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Objective: The objective of this study was to reveal the influence of acute and chronic heat stress (HS) on the abundance and function of rumen microbiome and host metabolism. Methods: The forty mid-lactation goats were randomly divided into two artificial environments a control group and a heat-stressed group. This study was collected from two periods, 1 day and 28 days. The first day was defined as control 1 (CT1) and HS 1 (acute HS), and the last day was defined as CT28 and HS28 (chronic HS). On the first and last day, 6 dairy goats in each group were randomly selected to collect rumen liquid after the morning feeding through oral stomach tubes. The barn temperature and humidity were recorded every day. Results: Disruption of the rumen microbiome was observed under chronic HS, represented by an increase in the abundance of Prevotella and Bacteroidales (p<0.05), and upregulation of carbohydrate transport and metabolism functions (p<0.05). Additionally, the abundance of Succinimonas and Ruminobacter in chronic HS is lower than in acute HS (p<0.05), and the functions of intracellular trafficking, secretion and vesicular transport, and the cytoskeleton were downregulated (p<0.05). Conclusion: HS affected the interaction between the microbiota and host, thereby regulating milk production in dairy goats. These findings increased understanding of the crosstalk between hosts and microorganisms.
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Ficus concinna is an important contributor to tropical forest biodiversity. Here, we provide the first report of the complete F. concinna chloroplast genome, thereby providing a basis for subsequent phylogenetic analyses of the Moraceae family. The final assembled chloroplast genome was 160,331 bp in size and included a 20,018 bp long small single-copy (SSC) region, an 88,541 bp long large single-copy (LSC) region, and two 25,886 bp inverted repeats (IRs). The total content of GC for this chloroplast genome was 42.6%, with respective frequencies of 33.6%, 35.9%, and 28.9% in the SSC, LSC, and IR regions, accordingly. Overall, the chloroplast genome was found to encode 131 genes, comprising 37 tRNAs, 86 protein-coding genes, and 8 rRNAs. Phylogenetic analyses revealed a close relationship between F. concinna and Ficus altissima, consistent with prior research results. Together, these data provide valuable insights regarding the evolution and conserved genetic features of F. concinna.
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Cardiac alternans closely linked to calcium dysregulation is a crucial risk factor for fatal arrhythmia causing especially sudden death. Calcium overload is well-known to activate Ca2+-dependent protein kinase C (PKC); however, the effects of PKC on arrhythmogenic cardiac alternans have not yet been investigated. This study aimed to determine the contributions of PKC activities in cardiac alternans associated with calcium cycling disturbances. In the present study, action potential duration alternans (APD-ALT) induced by high free intracellular calcium ([Ca2+]i) exerted not only in a calcium concentration-dependent manner but also in a frequency-dependent manner. High [Ca2+]i-induced APD-ALT was suppressed by not only BAPTA-AM but also nifedipine. On the other hand, PKC inhibitors BIM and Gö 6976 eliminated high [Ca2+]i-induced APD-ALT, and PKC activator PMA was found to induce APD-ALT at normal [Ca2+]i condition. Furthermore, BIM effectively prevented calcium transient alternans (CaT-ALT) and even CaT disorders caused by calcium overload. Moreover, BIM not only eliminated electrocardiographic T-wave alternans (TWA) caused by calcium dysregulation, but also lowered the incidence of ventricular arrhythmias in isolated hearts. What's more, BIM prevented the expression of PKC α upregulated by calcium overload in high calcium-perfused hearts. We firstly found that pharmacologically inhibiting Ca2+-dependent PKC over-activation suppressed high [Ca2+]i-induced cardiac alternans. This recognition indicates that inhibition of PKC activities may become a therapeutic target for the prevention of pro-arrhythmogenic cardiac alternans associated with calcium dysregulation.
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Arritmias Cardíacas/etiología , Calcio/metabolismo , Miocitos Cardíacos/fisiología , Proteína Quinasa C/metabolismo , Potenciales de Acción , Animales , Arritmias Cardíacas/enzimología , Arritmias Cardíacas/prevención & control , Sistema de Conducción Cardíaco/fisiopatología , Terapia Molecular Dirigida , Técnicas de Placa-Clamp , Cultivo Primario de Células , Proteína Quinasa C/antagonistas & inhibidores , Proteínas Quinasas/metabolismo , ConejosRESUMEN
Eleutheroside B (EB) is the main active constituent derived from the Chinese herb Acanthopanax senticosus (AS) that has been reported to possess cardioprotective effects. In this study we investigated the effects of EB on cardiac electrophysiology and its suppression on atrial fibrillation (AF). Whole-cell recording was conducted in isolated rabbit atrial myocytes. The intracellular calcium ([Ca2+]i) concentration was measured using calcium indicator Fura-2/AM fluorescence. Monophasic action potential (MAP) and electrocardiogram (ECG) synchronous recordings were conducted in Langendorff-perfused rabbit hearts using ECG signal sampling and analysis system. We showed that EB dose-dependently inhibited late sodium current (INaL), transient sodium current (INaT), and sea anemone toxin II (ATX II)-increased INaL with IC50 values of 167, 1582, and 181 µM, respectively. On the other hand, EB (800 µM) did not affect L-type calcium current (ICaL), inward rectifier potassium channel current (IK), and action potential duration (APD). Furthermore, EB (300 µM) markedly decreased ATX II-prolonged the APD at 90% repolarization (APD90) and eliminated ATX II-induced early afterdepolarizations (EADs), delayed afterdepolarizations (DADs), and triggered activities (TAs). Moreover, EB (200 µM) significantly suppressed ATX II-induced Na+-dependent [Ca2+]i overload in atrial myocytes. In the Langendorff-perfused rabbit hearts, application of EB (200 µM) or TTX (2 µM) substantially decreased ATX II-induced incidences of atrial fibrillation (AF), ventricular fibrillation (VF), and heart death. These results suggest that augmented INaL alone is sufficient to induce AF, and EB exerts anti-AF actions mainly via blocking INaL, which put forward the basis of pharmacology for new clinical application of EB.
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Fibrilación Atrial/prevención & control , Cardiotónicos/farmacología , Glucósidos/farmacología , Miocitos Cardíacos/efectos de los fármacos , Fenilpropionatos/farmacología , Potenciales de Acción/efectos de los fármacos , Animales , Calcio/metabolismo , Cardiotónicos/administración & dosificación , Venenos de Cnidarios/toxicidad , Relación Dosis-Respuesta a Droga , Electrocardiografía , Glucósidos/administración & dosificación , Miocitos Cardíacos/metabolismo , Técnicas de Placa-Clamp , Fenilpropionatos/administración & dosificación , Conejos , Bloqueadores de los Canales de Sodio/administración & dosificación , Bloqueadores de los Canales de Sodio/farmacologíaRESUMEN
Increasing evidence shows that Curcumin (Cur) has a protective effect against cardiovascular diseases. However, the role of Cur in the electrophysiology of cardiomyocytes is currently not entirely understood. Therefore, the present study was conducted to investigate the effects of Cur on the action potential and transmembrane ion currents in rabbit ventricular myocytes to explore its antiarrhythmic property. The whole-cell patch clamp was used to record the action potential and ion currents, while the multichannel acquisition and analysis system was used to synchronously record the electrocardiogram and monophasic action potential. The results showed that 30 µmol/L Cur shortened the 50 and 90% repolarization of action potential by 17 and 7%, respectively. In addition, Cur concentration dependently inhibited the Late-sodium current (I Na.L), Transient-sodium current (I Na.T), L-type calcium current (I Ca.L), and Rapidly delayed rectifying potassium current (I Kr), with IC50 values of 7.53, 398.88, 16.66, and 9.96 µmol/L, respectively. Importantly, the inhibitory effect of Cur on I Na.L was 52.97-fold higher than that of I Na.T. Moreover, Cur decreased ATX II-prolonged APD, suppressed the ATX II-induced early afterdepolarization (EAD) and Ca2+-induced delayed afterdepolarization (DAD) in ventricular myocytes, and reduced the occurrence and average duration of ventricular tachycardias and ventricular fibrillations induced by ischemia-reperfusion injury. In conclusion, Cur inhibited I Na.L, I Na.T, I Ca.L, and I Kr; shortened APD; significantly suppressed EAD and DAD-like arrhythmogenic activities at the cellular level; and exhibited antiarrhythmic effect at the organ level. It is first revealed that Cur is a multi-ion channel blocker that preferentially blocks I Na.L and may have potential antiarrhythmic property.
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PURPOSE: Inflammation is closely associated with prognosis in gastric cancer (GC). We aimed to assess the predictive value of existing inflammatory and tumor markers in GC, to establish a systemic score based on valuable predictors for early risk stratification of patients, and to create a nomogram for individual risk prediction. PATIENTS AND METHODS: We retrospectively analyzed 401 GC patients who underwent curative gastrectomy from 2007 to 2016. RESULTS: Through univariate and multivariate survival analysis, age (>60 years), depth of invasion (pT3-4), lymph node invasion (pN1-3), histologic classification (poor), adjuvant chemotherapy (no), albumin fibrinogen ratio (AFR) (<13.33), and carbohydrate antigen 19-9 (CA19-9) (>27 U/mL) independently indicated inferior disease-free survival (DFS). In addition, depth of invasion, lymph node invasion, histologic classification, adjuvant chemotherapy, AFR, and CA19-9 were incorporated in the prediction of cancer-specific survival (CSS). A combined AFR and CA19-9 prognostic score (CACPS) was established. Lower AFR (<13.33) and higher CA19-9 (>27 U/mL) were allocated 1 point each in the CACPS (range, 0-2). CACPS can be used as an independent predictor for DFS and CSS in multivariate analysis (for DFS: CACPS 1: HR=2.039, 95% CI: 1.357-3.065, P=0.001; CACPS 2: HR=2.419, 95% CI: 1.397-4.186, P=0.002; for CSS: CACPS 1: HR=2.035, 95% CI: 1.292-3.205, P=0.002; CACPS 2: HR=2.255, 95% CI: 1.252-4.059, P=0.007), with a higher CACPS indicating poor survival according to Kaplan-Meier curves (both P<0.001). Moreover, a nomogram for DFS and CSS was generated using the significant characteristics in the multivariate analysis, which exhibited high accuracy (for DFS: C-index=0.743, 95% CI: 0.698-0.788; for CSS: C-index=0.766, 95% CI: 0.718-0.814) versus tumor-node-metastasis staging (for DFS: C-index=0.692, 95% CI: 0.650-0.734; for CSS: C-index=0.720, 95% CI: 0.675-0.764). CONCLUSION: Preoperative CACPS exhibited high accuracy in predicting prognosis for GC patients who underwent curative resection.
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BACKGROUND: The aim of this study was to detect the expression of long noncoding RNA small nucleolar RNA host gene 18 (SNHG18) andsemaphorin 5A (SEMA5A) genes in multiple myeloma (MM) patients and to explore the correlation of the expression of these genes with the clinical characteristics and prognosis of MM patients. METHODS: Forty-seven newly diagnosed MM, 18 complete remission MM, 13 refractory/relapse MM, and 22 iron deficiency anemia (serving as control) samples were extracted at the Department of Hematology, Second Aï¬liated Hospital of Xian Jiaotong University between January 2015 and December 2016. The clinical features of the MM patients are summarized. Real-time quantitative PCR was performed to analyze the relative expression levels of the SNHG18 and SEMA5Agenes. The clinical characteristics and overall survival (OS) of the MM patients were statistically analyzed while measuring different levels of SNHG18 and SEMA5Agene expression. At the same time, the correlation between the expression of SNHG18 and SEMA5A was also analyzed. RESULTS: The analysis confirmed that SNHG18 and its possible target gene SEMA5A were both highly expressed in newly diagnosed MM patients. After analyzing the clinical signiï¬cance of SNHG18 and SEMA5A in MM patients, we found that the expression of SNHG18 and SEMA5A was related to the Durie-Salmon (DS), International Staging System (ISS), and Revised International Staging System (R-ISS) classification systems, and the Mayo Clinic Risk Stratification for Multiple Myeloma (mSMART; p < 0.05). Moreover, we observed a significant difference in OS between the SNHG18/SEMA5A high expression group and the low expression group. We found a positive correlation between SNHG18 and SEMA5A expression (r = 0.709, p < 0.01). Surprisingly, the expected median OS times of both the SNHG18 and SEMA5Ahigh expression groups were significantly decreased, which was in contrast to those of both the SNHG18 and SEMA5Alow expression groups and the single-gene high expression group (p < 0.05). CONCLUSION: High expression of both SNHG18 and SEMA5A is associated with poor prognosis in patients with MM.
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Regulación Neoplásica de la Expresión Génica , Mieloma Múltiple/sangre , Proteínas de Neoplasias/sangre , ARN Largo no Codificante/sangre , ARN Neoplásico/sangre , Semaforinas/sangre , Adulto , Anciano , Anemia Ferropénica/sangre , Anemia Ferropénica/genética , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Mieloma Múltiple/genética , Mieloma Múltiple/mortalidad , Mieloma Múltiple/patología , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genética , Estadificación de Neoplasias , Pronóstico , ARN Largo no Codificante/biosíntesis , ARN Largo no Codificante/genética , ARN Neoplásico/biosíntesis , ARN Neoplásico/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Recurrencia , Inducción de Remisión , Semaforinas/biosíntesis , Semaforinas/genéticaRESUMEN
Exosomes are small secretory vesicles that are involved in intercellular communication. Exosomes are secreted by many types of cells and exert important functions in plasma-membrane exchange as well as the transport of bioactive substances, such as proteins, messenger ribonucleic acids (mRNAs), micro ribonucleic acids (miRNAs) and organelles. Exosomes may regulate physiological processes by altering gene regulatory networks or epigenetic recombination. Recent studies have shown that exosomes secreted by stem cells can effectively transport proteins, mRNAs and miRNAs and play important roles in the regulation of tissue regeneration. This report reviews current progress in exosome studies as well as their emerging roles in stem cell research and potential clinical use.
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Exosomas/fisiología , Transporte de Proteínas , Transporte de ARN , Investigación con Células Madre , Humanos , MicroARNs/metabolismo , ARN Mensajero/metabolismoRESUMEN
Barbaloin (10-ß-D-glucopyranosyl-1,8-dihydroxy-3-(hydroxymethyl)-9(10H)-anthracenone) is extracted from the aloe plant and has been reported to have anti-inflammatory, antitumor, antibacterial, and other biological activities. Here, we investigated the effects of barbaloin on cardiac electrophysiology, which has not been reported thus far. Cardiac action potentials (APs) and ionic currents were recorded in isolated rabbit ventricular myocytes using whole-cell patch-clamp technique. Additionally, the antiarrhythmic effect of barbaloin was examined in Langendorff-perfused rabbit hearts. In current-clamp recording, application of barbaloin (100 and 200 µmol/L) dose-dependently reduced the action potential duration (APD) and the maximum depolarization velocity (Vmax), and attenuated APD reverse-rate dependence (RRD) in ventricular myocytes. Furthermore, barbaloin (100 and 200 µmol/L) effectively eliminated ATX II-induced early afterdepolarizations (EADs) and Ca2+-induced delayed afterdepolarizations (DADs) in ventricular myocytes. In voltage-clamp recording, barbaloin (10-200 µmol/L) dose-dependently inhibited L-type calcium current (ICa.L) and peak sodium current (INa.P) with IC50 values of 137.06 and 559.80 µmol/L, respectively. Application of barbaloin (100, 200 µmol/L) decreased ATX II-enhanced late sodium current (INa.L) by 36.6%±3.3% and 71.8%±6.5%, respectively. However, barbaloin up to 800 µmol/L did not affect the inward rectifier potassium current (IK1) or the rapidly activated delayed rectifier potassium current (IKr) in ventricular myocytes. In Langendorff-perfused rabbit hearts, barbaloin (200 µmol/L) significantly inhibited aconitine-induced ventricular arrhythmias. These results demonstrate that barbaloin has potential as an antiarrhythmic drug.
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Antracenos/farmacología , Arritmias Cardíacas/prevención & control , Canales de Potasio con Entrada de Voltaje/metabolismo , Aconitina/antagonistas & inhibidores , Aconitina/farmacología , Potenciales de Acción/efectos de los fármacos , Animales , Arritmias Cardíacas/inducido químicamente , Calcio/farmacología , Relación Dosis-Respuesta a Droga , Preparación de Corazón Aislado , Miocitos Cardíacos/fisiología , Técnicas de Placa-Clamp , Conejos , Venenos de Escorpión/antagonistas & inhibidores , Venenos de Escorpión/farmacologíaRESUMEN
BACKGROUND: The mixed lineage kinase domain-like protein has recently been identified as a key downstream component of tumor necrosis factor-induced necroptosis, which is an important pathway of cancer cell death. The goal of the current study is to explore the expression of mixed lineage kinase domain-like protein in colon cancer tissues and evaluate the prognostic value in patients with colon cancer. METHODS: We collected normal and cancer colon tissues from 135 patients diagnosed with colon cancer after radical operation during July 2007 to April 2009 at The Affiliated Hospital of Qingdao University. Immunohistochemistry analysis was scored using an established scoring system. Kaplan-Meier survival curves were generated for recurrence-free survival and overall survival for all patients and 2 subsets of patients. The relationship between mixed lineage kinase domain-like protein expression and prognosis parameter (recurrence-free survival, overall survival) was analyzed by univariate and multivariate Cox regression analyses. RESULTS: The median age of all patients was 67 years and 56.3% were male. Low expression of mixed lineage kinase domain-like protein was associated with decreased overall survival (78.6 vs 81.2 months; P = .011) in all patients. In the subset of 79 patients who received adjuvant chemotherapy, low expression of mixed lineage kinase domain-like protein was associated with decreased recurrence-free survival (60.4 vs 72.8 months; P = .032) and decreased overall survival (66.3 vs 72.9 months; P = .005). Low expression of mixed lineage kinase domain-like protein was associated with decreased overall survival (74.9 vs 79.8 months; P = .006) and recurrence-free survival (69.6 vs 78.8 months; P = .005) among patients with Tumor Node Metastasis (TNM) stage II colon cancer. CONCLUSIONS: Low expression of mixed lineage kinase domain-like protein was associated with decreased overall survival in all patient-group with resected colon cancer. It is associated with decreased recurrence-free survival and overall survival in the subset of patients who receive adjuvant chemotherapy and patients who were TNM stage II. Mixed lineage kinase domain-like protein may provide important prognostic information in patients with colon cancer.
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Neoplasias del Colon/enzimología , Proteínas Quinasas/metabolismo , Anciano , Biomarcadores de Tumor/metabolismo , Quimioterapia Adyuvante , Neoplasias del Colon/mortalidad , Neoplasias del Colon/terapia , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Análisis Multivariante , Pronóstico , Modelos de Riesgos Proporcionales , Resultado del TratamientoRESUMEN
AIM: The augmentation of late sodium current (INa.L) not only causes intracellular Na+ accumulation, which results in intracellular Ca2+ overload via the reverse mode of the Na+/Ca2+ exchange current (reverse-INCX), but also prolongs APD and induces early afterdepolarizations (EAD), which can lead to arrhythmia and cardiac dysfunction. Thus, the inhibition of INa.L is considered to be a potential way for therapeutic intervention in ischemia and heart failure. In this study we investigated the effects of tolterodine (Tol), a competitive muscarinic receptor antagonist, on normal and veratridine (Ver)-augmented INa.L, reverse-INCX and APD in isolated rabbit ventricular myocytes, which might contribute to its cardioprotective activity. METHODS: Rabbit ventricular myocytes were prepared. The INa.L and reverse-INCX were recorded in voltage clamp mode, whereas action potentials and Ver-induced early afterdepolarizations (EADs) were recorded in current clamp mode. Drugs were applied via superfusion. RESULTS: Tol (3-120 nmol/L) concentration-dependently inhibited the normal and Ver-augmented INa.L with IC50 values of 32.08 nmol/L and 42.47 nmol/L, respectively. Atropine (100 µmol/L) did not affect the inhibitory effects of Tol (30 nmol/L) on Ver-augmented INa.L. In contrast, much high concentrations of Tol was needed to inhibit the transient sodium current (INa.T) with an IC50 value of 183.03 µmol/L. In addition, Tol (30 nmol/L) significantly shifted the inactivation curve of INa.T toward a more depolarizing membrane potential without affecting its activation characteristics. Moreover, Tol (30 nmol/L) significantly decreased Ver-augmented reverse-INCX. Tol (30 nmol/L) increased the action potential duration (APD) by 16% under the basal conditions. Ver (20 µmol/L) considerably extended the APD and evoked EADs in 18/24 cells (75%). In the presence of Ver, Tol (30 nmol/L) markedly decreased the APD and eliminated EADs (0/24 cells). CONCLUSION: Tol inhibits normal and Ver-augmented INaL and decreases Ver-augmented reverse-INCX. In addition, Tol reverses the prolongation of the APD and eliminates the EADs induced by Ver, thus prevents Ver-induced arrhythmia.
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Antiarrítmicos/farmacología , Antagonistas Muscarínicos/farmacología , Miocitos Cardíacos/efectos de los fármacos , Bloqueadores de los Canales de Sodio/farmacología , Canales de Sodio/fisiología , Intercambiador de Sodio-Calcio/metabolismo , Tartrato de Tolterodina/farmacología , Veratridina/farmacología , Potenciales de Acción , Animales , Femenino , Ventrículos Cardíacos/citología , Técnicas In Vitro , Masculino , Miocitos Cardíacos/fisiología , Técnicas de Placa-Clamp , ConejosRESUMEN
AIM: Intracellular Ca(2+) ([Ca(2+)]i) overload occurs in myocardial ischemia. An increase in the late sodium current (INaL) causes intracellular Na(+) overload and subsequently [Ca(2+)]i overload via the reverse-mode sodium-calcium exchanger (NCX). Thus, inhibition of INaL is a potential therapeutic target for cardiac diseases associated with [Ca(2+)]i overload. The aim of this study was to investigate the effects of ketamine on Na(+)-dependent Ca(2+) overload in ventricular myocytes in vitro. METHODS: Ventricular myocytes were enzymatically isolated from hearts of rabbits. INaL, NCX current (INCX) and L-type Ca(2+) current (ICaL) were recorded using whole-cell patch-clamp technique. Myocyte shortening and [Ca(2+)]i transients were measured simultaneously using a video-based edge detection and dual excitation fluorescence photomultiplier system. RESULTS: Ketamine (20, 40, 80 µmol/L) inhibited INaL in a concentration-dependent manner. In the presence of sea anemone toxin II (ATX, 30 nmol/L), INaL was augmented by more than 3-fold, while ketamine concentration-dependently suppressed the ATX-augmented INaL. Ketamine (40 µmol/L) also significantly suppressed hypoxia or H2O2-induced enhancement of INaL. Furthermore, ketamine concentration-dependently attenuated ATX-induced enhancement of reverse-mode INCX. In addition, ketamine (40 µmol/L) inhibited ICaL by 33.4%. In the presence of ATX (3 nmol/L), the rate and amplitude of cell shortening and relaxation, the diastolic [Ca(2+)]i, and the rate and amplitude of [Ca(2+)]i rise and decay were significantly increased, which were reverted to control levels by tetrodotoxin (TTX, 2 µmol/L) or by ketamine (40 µmol/L). CONCLUSION: Ketamine protects isolated rabbit ventricular myocytes against [Ca(2+)]i overload by inhibiting INaL and ICaL.
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Calcio/metabolismo , Antagonistas de Aminoácidos Excitadores/farmacología , Ketamina/farmacología , Miocitos Cardíacos/efectos de los fármacos , Intercambiador de Sodio-Calcio/metabolismo , Sodio/metabolismo , Animales , Hipoxia de la Célula/efectos de los fármacos , Células Cultivadas , Femenino , Peróxido de Hidrógeno/metabolismo , Masculino , Miocitos Cardíacos/citología , Miocitos Cardíacos/metabolismo , Técnicas de Placa-Clamp , ConejosRESUMEN
Ranolazine (RAN), a novel antianginal agent, inhibits the increased late sodium current (INa.L) under many pathological conditions. In this study, the whole-cell patch-clamp technique was used to explore the effects of RAN on INa.L and reverse Na(+)/Ca(2+) exchange current (INCX) in rabbit ventricular myocytes during hypoxia.Tetrodotoxin (TTX) at 2 µM or RAN at 9 µM decreased significantly INa.L and reverse INCX under normoxia and RAN had no further effects on both currents in the presence of TTX. RAN (3, 6, and 9 µM) attenuated hypoxia-increased INa.L and reverse INCX in a concentration-dependent manner. Hypoxia-increased INa.L and reverse INCX were inhibited by 2 µM TTX, whereas 9 µM RAN applied sequentially did not further decrease both currents. In another group, after both currents were decreased by 9 µM RAN, 2 µM TTX had no further effects in the presence of Ran. In monophasic action potential (MAP) recording, early after-depolarizations (EADs) were suppressed by RAN (9 µM) during hypoxia. In conclusion, RAN decreased reverse INCX by inhibiting INa.L in normoxia, concentration-dependently attenuated the increase of INa.L, which thereby decreased the reverse INCX, and obviously relieved EADs during hypoxia.
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Acetanilidas/farmacología , Potenciales de Acción/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Ventrículos Cardíacos/citología , Hipoxia/metabolismo , Hipoxia/fisiopatología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/fisiología , Piperazinas/farmacología , Intercambiador de Sodio-Calcio/metabolismo , Sodio/metabolismo , Animales , Células Cultivadas , Relación Dosis-Respuesta a Droga , Técnicas de Placa-Clamp , Conejos , Ranolazina , Tetrodotoxina/farmacologíaRESUMEN
OBJECTIVE: To screen out related microRNAs in keloid tissue, and identify their effect on the proliferation of keloid fibroblasts. METHODS: 8 cases of keloid tissue and 8 cases of normal skin tissue were collected as specimens. The differently expressed miRNA in keloid tissue from normal skin tissue were screened out with gene chip( Exiqon company), which was validated with quantitative real-time PCR. Then miRNA mimics was transfected into keloid fibroblasts line to stimulate high expression of mature miRNA in cells. The effect on the proliferation of fibroblasts in keloid was tested by Edu. RESULTS: (1) A total of 17 differently expressed microRNAs were found, including miR-199a-5p. (2) The expression of miR-199a-5p had been verified by qRT-PCR to be down-regulated in keloid, which was consistent with the result of array. (3) The positive rate of EdU in miR-199a-5p mimics transfected group and negative control group was (20.72 +/- 2.50)% and (27.68 +/- 4.92)%, respectively. The proliferative rate of keloid fibroblasts turned down in miR-199a-5p-transfected group (t = 2.183, P = 0.047). Besides that, the cell cycle changed after transfection. The percentage of S and G2/M phase in miR-199a-5p mimics transfected group was 33.93 +/- 1.30 and 10.87 +/- 0.80, respectively, while it was 31.39 +/- 0.79 and 9.27 +/- 0.46 in negative control group, and the difference was statistically significant. CONCLUSIONS: (1) The miRNA expression profile is different between keloid and normal skin; (2) The expression of miR-199a-5p is down-regulated in keloid and miR-199a-5p can affect the cell cycle and suppress proliferation of keloid fibroblasts. It indicateds that miR-199a-5p may be involved in regulating fibroblastic proliferation.
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Queloide/metabolismo , Queloide/patología , MicroARNs/metabolismo , Proliferación Celular , Células Cultivadas , Regulación hacia Abajo , Femenino , Fibroblastos/metabolismo , Perfilación de la Expresión Génica , Humanos , Queloide/genética , Masculino , MicroARNs/genéticaRESUMEN
The article evaluated the inhibitory action of procaine on wild-type and mutated HERG potassium channel current (I(HERG)) to determine whether mutations in the S6 region are important for the inhibition of I(HERG) by procaine. HERG channels (WT, Y652A, and F656A) were expressed in Xenopus laevis oocytes and studied using the standard two-microelectrode voltage-clamp technique. The results revealed that WT HERG is blocked in a concentration-, voltage-, and state-dependent manner by procaine ([IC50] = 34.79 µM). The steady state activation curves slightly move to the negative, while inactivation parameters move to the positive in the presence of procaine. Time-dependent test reveals that voltage-dependent I(HERG) blockade occurs extremely rapidly. Furthermore, the mutation to Ala of Y652 and F656 produce about 11-fold and 18-fold increases in IC50 for I(HERG) blockade, respectively. Simultaneously, for Y652A, the steady state activation and inactivation parameters are shifted to more positive values after perfusion of procaine. Conclusively, procaine state-dependently inhibits HERG channels (WT, Y652A, and F656A). The helix residues Y652 and F656 in the S6 transmembrane domain might play a role in interaction of the drug with the channel.
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Anestésicos Locales/farmacología , Canales de Potasio Éter-A-Go-Go/antagonistas & inhibidores , Canales de Potasio Éter-A-Go-Go/química , Canales de Potasio Éter-A-Go-Go/genética , Secuencias Hélice-Asa-Hélice/genética , Proteínas de la Membrana/genética , Mutación , Procaína/farmacología , Animales , Células Cultivadas , Relación Dosis-Respuesta a Droga , Femenino , Oocitos , Técnicas de Placa-Clamp , Estructura Terciaria de Proteína/genética , Factores de Tiempo , Xenopus laevisRESUMEN
The protective effects of puerarin on liver damage were evaluated by carbon tetrachloride (CCl4)-induced hepatotoxicity in rats. Male rats were orally treated with puerarin daily, and received CCl4 intraperitoneally twice a week for 4 weeks. Our results showed that puerarin at doses of 50, 100, and 200 mg/kg b. w. significantly reduced the elevated activities of serum alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase at least 15%, 17%, 14% and 18%, respectively. In addition, puerarin at different doses significantly decreased (p<0.05) the level of hepatic thiobarbituric acid reactive substances compared to the CCl4-treated group. Furthermore, the treatment of puerarin was also found to significantly increase the activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione-S-transferase, and glutathione content at least 40%, 12%, 25%, 52%, 17% and 44% in the liver of CCl4-treated rats, respectively. Liver histopathology also showed that puerarin reduced the incidence of liver lesions induced by CCl4. The results suggest that puerarin exhibits potent hepatoprotective effects on CCl4-induced liver damages in rats, and that the hepatoprotective effects of puerarin may be due to both the inhibition of lipid peroxidation and to increase of antioxidant enzymes activity.
Asunto(s)
Antioxidantes/uso terapéutico , Intoxicación por Tetracloruro de Carbono/prevención & control , Suplementos Dietéticos , Isoflavonas/uso terapéutico , Peroxidación de Lípido , Hígado/metabolismo , Estrés Oxidativo , Animales , Antioxidantes/administración & dosificación , Biomarcadores/sangre , Biomarcadores/metabolismo , Intoxicación por Tetracloruro de Carbono/sangre , Intoxicación por Tetracloruro de Carbono/metabolismo , Intoxicación por Tetracloruro de Carbono/patología , Glutatión/metabolismo , Isoflavonas/administración & dosificación , Hígado/efectos de los fármacos , Hígado/patología , Masculino , Oxidación-Reducción , Oxidorreductasas/sangre , Oxidorreductasas/metabolismo , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismoRESUMEN
Many studies indicate that an increase in late sodium current (I(Na.L)) of cardiomyocytes causes intracellular Na overload and subsequently raises the reverse Na/Ca exchanger current (INCX), ultimately resulting in intracellular Ca overload. Therefore, using drugs to inhibit the increased INa.L under various pathological conditions can lower intracellular Ca overload. This study was intended to explore the effect of sophocarpine (SOP) on the increase in INa.L, INCX, calcium transient and contraction in rabbit ventricular myocytes induced by Anemonia sulcata toxin II (ATX II), an opener of sodium channel, with the application of whole-cell patch-clamp techniques, the video-based motion edge detection system, and the intracellular calcium concentration determination system. The results indicate that tetrodotoxin (TTX, 4 µM ) obviously decreased INa.L and INCX enlarged by ATX II (30 nM), and SOP (20, 40, and 80 µM) also inhibited both the parameters concentration dependently in rabbit ventricular myocytes. However, transient sodium current remained unaffected by the above-mentioned concentrations of ATX II, TTX, and SOP. In addition, SOP also reversed diastolic calcium concentration, calcium transient amplitude, and ventricular muscle contractility augmented by ATX II. Its effects were similar to those of TTX, a specific inhibitor of the sodium channel. In conclusion, SOP inhibits INa.L, INCX, diastolic Ca concentration, and contractility in rabbit ventricular myocytes, which suggests that relief of intracellular Ca overload through inhibiting INa.L is likely to become a new therapeutic mechanism of SOP against arrhythmia and myocyte damage associated with intracellular Ca overload.
Asunto(s)
Alcaloides/farmacología , Calcio/metabolismo , Venenos de Cnidarios/farmacología , Sodio/metabolismo , Alcaloides/administración & dosificación , Animales , Arritmias Cardíacas/tratamiento farmacológico , Arritmias Cardíacas/etiología , Arritmias Cardíacas/fisiopatología , Relación Dosis-Respuesta a Droga , Femenino , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/patología , Masculino , Contracción Miocárdica/efectos de los fármacos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Técnicas de Placa-Clamp , Conejos , Canales de Sodio/metabolismo , Intercambiador de Sodio-Calcio/metabolismo , Tetrodotoxina/farmacologíaRESUMEN
The objectives of this study were to investigate the effects of veratridine (VER) on persistent sodium current (I(Na.P)), Na(+)/Ca(2+) exchange current (I(NCX)), calcium transients and the action potential (AP) in rabbit ventricular myocytes, and to explore the mechanism in intracellular calcium overload and myocardial contraction enhancement by using whole-cell patch clamp recording technique, visual motion edge detection system, intracellular calcium measurement system and multi-channel physiological signal acquisition and processing system. The results showed that I(Na.P) and reverse I(NCX) in ventricular myocytes were obviously increased after giving 10, 20 µmol/L VER, with the current density of I(Na.P) increasing from (-0.22 ± 0.12) to (-0.61 ± 0.13) and (-2.15 ± 0.14) pA/pF (P < 0.01, n = 10) at -20 mV, and that of reverse I(NCX) increasing from (1.62 ± 0.12) to (2.19 ± 0.09) and (2.58 ± 0.11) pA/pF (P < 0.05, n = 10) at +50 mV. After adding 4 µmol/L tetrodotoxin (TTX), current density of I(Na.P) and reverse I(NCX) returned to (-0.07 ± 0.14) and (1.69 ± 0.15) pA/pF (P < 0.05, n = 10). Another specific blocker of I(Na.P), ranolazine (RAN), could obviously inhibit VER-increased I(Na.P) and reverse I(NCX). After giving 2.5 µmol/L VER, the maximal contraction rate of ventricular myocytes increased from (-0.91 ± 0.29) to (-1.53 ± 0.29) µm/s (P < 0.01, n = 7), the amplitude of contraction increased from (0.10 ± 0.04) to (0.16 ± 0.04) µm (P < 0.05, n = 7), and the baseline of calcium transients (diastolic calcium concentration) increased from (1.21 ± 0.08) to (1.37 ± 0.12) (P < 0.05, n = 7). After adding 2 µmol/L TTX, the maximal contraction rate and amplitude of ventricular myocytes decreased to (-0.86 ± 0.24) µm/s and (0.09 ± 0.03) µm (P < 0.01, n = 7) respectively. And the baseline of calcium transients reduced to (1.17 ± 0.09) (P < 0.05, n = 7). VER (20 µmol/L) could extend action potential duration at 50% repolarization (APD(50)) and at 90% repolarization (APD(90)) in ventricular myocytes from (123.18 ± 23.70) to (271.90 ± 32.81) and from (146.94 ± 24.15) to (429.79 ± 32.04) ms (P < 0.01, n = 6) respectively. Early afterdepolarizations (EADs) appeared in 3 out of the 6 cases. After adding 4 µmol/L TTX, APD(50) and APD(90) were reduced to (99.07 ± 22.81) and (163.84 ± 26.06) ms (P < 0.01, n = 6) respectively, and EADs disappeared accordingly in 3 cases. It could be suggested that: (1) As a specific agonist of the I(Na.P), VER could result in I(Na.P) increase and intracellular Na(+) overload, and subsequently intracellular Ca(2+) overload with the increase of reverse I(NCX). (2) The VER-increased I(Na.P) could further extend the action potential duration (APD) and induce EADs. (3) TTX could restrain the abnormal VER-induced changes of the above-mentioned indexes, indicating that these abnormal changes were caused by the increase of I(Na.P). Based on this study, it is concluded that as the I(Na.P) agonist, VER can enhance reverse I(NCX) by increasing I(Na.P), leading to intracellular Ca(2+) overload and APD abnormal extension.
Asunto(s)
Potenciales de Acción , Miocitos Cardíacos/efectos de los fármacos , Intercambiador de Sodio-Calcio/metabolismo , Veratridina/farmacología , Acetanilidas/farmacología , Animales , Calcio/metabolismo , Contracción Miocárdica , Miocitos Cardíacos/citología , Técnicas de Placa-Clamp , Piperazinas/farmacología , Conejos , Ranolazina , Tetrodotoxina/farmacologíaRESUMEN
Keloids are fibroproliferative dermal lesions characterized by the proliferation of fibroblasts and the formation of excess scar tissue, for which no effective treatment exists. We transfected a replication-incompetent adenovirus vector expressing green fluorescent protein and interleukin-24 gene (Ad-GFP/IL-24) into keloid fibroblasts (KF) and normal dermal fibroblasts (NDF) in vitro to investigate the suppression effects by observation on cell lines growth, apoptosis, mitosis cycle, etc. The expression of GFP and IL-24 mRNA confirmed that Ad-GFP/IL-24 was transfected into KF and NDF successfully. The expression level of secreting IL-24 protein detected by enzyme-linked immunosorbent assay in Ad-GFP/IL-24-treated KF and PBS-treated NDF was higher than controls; treatment with Ad-GFP/IL-24 in KF induced growth suppression (71.83% ± 6.67%, P < 0.05 to 9.79% ± 3.34%, P < 0.01), apoptosis (24.2% ± 3.08% to 66.51% ± 5.29%, P < 0.01) and increased the percentage of the G2/M phase (42.26% ± 6.44%, P < 0.01) in KF but not in NDF. The data showed that the exogenous IL-24 gene could selectively inhibit human KF proliferation and induce significant apoptosis.
Asunto(s)
Apoptosis/fisiología , Proliferación Celular , Interleucinas/fisiología , Queloide/fisiopatología , Adenoviridae/genética , Adolescente , Adulto , Línea Celular , Supervivencia Celular/fisiología , Células Cultivadas , Niño , Ensayo de Inmunoadsorción Enzimática , Femenino , Fibroblastos/metabolismo , Fibroblastos/fisiología , Expresión Génica , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Interleucinas/genética , Interleucinas/metabolismo , Queloide/metabolismo , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Piel/citología , Transfección , Adulto JovenRESUMEN
A variety of nerve conduits incorporated with chemical and biological factors have been developed to further stimulate nerve regeneration. Although most of the nerve guides in studies are basically limited to bridge a short gap of nerve defect in rat models, it is vital to evaluate effects of conduits on nerve regeneration over distance greater than 20 mm, or more clinically relevant nerve gap lengths in higher mammals. In this study, a poly(lactide-co-glycolide) (PLGA) nerve conduit, treated with pulsed plasma and coated with ciliary neurotrophic factor (CNTF) as well as chitosan, was used to repair 25-mm-long canine tibial nerve defects in eighteen cross-bred dogs. The canines were randomly divided into three groups (n = 6), a 25-mm segment of the tibial nerve was removed and replaced by a PLGA/chitosan-CNTF nerve conduit, PLGA/chitosan conduit and autologous nerve grafts were performed as the control. The results were evaluated by general observation, electromyogram testing, S-100 histological immunostaining, and image analysis at 3 months after operation. The histological results demonstrated that the PLGA/chitosan-CNTF conduits and PLGA/chitosan conduits were capable of leading the damaged axons through the lesioned area. Through the comparison of the three groups, the results in PLGA/chitosan-CNTF conduits group were better than that of PLGA/chitosan conduits group, while they were similar to autologous nerve grafts group. Therefore, CNTF-coated PLGA/chitosan nerve conduits could be an alternative artificial nerve conduit for nerve regeneration.