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Adrenal incidentalomas are a frequent cause for consultation in endocrinology. Current guidelines provide an algorithm for their evaluation to determinate the risk of hormonally active or malignant condition. We report a unique case of benign adrenal incidentaloma in a 53-year-old woman with multiple malignancy criteria on contrast-free computed tomography and [18F]fluorodeoxyglucose positron emission tomography-computed tomography, leading to a left adrenalectomy. Histopathologic analysis showed a 15-mm intra-adrenal lymphoid organ localized in the medulla, without any cellular atypia and organized with a capsule and multiple germinal centers. The surrounding adrenal tissue was unremarkable, and the patient did not develop any inflammatory, infectious, or neoplastic disease during the 2 following years. This is the first described case of a well-organized intra-adrenal ectopic lymphoid organ in the absence of any evident stimulus.
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Biological membranes play a crucial role in actively hosting, modulating and coordinating a wide range of molecular events essential for cellular function. Membranes are organized into diverse domains giving rise to dynamic molecular patchworks. However, the very definition of membrane domains has been the subject of continuous debate. For example, in the plant field, membrane domains are often referred to as nanodomains, nanoclusters, microdomains, lipid rafts, membrane rafts, signalling platforms, foci or liquid-ordered membranes without any clear rationale. In the context of plant-microbe interactions, microdomains have sometimes been used to refer to the large area at the plant-microbe interface. Some of these terms have partially overlapping meanings at best, but they are often used interchangeably in the literature. This situation generates much confusion and limits conceptual progress. There is thus an urgent need for us as a scientific community to resolve these semantic and conceptual controversies by defining an unambiguous nomenclature of membrane domains. In this Review, experts in the field get together to provide explicit definitions of plasma membrane domains in plant systems and experimental guidelines for their study. We propose that plasma membrane domains should not be considered on the basis of their size alone but rather according to the biological system being considered, such as the local membrane environment or the entire cell.
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Membrana Celular , Microdomínios da Membrana , Plantas , Terminologia como Assunto , Microdomínios da Membrana/metabolismo , Membrana Celular/metabolismoRESUMO
Direct current stimulation (DCS) is a non-invasive approach to stimulate the nervous system that is now considered a powerful tool for treating neurological diseases such as those affecting cognitive or locomotor functions. DCS, as applied clinically today, is an approach built on early uses in antiquity and knowledge gained over time. Its current use makes use of specific devices and takes into account knowledge of the mechanisms by which this approach modulates functioning of the nervous system at the cellular level. Over the last 20 years, although there are few studies, it has been shown that DCS can also modulate the breathing autonomic function. In this narrative review, after briefly providing the historical perspective and describing the principles and the main cellular and molecular effects, we summarize the currently available data regarding the modulation of ventilation, and propose that DCS could be used to treat autonomic or non-autonomic neurological disorders affecting breathing.
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OBJECTIVE: The objective of this study was to retrospectively assess the effect of Radiofrequency Volumetric Tissue Reduction (RFVTR) on hypertrophic turbinates and clinical outcome in brachycephalic dogs when included in multi-level surgery (MLS). STUDY DESIGN: Clinical retrospective multicenter study. ANIMALS: 132 client-owned brachycephalic dogs. METHODS: 132 brachycephalic dogs with high-grade Brachycephalic Obstructive Airway Ayndrome (BOAS) and hypertrophic turbinates were treated with RFVTR as part of MLS of the upper airways. Intranasal obstruction was evaluated by computer tomography (CT) and antero-/retrograde rhinoscopy before and 6 months after RFVTR. The clinical records, the CT images and the rhinoscopy videos were reviewed and clinical evolution was evaluated using a standardized questionnaire. The data was scored semi-quantitatively. RESULTS: In this study, 132 patients were included for a follow-up period of 120 weeks. RFVTR resulted in minor complications, including serous nasal discharge within the first postoperative week in all dogs, and intermittent nasal congestion between 3-8 weeks after treatment in 24.3% of the patients. Rhinoscopy and CT follow-ups were available for 33 patients. Six months after treatment intranasal airspace was increased (p = 0.002) and the presence and overall amount of mucosal contact points was reduced (p = 0.039). CONCLUSION: MLS with RFVTR led to a significant reduction in turbinate volume at the 6-month follow-up examination and significant clinical improvement over a long-term period of 120 weeks. This suggests the viability of RFVTR as a turbinate-preserving treatment for intranasal obstruction in dogs with BOAS. CLINICAL SIGNIFICANCE: RFVTR is a minimally invasive turbinoplasty technique for intranasal obstruction in dogs with BOAS and can be included in MLS without increasing complication rates.
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Doenças do Cão , Conchas Nasais , Animais , Cães , Conchas Nasais/cirurgia , Conchas Nasais/patologia , Conchas Nasais/diagnóstico por imagem , Estudos Retrospectivos , Doenças do Cão/cirurgia , Doenças do Cão/diagnóstico por imagem , Masculino , Feminino , Obstrução Nasal/cirurgia , Obstrução Nasal/veterinária , Obstrução Nasal/patologia , Hipertrofia , Tomografia Computadorizada por Raios X , Resultado do Tratamento , Obstrução das Vias Respiratórias/veterinária , Obstrução das Vias Respiratórias/cirurgia , Obstrução das Vias Respiratórias/diagnóstico por imagemRESUMO
Endophytic fungal-based biopesticides are sustainable and ecologically-friendly biocontrol agents of several pests and diseases. However, their potential in managing tomato fusarium wilt disease (FWD) remains unexploited. This study therefore evaluated effectiveness of nine fungal isolates against tomato fusarium wilt pathogen, Fusarium oxysporum f. sp. lycopersici (FOL) in vitro using dual culture and co-culture assays. The efficacy of three potent endophytes that inhibited the pathogen in vitro was assessed against FWD incidence, severity, and ability to enhance growth and yield of tomatoes in planta. The ability of endophytically-colonized tomato (Solanum lycopersicum L.) plants to systemically defend themselves upon exposure to FOL were also assessed through defence genes expression using qPCR. In vitro assays showed that endophytes inhibited and suppressed FOL mycelial growth better than entomopathogenic fungi (EPF). Endophytes Trichoderma asperellum M2RT4, Hypocrea lixii F3ST1, Trichoderma harzianum KF2R41, and Trichoderma atroviride ICIPE 710 had the highest (68.84-99.61%) suppression and FOL radial growth inhibition rates compared to EPF which exhibited lowest (27.05-40.63%) inhibition rates. Endophytes T. asperellum M2RT4, H. lixii F3ST1 and T. harzianum KF2R41 colonized all tomato plant parts. During the in planta experiment, endophytically-colonized and FOL-infected tomato plants showed significant reduction of FWD incidence and severity compared to non-inoculated plants. In addition, these endophytes contributed to improved growth promotion parameters and yield. Moreover, there was significantly higher expression of tomato defence genes in T. asperellum M2RT4 colonized than in un-inoculated tomato plants. These findings demonstrated that H. lixii F3ST1 and T. asperellum M2RT4 are effective biocontrol agents against FWD and could sustainably mitigate tomato yield losses associated with fusarium wilt.
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Endófitos , Fusarium , Doenças das Plantas , Solanum lycopersicum , Fusarium/patogenicidade , Fusarium/fisiologia , Solanum lycopersicum/microbiologia , Solanum lycopersicum/crescimento & desenvolvimento , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Endófitos/fisiologia , Hypocreales/fisiologia , Hypocreales/patogenicidade , Antibiose , Controle Biológico de Vetores/métodos , Agentes de Controle BiológicoRESUMO
QUESTION: Dyspnoea persisting despite treatments of underlying causes requires symptomatic approaches. Medical hypnosis could provide relief without the untoward effects of pharmacological approaches. We addressed this question through experimentally induced dyspnoea in healthy humans (inspiratory threshold loading -excessive inspiratory effort- and carbon dioxide stimulation -air hunger-) MATERIAL AND METHODS: 20 volunteers (10 women, 21-40) were studied on 4 separate days. The order of the visits was randomised in two steps, firstly "inspiratory threshold loading first" versus "carbon dioxide first" group (n=10 in each group), secondly "medical hypnosis first" versus "visual distraction first" subgroup (n=5 in each subgroup). Each visit comprised three 5-minutes periods (reference, intervention, washout) during which participants used visual analog scales to rate dyspnoea's sensory and affective dimensions and after which they completed the Multidimensional Dyspnea Profile. RESULTS: Medical hypnosis reduced both dimensions of dyspnoea significantly more than visual distraction (inspiratory threshold loading: sensory reduction after 5â min 34% of full VAS versus 8% -p=0.0042-; affective reduction 17.6% versus 2.4% -p=0.044-; carbon dioxide; sensory reduction after 5â min 36.9% versus 3% -p=0.0015-; affective reduction 29.1% versus 8.7% -p=0.0023-). The Multidimensional Dyspnea Profile showed more marked sensory effects during inspiratory threshold loading and more marked affective effects during carbon dioxide stimulation. ANSWER TO THE QUESTION: Medical hypnosis was more effective than visual distraction at attenuating the sensory and affective dimensions of experimentally induced dyspnoea. This provides a strong rationale for clinical studies of hypnosis in persistent dyspnoea patients.
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We report discovery of a new bacterial genus and species of the family Pasteurellaceae by using phylogenetic and metabolic analysis. The bacterium, Emayella augustorita, was isolated from blood cultures of a patient in France diagnosed with an adenocarcinoma of the intestines and who was treated with a biliary prosthesis placement.
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Hemocultura , Infecções por Pasteurellaceae , Pasteurellaceae , Filogenia , Sepse , Humanos , Pasteurellaceae/isolamento & purificação , Pasteurellaceae/genética , Pasteurellaceae/classificação , Infecções por Pasteurellaceae/microbiologia , Infecções por Pasteurellaceae/diagnóstico , Sepse/microbiologia , Sepse/diagnóstico , RNA Ribossômico 16S/genética , Masculino , Antibacterianos/uso terapêutico , Antibacterianos/farmacologia , Bacteriemia/microbiologia , Bacteriemia/diagnóstico , França , IdosoRESUMO
Septic patients with worst clinical prognosis have increased circulating immature granulocytes (IG), displaying limited phagocytosis and reactive oxygen species (ROS) production. Here, we developed an ex vivo model of incubation of human granulocytes, from septic patients or healthy donors, with Escherichia coli. We showed that the ROS production in Sepsis-IG is lower due to decreased activation and protein expression of the NADPH oxidase complex. We also demonstrated that the low level of ROS production and lower phagocytosis of IG in sepsis induce the bacterial SOS response, leading to the expression of the SOS-regulated quinolone resistance gene qnrB2. Without antimicrobial pressure, the sepsis immune response alone may promote antibiotic resistance expression.
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Background: Better cardiovascular health is associated with lower risk of various chronic diseases, but its association with multimorbidity is poorly understood. We aimed to examine whether change in cardiovascular health is associated with multimorbidity risk. Methods: The primary analysis was conducted in the Whitehall II multiwave prospective cohort study (UK) and the validation analysis in the Finnish Public Sector cohort study (Finland). Change in cardiovascular health was assessed using the American Heart Association Life's Simple 7 (LS7) and Life's Essential 8 (LE8) at baseline and re-assessments, using objective measures in Whitehall II and self-reports and pharmacy claims in the Finnish Public Sector cohort study, respectively. Multimorbidity was defined as the presence of two or more of 12 chronic diseases during follow-up. We estimated hazard ratios (HR) and 95% confidence intervals (CI) using Cox's proportional hazard models with age as time scale, adjusting for sex, education, occupation, marital status, and ethnicity. Findings: In the primary analysis among 9715 participants, mean age was 44.8 (standard deviation 6.0) years and 67.6% participants were men at baseline. During the median follow-up of 31.4 (interquartile range 26.8-32.3) years, 2751 participants developed multimorbidity. The hazard of multimorbidity decreased by 8% (HR 0.92, 95% CI 0.88-0.96) per ideal LS7 metric increment over 5 years and by 14% (HR 0.86, 95% CI 0.80-0.93) per ten points increase in LE8 score over 10 years. These findings were replicated in the validation analysis among 75,377 participants in terms of 4-year change in cardiovascular health. Interpretation: Improvement in cardiovascular health was associated with lower multimorbidity risk in two community-based cohort studies. Interventions improving cardiovascular health of the community may contribute to multimorbidity prevention. Funding: None.
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Empagliflozin has been successfully repurposed for treating neutropenia and neutrophil dysfunction in patients with glycogen storage disease type 1b (GSD 1b), however, data in infants are missing. We report on efficacy and safety of empagliflozin in infants with GSD 1b. This is an international retrospective case series on 21 GSD 1b infants treated with empagliflozin (total treatment time 20.6 years). Before starting empagliflozin (at a median age of 8.1 months with a median dose of 0.3 mg/kg/day) 12 patients had clinical signs and symptoms of neutrophil dysfunction. Six of these previously symptomatic patients had no further neutropenia/neutrophil dysfunction-associated findings on empagliflozin. Eight patients had no signs and symptoms of neutropenia/neutrophil dysfunction before start and during empagliflozin treatment. One previously asymptomatic individual with a horseshoe kidney developed a central line infection with pyelonephritis and urosepsis during empagliflozin treatment. Of the 10 patients who were treated with G-CSF before starting empagliflozin, this was stopped in four and decreased in another four. Eleven individuals were never treated with G-CSF. While in 17 patients glucose homeostasis remained stable on empagliflozin, four showed glucose homeostasis instability in the introductory phase. In 17 patients, no other side effects were reported, while genital (n = 2) or oral (n = 1) candidiasis and skin infection (n = 1) were reported in the remaining four. Empagliflozin had a good effect on neutropenia/neutrophil dysfunction-related signs and symptoms and a favourable safety profile in infants with GSD 1b and therefore qualifies for further exploration as first line treatment.
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Compostos Benzidrílicos , Glucosídeos , Doença de Depósito de Glicogênio Tipo I , Neutropenia , Neutrófilos , Humanos , Doença de Depósito de Glicogênio Tipo I/tratamento farmacológico , Doença de Depósito de Glicogênio Tipo I/complicações , Neutropenia/tratamento farmacológico , Masculino , Feminino , Lactente , Compostos Benzidrílicos/uso terapêutico , Compostos Benzidrílicos/administração & dosagem , Estudos Retrospectivos , Neutrófilos/efeitos dos fármacos , Glucosídeos/uso terapêutico , Glucosídeos/farmacologia , Glucosídeos/administração & dosagem , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Resultado do Tratamento , Fator Estimulador de Colônias de Granulócitos/uso terapêuticoRESUMO
In recent years, European Union member states have hastened energy policy deliberations to address supply and sustainability concerns, placing a significant emphasis on nuclear energy as a means to achieve decarbonization goals. However, despite its significant role in power generation, nuclear energy faces significant challenges linked to fuel reprocessing and waste disposal, that hinder its broader expansion. In this context, the separation of technetium represents a concerning issue. Indeed, technetium's catalytic activity can impede the extraction of uranium, neptunium, and plutonium, affecting waste reprocessing efficiency. Additionally, the stable form of technetium in aerobic conditions, pertechnetate (TcO4 -), poses risks of groundwater contamination due to its mobility and solubility. Hence, sensing and separation of TcO4 - is imperative for both nuclear fuel processing and minimising radioactive contamination in the environment. However, the binding of TcO4 - and its separation from contaminated solutions present challenges due to the acidic (or basic) waste components and the high ionic strength in real matrices. Supramolecular chemists have addressed these issues by designing receptors inspired by molecular recognition principles. This article explores recent advancements and future directions in TcO4 - sensing and separation (using extraction and sorption) with a focus on molecular hosts. Metal-organic receptors will also be discussed.
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BACKGROUND: The increase in antibiotic resistance is a major public health issue. The development of rapid antimicrobial susceptibility testing (AST) methods is becoming a priority to ensure early and appropriate antibiotic therapy. OBJECTIVES: To evaluate sedimentation field-flow fractionation (SdFFF) as a method for performing AST in less than 3â h. METHODS: SdFFF is based on the detection of early biophysical changes in bacteria, using a chromatographic-type technology. One hundred clinical Escherichia coli strains were studied. A calibrated bacterial suspension was incubated for 2â h at 37°C in the absence (untreated) or presence (treated) of five antibiotics used at EUCAST breakpoint concentrations. Bacterial suspensions were then injected into the SdFFF machine. For each E. coli isolate, retention times and elution profiles of antibiotic-treated bacteria were compared with retention times and elution profiles of untreated bacteria. Algorithms comparing retention times and elution profiles were used to determine if the strain was susceptible or resistant. Performance evaluation was done according to CLSI and the ISO standard 20776-2:2021 with broth microdilution used as the reference method. RESULTS: AST results from SdFFF were obtained in less than 3â h. SdFFF showed high categorical agreement (99.8%), sensitivity (99.5%) and specificity (100.0%) with broth microdilution. Results for each antimicrobial were also in agreement with the ISO 20776-2 recommendations, with sensitivity and specificity of ≥95.0%. CONCLUSIONS: This study showed that SdFFF can be used as a rapid, accurate and reliable phenotypic AST method with a turnaround time of less than 3â h.
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Antibacterianos , Escherichia coli , Fracionamento por Campo e Fluxo , Testes de Sensibilidade Microbiana , Testes de Sensibilidade Microbiana/métodos , Testes de Sensibilidade Microbiana/normas , Projetos Piloto , Escherichia coli/efeitos dos fármacos , Antibacterianos/farmacologia , Humanos , Fracionamento por Campo e Fluxo/métodos , Infecções por Escherichia coli/microbiologia , Fatores de TempoRESUMO
Introduction: The Profamille V3.2 multi-family psycho-educational program directed at caregivers of relatives with schizophrenia or schizophrenia related disorder has been shown to decrease the annual prevalence of suicide attempts. It has been reported that psychoeducation of families can sometimes improve compliance with treatment. This study investigates whether the Profamille program improves compliance and thus reduces the risk of suicide among patients. Method: This is a retrospective study of 179 groups of family caregivers, encompassing 1946 participants enrolled in Module 1 of the Profamille program and followed up one year after completion of the module. Evaluations were conducted using questionnaires filled out by family caregivers at three distinct times: prior to beginning the program, upon its completion, and again one year following its conclusion. The annual prevalence of suicide attempts was measured both before the program began and one year after its conclusion, while compliance to treatment was evaluated at the start and end of the program. Result: After the Profamille program, the annual prevalence of suicide attempts fell by a factor of 2 (p-value = 0.00002) and patient compliance improved (p-value <0.000001). This reduction in suicide attempts was observed independently of improved compliance. Compliance seems to have an additional effect, but only after participation in the program. Conclusion: The Profamille program reduces patients' risk of suicide even when patients are not taking the treatment. When family psychoeducation is not proposed in schizophrenia or schizophrenia related disorder, this can represent a loss of chance for patients.
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OBJECTIVE: The present study aimed to evaluate low-dose oral misoprostol induction, and compare different methods used in second-line induction in patients with a Bishop score less than 6. METHODS: This retrospective study analyzed the medical history and courses of pregnancy of all patients induced with first-line of low-dose oral misoprostol (50 µg every 4 h with a total of 200 µg/24 h) from April 2021 to June 2022 in a university hospital center, and reported outcomes according to the second-line method of induction. RESULTS: Among 437 labor inductions with low-dose oral misoprostol, 120 patients required a second-line induction. Predictive factors of first-line failure were higher body mass index (P = 0.011), absence of premature rupture of membranes (P = 0.021) and earlier term of pregnancy (P < 0.001). Regarding second methods of induction of labor, time from induction to delivery was shorter in the oxytocin group than the dinoprostone and misoprostol groups (24.0 vs. 41 and 51.0 h, respectively; P < 0.001), and was also significantly shorter in the dinoprostone than the misoprostol group (P = 0.048). Cesarean section rates did not differ between the three groups (P = 0.651). There were no clinically significant differences in adverse events between the groups. CONCLUSION: Normal body mass index, previous rupture of membranes and later term of induction of labor were the three favoring success factors during first-line oral misoprostol. In cases of a Bishop score <6, oxytocin may be the best option to reduce duration to delivery, with the same maternal-fetal outcomes, including a similar rate of vaginal delivery.
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H chain-only Igs are naturally produced in camelids and sharks. Because these Abs lack the L chain, the Ag-binding domain is half the size of a traditional Ab, allowing this type of Ig to bind to targets in novel ways. Consequently, the H chain-only single-domain Ab (sdAb) structure has the potential to increase the repertoire and functional range of an active humoral immune system. The majority of vertebrates use the standard heterodimeric (both H and L chains) structure and do not produce sdAb format Igs. To investigate if other animals are able to support sdAb development and function, transgenic chickens (Gallus gallus) were designed to produce H chain-only Abs by omitting the L chain V region and maintaining only the LC region to serve as a chaperone for Ab secretion from the cell. These birds produced 30-50% normal B cell populations within PBMCs and readily expressed chicken sequence sdAbs. Interestingly, the H chains contained a spontaneous CH1 deletion. Although no isotype switching to IgY or IgA occurred, the IgM repertoire was diverse, and immunization with a variety of protein immunogens rapidly produced high and specific serum titers. mAbs of high affinity were efficiently recovered by single B cell screening. In in vitro functional assays, the sdAbs produced by birds immunized against SARS-CoV-2 were also able to strongly neutralize and prevent viral replication. These data suggest that the truncated L chain design successfully supported sdAb development and expression in chickens.
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Animais Geneticamente Modificados , Galinhas , Cadeias Pesadas de Imunoglobulinas , Anticorpos de Domínio Único , Animais , Galinhas/imunologia , Anticorpos de Domínio Único/imunologia , Anticorpos de Domínio Único/genética , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Pesadas de Imunoglobulinas/imunologia , SARS-CoV-2/imunologia , SARS-CoV-2/genética , COVID-19/imunologia , Transgenes/genética , Linfócitos B/imunologia , Anticorpos Antivirais/imunologia , Cadeias Leves de Imunoglobulina/genética , Cadeias Leves de Imunoglobulina/imunologia , HumanosRESUMO
BACKGROUND: This study was a secondary analysis of the ROBOGYN-1004 trial conducted between 2010 and 2015. The study aimed to identify factors that affect postoperative morbidity after either robot-assisted laparoscopy (RL) or conventional laparoscopy (CL) in gynecologic oncology. METHODS: The study used two-level logistic regression analyses to evaluate the prognostic and predictive value of patient, surgery, and center characteristics in predicting severe postoperative morbidity 6 months after surgery. RESULTS: This analysis included 368 patients. Severe morbidity occurred in 49 (28 %) of 176 patients who underwent RL versus 41 (21 %) of 192 patients who underwent CL (p = 0.15). In the multivariate analysis, after adjustment for the treatment group (RL vs CL), the risk of severe morbidity increased significantly for patients who had poorer performance status, with an odds ratio (OR) of 1.62 for the 1-point difference in the WHO performance score (95 % CI 1.06-2.47; p = 0.027) and according to the type of surgery (p < 0.001). A focus on complex surgical acts showed significant more morbidity in the RL group than in the CL group at the less experienced centers (OR, 3.31; 95 % CI 1.0-11; p = 0.05) compared with no impact at the experienced centers (OR, 0.87; 95 % CI 0.38-1.99; p = 0.75). CONCLUSION: The findings suggest that the center's experience may have an impact on the risk of morbidity for patients undergoing complex robot-assisted surgical procedures.
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Neoplasias dos Genitais Femininos , Laparoscopia , Complicações Pós-Operatórias , Procedimentos Cirúrgicos Robóticos , Adulto , Idoso , Feminino , Humanos , Pessoa de Meia-Idade , Seguimentos , Neoplasias dos Genitais Femininos/cirurgia , Procedimentos Cirúrgicos em Ginecologia/métodos , Procedimentos Cirúrgicos em Ginecologia/efeitos adversos , Laparoscopia/efeitos adversos , Laparoscopia/métodos , Procedimentos Cirúrgicos Minimamente Invasivos/efeitos adversos , Procedimentos Cirúrgicos Minimamente Invasivos/métodos , Morbidade , Complicações Pós-Operatórias/etiologia , Prognóstico , Procedimentos Cirúrgicos Robóticos/efeitos adversos , Procedimentos Cirúrgicos Robóticos/métodosRESUMO
Signaling of G protein-activated inwardly rectifying K+ (GIRK) channels is an important mechanism of the parasympathetic regulation of the heart rate and cardiac excitability. GIRK channels are inhibited during stimulation of Gq-coupled receptors (GqPCRs) by depletion of phosphatidyl-4,5-bisphosphate (PIP2) and/or channel phosphorylation by protein kinase C (PKC). The GqPCR-dependent modulation of GIRK currents in terms of specific PKC isoform activation was analyzed in voltage-clamp experiments in rat atrial myocytes and in CHO or HEK 293 cells. By using specific PKC inhibitors, we identified the receptor-activated PKC isoforms that contribute to phenylephrine- and angiotensin-induced GIRK channel inhibition. We demonstrate that the cPKC isoform PKCα significantly contributes to GIRK inhibition during stimulation of wildtype α1B-adrenergic receptors (α1B-ARs). Deletion of the α1B-AR serine residues S396 and S400 results in a preferential regulation of GIRK activity by PKCß. As a novel finding, we report that the AT1-receptor-induced GIRK inhibition depends on the activation of the nPKC isoform PKCε whereas PKCα and PKCß do not mainly participate in the angiotensin-mediated GIRK reduction. Expression of the dominant negative (DN) PKCε prolonged the onset of GIRK inhibition and significantly reduced AT1-R desensitization, indicating that PKCε regulates both GIRK channel activity and the strength of the receptor signal via a negative feedback mechanism. The serine residue S418 represents an important phosphorylation site for PKCε in the GIRK4 subunit. To analyze the functional impact of this PKC phosphorylation site for receptor-specific GIRK channel modulation, we monitored the activity of a phosphorylation-deficient (GIRK4 (S418A)) GIRK4 channel mutant during stimulation of α1B-ARs or AT1-receptors. Mutation of S418 did not impede α1B-AR-mediated GIRK inhibition, suggesting that S418 within the GIRK4 subunit is not subject to PKCα-induced phosphorylation. Furthermore, activation of angiotensin receptors induced pronounced GIRK4 (S418A) channel inhibition, excluding that this phosphorylation site contributes to the AT1-R-induced GIRK reduction. Instead, phosphorylation of S418 has a facilitative effect on GIRK activity that was abolished in the GIRK4 (S418A) mutant. To summarize, the present study shows that the receptor-dependent regulation of atrial GIRK channels is attributed to the GqPCR-specific activation of different PKC isoforms. Receptor-specific activated PKC isoforms target distinct phosphorylation sites within the GIRK4 subunit, resulting in differential regulation of GIRK channel activity with either facilitative or inhibitory effects on GIRK currents.
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Cricetulus , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G , Proteína Quinase C , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Animais , Fosforilação , Células HEK293 , Humanos , Ratos , Proteína Quinase C/metabolismo , Células CHO , Receptores Adrenérgicos alfa 1/metabolismo , Miócitos Cardíacos/metabolismo , Masculino , Ratos Wistar , Proteína Quinase C-alfa/metabolismo , Isoenzimas/metabolismoRESUMO
Around 60% of individuals with neurodevelopmental disorders (NDD) remain undiagnosed after comprehensive genetic testing, primarily of protein-coding genes1. Increasingly, large genome-sequenced cohorts are improving our ability to discover new diagnoses in the non-coding genome. Here, we identify the non-coding RNA RNU4-2 as a novel syndromic NDD gene. RNU4-2 encodes the U4 small nuclear RNA (snRNA), which is a critical component of the U4/U6.U5 tri-snRNP complex of the major spliceosome2. We identify an 18 bp region of RNU4-2 mapping to two structural elements in the U4/U6 snRNA duplex (the T-loop and Stem III) that is severely depleted of variation in the general population, but in which we identify heterozygous variants in 119 individuals with NDD. The vast majority of individuals (77.3%) have the same highly recurrent single base-pair insertion (n.64_65insT). We estimate that variants in this region explain 0.41% of individuals with NDD. We demonstrate that RNU4-2 is highly expressed in the developing human brain, in contrast to its contiguous counterpart RNU4-1 and other U4 homologs, supporting RNU4-2's role as the primary U4 transcript in the brain. Overall, this work underscores the importance of non-coding genes in rare disorders. It will provide a diagnosis to thousands of individuals with NDD worldwide and pave the way for the development of effective treatments for these individuals.
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Background and purpose: Image-guided adapted brachytherapy (IGABT) is superior to other radiotherapy techniques in the treatment of locally advanced cervical cancer (LACC). We aimed to investigate the benefit of interstitial needles (IN) for a combined intracavitary/interstitial (IC/IS) approach using IGABT over the intracavitary approach (IC) alone in patients with LACC after concomitant external beam radiotherapy (EBRT) and chemotherapy. Materials and methods: We included consecutive patients with LACC who were treated with IC/IS IGABT after radiochemotherapy (RCT) in our retrospective, observational study. Dosimetric gain and sparing of organs at risk (OAR) were investigated by comparing the IC/IS IGABT plan with a simulated plan without needle use (IC IGABT plan) and the impact of other clinical factors on the benefit of IC/IS IGABT. Results: Ninety-nine patients were analyzed, with a mean EBRT dose of 45.5 ± 1.7 Gy; 97 patients received concurrent chemotherapy. A significant increase in median D90% High Risk Clinical target volume (HR-CTV) was found for IC/IS (82.8 Gy) vs IC (76.2 Gy) (p < 10-4). A significant decrease of the delivered dose for all OAR was found for IC/IS vs IC for median D2cc to the bladder (77.2 Gy), rectum (68 Gy), sigmoid (53.2 Gy), and small bowel (47 Gy) (all p < 10-4). Conclusion: HR-CTV coverage was higher with IC/IS IGABT than with IC IGABT, with lower doses to the OAR in patients managed for LACC after RCT. Interstitial brachytherapy in the management of LACC after radiotherapy provides better coverage of the target volumes, this could contribute to better local control and improved survival of patients.