Routing of Kv7.1 to endoplasmic reticulum plasma membrane junctions.

AIM: The voltage-gated Kv7.1 channel, in association with the regulatory subunit KCNE1, contributes to the IKs current in the heart. However, both proteins travel to the plasma membrane using different routes. While KCNE1 follows a classical Golgi-mediated anterograde pathway, Kv7.1 is located in endoplasmic reticulum-plasma membrane junctions (ER-PMjs), where it associates with KCNE1 before being delivered to the plasma membrane. METHODS: To characterize the channel routing to these spots we used a wide repertoire of methodologies, such as protein expression analysis (i.e. protein association and biotin labeling), confocal (i.e. immunocytochemistry, FRET, and FRAP), and dSTORM microscopy, transmission electron microscopy, proteomics, and electrophysiology. RESULTS: We demonstrated that Kv7.1 targeted ER-PMjs regardless of the origin or architecture of these structures. Kv2.1, a neuronal channel that also contributes to a cardiac action potential, and JPHs, involved in cardiac dyads, increased the number of ER-PMjs in nonexcitable cells, driving and increasing the level of Kv7.1 at the cell surface. Both ER-PMj inducers influenced channel function and dynamics, suggesting that different protein structures are formed. Although exhibiting no physical interaction, Kv7.1 resided in more condensed clusters (ring-shaped) with Kv2.1 than with JPH4. Moreover, we found that VAMPs and AMIGO, which are Kv2.1 ancillary proteins also associated with Kv7.1. Specially, VAP B, showed higher interaction with the channel when ER-PMjs were stimulated by Kv2.1. CONCLUSION: Our results indicated that Kv7.1 may bind to different structures of ER-PMjs that are induced by different mechanisms. This variable architecture can differentially affect the fate of cardiac Kv7.1 channels.

Implementation of the Baby-Friendly Hospital Initiative in Mexico: a systematic literature review using the RE-AIM framework.

The Baby-Friendly Hospital Initiative (BFHI) is a global strategy to encourage health facilities to promote, support, and protect breastfeeding by implementing a package of policies and practices known as the Ten Steps to Successful Breastfeeding. Prior studies have found that implementing the Ten Steps has a positive impact on breastfeeding outcomes. Yet, little is known about the implementation of the Ten Steps in Mexico. The objective of this study was to conduct a systematic review to evaluate the reach, efficacy/effectiveness, adoption, implementation, and maintenance of the Ten Steps in Mexico, using the RE-AIM framework. The systematic literature review included studies published in English or Spanish without date restrictions. Two of the authors coded each of the articles through a harmonized data extraction tool, and group meetings were used to discuss any discrepancies. The reviewed data were managed in the Rayyan platform. The risk of study bias was assessed through the Johanna Briggs Institute critical appraisal checklists. Of the 1,123 articles initially identified, 6 met the review inclusion criteria. None of the articles evaluated the reach and maintenance of the Ten Steps. The articles identified major gaps in the implementation of the Ten Steps. Most of the articles had important limitations in terms of their quality. In Mexico, it is necessary to rethink the BFHI and employ multiple strategies to improve implementation of the Ten Steps, including developing transparent BFHI monitoring mechanisms that produce data on implementation and that are publicly available, as well as investing in implementation research and evaluation to generate strong evidence to support the adoption and efficient maintenance of the Ten Steps in health facilities in Mexico. When properly implemented, BFHI becomes central to promote, protect, and support breastfeeding. Therefore, it is essential for Mexico to position BFHI as a top priority of the country's public policy agenda. Systematic Review Registration: identifier: CRD42021248118.

Sigma-1 receptor modulation fine-tunes KV1.5 channels and impacts pulmonary vascular function.

KV1.5 channels are key players in the regulation of vascular tone and atrial excitability and their impairment is associated with cardiovascular diseases including pulmonary arterial hypertension (PAH) and atrial fibrillation (AF). Unfortunately, pharmacological strategies to improve KV1.5 channel function are missing. Herein, we aimed to study whether the chaperone sigma-1 receptor (S1R) is able to regulate these channels and represent a new strategy to enhance their function. By using different electrophysiological and molecular techniques in X. laevis oocytes and HEK293 cells, we demonstrate that S1R physically interacts with KV1.5 channels and regulate their expression and function. S1R induced a bimodal regulation of KV1.5 channel expression/activity, increasing it at low concentrations and decreasing it at high concentrations. Of note, S1R agonists (PRE084 and SKF10047) increased, whereas the S1R antagonist BD1047 decreased, KV1.5 expression and activity. Moreover, PRE084 markedly increased KV1.5 currents in pulmonary artery smooth muscle cells and attenuated vasoconstriction and proliferation in pulmonary arteries. We also show that both KV1.5 channels and S1R, at mRNA and protein levels, are clearly downregulated in samples from PAH and AF patients. Moreover, the expression of both genes showed a positive correlation. Finally, the ability of PRE084 to increase KV1.5 function was preserved under sustained hypoxic conditions, as an in vitro PAH model. Our study provides insight into the key role of S1R in modulating the expression and activity of KV1.5 channels and highlights the potential role of this chaperone as a novel pharmacological target for pathological conditions associated with KV1.5 channel dysfunction.

Resolvin D2 Attenuates Cardiovascular Damage in Angiotensin II-Induced Hypertension.

BACKGROUND: Resolution of inflammation is orchestrated by specialized proresolving lipid mediators (SPMs), and this would be impaired in some cardiovascular diseases. Among SPMs, resolvins (Rv) have beneficial effects in cardiovascular pathologies, but little is known about their effect on cardiovascular damage in hypertension. METHODS: Aorta, small mesenteric arteries, heart, and peritoneal macrophages were taken from C57BL/6J mice, infused or not with angiotensin II (AngII; 1.44 mg/kg/day, 14 days) in presence or absence of resolvin D2 (RvD2) (100 ng/mice, every second day) starting 1 day before or 7 days after AngII infusion. RESULTS: Enzymes and receptors involved in SPMs biosynthesis and signaling were increased in aorta or heart from AngII-infused mice. We also observed a differential regulation of SPMs in heart from these mice. Preventive treatment with RvD2 partially avoided AngII-induced hypertension and protected the heart and large and small vessels against functional and structural alterations induced by AngII. RvD2 increased the availability of vasoprotective factors, modified SPMs profile, decreased cardiovascular fibrosis, and increased the infiltration of pro-resolving macrophages. When administered in hypertensive animals with established cardiovascular damage, RvD2 partially improved cardiovascular function and structure, decreased fibrosis, reduced the infiltration of neutrophils, and shifted macrophage phenotype toward a pro-resolving phenotype. CONCLUSIONS: There is a disbalance between proinflammatory and resolution mediators in hypertension. RvD2 protects cardiovascular function and structure when administered before and after the development of hypertension by modulating vascular factors, fibrosis and inflammation. Activating resolution mechanisms by treatment with RvD2 may represent a novel therapeutic strategy for the treatment of hypertensive cardiovascular disease.

Síndrome de Frasier: ¿Cómo diagnosticar una enfermedad rara en la consulta de Ginecología? Reporte de caso / Frasier Syndrome: How to diagnose a rare disease in a Gynecology consultation? A case report

Resumen INTRODUCCIÓN: El síndrome de Frasier es una enfermedad de herencia autosómica dominante con una prevalencia inferior a 1 caso por cada millón de recién nacidos vivos. Hasta la fecha se han descrito unos 150 casos. Este síndrome se caracteriza por pseudohermafroditismo masculino, disgenesia gonadal 46, XY y enfermedad glomerular, todo ello producido por una mutación del gen WT1. CASO CLÍNICO: Paciente de 16 años que consultó por amenorrea primaria y ausencia de caracteres sexuales secundarios. Antecedentes: glomerulonefritis focal segmentaria corticorresistente desde la infancia. En la exploración física se objetivó un estadio Tanner 1. Las pruebas complementarias pusieron de manifiesto la ausencia de útero y anejos y un hipogonadismo hipergonadotrópico con cariotipo 46, XY. Ante los hallazgos se decidió la laparoscopia exploradora y salpingooforectomía bilateral. El informe anatomopatológico fue de disgerminoma de ovario derecho. La sospecha clínica se confirmó en el estudio genético, que reportó una mutación del gen WT1, diagnóstica de síndrome de Frasier. En la actualidad, la paciente recibe tratamiento inmunosupresor y hormonal sustitutivo, con una evolución favorable. CONCLUSIÓN: El diagnóstico temprano del síndrome de Frasier es fundamental en virtud del riesgo asociado de malignidad. La baja frecuencia de la enfermedad y la asociación común con retraso puberal en pacientes con enfermedades crónicas puede favorecer el retraso del diagnóstico. El reporte de los casos diagnosticados de este síndrome, y el tratamiento multidisciplinario son decisivos para mejorar el conocimiento de esta rara enfermedad.

Abstract INTRODUCTION: Frasier Syndrome is an autosomal dominant inherited disease with a prevalence of less than 1 per million live births. To date, about 150 cases have been described. This syndrome is characterized by male pseudohermaphroditism, 46, XY gonadal dysgenesis, and glomerular disease, all caused by a mutation of the WT1 gene. It is essential to learn more about this disease, not only because of the high risk of ovarian neoplasia, but also because its early diagnosis will improve the prognosis. CLINICAL CASES: We report the case of a 16-year-old woman who consulted for primary amenorrhea and absence of secondary sexual characteristics. As medical history, she highlighted steroid-resistant focal segmental glomerulonephritis since childhood. The examination revealed Tanner stage 1. Complementary tests revealed the absence of the uterus and adnexa and hypergonadotropic hypogonadism with a 46, XY karyotype. Given the findings, it was decided to perform an exploratory laparoscopy and bilateral salpingo-oophorectomy. The anatomopathological result reported dysgerminoma of the right ovary. The clinical suspicion was confirmed by genetic study, which reported a mutation of the WT1 gene, diagnostic of Frasier Syndrome. Currently, the patient undergoes, along with immunosuppressive treatment, hormone replacement therapy, with a favorable evolution. CONCLUSION: Early diagnosis of Frasier Syndrome is essential given the associated risk of malignancy. The low frequency of the disease and the usual association of delayed puberty in patients with chronic diseases may lead to a diagnostic delay. Therefore, reporting the diagnosed cases of this syndrome, as well as its multidisciplinary management, is essential to improve knowledge about this rare disease.

Modulation of KV4.3-KChIP2 Channels by IQM-266: Role of DPP6 and KCNE2.

The transient outward potassium current (Itof) is generated by the activation of KV4 channels assembled with KChIP2 and other accessory subunits (DPP6 and KCNE2). To test the hypothesis that these subunits modify the channel pharmacology, we analyzed the electrophysiological effects of (3-(2-(3-phenoxyphenyl)acetamido)-2-naphthoic acid) (IQM-266), a new KChIP2 ligand, on the currents generated by KV4.3/KChIP2, KV4.3/KChIP2/DPP6 and KV4.3/KChIP2/KCNE2 channels. CHO cells were transiently transfected with cDNAs codifying for different proteins (KV4.3/KChIP2, KV4.3/KChIP2/DPP6 or KV4.3/KChIP2/KCNE2), and the potassium currents were recorded using the whole-cell patch-clamp technique. IQM-266 decreased the maximum peak of KV4.3/KChIP2, KV4.3/KChIP2/DPP6 and KV4.3/KChIP2/KCNE2 currents, slowing their time course of inactivation in a concentration-, voltage-, time- and use-dependent manner. IQM-266 produced an increase in the charge in KV4.3/KChIP2 channels that was intensified when DPP6 was present and abolished in the presence of KCNE2. IQM-266 induced an activation unblocking effect during the application of trains of pulses to cells expressing KV4.3/KChIP2 and KV4.3/KChIP2/KCNE2, but not in KV4.3/KChIP2/DPP6 channels. Overall, all these results are consistent with a preferential IQM-266 binding to an active closed state of Kv4.3/KChIP2 and Kv4.3/KChIP2/KCNE2 channels, whereas in the presence of DPP6, IQM-266 binds preferentially to an inactivated state. In conclusion, DPP6 and KCNE2 modify the pharmacological response of KV4.3/KChIP2 channels to IQM-266.

Pharmacological Approaches for the Modulation of the Potassium Channel KV4.x and KChIPs.

Ion channels are macromolecular complexes present in the plasma membrane and intracellular organelles of cells. Dysfunction of ion channels results in a group of disorders named channelopathies, which represent an extraordinary challenge for study and treatment. In this review, we will focus on voltage-gated potassium channels (KV), specifically on the KV4-family. The activation of these channels generates outward currents operating at subthreshold membrane potentials as recorded from myocardial cells (ITO, transient outward current) and from the somata of hippocampal neurons (ISA). In the heart, KV4 dysfunctions are related to Brugada syndrome, atrial fibrillation, hypertrophy, and heart failure. In hippocampus, KV4.x channelopathies are linked to schizophrenia, epilepsy, and Alzheimer's disease. KV4.x channels need to assemble with other accessory subunits (ß) to fully reproduce the ITO and ISA currents. ß Subunits affect channel gating and/or the traffic to the plasma membrane, and their dysfunctions may influence channel pharmacology. Among KV4 regulatory subunits, this review aims to analyze the KV4/KChIPs interaction and the effect of small molecule KChIP ligands in the A-type currents generated by the modulation of the KV4/KChIP channel complex. Knowledge gained from structural and functional studies using activators or inhibitors of the potassium current mediated by KV4/KChIPs will better help understand the underlying mechanism involving KV4-mediated-channelopathies, establishing the foundations for drug discovery, and hence their treatments.

KV1.5-KVß1.3 Recycling Is PKC-Dependent.

KV1.5 channel function is modified by different regulatory subunits. KVß1.3 subunits assemble with KV1.5 channels and induce a fast and incomplete inactivation. Inhibition of PKC abolishes the KVß1.3-induced fast inactivation, decreases the amplitude of the current KV1.5-KVß1.3 and modifies their pharmacology likely due to changes in the traffic of KV1.5-KVß1.3 channels in a PKC-dependent manner. In order to analyze this hypothesis, HEK293 cells were transfected with KV1.5-KVß1.3 channels, and currents were recorded by whole-cell configuration of the patch-clamp technique. The presence of KV1.5 in the membrane was analyzed by biotinylation techniques, live cell imaging and confocal microscopy approaches. PKC inhibition resulted in a decrease of 33 ± 7% of channels in the cell surface due to reduced recycling to the plasma membrane, as was confirmed by confocal microscopy. Live cell imaging indicated that PKC inhibition almost abolished the recycling of the KV1.5-KVß1.3 channels, generating an accumulation of channels into the cytoplasm. All these results suggest that the trafficking regulation of KV1.5-KVß1.3 channels is dependent on phosphorylation by PKC and, therefore, they could represent a clinically relevant issue, mainly in those diseases that exhibit modifications in PKC activity.

Genomic path to pandrug resistance in a clinical isolate of Klebsiella pneumoniae.

Carbapenem-resistant Klebsiella pneumoniae have spread globally throughout tertiary hospitals. Many Carbapenem-resistant K. pneumoniae clinical isolates are multidrug-resistant (MDR) and may become eventually pandrug-resistant (PDR). Here we present the closed genome of a PDR VIM-1-producing K. pneumoniae strain (KP1050) obtained in a tertiary hospital. The isolate belonged to sequence type 54 (ST54) and had five extrachromosomal elements (four plasmids and a circular phage genome). Most of the antimicrobial resistance genes (ARGs) were located in two clusters borne by two of the plasmids, comprising a class 1 integron that contained up to 14 ARGs including a VIM-1 metallo-ß-lactamase gene, and an IS26 transposon that contained a mobile element from Acinetobacter baumannii encoding the amikacin resistance gene aac(6')-Ian. A MDR isolate obtained 6 years previously was identified (KP1048) retrospectively and was sequenced. Comparison of the two genomes showed that chromosomal mutations in outer membrane porins as well as mutations in the ramR and phoQ genes contributed to increase the resistance spectrum.

Litiasis biliar complicada tratada satisfactoriamente mediante colangiografía percutánea y drenaje biliar en lactante de 2 meses / Complicated bile duct stones successfully treated by percutaneous cholangiography and biliary drainage catheter in a 2-month-old infant

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Usefulness of clinical, ultrasonographic, hysteroscopic, and immunohistochemical parameters in differentiating endometrial polyps from endometrial cancer.

STUDY OBJECTIVE: To evaluate the usefulness of clinical, ultrasonographic, hysteroscopic, and immunohistochemical parameters in differentiating endometrial polyps from endometrial cancer. DESIGN: Cross-sectional study (Canadian Task Force classification II-2). SETTING: Tertiary public hospital, university teaching center. PATIENTS: Eighty-two women who underwent hysteroscopic polypectomy and 20 women who underwent surgery to treat endometrial cancer. INTERVENTIONS: Analysis of medical records and immunohistochemical assessment of estrogen receptors, progesterone receptors, and endothelial markers CD34 and CD105. MEASUREMENTS AND MAIN RESULTS: Among women with endometrial cancer and endometrial polyps, respectively, mean age was 63 and 57 years (p = .01), 89% and 67% were postmenopausal (p < .05), and 85% and 30.5% had postmenopausal bleeding (p < .01). No sonographic parameter enabled differentiation of endometrial polyp from cancer. Of patients with endometrial cancer, 72% exhibited signs suggestive of hyperplasia, and endometrial polyps were diagnosed during hysteroscopy. Estrogen receptors (≥ 2 vs ≥ 1; p < .001) and progesterone receptors (≥ 3 vs ≥ 2; p = .07) were greater in endometrial polyps. There was no significant difference in microvessel density (p > .05). CONCLUSIONS: Ultrasonographic parameters and endothelial markers did not enable differentiation of polyps from endometrial neoplasia. Postmenopausal bleeding and endometrial hypervascularization along with vascular atypia at diagnostic hysteroscopy showed a greater association with endometrial cancer.

Fine needle aspirate cell blocks are reliable for detection of hormone receptors and HER-2 by immunohistochemistry in breast carcinoma.

AIMS: To evaluate the reliability of fine needle aspirate cell blocks in the assessment of oestrogen receptor (ER), progesterone receptor (PR) and HER-2/neu proteins by immunohistochemistry in comparison with surgical specimens. MATERIALS AND METHODS: This is a retrospective study of 62 cases of breast carcinoma diagnosed by fine needle aspiration cytology (FNAC) and confirmed using the surgical specimen. Immunohistochemical tests were performed to assess the presence of oestrogen receptor (ER), progesterone receptor (PR) and HER-2/neu proteins in cell blocks and the corresponding surgical specimens. The cell block method used alcohol prior to formalin fixation. Cases with 10% or more stained cells were considered positive for ER and PR. Positivity for HER-2/neu was assessed on a scale of 0-3+. The criterion for positivity was a score of 3+. RESULTS: Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of the cell blocks in the investigation of ER, PR and HER-2/neu protein (3+) were (%): ER, 92.7, 85.7, 92.7, 85.7 and 90.3; PR, 92.7, 94.7, 97.4, 87.0 and 93.5; HER-2/neu, 70.0, 100.0, 100.0, 94.5 and 95.2. Discrepancies were seen in cell blocks in the 1+ and 2+ HER-2/neu staining scores: two of 12 cases scoring 2+ and one case of 26 scoring 1+ on cell blocks scored 3+ on surgical specimens. The correlation index between cell block and corresponding surgical specimen varied from 90% to 94%. CONCLUSION: Cell blocks provide a useful method of assessing ER, PR and HER-2/neu, mainly for inoperable and recurrent cases, but consideration should be given to carrying out FISH analysis on 1+ as well as 2+ HER-2/neu results.

Ventriculectomia parcial esquerda: ponte para transplante? / Partial left ventriculectomy: bridge to transplantation?

OBJETIVO: Analisar os resultados e a viabilidade da ventriculectomia parcial esquerda (VPE) como ponte para transplante cardíaco (TX). DELINEAMENTO: Estudo de coorte histórica e prospectivo. CASUISTICA E MÉTODOS: Cinquenta e três pacientes (pts) foram submetidos a VPE em um período de 5 anos. Destes, 7 pts com contra-indicação inicial ao TX, idades variando de 37 a 64 anos, 5 homens e 2 mulheres, com miocardiopatia dilatada, foram subseqüentemente relistados e transplantados. Foram analisados a fração de ejeção (FE), o diâmetro diastólico final do ventrículo esquerdo (DDFVE), a CF da NYHA, o consumo máximo de oxigênio (VO2 máx) e os escores de qualidade de vida (QV) antes da VPE, aos 3 e 6 meses, e pré-transplante...

Cirurgia cardíaca em pacientes com insuficiência renal crônica em hemodiálise / Heart surgery in patients with chronic renal failure in haemodyalisis

A principal causa de óbito dos pacientes renais crônicos é de origem cardiovascular e muitos necessitam cirurgia cardíaca para correçäo de suas patologias. O presente trabalho apresenta os resultados de um grupo de pacientes portadores de insuficiência renal crônica, em programa de hemodiálise, submetidos a cirurgia cardíaca. Foram estudados retrospectivamente 9 pacientes operados com circulaçäo extracorpórea no Hospital Säo Francisco-Santa Casa de Misericórdia de Porto Alegre, RS, no período de 1990/96...

Doença de Chagas: revisäo de 8 casos simultâneos de miocardite aguda chagásica; 25 anos após / Chagas disease: review of 8 simultaneous cases of acute Chagas myocarditis; 25 years after

Objetivo - Relatar a evoluçäo de pacientes após 25 anos após uma miciroepidemia de doença de Chagas, surgida em Teotônia-RS em 1965 e discutir a importância da transnissäo via oral (VO) como porta de entrada. Métodos - Oito pacientes de um grupo de 17 pessoas acometidas de infecçäo chagásica aguda, em 1965, estäo fichadas no IC/FUC do RS com diagnóstico de doença de Chagas. Foram examinadas clinicamente e se submeteram a exames laboratoriais de rotina, provas específicas para o Trypanozoma cruzi, eletrocardiograma (ECG), radiografia de tórax (RX) e ecocardiograma (ECO) em 1991. Resultados - Nenhum dos 8 pacientes apresentou manifestaçöes orgânicas da doença de Chagas, os exames laboratoriais, ECG, RX e ECO foram normais. Na imunofluorescência somente um paciente apresentou indice 1/20. Conclusäo - 1) A evoluçäo dos oito casos nestes 25 anos foi benigna, näo apresentando nenhum sinal ou manifestaçäo orgânica de doença de Chagas e, especificamente, sem miocardiopatia; 2) a transmissäo VO deve ser considerada definitivamente como porta de entrada para a doença de Chagas e, provavelmente, influi na evoluçäo da fase crônica

Purpose - To analyse the evolution of 8 cases of Chagas' disease after a microepidemy of 17 cases which ocurred in Teotonia - RS - in 1965 and to discuss the importance of oral contamination in Chagas' disease. Methods - All 8 patients are followed at the IC/ FUC - RS - Brazil since 1980. Clinical examinations, blood tests, ECG, Cx-Ray and Echo were made in 1991. Results - None of the eight cases presented any sign of chronic Chagas' disease manifestation. The blood tests, ECG, Cx-Ray and Echo were normais. Only one case showed a imunofluorescence of 1/20. Conclusion - 1) The follow-up after 25 years in all the 8 cases did not show clinical alterations or any chronic manifestation of Chagas' disease. All the tests performed had normal results; 2) the oral transmission must be considered definitive in Chagas' disease