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1.
Mol Cell Biochem ; 396(1-2): 87-98, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25063217

RESUMEN

Phenotype modulation of pulmonary artery smooth muscle cells (PASMCs) plays an important role during hypoxia-induced vascular remodeling and pulmonary hypertension (PAH). We had previously shown that calcium-sensing receptor (CaSR) is expressed in rat PASMCs. However, little is known about the role of CaSR in phenotypic modulation of PASMCs in hypoxia-induced PAH as well as the underlying mechanisms. In this study, we investigated whether CaSR induces the proliferation of PASMCs in small pulmonary arteries from both rats and human with PAH. PAH was induced by exposing rats to hypoxia for 7-21 days. The mean pulmonary arterial pressure (mPAP), right ventricular hypertrophy index (RVI), the percentage of medial wall thickness to the external diameter (WT %), and cross-sectional total vessel wall area to the total area (WA %) of small pulmonary arteries were determined by hematoxylin and eosin (HE), masson trichrome and Weigert's staining. The protein expressions of matrix metalloproteinase (MMP)-2 and MMP-9, the tissue inhibitors of metalloproteinase (TIMP)-3, CaSR, proliferating cell nuclear antigen (PCNA), phosphorylated extracellular signal-regulated kinase (p-ERK), and smooth muscle cell (SMC) phenotype marker proteins in rat small pulmonary arteries, including calponin, SMα-actin (SMAα), and osteopontin (OPN), were analyzed by immunohistochemistry and Western blotting, respectively. In addition, immunohistochemistry was applied to paraffin-embedded human tissues from lungs of normal human and PAH patients with chronic heart failure (PAH/CHF). Compared with the control group, mPAP, RVI, WT % and WA % in PAH rats were gradually increased with the prolonged hypoxia. At the same time, the expressions of CaSR, MMP-2, MMP-9, TIMP-3, PCNA, OPN, and p-ERK were markedly increased, while the expressions of SMAα and calponin were significantly reduced in lung tissues or small pulmonary arteries of PAH rats. Neomycin (an agonist of CaSR) enhanced but NPS2390 (an antagonist of CaSR) weakened these hypoxic effects. We further found that the expression change of CaSR, PCNA, and SMC phenotypic marker proteins in PAH/CHF lungs was similar to those in PAH rats. Our data suggest that CaSR is involved in the pulmonary vascular remodeling and PAH by promoting phenotypic modulation of small pulmonary arteries.


Asunto(s)
Hipertensión Pulmonar/metabolismo , Arteria Pulmonar/metabolismo , Receptores Sensibles al Calcio/metabolismo , Remodelación Vascular/fisiología , Animales , Modelos Animales de Enfermedad , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/patología , Humanos , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/patología , Hipertrofia Ventricular Derecha/fisiopatología , Hipoxia/complicaciones , Hipoxia/metabolismo , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Antígeno Nuclear de Célula en Proliferación/metabolismo , Arteria Pulmonar/patología , Ratas Wistar , Valores de Referencia , Inhibidor Tisular de Metaloproteinasa-3/metabolismo
2.
Differentiation ; 85(1-2): 32-40, 2013 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-23314289

RESUMEN

The calcium-sensing receptor (CaSR), a G protein coupled receptor, is involved in a number of physiological and pathological processes. Embryonic stem cells (ESCs) have a potential role to differentiate into all types of cells. Whether CaSR is functionally expressed in ESCs is unclear. In this study, the expression and distribution of CaSR in 129 mouse ES-D3 cell lines were detected by Western blotting and immunofluorescence; and the intracellular calcium concentration ([Ca(2+)]i) was measured using Laser Confocal Scanning Microscopy. Mouse embryonic stem cells (mESCs) were cultured to embryoid bodies (EBs) and the differentiation of EBs into cardiomyocytes was induced by icariin (ICA). The cardiac specific proteins, a-Actinin and cardiac troponin-I (cTnI), were analyzed by immunofluorescence, and the differentiation rate was analyzed by flow cytometry. The expression of cardiac-specific transcription factors, Nkx2.5 and GATA-4, was detected by Western blotting. We found that the CaSR protein exists in both mESCs and mESC-derived cardiomyocytes (mESC-CMs). Increasing extracellular calcium or neomycin (an agonist of CaSR) increased [Ca(2+)]i and the differentiation rate. These effects were abolished by inhibition of CaSR, phospholipase C, IP3 receptor and Ca(2+) ATPase, or by depletion of the sarcoplasmic reticulum Ca(2+) store, respectively. Activation of CaSR up-regulated protein expression of Nkx2.5 and GATA4 in EBs at an early stage of ICA-induced differentiation. In conclusion, CaSR is functionally expressed in mESCs, and activation of CaSR is involved in the differentiation of mESCs into cardiomyocytes by facilitating the expression of NKx2.5 and GATA-4.


Asunto(s)
Diferenciación Celular , Células Madre Embrionarias/metabolismo , Miocitos Cardíacos/metabolismo , Receptores Sensibles al Calcio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Actinina/genética , Actinina/metabolismo , Animales , Calcio/metabolismo , Línea Celular , Cuerpos Embrioides/citología , Cuerpos Embrioides/metabolismo , Células Madre Embrionarias/citología , Flavonoides/farmacología , Factor de Transcripción GATA4/genética , Factor de Transcripción GATA4/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteína Homeótica Nkx-2.5 , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Ratones , Miocitos Cardíacos/citología , Neomicina/farmacología , Receptores Sensibles al Calcio/genética , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Troponina I/genética , Troponina I/metabolismo , Regulación hacia Arriba
3.
Mol Cell Biochem ; 362(1-2): 115-22, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22083546

RESUMEN

Matrix metalloproteinase-2 (MMP-2) is constitutively expressed in vascular smooth muscle cells (VSMCs) and up-regulated in atherosclerotic lesion by various stimuli, such as oxidized low-density lipoprotein (oxLDL). Calcium-sensing receptor (CaSR) is also expressed in VSMCs, but it remains unclear whether CaSR is associated with overproduction of MMP-2 in VSMCs. In this study, the expression of MMP-2 was detected by real-time PCR and Western blot analysis, and the gelatinolytic activity of MMP-2 was measured using gelatin zymography. Our results showed that oxLDL enhanced MMP-2 expression and activity in rat aortic VSMCs in a time- and dose-dependent manner. In addition, CaSR expression was up-regulated by oxLDL. Manipulating CaSR function in these cells by NPS2390 (an antagonist of CaSR) or GdCl(3) (an agonist of CaSR) affected the oxLDL-induced MMP-2 production. In VSMCs, oxLDL stimulated the rapid activation of phosphatidylinositol 3-kinase (PI3K)/Akt signal pathway, as determined by Western blot analysis. Phosphorylation of Akt and MMP-2 production stimulated by oxLDL were attenuated by LY294002 (a specific inhibitor of PI3K). Activation of Akt was suppressed by NPS2390 but enhanced by GdCl(3). In contrast, oxLDL had no stimulatory effect on the phosphorylation of JNK, and pretreatment with SP600125 (an inhibitor of JNK) produced no significant effect on oxLDL-induced MMP-2 production. These results suggest that CaSR mediates oxLDL-induced MMP-2 production in VSMCs via PI3K/Akt signal pathway.


Asunto(s)
Lipoproteínas LDL/metabolismo , Metaloproteinasa 2 de la Matriz/biosíntesis , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores Sensibles al Calcio/metabolismo , Adamantano/análogos & derivados , Adamantano/farmacología , Animales , Antracenos/farmacología , Aorta/metabolismo , Aterosclerosis/metabolismo , Células Cultivadas , Cromonas/farmacología , Gadolinio/farmacología , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Sistema de Señalización de MAP Quinasas , Masculino , Metaloproteinasa 2 de la Matriz/metabolismo , Morfolinas/farmacología , Músculo Liso Vascular/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Quinoxalinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores Sensibles al Calcio/biosíntesis
4.
J Biomed Sci ; 18: 16, 2011 Feb 11.
Artículo en Inglés | MEDLINE | ID: mdl-21314926

RESUMEN

BACKGROUND: The extracellular calcium-sensing receptor (CaSR) belongs to family C of the G protein coupled receptors. Whether the CaSR is expressed in the pulmonary artery (PA) is unknown. METHODS: The expression and distribution of CaSR were detected by RT-PCR, Western blotting and immunofluorescence. PA tension was detected by the pulmonary arterial ring technique, and the intracellular calcium concentration ([Ca2+]i) was detected by a laser-scanning confocal microscope. RESULTS: The expressions of CaSR mRNA and protein were found in both rat pulmonary artery smooth muscle cells (PASMCs) and PAs. Increased levels of [Ca2+]o (extracellular calcium concentration) or Gd3+ (an agonist of CaSR) induced an increase of [Ca2+]i and PAs constriction in a concentration-dependent manner. In addition, the above-mentioned effects of Ca2+ and Gd3+ were inhibited by U73122 (specific inhibitor of PLC), 2-APB (specific antagonist of IP3 receptor), and thapsigargin (blocker of sarcoplasmic reticulum calcium ATPase). CONCLUSIONS: CaSR is expressed in rat PASMCs, and is involved in regulation of PA tension by increasing [Ca2+]i through G-PLC-IP3 pathway.


Asunto(s)
Arteria Pulmonar/metabolismo , Receptores Sensibles al Calcio/genética , Animales , Secuencia de Bases , Western Blotting , Compuestos de Boro/farmacología , Calcio/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Inhibidores Enzimáticos/farmacología , Estrenos/farmacología , Técnica del Anticuerpo Fluorescente , Técnicas In Vitro , Masculino , Datos de Secuencia Molecular , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/metabolismo , Arteria Pulmonar/citología , Pirrolidinonas/farmacología , ARN Mensajero/genética , Ratas , Ratas Wistar , Receptores Sensibles al Calcio/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tapsigargina/farmacología , Fosfolipasas de Tipo C/antagonistas & inhibidores
5.
J Biomed Sci ; 18: 18, 2011 Feb 16.
Artículo en Inglés | MEDLINE | ID: mdl-21324201

RESUMEN

BACKGROUND: Myocardial ischemia/reperfusion injury is the major cause of morbidity and mortality for cardiovascular diseases. Dopamine D2 receptors are expressed in cardiac tissues. However, the roles of dopamine D2 receptors in myocardial ischemia/reperfusion injury and cardiomyocyte apoptosis are unclear. Here we investigated the effects of both dopamine D2 receptors agonist (bromocriptine) and antagonist (haloperidol) on apoptosis of cultured neonatal rat ventricular myocytes induced by ischemia/reperfusion injury. METHODS: Myocardial ischemia/reperfusion injury was simulated by incubating primarily cultured neonatal rat cardiomyocytes in ischemic (hypoxic) buffer solution for 2 h. Thereafter, these cells were incubated for 24 h in normal culture medium. RESULTS: Treatment of the cardiomyocytes with 10 µM bromocriptine significantly decreased lactate dehydrogenase activity, increased superoxide dismutase activity, and decreased malondialdehyde content in the culture medium. Bromocriptine significantly inhibited the release of cytochrome c, accumulation of [Ca2+]i, and apoptosis induced by ischemia/reperfusion injury. Bromocriptine also down-regulated the expression of caspase-3 and -9, Fas and Fas ligand, and up-regulated Bcl-2 expression. In contrast, haloperidol (10 µM) had no significant effects on the apoptosis of cultured cardiomyocytes under the aforementioned conditions. CONCLUSIONS: These data suggest that activation of dopamine D2 receptors can inhibit apoptosis of cardiomyocytes encountered during ischemia/reperfusion damage through various pathways.


Asunto(s)
Daño por Reperfusión Miocárdica/metabolismo , Miocitos Cardíacos/metabolismo , Receptores de Dopamina D2/metabolismo , Animales , Animales Recién Nacidos , Apoptosis , Bromocriptina/farmacología , Calcio/metabolismo , Células Cultivadas , Antagonistas de los Receptores de Dopamina D2 , Haloperidol/farmacología , Masculino , Isquemia Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Ratas , Ratas Wistar , Receptores de Dopamina D2/agonistas
6.
EBioMedicine ; 40: 276-289, 2019 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-30594556

RESUMEN

BACKGROUND: Endometrial cancer is one of the most common gynecological malignancies and has exhibited an increasing incidence rate in recent years. Cancer stem cells (CSCs), which are responsible for tumor growth and chemoresistance, have been confirmed in endometrial cancer. However, it is still challenging to identify endometrial cancer stem cells to then target for therapy. METHODS: Flow cytometry was used to identify the endometrial cancer stem cells. Sphere formation assay, western blotting, qRT-PCR assay, cell viability assay, xenograft assay and immunohistochemistry staining analysis were utilized to evaluate the effect of SPARC-related modular calcium binding 2 (SMOC-2) on the cells proliferation and drug resistance. Cell viability assay, qRT-PCR assay, immunofluorescence staining, Co-IP assay and luciferase reporter gene assay were performed to explore the possible molecular mechanism by which SMOC-2 activates WNT/ß-catenin pathway. FINDINGS: We found the expression of SPARC-related modular calcium binding 2 (SMOC-2), a member of SPARC family, was higher in endometrial CSCs than that in non-CSCs. SMOC-2 was also more highly expressed in spheres than in monolayer cultures. The silencing of SMOC-2 suppressed cell sphere ability; reduced the expression of the stemness-associated genes SOX2, OCT4 and NANOG; and enhanced chemosensitivity in endometrial cancer cells. By co-culture IP assay, we demonstrated that SMOC-2 directly interacted with WNT receptors (Fzd6 and LRP6), enhanced ligand-receptor interaction with canonical WNT ligands (Wnt3a and Wnt10b), and finally, activated the WNT/ß-catenin pathway in endometrial cancer. SMOC-2 expression was closely correlated with CSC markers CD133 and CD44 expression in endometrial cancer tissue. INTERPRETATION: Taken together, we conclude that SMOC-2 might be a novel endometrial cancer stem cell signature gene and therapeutic target for endometrial cancer. FUND: National Natural Science Foundation of China, Scientific and Technological Innovation Act Program of Shanghai Science and Technology Commission, Scientific and Technological Innovation Act Program of Fengxian Science and Technology Commission, Natural Science Foundation of Shanghai.


Asunto(s)
Proteínas de Unión al Calcio/genética , Resistencia a Antineoplásicos/genética , Neoplasias Endometriales/genética , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Animales , Biomarcadores , Línea Celular Tumoral , Supervivencia Celular/genética , Modelos Animales de Enfermedad , Neoplasias Endometriales/metabolismo , Neoplasias Endometriales/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Genes Reporteros , Humanos , Ratones , Clasificación del Tumor , Estadificación de Neoplasias , Paclitaxel/farmacología , Vía de Señalización Wnt/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
7.
Sheng Li Xue Bao ; 59(2): 133-40, 2007 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-17437034

RESUMEN

Hydrogen sulfide (H(2)S) is among a family of endogenous molecules of gas, defined as gasotransmitters. In recent years, endogenous production of H(2)S and its physiological importance have been realized. Abnormal metabolism and functions of H(2)S contribute to or participate in the pathogenesis of many diseases. This article reviews recent discoveries on the roles of H(2)S in the regulation of cell proliferation and apoptosis. The molecular mechanisms for the cellular effects of H(2)S are also recapitulated, including changes in mitogen-activated protein kinase, cell cycle-related kinase, cell death-related gene and ion channels. A better understanding of H(2)S-regualted cell growth or death will pave way for future design of novel pharmacological and therapeutic interventions for various diseases.


Asunto(s)
Apoptosis/fisiología , Proliferación Celular , Sulfuro de Hidrógeno/metabolismo , Animales , Quinasas Ciclina-Dependientes/metabolismo , Humanos , Proteínas Quinasas Activadas por Mitógenos/metabolismo
8.
Sci Rep ; 7: 44744, 2017 03 17.
Artículo en Inglés | MEDLINE | ID: mdl-28303973

RESUMEN

Cervical cancer is an infectious cancer and the most common gynecologic cancer worldwide. E6/E7, the early genes of the high-risk mucosal human papillomavirus type, play key roles in the carcinogenic process of cervical cancer. However, little was known about its roles in modulating tumor microenvironment, particular extracellular matrix (ECM). In this study, we found that E6/E7 could regulate multiple ECM proteins, especially collagen triple helix repeat containing 1 (CTHRC1). CTHRC1 is highly expressed in cervical cancer tissue and serum and closely correlated with clinicopathological parameters. CTHRC1 promotes cervical cancer cell migration and invasion in vitro and metastasis in vivo. E6/E7 regulates the expression of CTHRC1 in cervical cancer by E6/E7-p53-POU2F1 (POU class 2 homeobox 1) axis. Futhermore, CTHRC1 activates Wnt/PCP signaling pathway. Take together, E6/E7-p53-POU2F1-CTHRC1 axis promotes cervical cancer cell invasion and metastasis and may act as a potential therapeutic target for interventions against cervical cancer invasion and metastasis.


Asunto(s)
Polaridad Celular , Proteínas de la Matriz Extracelular/metabolismo , Factor 1 de Transcripción de Unión a Octámeros/metabolismo , Proteínas Oncogénicas Virales/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Neoplasias del Cuello Uterino/metabolismo , Neoplasias del Cuello Uterino/patología , Vía de Señalización Wnt , Movimiento Celular/genética , Proliferación Celular/genética , Proteínas de la Matriz Extracelular/sangre , Proteínas de la Matriz Extracelular/genética , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Silenciador del Gen , Humanos , Persona de Mediana Edad , Metástasis de la Neoplasia , Curva ROC , Regulación hacia Arriba/genética , Neoplasias del Cuello Uterino/sangre , Neoplasias del Cuello Uterino/genética
9.
Cancer Lett ; 363(1): 71-82, 2015 Jul 10.
Artículo en Inglés | MEDLINE | ID: mdl-25864591

RESUMEN

Ovarian cancer remains the disease with the highest associated mortality rate of gynecologic malignancy due to cancer metastasis. Rearrangement of actin cytoskeleton by cytoskeleton protein plays a critical role in tumor cell metastasis. MICAL-L2, a member of MICAL family, can interact with actin-binding proteins, regulate actin cross-linking and coordinate the assembly of adherens junctions and tight junctions. However, the roles of MICAL-L2 in tumors and diseases have not been explored. In this study, we found that MICAL-L2 protein is significantly up-regulated in ovarian cancer tissues along with FIGO stage and associated with histologic subgroups of ovarian cancer. Silencing of MICAL-L2 suppressed ovarian cancer cell proliferation, migration and invasion ability. Moreover, silencing of MICAL-L2 prevented nuclear translocation of ß-catenin, inhibited canonical wnt/ß-catenin signaling and induced the mesenchymal-epithelial transition (MET). Taken together, our data indicated that MICAL-L2 may be an important regulator of epithelial-mesenchymal transition (EMT) in ovarian cancer cells and a new therapeutic target for interventions against ovarian cancer invasion and metastasis.


Asunto(s)
Transición Epitelial-Mesenquimal/genética , Regulación Neoplásica de la Expresión Génica , Proteínas de Microfilamentos/genética , Neoplasias Ováricas/genética , Interferencia de ARN , Animales , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Femenino , Humanos , Ratones Desnudos , Proteínas de Microfilamentos/metabolismo , Invasividad Neoplásica , Estadificación de Neoplasias , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Fenotipo , Factores de Tiempo , Transfección , Vía de Señalización Wnt , beta Catenina/metabolismo
10.
Int J Clin Exp Pathol ; 7(4): 1348-58, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24817931

RESUMEN

Endometrial carcinoma (EC) is the most common gynecologic cancer worldwide and is one of the leading causes of death in women. Therefore, it is urgent to elucidate the pathological mechanisms of EC. SERPINA3 is a member of the serpin super-family of protease inhibitors. Its aberrant expression has been observed in various tumor cells. However, its clinical significance and biological function in endometrial cancer remains unknown. In the present study, we demonstrated that SERPINA3 expression was significantly up-regulated in EC samples and was closely correlated with lower differentiation, higher stage, positive lymph node or vascular thrombosis and negative estrogen receptor (ER), indicating a poor prognosis. We then demonstrated that SERPINA3 promoted EC cells proliferation by regulating G2/M checkpoint in cell cycle and inhibited cells apoptosis, and we further uncovered that the pro-proliferative effect of SERPINA3 on EC was likely ascribed to the activation of MAPK/ERK1/2 and PI3K/AKT signaling. The results of our study may provide insight into the application of SERPINA3 as a novel predictor of clinical outcomes and a potential therapeutic target of EC.


Asunto(s)
Apoptosis/fisiología , Puntos de Control del Ciclo Celular/fisiología , Proliferación Celular/fisiología , Neoplasias Endometriales/fisiopatología , Puntos de Control de la Fase G2 del Ciclo Celular/fisiología , Puntos de Control de la Fase M del Ciclo Celular/fisiología , Serpinas/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Neoplasias Endometriales/patología , Femenino , Humanos , Técnicas In Vitro , Sistema de Señalización de MAP Quinasas/fisiología , Persona de Mediana Edad , Quinasas de Proteína Quinasa Activadas por Mitógenos/fisiología , Estadificación de Neoplasias , Fosfatidilinositol 3-Quinasas/fisiología , Proteínas Proto-Oncogénicas c-akt/fisiología , Transducción de Señal/fisiología , Regulación hacia Arriba/fisiología
11.
Diabetes Res Clin Pract ; 95(3): 378-85, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22137362

RESUMEN

To observe the dynamic expression of calcium-sensing receptor (CaSR) in myocardium of diabetic rats and explore its role in diabetic cardiomyopathy (DCM), 40 male Wistar rats were randomly divided into 4 groups including control, diabetic-4 weeks, diabetic-8 weeks and spermine treatment groups (240 µM of spermine in drinking water). The type 2 Diabetes mellitus (DM) models were established by intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) after high-fat and high-sugar diet for one month. The echocardiographic parameters were measured, cardiac morphology was observed by electron microscope and HE staining. The intracellular calcium concentration ([Ca(2+)](i)) was detected by laser-scanning confocal microscope. Western blot analyzed the expression of CaSR, protein kinase C α(PKC-α) and calcium handling regulators, such as phospholamban (PLN), Ca(2+)-ATPase (SERCA), and ryanodine receptor (RyR). Compared with control group, [Ca(2+)](i) and the expression of CaSR, RyR and SERCA/PLN were decreased, while PKC-α and PLN were significantly increased in a time-dependent manner in diabetic groups. Meanwhile diabetic rats displayed abnormal cardiac structure and systolic and diastolic dysfunction, and spermine (CaSR agonist) could prevent or slow its progression. These results indicate that the CaSR expression of myocardium is reduced in the progress of DCM, and its potential mechanism is related to the impaired intracellular calcium homeostasis.


Asunto(s)
Cardiomiopatías Diabéticas/metabolismo , Receptores Sensibles al Calcio/metabolismo , Animales , Calcio/análisis , Diabetes Mellitus Experimental , Homeostasis , Masculino , Miocardio/metabolismo , Ratas , Ratas Wistar , Receptores Sensibles al Calcio/análisis , Espermina/uso terapéutico
12.
Basic Clin Pharmacol Toxicol ; 108(3): 185-93, 2011 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-21073657

RESUMEN

Activation of the calcium-sensing receptor (CaSR) leads to an increase of intracellular calcium concentration and alteration of cellular activities. High level of intracellular calcium is involved in hypoxia-induced proliferation of pulmonary arterial smooth muscle cells (PASMCs). However, whether the CaSR is expressed in PAMSCs and is related to the hypoxia-induced proliferation of PASMCs is unclear. In this study, the expression and distribution of CaSRs were detected by RT-PCR, western blotting and immunofluorescence; the intracellular concentration of free calcium ([Ca(2+) ](i) ) was determined by confocal laser scanning microscopy; cell proliferation was tested using an MTT and BrdU incorporation assay; cell cycle analysis was carried out using a flow cytometric assay; and the expression of proliferating cell nuclear antigen (PCNA), extracellular signal-regulated protein kinase 1,2 (ERK1,2) and AKT were analysed by western blotting. We observed that both CaSR mRNA and protein were expressed in rat PASMCs. Lowering of oxygen from 21% to 2.5% led to increased [Ca(2+) ](i) and CaSR expression. This condition of hypoxia also stimulated PASMCs proliferation accompanying with increased phosphorylation of ERK1,2 and AKT. GdCl(3) (an agonist of CaSR) or NPS2390 (an antagonist of CaSR) amplified or weakened the effect of hypoxia, respectively. PD98059 (a MEK1 inhibitor) or LY294002 (a PI3K inhibitors) decreased the up-regulation of PCNA expression and the increase of the cell proliferation index induced by hypoxia and GdCl(3) in PASMCs. Our results suggest that CaSR is expressed in rat PASMCs, and that CaSR activation through MEK1/ERK1,2 and PI3 kinase pathways is involved in hypoxia-induced proliferation of PASMCs.


Asunto(s)
Hipoxia de la Célula , Proliferación Celular , Sistema de Señalización de MAP Quinasas , Músculo Liso Vascular/metabolismo , Fosfatidilinositol 3-Quinasa/metabolismo , Arteria Pulmonar/metabolismo , Receptores Sensibles al Calcio/metabolismo , Animales , Señalización del Calcio/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Inhibidores Enzimáticos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Masculino , Músculo Liso Vascular/citología , Músculo Liso Vascular/efectos de los fármacos , Inhibidores de las Quinasa Fosfoinosítidos-3 , Fosforilación/efectos de los fármacos , Antígeno Nuclear de Célula en Proliferación/metabolismo , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Arteria Pulmonar/citología , Arteria Pulmonar/efectos de los fármacos , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptores Sensibles al Calcio/agonistas , Receptores Sensibles al Calcio/antagonistas & inhibidores , Receptores Sensibles al Calcio/genética
13.
Shi Yan Sheng Wu Xue Bao ; 35(2): 117-22, 2002 Jun.
Artículo en Zh | MEDLINE | ID: mdl-15344329

RESUMEN

Bt-CpTI fusion protein gene was transferred to the explants of hypocotyl, cotyledon with petiole and shoot apex of cabbage (Brassica oleracea L. var. capitata) variety "Zhonggan No 8" via Agrobacterium tumefaciens and particle bombardment, and 13 kanamycin-resistant plants were obtained. PCR and Southern blotting hybridization verified that all these plants of the kanr plants of type I regenerated from hypocotyl and cotyledon with petiole mediated by A. tumefaciens were transgenic plants, 2 karr plants of type II stemed from shoot apex mediated by A. tumefaciens were "false-positive"plants and one of 2 kanr plants of type III regenerated from shoot apex via particle bombardment was non-transformed plants. It was showed that part of transgenic plants had high activity of trypsin inhibitor and strong resistance to resist common cabbage worm through the analysis of CpTI relative capacity and insect-resistant test.


Asunto(s)
Agrobacterium tumefaciens/genética , Proteínas Bacterianas/genética , Brassica/genética , Endotoxinas/genética , Proteínas Hemolisinas/genética , Plantas Modificadas Genéticamente/genética , Proteínas Recombinantes de Fusión/genética , Inhibidores de Tripsina/genética , Toxinas de Bacillus thuringiensis , Southern Blotting , Reacción en Cadena de la Polimerasa
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