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1.
Invest Ophthalmol Vis Sci ; 60(15): 5104-5111, 2019 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-31826237

RESUMO

Purpose: Cell-cell contact in retinal pigment epithelium (RPE) involves adherent junctions, gap junctions, and tight junctions, which are primarily composed by E-cadherin, zona occludens 1 (ZO-1), and connexin 43, respectively. Here, we aimed to explore the relationship and interplay between these junction-associated proteins. Methods: E-cadherin, connexin 43, and ZO-1 expression in human primary RPE in the early phase after TGF-ß1 stimulation was detected. The knockdown of E-cadherin, ZO-1, and connexin 43 was performed to characterize the regulatory network involving these three proteins. Dye transfer and FITC-dextran permeability assays were conducted to observe the epithelial functional alterations. Transmission electron microscopy (TEM) was used to observe the ultrastructure of the cell-cell junctions in mouse RPE. The immunofluorescence staining and coimmunoprecipitation were performed to observe the colocalization and the physical association of E-cadherin, ZO-1, and connexin 43. Results: Among these three components, E-cadherin appeared to be the first protein that was downregulated after TGF-ß1 treatment. The ultrastructures of adherent junctions, gap junctions, and tight junctions could be observed in mouse RPE by TEM. E-cadherin, ZO-1, and connexin 43 were colocalized and physically bound to each other. The knockdown of one of these three proteins led to downregulation of the other two proteins and compromised epithelial function. Conclusions: E-cadherin, ZO-1, and connexin 43 were physically associated with each other and were mutually regulated. To enhance the understanding of cell-cell contacts, a holistic view is needed. Our results provide new insights in RPE disorders such as proliferative vitreoretinopathy.


Assuntos
Caderinas/genética , Conexina 43/genética , Regulação da Expressão Gênica , Epitélio Pigmentado da Retina/metabolismo , Vitreorretinopatia Proliferativa/genética , Proteína da Zônula de Oclusão-1/genética , Animais , Caderinas/biossíntese , Células Cultivadas , Conexina 43/biossíntese , Humanos , Junções Intercelulares , Camundongos , Microscopia Eletrônica de Transmissão , RNA/genética , Epitélio Pigmentado da Retina/ultraestrutura , Junções Íntimas , Vitreorretinopatia Proliferativa/metabolismo , Vitreorretinopatia Proliferativa/patologia , Proteína da Zônula de Oclusão-1/biossíntese
2.
Oxid Med Cell Longev ; 2019: 5703764, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178968

RESUMO

MicroRNAs (miRNAs) are regarded as a potential method for the treatment of atrial fibrillation (AF) although its molecular mechanism remains unknown. We found in our previous study that the level of peripheral blood miR-27b-3p and the expression of atrial tissue CX43 were both significantly downregulated in AF patients. In the present study, we propose and test this hypothesis that overexpression of miR-27b-3p attenuates atrial fibrosis, increases CX43 expression, and regulates the signaling pathway of Wnt/ß-Catenin by targeting Wnt3a. miR-27b-3p overexpression was induced by rat tail vein injection of adeno-associated virus. Two weeks after transfection of adeno-associated virus, the rat AF model was established by tail vein injection of acetylcholine- (ACh-) CaCl2 for 7 days, and 1 ml/kg was injected daily. The incidence and duration of AF were recorded with an electrocardiogram. Cardiac function was monitored by cardiac ultrasound. Serum cardiac enzyme was detected by ELISA. The expression of atrial miR-27b-3 and Wnt3a was assayed by quantitative RT-PCR. Atrial fibrosis was determined by Masson's trichrome staining. Expression of atrial Collagen-I and Collagen-III was tested by the immunohistochemical method. Expression of CX43 was measured by immunofluorescence. The expression of Collagen-I, a-SMA, Collagen-III, TGF-ß1, CX43, Wnt3a, ß-Catenin, and p-ß-Catenin was assayed by western blot. Our results showed that miR-27b-3p overexpression could reduce the incidence and duration of AF, alleviate atrial fibrosis, increase atrial CX43 expression, and decrease the expression of Collagen-I, a-SMA, Collagen-III, TGF-ß1, Wnt3a, and p-ß-Catenin. In addition, the results of luciferase activity assay showed that Wnt3a is a validated miR-27b-3p target in HEK 293T cells. Our results provide a new evidence that miR-27b-3p regulates the signaling pathway of Wnt/ß-Catenin by targeting Wnt3a, which may play an important role in the development of atrial fibrosis and AF.


Assuntos
Fibrilação Atrial/metabolismo , MicroRNAs/metabolismo , Via de Sinalização Wnt , Proteína Wnt3A/metabolismo , beta Catenina/metabolismo , Animais , Fibrilação Atrial/genética , Fibrilação Atrial/patologia , Fibrilação Atrial/fisiopatologia , Conexina 43/biossíntese , Conexina 43/genética , Fibrose , Células HEK293 , Átrios do Coração/metabolismo , Átrios do Coração/patologia , Átrios do Coração/fisiopatologia , Humanos , Masculino , MicroRNAs/biossíntese , MicroRNAs/genética , Ratos , Ratos Sprague-Dawley , Proteína Wnt3A/genética , beta Catenina/genética
3.
World Neurosurg ; 126: e392-e401, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30822595

RESUMO

OBJECTIVE: To develop a method to distinguish atypical meningiomas (AMs) with malignant progression (MP) from primary AMs without a clinical history. METHODS: The clinical, radiologic, and pathologic data of 33 previously Simpson grade I resected (if any) as well as no radiotherapy treated intracranial AMs between January 2008 and December 2015 were reviewed. Immunohistochemical staining for connexin 43 (Cx43) and Ki-67 was performed. Descriptive analysis and univariate and multivariate logistic regression analyses were used to explore independent predictors of MP. A multivariable logistic model was developed to estimate the risk of MP, and its diagnostic value was determined from a receiver operating characteristic curve. RESULTS: There were 11 AMs (33.3%) with histopathologically confirmed MP from benign meningiomas. The other 22 (66.7%) were initially diagnosed AMs with no histopathologically confirmed MP during a median 60.5 months (range, 42-126 months) of follow-up. Univariate and multivariate logistic analyses showed that irregular tumor shape (P = 0.010) and low Cx43 expression (P = 0.010) were independent predictors of the presence of MP, and the predicted probability was calculated by the following formula: P = 1/[1+exp.{1.218-(3.202×Shape)+(3.814×Cx43)}]. P > 0.5 for an irregularly shaped (score 1) AM with low Cx43 expression (score 0) indicated a high probability of MP. The sensitivity, specificity, positive predictive value, negative predictive value, and overall predictive accuracy were 63.6, 95.6, 87.5, 84.0, and 84.8%, respectively. CONCLUSIONS: Low Cx43 expression and irregular tumor shape were independent predictors of the presence of MP. The relevant logistic regression model was found to be effective in distinguishing MP-AMs from primary AMs.


Assuntos
Modelos Logísticos , Neoplasias Meníngeas/patologia , Meningioma/patologia , Adulto , Idoso , Biomarcadores Tumorais/análise , Conexina 43/biossíntese , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade
4.
Am J Dermatopathol ; 41(11): 810-818, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30839347

RESUMO

INTRODUCTION: Benign cutaneous tumors with follicular differentiation are alleged to differentiate toward parts of the hair follicle. Connexin 43 (Cx43) is a gap junction protein, the tumoral role of which has been investigated in several types of tumors. OBJECTIVE: To study the pattern of expression of Cx43 in benign cutaneous tumors with follicular differentiation and to compare it with that shown by their alleged anatomical counterparts of the hair follicle. MATERIALS AND METHODS: Five cases each of trichofolliculoma, trichilemmoma, fibrofolliculoma/trichodiscoma, trichoblastoma, trichoepithelioma, pilomatrixoma, and proliferating trichilemmal tumor, 3 cases of pilar sheath acanthoma, and 1 case of tumor of the follicular infundibulum were examined. Anti-Cx43 antibody was used. RESULTS: Cx43 was expressed by all follicular tumors studied. Comparisons between trichoblastoma and trichoepithelioma and their respective normal counterparts could not be made. In 3 tumors (trichofolliculoma, pilomatrixoma, and the spectrum fibrofolliculoma/trichodiscoma), there was a parallelism between their Cx43 expression pattern and that of their alleged anatomical counterparts. In pilar sheath acanthoma, trichilemmoma, and the tumor of the follicular infundibulum, we only found partial similarities in Cx43 expression. Only the proliferating trichilemmal tumor showed a discordant pattern of expression. CONCLUSIONS: Cx43 expression is preserved in benign cutaneous tumors with follicular differentiation and the patterns of Cx43 expression in benign cutaneous tumors with follicular differentiation parallel those of their alleged anatomical counterparts in 5 types (either totally or partially). This preservation might be related to the good behavior of the entities studied.


Assuntos
Conexina 43/biossíntese , Doenças do Cabelo/metabolismo , Folículo Piloso/patologia , Neoplasias Cutâneas/metabolismo , Conexina 43/análise , Doenças do Cabelo/patologia , Folículo Piloso/metabolismo , Humanos , Neoplasias Cutâneas/patologia
5.
J Am Heart Assoc ; 8(4): e010254, 2019 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-30757948

RESUMO

Background Chronic intermittent hypoxia ( CIH ) is a distinct pathological mechanism of obstructive sleep apnea ( OSA ), which is recognized as an independent risk factor for cardiovascular diseases. The aims of this study were to ascertain whether CIH induces atrial fibrillation ( AF ), to determine whether cardiac sympathetic denervation ( CSD ) can prevent it and suppress blood pressure, and to explore the potential molecular mechanisms involved. Methods and Results Sixty Sprague-Dawley male rats were randomly divided into 4 groups: sham, CSD , CIH , CIH + CSD . The rats were exposed either to CIH 8 hours daily or normoxia for 6 weeks. Cardiac pathology and structure were analyzed by hematoxylin and eosin staining and echocardiogram. ECG, blood pressure, body weight, and blood gas were recorded. Connexin 43 and tyrosine hydroxylase were detected by western blot, immunohistochemistry, and immunofluorescence. CIH induced atrial remodeling, and increased AF inducibility. CSD treatment reduced postapneic blood pressure rises and AF susceptibility, which could attenuate CIH -associated structural atrial arrhythmogenic remodeling. In addition, CIH -induced sympathetic nerve hyperinnervation and CSD treatment reduced sympathetic innervation, which may affect CIH -induced AF -associated sympathovagal imbalance. Connexin 43 was specifically downregulated in CIH , whereas CSD treatment increased its expression. Conclusions These results suggested CIH induces atrial remodeling, increases AF inducibility, results in sympathetic nerve hyperinnervation, and decreases connexin 43 expression, but CSD treatment reduces AF susceptibility, postapneic blood pressure increase, sympathetic innervation, and the alteration of Cx43, which may be a key point in the genesis of CIH -induced AF .


Assuntos
Fibrilação Atrial/terapia , Pressão Sanguínea/fisiologia , Hipóxia/complicações , Apneia Obstrutiva do Sono/complicações , Simpatectomia/métodos , Animais , Fibrilação Atrial/etiologia , Fibrilação Atrial/fisiopatologia , Remodelamento Atrial/fisiologia , Western Blotting , Doença Crônica , Conexina 43/biossíntese , Modelos Animais de Doenças , Eletrocardiografia , Hipóxia/sangue , Imuno-Histoquímica , Masculino , Ratos , Ratos Sprague-Dawley , Apneia Obstrutiva do Sono/sangue
6.
Breast Cancer Res Treat ; 174(1): 93-102, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30474779

RESUMO

PURPOSE: Gap junctions are specialized membrane structures that form channels between adjacent cells allowing cell communication. Gap junctions and specifically Connexin 43 (Cx43) are down-regulated in cancer; however, there are contrasting reports on how this effects breast cancer patient survival. This paper is the first large-scale tissue microarray analysis of Cx43 expression in breast cancer patients with an associated clinical long-term follow-up. METHODS: Using a validated TMA of 1118 primary breast cancers, coupled to a comprehensive database of clinicopathological variables, the expression levels and subcellular localisation of Cx43 was assessed by immunohistochemistry. Its impact in terms of survival, distant metastasis-free survival, and clinicopathological variables was determined. RESULTS: Patients whose tumors expressed high levels of Cx43 had significantly better survival (p < 0.001) than patients with low levels. High Cx43 expression within tumors was associated with an 18-month survival advantage. Loss of Cx43 expression was associated with markers of poor prognosis, namely large tumor size, high grade, high proliferation status, high pleomorphism, high mitosis, poor Nottingham Prognostic Index (NPI), and triple negative tumors. Cx43 expression was independent of tumor size, grade, stage and ER-status in predicting poor survival on multivariate analysis (p = 0.004). CONCLUSION: Connexin 43 (Cx43) is an independent predictor of breast cancer survival and distant metastasis-free survival. High expression of Cx43 was seen in only 13% of tumors, suggesting that drugs to increase Cx43 expression may result in prolonged patients survival.


Assuntos
Biomarcadores Tumorais/análise , Neoplasias da Mama/patologia , Conexina 43/biossíntese , Adulto , Idoso , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Conexina 43/análise , Feminino , Humanos , Estimativa de Kaplan-Meier , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos
7.
Heart Lung Circ ; 28(11): 1755-1761, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30322759

RESUMO

BACKGROUND: MicroRNAs (miRNAs) are critical modulators of various physiological and pathological processes, but their role in cardiac arrhythmias remains yet to be completely understood. Connexin43 (Cx43) is an important cardiac gap junction protein and a potential target of miR-206, and downregulation of Cx43 induces ventricular tachyarrhythmias. METHODS: We investigated the effects of miR-206 overexpression on the adult mouse heart and in cardiac arrhythmias. Luciferase activity assay was employed to validate Cx43 as a direct target of miR-206. Expression of Cx43 was measured in cardiac muscle cell line HL-1 securely expressing miR-206. An inducible miR-206 overexpression mouse model was established to evaluate the in vivo effect of miR-206 on Cx43 expression and cardiac rhythm. RESULTS: MiR-206 directly recognised 3'-untranslated region of Cx43 mRNA to inhibit its expression in HL-1 cells. Induction of miR-206 in the adult mouse heart suppressed Cx43 expression, particularly in the atria and ventricle. Importantly, miR-206 overexpression also induced abnormal heart-rate and PR interval, and shortened life-span in the experimental mice. CONCLUSIONS: In cardiomyocytes, miR-206 is a upstream regulator of Cx43, and its overexpression downregulates Cx43 to induce abnormal heart-rate and PR interval.


Assuntos
Arritmias Cardíacas/genética , Conexina 43/genética , Regulação para Baixo , Regulação da Expressão Gênica , MicroRNAs/genética , Miócitos Cardíacos/metabolismo , Animais , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/patologia , Western Blotting , Linhagem Celular , Conexina 43/biossíntese , Modelos Animais de Doenças , Camundongos , Camundongos Transgênicos , MicroRNAs/biossíntese , Miócitos Cardíacos/patologia
8.
Biomed Pharmacother ; 108: 1469-1476, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30372849

RESUMO

As one of the most popular anesthetics, sevoflurane is widely used in pediatric anesthesia. Unfortunately, an increasing number of studies have demonstrated that sevoflurane has potential neurotoxic effects on the developing brain and cognition, even in adolescence. Connexin 43 (Cx43) has been documented to contribute to cognitive dysfunction. The present study hypothesized that Cx43 may participate in sevoflurane-induced neuroinjury and investigated the underlying mechanisms in young Sprague Dawley (SD) rats. Seven-day-old SD rats (P7) were exposed to 3% sevoflurane for 4 h. The levels of Cx43,mitogen-activated protein kinase (MAPK) signaling pathway components(including total and phosphorylated p38, extracellular signal-regulated kinase (ERK), and c-Jun n-terminal kinase (JNK) and activator protein 1(AP-1) transcription factors (including total and phosphorylated c-Fos, and c-Jun) were assessed by Western blot analysis. Neuronal apoptosis was detected using immunohistochemistry (IHC). The Morris water maze (MWM) was performed to evaluate cognitive function from P28 to P33. The results showed that anesthesia with 3% sevoflurane for 4 h increased Cx43 levels in the rat hippocampus from 6 h to 3 d, and compared with sevoflurane exposure in the control group rats, exposure in P7 SD rats also increased the ratios of phosphorylated JNK to JNK and, phosphorylated c-Jun to c-Jun in the hippocampus from 6 h to 3 d. All these effects could be alleviated by pretreatment with the JNK inhibitor SP600125 (10 mg/kg). Neuroapoptosis was similarly increased from 6 h to 1 d after inhaled sevoflurane exposure. Finally, the MWM indicated that sevoflurane could increase the escape latency and, decrease the number of platform crossings from P28 to P33. Overall, our findings suggested that sevoflurane increased Cx43 expression and induced to apoptosis by activating the JNK/c-Jun signaling pathway in the hippocampus of P7 rats. This finding may reveal a new strategy for preventing sevoflurane-induced neuronal dysfunction.


Assuntos
Conexina 43/biossíntese , Hipocampo/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/biossíntese , Sistema de Sinalização das MAP Quinases/fisiologia , Sevoflurano/toxicidade , Fator de Transcrição AP-1/biossíntese , Animais , Animais Recém-Nascidos , Feminino , Hipocampo/efeitos dos fármacos , Hipocampo/crescimento & desenvolvimento , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Inibidores da Agregação de Plaquetas/toxicidade , Ratos , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia
9.
Artif Cells Nanomed Biotechnol ; 46(sup3): S717-S724, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30289284

RESUMO

Diaphragmatic myoblasts (DM) are stem cells of the diaphragm, a muscle displaying high resistance to stress and exhaustion. We hypothesized that DM modified to overexpress connexin-43 (cx43), seeded on aligned poly (l-lactic acid) (PLLA) sheets would decrease infarct size and improve ventricular function in sheep with acute myocardial infarction (AMI). Sheep with AMI received PLLA sheets without DM (PLLA group), sheets with DM (PLLA-DM group), sheets with DM overexpressing cx43 (PLLA-DMcx43) or no treatment (control group, n = 6 per group). Infarct size (cardiac magnetic resonance) decreased ∼25% in PLLA-DMcx43 [from 8.2 ± 0.6 ml (day 2) to 6.5 ± 0.7 ml (day 45), p < .01, ANOVA-Bonferroni] but not in the other groups. Ejection fraction (EF%) (echocardiography) at 3 days post-AMI fell significantly in all groups. At 45 days, PLLA-DM y PLLA-DMcx43 recovered their EF% to pre-AMI values (PLLA-DM: 61.1 ± 0.5% vs. 58.9 ± 3.3%, p = NS; PLLA-DMcx43: 64.6 ± 2.9% vs. 56.9 ± 2.4%, p = NS), but not in control (56.8 ± 2.0% vs. 43.8 ± 1.1%, p < .01) and PLLA (65.7 ± 2.1% vs. 56.6 ± 4.8%, p < .01). Capillary density was higher (p < .05) in PLLA-DMcx43 group than in the remaining groups. In conclusion, PLLA-DMcx43 reduces infarct size in sheep with AMI. PLLA-DMcx43 and PLLA-DM improve ventricular function similarly. Given its safety and feasibility, this novel approach may prove beneficial in the clinic.


Assuntos
Conexina 43/biossíntese , Oclusão Coronária , Diafragma/metabolismo , Mioblastos , Infarto do Miocárdio , Poliésteres/química , Tecidos Suporte/química , Função Ventricular , Animais , Oclusão Coronária/metabolismo , Oclusão Coronária/patologia , Oclusão Coronária/fisiopatologia , Oclusão Coronária/terapia , Diafragma/patologia , Masculino , Mioblastos/metabolismo , Mioblastos/patologia , Mioblastos/transplante , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Infarto do Miocárdio/terapia , Ovinos
10.
J Immunol ; 201(10): 2986-2997, 2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30341186

RESUMO

Connexin 43 (Cx43) deficiency was found to increase mortality in a mouse model of bacterial peritonitis, and Cx43 is upregulated in macrophages by LPS treatment. In this study, we characterized a novel signaling pathway for LPS-induced Cx43 expression in RAW264.7 cells and thioglycolate-elicited peritoneal macrophages (TGEMs). LPS alone or LPS-containing conditioned medium (CM) upregulated Cx43. Overexpression or silencing of Cx43 led to the enhancement or inhibition, respectively, of CM-induced TGEM migration. This response involved the inducible NO synthase (iNOS)/focal adhesion kinase (FAK)/Src pathways. Moreover, CM-induced migration was compromised in TGEMs from Cx43+/- mice compared with TGEMs from Cx43+/+ littermates. Cx43 was upregulated by a serum/glucocorticoid-regulated kinase 1 (SGK) activator and downregulated, along with inhibition of CM-induced TGEM migration, by knockdown of the SGK gene or blockade of the SGK pathway. LPS-induced SGK activation was abrogated by Torin2, whereas LPS-induced Cx43 was downregulated by both Torin2 and rapamycin. Analysis of the effects of FK506 and methylprednisolone, common immunosuppressive agents following organ transplantation, suggested a link between these immunosuppressive drugs and impaired macrophage migration via the Cx43/iNOS/Src/FAK pathway. In a model of Escherichia coli infectious peritonitis, GSK650349-, an SGK inhibitor, or Torin2-treated mice showed less accumulation of F4/80+CD11b+ macrophages in the peritoneal cavity, with a delay in the elimination of bacteria. Furthermore, following pretreatment with Gap19, a selective Cx43 hemichannel blocker, the survival of model mice was significantly reduced. Taken together, our study suggested that Cx43 in macrophages was associated with macrophage migration, an important immune process in host defense to infection.


Assuntos
Movimento Celular/imunologia , Conexina 43/biossíntese , Macrófagos/imunologia , Transdução de Sinais/imunologia , Animais , Conexina 43/imunologia , Quinase 1 de Adesão Focal/imunologia , Quinase 1 de Adesão Focal/metabolismo , Regulação da Expressão Gênica/imunologia , Proteínas Imediatamente Precoces/imunologia , Proteínas Imediatamente Precoces/metabolismo , Lipopolissacarídeos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/imunologia , Óxido Nítrico Sintase Tipo II/metabolismo , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Células RAW 264.7 , Serina-Treonina Quinases TOR/imunologia , Serina-Treonina Quinases TOR/metabolismo , Quinases da Família src/imunologia , Quinases da Família src/metabolismo
11.
Neurourol Urodyn ; 37(8): 2502-2509, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30070388

RESUMO

AIMS: Sacral spinal cord injury (SCI) could induce underactive bladder (UAB). Malfunction of connexin 43 (CX43) regulated by TGF-ß1 might involve in urinary bladder dysfunction. We studied the changes of CX43 and TGF-ß1/Smad3 signaling in detrusor of neurogenic bladder (NB) in sacral SCI rats. METHODS: Sacral SCI was produced by hemisection (SSCH) or transection (SSCT) of spinal cord between L4 and L5 in female Wistar rats. BBB scores, residual urine volume and bladder weight as well as characteristic cystometric parameters at 6th week were used to confirm the successful establishment of NB. Western blotting and qRT-PCR were used to exam the protein and mRNA expression levels of CX43, CX45, TGF-ß1, and Smad3 in detrusor. RESULTS: BBB scores were significantly decreased, with the lowest in SSCT rats (P < 0.01). The residual urine volume, mean bladder weight, and cystometric parameters were increased, with the highest in SSCT rats. CX43 and phospho-CX43 protein levels were significantly decreased, but those of TGF-ß1, Smad3, and phospho-Smad3 were significantly increased. It was the protein and mRNA levels of CX43 but not those of CX45 which were decreased in negative accordance with those of TGF-ß1 and Smad3. Those changes were more significant in SSCT than in SSCH rats. CONCLUSIONS: This study indicates that voiding dysfunction is related to the decreased CX43 function in detrusor from NB. TGF-ß1/Smad3 signaling might be involved in the down-regulation of CX43 in SCI rats. Early regulation of CX43 might be beneficial to patients with voiding dysfunction.


Assuntos
Conexina 43/biossíntese , Traumatismos da Medula Espinal/fisiopatologia , Fator de Crescimento Transformador beta1/biossíntese , Bexiga Urinaria Neurogênica/fisiopatologia , Animais , Conexina 43/genética , Estado de Descerebração/fisiopatologia , Feminino , Tamanho do Órgão , Ratos , Ratos Wistar , Proteína Smad3/biossíntese , Proteína Smad3/genética , Traumatismos da Medula Espinal/complicações , Fator de Crescimento Transformador beta1/genética , Bexiga Urinária/patologia , Bexiga Urinaria Neurogênica/etiologia , Urodinâmica
12.
Med Sci Monit ; 24: 5008-5014, 2018 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-30022020

RESUMO

BACKGROUND Cardiac infarction frequently leads to arrhythmia and ischemia/reperfusion (I/R) aggravates cardiac injury. Pinocembrin can resist cerebral ischemia and decrease cardiac infarction area. This study thus generated a rat myocardial I/R model to assess the effect on ventricular rhythm and expression of gap junction connexin (Cx43). MATERIAL AND METHODS Male SD rats were randomly assigned into sham, model, and pinocembrin (30 mg/kg) pretreatment groups (N=15 each). The I/R model was generated by ligation of the left anterior descending coronary artery for 30 min. The pinocembrin group received intravenous injection 10 min before surgery. Heart rate (HR), mean artery pressure (MAP), rate pressure product (RPP), and arrhythmia were observed at 10 min before ischemia, 30 min after ischemia, and at 30, 60, and 120 min after reperfusion. ELISA was used to assess serum CK-MB and cTnI levels. Na+-K+ATPase and Ca+-Mg2+ATPase levels were quantified by spectrometry, followed by HE staining, IHC approach for Cx43 expression, and Western blot for Kir2.1 protein expression. RESULTS Model rats had significantly lower HR, MAP, and RPP than in the sham group, and the pinocembrin pretreatment group had higher serum indexes. Arrhythmia index, CK-MB, and cTnI were higher in the model and pinocembrin groups, while Na+-K+ATPase, Ca+-Mg2+ATPase, Cx43, and Kir2.1 proteins were lower (p<0.05). CONCLUSIONS Pinocembrin alleviated ventricular arrhythmia in I/R rats via enhancing Na+-K+ATPase and Ca+-Mg2+ATPase activity and upregulating Cx43 and Kir2.1 protein expression.


Assuntos
Conexina 43/biossíntese , Flavanonas/farmacologia , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/metabolismo , Animais , Arritmias Cardíacas/tratamento farmacológico , Arritmias Cardíacas/metabolismo , Modelos Animais de Doenças , Masculino , Infarto do Miocárdio/metabolismo , Isquemia Miocárdica/tratamento farmacológico , Isquemia Miocárdica/metabolismo , Ratos , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacos
13.
Exp Mol Pathol ; 104(1): 12-18, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29246788

RESUMO

It is accepted that alteration of connexin43 (Cx43) expression in glomeruli is a common pathological response in several forms of kidney diseases. To date, however the change of the Cx43 expression in obesity-related glomerulopathy (ORG) has not been reported. In this study, the alteration of Cx43 expression in the glomeruli of rat with ORG was defined. Five-week-old rats were fed with high-fat diet for 18weeks to establish the ORG model, then the histological change of glomeruli, the foot process effacement of podocyte, the markers for podocyte injury (nephrin,podocin and WT1) and Cx43 expression in glomeruli were examined respectively. The results demonstrated metabolic disorder, hyperinsulinemia, systemic inflammation and microalbuminuria in ORG rats. There was significant hypertrophy, glomerular expansion and inflammatory cell infiltration in the kidney of ORG rats compared to the control group. Significant foot process effacement of the podocyte in the glomeruli, nephrin loss and density reduction were shown in the ORG rats, and Cx43 expression was significant upregulated in glomeruli of ORG rats compared to the control group. The results indicate the correlation of overexpressed Cx43 with the obesity related renal inflammation and suggest that Cx43 might be a potential target in the development of obesity related glomerulopathy.


Assuntos
Conexina 43/biossíntese , Glomerulonefrite/metabolismo , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Obesidade/metabolismo , Animais , Dieta Hiperlipídica , Modelos Animais de Doenças , Glomerulonefrite/patologia , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Masculino , Proteínas de Membrana/biossíntese , Obesidade/patologia , Podócitos/patologia , Ratos , Ratos Sprague-Dawley , Proteínas WT1/biossíntese
14.
Mol Reprod Dev ; 85(1): 62-71, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29226557

RESUMO

Culture media modifications, including the addition of various factors, are important for the in vitro production of oocytes and embryos. In this study, we investigated the effects of lysophosphatidic acid (LPA) on porcine embryo development. Porcine parthenogenetic embryos were cultured with 0, 0.1, 1, and 10 µM LPA for 7 days, or cultured in basic medium until Day 4 and then treated with LPA from Days 4 to 7. No difference in the in vitro development of embryos cultured with LPA for 7 days was observed. Conversely, rates of blastocyst and over-expanded blastocyst formation were higher in the 0.1 and 1 µM LPA-treated versus the other groups of embryos treated from Days 4 to 7. Moreover, formation of early blastocysts occurred earlier and embryo size was larger in LPA-treated compared to control embryos. Expression of Connexin 43 and gap junction and cell adhesion-related genes (GJC1 and CDH1, respectively) was also higher in LPA-treated compared to control embryos. Despite no difference in the blastocyst total cell number between groups, the apoptotic index was lower in the LPA-treated group than in the control group; indeed, BCL2L1 (B-cell lymphoma 2-like protein 1) expression increased while BAK (Bcl-2 homologous antagonist killer) decreased in the LPA-treated group. Thus, addition of LPA to the medium from Days 4 to 7 of culture improves blastocyst formation and aids the development of preimplantation embryos.


Assuntos
Blastocisto/citologia , Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Animais , Proteínas Cdh1/biossíntese , Conexina 43/biossíntese , Técnicas de Cultura Embrionária , Partenogênese , Suínos , Proteína Killer-Antagonista Homóloga a bcl-2/biossíntese , Proteína bcl-X/biossíntese
15.
Eur Rev Med Pharmacol Sci ; 21(22): 5283-5289, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29228446

RESUMO

OBJECTIVE: To observe the expressions of myocardial connexin43 (Cx43) and interleukin-17 (IL-17) in acute myocardial infarction (AMI) rats and investigate its possible mechanism of telmisartan in the prevention and treatment of arrhythmia in AMI. MATERIALS AND METHODS: Sprague Dawley (SD) rats were selected and myocardial infarction model was established. After the successful modeling, the rats were randomly divided into three groups: Sham group, MI group, Telm group. Ventricular arrhythmias was induced by the programmed electrical stimulation at 2, 4, 8 weeks. After 8 weeks, rats were sacrificed and heart tissues were collected for immunohistochemistry and Western blot detection. RESULTS: Telmisartan reduced the induction rate of ventricular arrhythmia after myocardial infarction in rats. Telmisartan increased the Cx43 expression while reduced the IL-17 expression in myocardial infarction in rats. Moreover, there was a negative correlation between the expressions of Cx43 and IL-17 after myocardial infarction. CONCLUSIONS: Telmisartan can reduce the occurrence rate of malignant arrhythmias after myocardial infarction, whose mechanism may be increasing the Cx43 expression through inhibition of IL-17 expression.


Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Arritmias Cardíacas/tratamento farmacológico , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Conexina 43/biossíntese , Coração/efeitos dos fármacos , Interleucina-17/antagonistas & inibidores , Infarto do Miocárdio/complicações , Miocárdio/metabolismo , Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Animais , Arritmias Cardíacas/etiologia , Benzimidazóis/uso terapêutico , Benzoatos/uso terapêutico , Estimulação Elétrica , Imuno-Histoquímica , Masculino , Ratos , Ratos Sprague-Dawley , Telmisartan
16.
Biol Reprod ; 97(3): 466-477, 2017 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-29025060

RESUMO

Implantation is a complex event demanding contributions from both embryo and endometrium. Despite advances in assisted reproduction, endometrial receptivity defects persist as a barrier to successful implantation in women with infertility. We previously demonstrated that maternal haploinsufficiency for the endocrine peptide adrenomedullin (AM) in mice confers a subfertility phenotype characterized by defective uterine receptivity and sparse epithelial pinopode coverage. The strong link between AM and implantation suggested the compelling hypothesis that administration of AM prior to implantation may improve fertility, protect against pregnancy complications, and ultimately lead to better maternal and fetal outcomes. Here, we demonstrate that intrauterine delivery of AM prior to blastocyst transfer improves the embryo implantation rate and spacing within the uterus. We then use genetic decrease-of-function and pharmacologic gain-of-function mouse models to identify potential mechanisms by which AM confers enhanced implantation success. In epithelium, we find that AM accelerates the kinetics of pinopode formation and water transport and that, in stroma, AM promotes connexin 43 expression, gap junction communication, and barrier integrity of the primary decidual zone. Ultimately, our findings advance our understanding of the contributions of AM to uterine receptivity and suggest potential broad use for AM as therapy to encourage healthy embryo implantation, for example, in combination with in vitro fertilization.


Assuntos
Adrenomedulina/farmacologia , Endométrio/citologia , Endométrio/efeitos dos fármacos , Fármacos para a Fertilidade Feminina/farmacologia , Fertilidade/efeitos dos fármacos , Junções Intercelulares/efeitos dos fármacos , Útero/citologia , Útero/efeitos dos fármacos , Animais , Comunicação Celular/efeitos dos fármacos , Conexina 43/biossíntese , Decídua/citologia , Decídua/efeitos dos fármacos , Implantação do Embrião/efeitos dos fármacos , Transferência Embrionária , Feminino , Junções Comunicantes/efeitos dos fármacos , Humanos , Camundongos , Camundongos Knockout , Água/metabolismo
17.
Int J Oncol ; 51(6): 1694-1704, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29075794

RESUMO

Interactions between bone marrow stromal cells (BMSCs) and multiple myeloma cells significantly contribute to the progression of multiple myeloma (MM). However, little is known about the molecular mechanisms that regulate these interactions. Connexin-43 (Cx-43) has been implicated in the interplay between BMSCs and MM cells. In this study, we hypothesized that the steroid receptor co-activator-3 (SRC3) expressed in BMSCs regulates the expression of Cx-43 to promote the proliferation and migration of myeloma cells. To address this, we co-cultured a human multiple myeloma cell line, RPMI-8226 transfected with either control BMSCs or sh-SRC3-BMSCs. We found that knocking down SRC3 expression in BMSCs inhibited the proliferation and migration of RPMI-8226 cells. In addition, we found that co-culturing RPMI 8266 cells with BMSCs increased Cx43 expression, while knocking down SRC3 expression in BMSCs decreased Cx43 expression. Moreover, our work revealed that SRC3 in BMSCs regulates Cx43 expression via the mitogen-activated protein kinase (MAPK) pathway. To validate this result in vivo, we knocked down SRC3 expression in BMSCs in nude mice and found that tumor growth and cell apoptosis were significantly decreased. In addition, mice treated with either RPMI 8266 cells overexpressing Cx43 or with a P38 MAPK inhibitor (SB202190) exhibited increased intratumoral leukocyte populations and promoted cell apoptosis in tumor tissue. Our findings demonstrate how SRC3 and Cx43 regulation between BMSCs and myeloma cells mediate cell growth and disease progression, with potential implications for prognosis and therapeutic interventions.


Assuntos
Conexina 43/biossíntese , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Coativador 3 de Receptor Nuclear/metabolismo , Animais , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Técnicas de Cocultura , Feminino , Técnicas de Silenciamento de Genes , Xenoenxertos , Humanos , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Coativador 3 de Receptor Nuclear/biossíntese , Coativador 3 de Receptor Nuclear/deficiência , Coativador 3 de Receptor Nuclear/genética
18.
Gene ; 635: 24-32, 2017 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-28903063

RESUMO

Within the testis, connexin43 encoded by Gja1 plays an important role in cell-to-cell communication between Leydig cells as well as between Sertoli cells and spermatogonia. In the adult male, Leydig cells are the principal producers of testosterone sustaining spermatogenesis, while Sertoli cells nourish, protect and support the differentiating germ cells. It has been shown previously that members of the AP-1 family regulate Gja1 expression in myometrial cells, suggesting that such regulatory mechanism may also be relevant within the testis. Thus, we performed cotransfections of AP-1 expression plasmids with different mouse Gja1 promoter/luciferase reporter constructs within TM3 Leydig and TM4 Sertoli cells. We showed that a functional cooperation between cJun and cFos activates Gja1 expression and requires an AP-1 DNA regulatory element located between -132 and -26 bp. In addition, such synergy relies on the recruitment of cFos to this region of the mouse Gja1 promoter. Hence, our data indicate that AP-1 members are important for optimal expression of Gja1 within Sertoli and Leydig cells from the testis.


Assuntos
Comunicação Celular/genética , Conexina 43/genética , Proteínas Oncogênicas v-fos/genética , Fator de Transcrição AP-1/genética , Animais , Conexina 43/biossíntese , Regulação da Expressão Gênica no Desenvolvimento , Células Intersticiais do Testículo/metabolismo , Masculino , Camundongos , Proteínas Oncogênicas v-fos/biossíntese , Regiões Promotoras Genéticas , Células de Sertoli/metabolismo , Espermatogênese/genética , Testículo/crescimento & desenvolvimento , Testículo/metabolismo , Testosterona/genética , Fator de Transcrição AP-1/biossíntese
19.
Am J Physiol Heart Circ Physiol ; 313(4): H810-H827, 2017 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-28710068

RESUMO

Cardiac fibroblasts (CFs) are known to regulate cardiomyocyte (CM) function in vivo and in two-dimensional in vitro cultures. This study examined the effect of CF activation on the regulation of CM electrical activity in a three-dimensional (3-D) microtissue environment. Using a scaffold-free 3-D platform with interspersed neonatal rat ventricular CMs and CFs, Gq-mediated signaling was selectively enhanced in CFs by Gαq adenoviral infection before coseeding with CMs in nonadhesive hydrogels. After 3 days, the microtissues were analyzed by signaling assay, histological staining, quantitative PCR, Western blots, optical mapping with voltage- or Ca2+-sensitive dyes, and microelectrode recordings of CF resting membrane potential (RMPCF). Enhanced Gq signaling in CFs increased microtissue size and profibrotic and prohypertrophic markers. Expression of constitutively active Gαq in CFs prolonged CM action potential duration (by 33%) and rise time (by 31%), prolonged Ca2+ transient duration (by 98%) and rise time (by 65%), and caused abnormal electrical activity based on depolarization-induced automaticity. Constitutive Gq activation in CFs also depolarized RMPCF from -33 to -20 mV and increased connexin 43 and connexin 45 expression. Computational modeling confers that elevated RMPCF and increased cell-cell coupling between CMs and CFs in a 3-D environment could lead to automaticity. In conclusion, our data demonstrate that CF activation alone is capable of altering action potential and Ca2+ transient characteristics of CMs, leading to proarrhythmic electrical activity. Our results also emphasize the importance of a 3-D environment where cell-cell interactions are prevalent, underscoring that CF activation in 3-D tissue plays a significant role in modulating CM electrophysiology and arrhythmias.NEW & NOTEWORTHY In a three-dimensional microtissue model, which lowers baseline activation of cardiac fibroblasts but enables cell-cell, paracrine, and cell-extracellular matrix interactions, we demonstrate that selective cardiac fibroblast activation by enhanced Gq signaling, a pathophysiological trigger in the diseased heart, modulates cardiomyocyte electrical activity, leading to proarrhythmogenic automaticity.


Assuntos
Potenciais de Ação/fisiologia , Fibroblastos/fisiologia , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/fisiologia , Miócitos Cardíacos/fisiologia , Animais , Animais Recém-Nascidos , Conexina 43/biossíntese , Conexinas/biossíntese , Junções Comunicantes/fisiologia , Potenciais da Membrana/fisiologia , Miócitos Cardíacos/ultraestrutura , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley
20.
Toxicol In Vitro ; 45(Pt 3): 366-373, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28576679

RESUMO

It has been shown that non-cytotoxic doses of Carbendazim (CBZ), a broad-spectrum benzimidazole fungicide, possess endocrine-disrupting (androgen-like) actions, ex vivo, on the pubertal rat seminiferous epithelium. Iprodione (IPR), a dicarboximide fungicide, is also known to be an endocrine-disrupter (anti-androgen). The effect of a mixture of these two pesticides was investigated in the validated rat seminiferous tubule culture model. Cultures were performed in the absence or presence of CBZ 50nM or IPR 50nM either alone or in mixture (Mix), over a 3-week period. Mix exerted a dramatic effect on two proteins (Connexin 43 and Claudin-11) of the blood-testis barrier and possessed similar effects to IPR on some germ cell populations. The presence of IPR together with CBZ (Mix) cancelled the effect of CBZ on the increase of the androgen-dependent TP1 and TP2 mRNAs and on the decrease of ERα, ERß mRNAs. Nevertheless, CBZ alone or IPR alone or Mix induced toxicity on spermatogenesis resulting in a decrease of round spermatids (the precursors of spermatozoa). These results strongly suggest that, even at these low concentrations, the effects of IPR and of CBZ are not solely dependent on their respective anti-androgenic and androgen-like effects and should involve several mechanisms of action.


Assuntos
Aminoimidazol Carboxamida/análogos & derivados , Benzimidazóis/toxicidade , Carbamatos/toxicidade , Disruptores Endócrinos/toxicidade , Fungicidas Industriais/toxicidade , Hidantoínas/toxicidade , Epitélio Seminífero/efeitos dos fármacos , Aminoimidazol Carboxamida/toxicidade , Animais , Barreira Hematotesticular/efeitos dos fármacos , Células Cultivadas , Claudinas/biossíntese , Claudinas/genética , Conexina 43/biossíntese , Conexina 43/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley , Maturidade Sexual , Espermatócitos/efeitos dos fármacos , Espermatogênese/efeitos dos fármacos
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