idoso , Canal Anal/fisiopatologia , Estudos de Coortes , Constipação Intestinal/diagnóstico , Endossonografia/métodos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sensibilidade e Especificidade , SoftwareRESUMO
Antipsychotic drugs are known to have sex-dependent effects on metabolic homeostasis. Liver plays a crucial role in drug degradation as well as in glucose and lipid metabolism. The present study examines the influence of clozapine and haloperidol on metabolic liver parameters. Over 12 weeks, male and female Sprague-Dawley rats were fed ground pellets containing clozapine or haloperidol. Liver mass was weighed and liver index calculated. Liver transaminases (ALAT, ALP), malondialdehyde, glucose, triglycerides, total cholesterol, HDL-cholesterol, and glycogen were determined. Finally, SREBP-1 and SREBP-2 as well as neutral fat deposits were examined. In male rats fed with clozapine, we found increased liver mass correlated with an increased liver index, high triglyceride levels, a high ratio of SREBP-1, and an elevated neutral fat distribution. Male and female haloperidol treated rats showed decreased liver mass and increased neutral fat deposition. Malondialdehyde was increased in all rats receiving antipsychotic medication indicating elevated oxidative stress. In both male and female clozapine treated rats, we found glycogen depletion related to decreased glucose levels in females. While liver transaminases were unchanged in the clozapine group, ALAT was elevated after haloperidol treatment in both sexes. Chronic clozapine intake exerts sex-dependent effects on hepatic metabolism. Although haloperidol has been shown to change fewer metabolic parameters, it causes oxidative stress and neutral fat deposits in liver tissue in both sexes.
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Clozapina/farmacologia , Haloperidol/farmacologia , Fígado/metabolismo , Caracteres Sexuais , Alanina Transaminase/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Biomarcadores/metabolismo , HDL-Colesterol/metabolismo , Feminino , Glucose/metabolismo , Glicogênio/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Malondialdeído/metabolismo , Tamanho do Órgão/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos Sprague-Dawley , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Triglicerídeos/metabolismoRESUMO
AIM: Three-dimensional high-resolution anorectal manometry (3DHRAM) is a new technique that can simultaneously provide physiological and topographical data on the terminal part of the digestive tract. Our object was to assess whether 3DHRAM is able to reliably diagnose excessive perineal descent already diagnosed with conventional defaecography, which is considered to be the gold standard. METHOD: All patients referred to our centre for anorectal manometry and conventional defaecography were evaluated with a maximum of 6 months between the two examinations. Anorectal manometry was performed using the 3D High-Resolution Given Imaging® probe. Excessive perineal descent was defined as the downward movement of the anal high-pressure zone during straining. At the end of the straining effort, the high-pressure zone regained its initial position, thereby indicating that the probe had not moved. RESULTS: Nineteen female patients of median age 53 (21-70) years were included in the study. All cases with excessive perineal descent diagnosed using defaecography were visualized with 3DHRAM. The degree of perineal descent determined by 3D and conventional defaecography was compared (Spearman correlation 0.726, P = 0.01). In contrast, the averages measured were significantly different; the average was 11.68 ± 3.3 mm for 3DHRAM but 34.21 ± 13.3 mm for conventional defaecography (P = 0.002). CONCLUSION: The results of the study demonstrate that 3DHRAM can diagnose excessive perineal descent with the same degree of reliability as defaecography. Quantitative measures were not correlated, however, possibly because of methodological differences. The study confirms the value of the morphological data provided by 3DHRAM.
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Canal Anal/fisiopatologia , Defecação/fisiologia , Manometria/métodos , Reto/fisiopatologia , Adulto , Idoso , Canal Anal/diagnóstico por imagem , Constipação Intestinal/fisiopatologia , Defecografia , Incontinência Fecal/fisiopatologia , Feminino , Humanos , Imageamento Tridimensional , Pessoa de Meia-Idade , Projetos Piloto , Pressão , Reto/diagnóstico por imagem , Estudos Retrospectivos , Transdutores de Pressão , Adulto JovemRESUMO
Faecal incontinence is common and significantly affects quality of life. Its treatment involves dietary manipulation, medical treatments, perineal rehabilitation or surgery. In this paper, the French National Society of Coloproctology offers recommendations based on the data in the current literature, including those on recently developed treatments. There is a lack of high quality data and most of the recommendations are therefore based either on grade of recommendation B or expert recommendation (Level 4). However, the literature supports the construction of an algorithm based on the available scientific evidence and expert recommendation which may be useful in clinical practice. The French National Society of Coloproctology proposes a decision-making algorithm that includes recent developments of treatment. The current recommendations support sacral nerve modulation as the key treatment for faecal incontinence. They do not support the use of sphincter substitutions except in certain circumstances. Transanal irrigation is a novel often successful treatment of faecal incontinence due to neurological disorders.
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Canal Anal/cirurgia , Antidiarreicos/uso terapêutico , Dietoterapia/métodos , Terapia por Estimulação Elétrica/métodos , Terapia por Exercício/métodos , Incontinência Fecal/terapia , Resina de Colestiramina/uso terapêutico , Fibras na Dieta/uso terapêutico , Medicina Baseada em Evidências , Humanos , Loperamida/uso terapêutico , Plexo Lombossacral , Mucilagem Vegetal/uso terapêutico , Irrigação Terapêutica/métodos , Nervo Tibial , Resultado do TratamentoRESUMO
AIM: The study aimed to assess, for the first time, the effectiveness of interferential therapy (IFT) in the treatment of slow transit constipation in adults and its impact on the quality of life. METHOD: All consecutive patients with slow transit constipation diagnosed by symptomology and a colonic transit time (CTT) of > 100 h measured with radiopaque markers were included in this prospective study. IFT was performed for 1 h/day over 3 months. Clinical improvement was based on the stool diary and the Knowles-Eccersley-Scott Symptom and Cleveland Clinic Constipation Scores. Quality of life was assessed with the Gastrointestinal Quality of Life Index questionnaire. RESULTS: Eleven patients with a median age of 51 years were included. At the end of the follow-up period, seven (63.6%) had significantly improved after IFT with a median of 0.66 stools per week [interquartile range (IQR) 0.33-0.66] before treatment and 1.66 (IQR 1.33-1.66) after (P = 0.007). The Knowles-Eccersley-Scott Symptom score changed from 30 (IQR 27-33) before treatment to 19 (IQR 17-26) after treatment (P = 0.005) and the Cleveland Clinic Constipation Score from 26 (IQR 25-28) to 17 (IQR 13-24; P = 0.005). The CTT improved from 103 h (IQR 101-113) to 98 h (IQR 94-107; P = 0.02). The Gastrointestinal Quality of Life Index score improved from 60 (IQR 57-63) to 95 (IQR 68-100; P = 0.005). CONCLUSION: IFT is a new non-invasive treatment for slow transit constipation. Further studies to confirm these results with longer follow-up are necessary.
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Constipação Intestinal/terapia , Defecação , Terapia por Estimulação Elétrica , Qualidade de Vida , Doença Crônica , Feminino , Trânsito Gastrointestinal , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Estatísticas não Paramétricas , Inquéritos e Questionários , Resultado do TratamentoRESUMO
AIM: Our aim was to compare for the first time measurements obtained with water-perfused catheter anorectal manometry and three-dimensional (3D) high-resolution manometry in patients with anorectal disorders. METHOD: Consecutive patients referred to our centre for anorectal manometry (ARM) were recruited to undergo the two procedures successively. Conventional manometry was carried out using a water-perfused catheter (WPAM) and high-resolution manometry was achieved with a 3D probe (3DHRAM). For each procedure, parameters recorded included the following: anal canal length, resting pressure, squeeze pressure and rectal sensitivity. RESULTS: Two hundred and one patients were included in this study. The mean values for resting and squeeze pressures were correlated and found to be significantly higher when measured with 3DHRAM than with WPAM. However, the length of the anal canal was not significantly different when measured by the two techniques without correlation between the two mean values obtained. The presence of the rectoanal inhibitory reflex was systematically assessed by both WPAM and 3DHRAM and anismus was also systematically diagnosed by both WPAM and 3DHRAM. CONCLUSION: The pressure values obtained with 3DHRAM are correlated with those measured with conventional manometry but are systematically higher. 3DHRAM has the advantage of providing a pressure recording over the entire length and circumference of the anal canal, allowing a more useful physiological assessment of anorectal function.
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Doenças do Ânus/diagnóstico , Manometria/métodos , Adulto , Idoso , Canal Anal/fisiopatologia , Doenças do Ânus/complicações , Doenças do Ânus/fisiopatologia , Constipação Intestinal/etiologia , Incontinência Fecal/etiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dor/etiologia , Pressão , Doenças Retais/complicações , Doenças Retais/diagnóstico , Doenças Retais/fisiopatologia , Reto/fisiopatologiaRESUMO
AIM: Three-dimensional high-resolution anorectal manometry (3D HRAM) is a new technique that can simultaneously provide physiological and topographical data. Our aim was to assess whether it can identify anal sphincter defects by comparing it with endoanal ultrasonography (EUS) considered as the gold standard. METHOD: An anal defect on 3D HRAM was defined as a continuous circumferential area over which the pressure was < 10 mmHg during the measurement of anal resting and voluntary contraction pressure. Inter-observer agreement was also assessed. RESULTS: A total of 100 patients (93 females) with a mean age of 53.5 ± 15.3 years were included. The positive diagnosis of an anal sphincter defect using 3D HRAM and EUS was in agreement (59.3%) (κ = 0.419) of the time for the internal anal sphincter (IAS) and (55.9%) (κ = 0.461) for the external anal sphincter (EAS). The inter-observer agreement for a diagnosis of an anal sphincter defect was (100%) (κ = 0.937) for the IAS and (95%) (κ = 0.751) for the EAS. The intra-class correlation coefficient for the extent of the defect was 0.853 for the IAS and 0.651 for the EAS. CONCLUSION: The preliminary results demonstrate some level of agreement in the diagnosis of anal sphincter defects between 3D HRAM and EUS but insufficient for 3D HRAM to be adequately reliable using the criteria chosen. The excellent inter-observer agreement, however, demonstrates that 3D HRAM is reproducible and provides a new dimension for the evaluation of sphincter function.
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Doenças do Ânus/diagnóstico por imagem , Doenças do Ânus/fisiopatologia , Endossonografia , Imageamento Tridimensional , Manometria/métodos , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , PressãoRESUMO
INTRODUCTION: An increased risk for metabolic syndrome has been described for patients with psychotic disorders. Antipsychotic drugs possibly contribute to metabolic changes. METHODS: Haloperidol or clozapine was orally fed to male and female Sprague Dawley rats for 12 weeks, and body weight gain, food and water intake were measured. The serum levels of fasting glucose, HbA1c, triglycerides, cholesterol, HDL and LDL, insulin, leptin, adiponectin and ghrelin were determined. Gonadal and perirenal fat pads were removed and weighed. RESULTS: We found increased body weight in the male clozapine group, but decreased ones in the male haloperidol group. Clozapine-treated male and female animals had higher fasting glucose, adiponectin, leptin, ghrelin, cholesterol, HDL and LDL levels, whereas haloperidol caused increased levels of insulin and decreased values of HbA1c, cholesterol, HDL and LDL. CONCLUSION: Both antipsychotic drugs cause sex-dependent metabolic changes, which are risk factors for the metabolic syndrome, be it hyperinsulinemia under haloperidol treatment or hyperglycemia, hyperleptinemia and hyperlipidemia under clozapine.
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Antipsicóticos/farmacologia , Clozapina/farmacologia , Haloperidol/farmacologia , Síndrome Metabólica/metabolismo , Adiposidade/efeitos dos fármacos , Animais , Antipsicóticos/sangue , Glicemia/metabolismo , Peso Corporal/efeitos dos fármacos , Clozapina/sangue , Ingestão de Líquidos/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Feminino , Haloperidol/sangue , Hormônios/sangue , Insulina/metabolismo , Lipídeos/sangue , Masculino , Atividade Motora/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Caracteres Sexuais , Aumento de Peso/efeitos dos fármacosRESUMO
OBJECTIVES: The objective of this study was to show that although transcutaneous electrical tibial nerve stimulation (TENS) is being increasingly used to treat fecal incontinence (FI), its efficacy has never been proved using controlled trials. METHODS: In this randomized, double-blind, sham-controlled trial, 144 patients aged 30-82 years from nine centers were randomly assigned to receive either active or sham stimulations for 3 months. The primary end point was the response to treatment based on the number of incontinence and urgency episodes. Secondary end points were severity scores, quality of life scores, delay to postpone defecation, patient self-assessment of treatment efficacy, physician assessment of TENS efficacy, anorectal manometry, and adverse events. RESULTS: No statistically significant difference was seen between active and sham TENS in terms of an improvement in the median number of FI/urgency episodes per week. Thirty-four patients (47%) who received the active TENS treatment exhibited a >30% decrease in the FI severity score compared with 19 patients (27%) who received the sham treatment (odds ratio 2.4, 95% confidence interval 1.1-5.1, P=0.02). No differences in delay to postpone defecation, patient self-assessment of treatment efficacy, or anorectal manometry were seen between the two groups. The evaluating physicians rated the active stimulations as more effective than the sham stimulations (P=0.01). One minor therapy-related adverse event was observed (1.5%) (see Supplementary Consort 1b). CONCLUSIONS: We failed to demonstrate any benefit of TENS on our primary end-point.
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Incontinência Fecal/fisiopatologia , Incontinência Fecal/terapia , Nervo Tibial , Estimulação Elétrica Nervosa Transcutânea , Adulto , Idoso , Idoso de 80 Anos ou mais , Canal Anal/fisiopatologia , Defecação , Método Duplo-Cego , Feminino , Humanos , Masculino , Manometria , Pessoa de Meia-Idade , Qualidade de Vida , Reto/fisiopatologia , Autorrelato , Índice de Gravidade de Doença , Inquéritos e Questionários , Fatores de Tempo , Estimulação Elétrica Nervosa Transcutânea/efeitos adversos , Estimulação Elétrica Nervosa Transcutânea/métodos , Resultado do TratamentoRESUMO
BACKGROUND: Molecular allergens enable the definition of sensitization profiles in allergic patients. AIM: To validate the most helpful allergens for the diagnosis of latex allergy in different clinical situations. METHODS: 130 patients suspected to be allergic to latex with positive IgE against natural rubber latex (NRL) have been studied: 97 were confirmed as latex allergic (among which 55 professionally exposed to latex and 35 with a peranaesthetic anaphylactic shock) and 33 were only sensitized to latex without clinical allergy. Each serum was tested for IgE against 9 recombinant latex allergens and bromelain using Phadia ImmunoCAP 250. RESULTS: rHev b 6.01, 6.02, 2 and 5 were the major allergens in the allergic population. An excellent correlation (94%) was observed between IgE against rHev b 6.01 and latex prick test positivities. IgE against rHev b 1, 3 and 5 were more frequent and their levels significantly higher in patients with peranaesthetic anaphylactic shock. Among the asymptomatic patients (29/33 allergic to pollen), NRL IgE positivity is explained by the presence of anti-rHev b 8 and/or anti-carbohydrate IgE. CONCLUSIONS: rHev b 6.01 and rHev b 5 specific IgE are of major interest to confirm latex allergy diagnosis. rHev b 5 is particularly useful in case of monosensitization where clinical symptoms and latex skin prick tests may be discordant, rHev b1 and rHev b 3 are interesting to document multi-operated and peranaesthetic latex allergy. Finally, rHev b 8 is a helpful marker to highlight latex/pollen cross-reactivity which improves the specificity of the serological tests.
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Alérgenos , Antígenos de Plantas , Hipersensibilidade ao Látex/diagnóstico , Látex/química , Adolescente , Adulto , Idoso , Alérgenos/imunologia , Antígenos de Plantas/imunologia , Criança , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina E/sangue , Látex/imunologia , Hipersensibilidade ao Látex/sangue , Hipersensibilidade ao Látex/imunologia , Masculino , Pessoa de Meia-Idade , Testes Cutâneos , Adulto JovemRESUMO
AIM: Sacral nerve stimulation (SNS) reduces incontinence episodes and improves the quality of life of patients treated for faecal incontinence. However, the exact mechanism of action of this technique remains unclear. The present article reviews the pertinent neuroanatomy and neurophysiology related to SNS and provides explanations for potential mechanisms of action. METHOD: A systematic review of the literature was performed for studies of the potential mechanisms of action of SNS, using MEDLINE, PubMed, Embase and the Cochrane Library. Articles dealing with the technique, adverse events and economic evaluations of SNS, as well as literature reviews, were excluded, except for reviews dealing with the mechanisms of action of SNS. The following inclusion criteria were used to select articles: (i) articles in English, (ii) randomized, double-blinded, sham-controlled studies, and (iii) cohort studies. Case-control studies or retrospective studies were cited only when randomized or cohort studies could not be found. RESULTS: We propose three hypotheses to explain the mechanism of action of SNS: (i) a somato-visceral reflex, (ii) a modulation of the perception of afferent information, and (iii) an increase in external anal sphincter activity. CONCLUSION: The mechanism of action of SNS in patients with faecal incontinence almost certainly depends on the modulation of spinal and/or supraspinal afferent inputs. Further research on humans and animals will be required to gain a better understanding of the mechanisms of action of SNS.
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Terapia por Estimulação Elétrica , Sistema Nervoso Entérico/fisiologia , Incontinência Fecal/fisiopatologia , Incontinência Fecal/terapia , Canal Anal/inervação , Colo/inervação , Defecação/fisiologia , Humanos , Plexo Lombossacral , Diafragma da Pelve/inervaçãoRESUMO
AIMS: Our main objective was to optimize the enrichment of Escherichia coli O26 in raw milk cheeses for their subsequent detection with a new automated immunological method. METHODS AND RESULTS: Ten enrichment broths were tested for the detection of E. coli O26. Two categories of experimentally inoculated raw milk cheeses, semi-hard uncooked cheese and 'Camembert' type cheese, were initially used to investigate the relative efficacy of the different enrichments. The enrichments that were considered optimal for the growth of E. coli O26 in these cheeses were then challenged with other types of raw milk cheeses. Buffered peptone water supplemented with cefixim-tellurite and acriflavin was shown to optimize the growth of E. coli O26 artificially inoculated in the cheeses tested. Despite the low inoculum level (1-10 CFU per 25 g) in the cheeses, E. coli O26 counts reached at least 5.10(4) CFU ml(-1) after 24-h incubation at 41.5 °C in this medium. CONCLUSIONS: All the experimentally inoculated cheeses were found positive by the immunological method in the enrichment broth selected. SIGNIFICANCE AND IMPACT OF THE STUDY: Optimized E. coli O26 enrichment and rapid detection constitute the first steps of a complete procedure that could be used in routine to detect E. coli O26 in raw milk cheeses.
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Técnicas Bacteriológicas , Queijo/microbiologia , Fluorescência , Técnicas Imunológicas , Escherichia coli Shiga Toxigênica/crescimento & desenvolvimento , Escherichia coli Shiga Toxigênica/isolamento & purificação , Animais , Meios de Cultura , Leite/microbiologiaRESUMO
Vasopressin type 2 receptor (V2R) exhibits mostly important properties for hydroosmotic equilibrium and, to a lesser extent, on vasomotricity. Drugs currently acting on this receptor are analogs of the natural neuropeptide, arginine vasopressin (AVP), and hence are competitive ligands. Peptides that reproduce specific sequences of a given receptor have lately been reported to interfere with its action, and if such molecules arise from regions remote from the binding site they would be anticipated to exhibit noncompetitive antagonism, but this has yet to be shown for V2R. Six peptides reproducing juxtamembranous regions of V2R were designed and screened; the most effective peptide, cravky (labeled VRQ397), was characterized. VRQ397 was potent (IC(50) = 0.69 +/- 0.25 nM) and fully effective in inhibiting V2R-dependent physiological function, specifically desmopressin-L-desamino-8-arginine-vasopressin (DDAVP)-induced cremasteric vasorelaxation; this physiological functional assay was utilized to avoid overlooking interference of specific signaling events. A dose-response profile revealed a noncompetitive property of VRQ397; correspondingly, VRQ397 bound specifically to V2R-expressing cells could not displace its natural ligand, AVP, but modulated AVP binding kinetics (dissociation rate). Specificity of VRQ397 was further confirmed by its inability to bind to homologous V1 and oxytocin receptors and its inefficacy to alter responses to stimulation of these receptors. VRQ397 exhibited pharmacological permissiveness on V2R-induced signals, as it inhibited DDAVP-induced PGI(2) generation but not that of cAMP or recruitment of beta-arrestin2. Consistent with in vitro and ex vivo effects as a V2R antagonist, VRQ397 displayed anticipated in vivo aquaretic efficacy. We hereby describe the discovery of a first potent noncompetitive antagonist of V2R, which exhibits functional selectivity, in line with properties of a negative allosteric modulator.
Assuntos
Antagonistas dos Receptores de Hormônios Antidiuréticos , Antagonistas de Hormônios/farmacologia , Músculo Liso/efeitos dos fármacos , Miométrio/efeitos dos fármacos , Oligopeptídeos/farmacologia , Bexiga Urinária/efeitos dos fármacos , 6-Cetoprostaglandina F1 alfa/metabolismo , Regulação Alostérica , Animais , Arginina Vasopressina/metabolismo , Linhagem Celular , AMP Cíclico/metabolismo , Desamino Arginina Vasopressina/metabolismo , Diurese/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Antagonistas de Hormônios/metabolismo , Humanos , Técnicas In Vitro , Ligantes , Masculino , Camundongos , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/metabolismo , Miométrio/metabolismo , Oligopeptídeos/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Vasopressinas/genética , Receptores de Vasopressinas/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transfecção , Bexiga Urinária/metabolismoRESUMO
An influenza virus matrix peptide in which either the charged amino or carboxyl terminus was substituted by methyl groups promoted folding of the class I human histocompatibility antigen (HLA-A2). A peptide modified at both termini did not promote stable folding. The thermal stability of HLA-A2 complexed with peptides that did not have either terminus was approximately 22 degrees C lower than that of the control peptide, whereas matrix peptide in which both anchor positions were substituted by alanines had its stability decreased by only 5.5 degrees C. Thus, the conserved major histocompatibility complex class I residues at both ends of the peptide binding site form energetically important sites for binding the termini of short peptides.
Assuntos
Antígeno HLA-A2/química , Peptídeos/química , Proteínas da Matriz Viral/química , Sequência de Aminoácidos , Sítios de Ligação , Antígeno HLA-A2/genética , Antígeno HLA-A2/metabolismo , Humanos , Ligação de Hidrogênio , Dados de Sequência Molecular , Mutação , Orthomyxoviridae , Peptídeos/metabolismo , Desnaturação Proteica , Dobramento de Proteína , Temperatura , Termodinâmica , Termolisina/química , Proteínas da Matriz Viral/metabolismo , Microglobulina beta-2/químicaRESUMO
G-protein-coupled receptor kinases (GRKs) comprise a family of seven mammalian serine/threonine protein kinases that phosphorylate and regulate agonist-bound, activated, G-protein-coupled receptors (GPCRs). GRKs and beta-arrestins are key participants in the canonical pathways leading to phosphorylation-dependent GPCR desensitization, endocytosis, intracellular trafficking and resensitization. Here we show that GRK4 isoforms are expressed in human breast cancer but not in normal epithelia. In addition, GRK4-over-expressing cells activated the mitogen-activated protein kinase (MAPK) mediated by ERK 1/2 and JNK phosphorylation in breast cancer-derived cell lines. Furthermore, suppression of beta-arrestins decreased GRK4-stimulated ERK 1/2 or JNK phosphorylations. These data indicate that high-level expression of GRK4 may activate MAPK signalling pathways mediated by beta-arrestins in breast cancer cells, suggesting that GRK4 may be implicated in breast cancer carcinogenesis.
Assuntos
Neoplasias da Mama/genética , Quinase 4 de Receptor Acoplado a Proteína G/genética , Isoformas de Proteínas/genética , Arrestinas/análise , Biomarcadores Tumorais/análise , Neoplasias da Mama/química , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Ativação Enzimática , Feminino , Quinase 4 de Receptor Acoplado a Proteína G/análise , Expressão Gênica , Humanos , Immunoblotting/métodos , Imuno-Histoquímica , MAP Quinase Quinase 4/metabolismo , Sistema de Sinalização das MAP Quinases , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosforilação , Isoformas de Proteínas/análise , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , beta-ArrestinasRESUMO
Over 150 mutations within the coding sequence of the V2 vasopressin receptor (V2R) gene are known to cause nephrogenic diabetes insipidus (NDI). A large number of these mutant receptors fail to fold properly and therefore are not routed to the cell surface. Here we show that selective, nonpeptidic V2R antagonists dramatically increase cell-surface expression and rescue the function of 8 mutant NDI-V2Rs by promoting their proper folding and maturation. A cell-impermeant V2R antagonist could not mimic these effects and was unable to block the rescue mediated by a permeant agent, indicating that the nonpeptidic antagonists act intracellularly, presumably by binding to and stabilizing partially folded mutants. In addition to opening new therapeutic avenues for NDI patients, these data demonstrate that by binding to newly synthesized mutant receptors, small ligands can act as pharmacological chaperones, promoting the proper folding and maturation of receptors and their targeting to the cell surface.