OBJECTIVETo examine whether calcineurin/NFAT signaling pathway mediates endothelin-1 (ET-1)-induced proliferation of pulmonary artery smooth muscle cells (PASMCs) by regulating phosphodiesterase-5 (PDE5) and the effect of the selective calcineurin inhibitor cyclosporine A and PDE5 inhibitor sildenafil on ET-1-induced PASMC proliferation.

METHODSPASMCs were treated with ET-1 to stimulate their proliferation with or without prior treatment of the cells with CsA or sildenafil. Calcineurin activity in the cells was measured using a calcineurin activity assay kit, PDE5 expression examined using immunoblotting, and cGMP level detected using a cGMP direct immunoassay kit. PASMC proliferation following the treatments was determined using [(3)H]thymidine incorporation assay.

RESULTSET-1 caused a 2.05-fold increase in the cellular calcineurin activity, a 1.80-fold increase in PDE5 expression, and a 3.20-fold increase in the DNA synthesis rate, and reduced the cGMP level by 67%. Pretreatment of the cells with Cyclosporine blocked the effects of ET-1, and PDE5 inhibition by sildenafil pretreatment also abolished ET-1-induced reduction of cGMP level in the cells. Both Cyclosporine and sildenafil suppressed ET-1-stimulated PASMC proliferation.

CONCLUSIONActivation of calcineurin/NFAT signaling pathway mediates ET-1-induced PASMC proliferation by stimulating PDE5 expression, which further degrades cGMP. Both Cyclosporine and sildenafil can suppress ET-1-stimulated PASMC proliferation in vitro.

Animals ; Calcineurin ; metabolism ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cyclic GMP ; metabolism ; Cyclic Nucleotide Phosphodiesterases, Type 5 ; metabolism ; Cyclosporine ; DNA ; biosynthesis ; Endothelin-1 ; pharmacology ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; enzymology ; NFATC Transcription Factors ; metabolism ; Piperazines ; Pulmonary Artery ; cytology ; Purines ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Sildenafil Citrate ; Sulfones

OBJECTIVE: To determine the effect of pregnancy-associated plasma protein A (PAPP-A) on the function of vascular endothelial cells (VEC). METHODS: Human umbilical vein endothelial cell (HUVEC) line, derived from human umbilical vein, was cultured in vitro with PAPP-A at 0, 50, 100, and 200 ng/mL for 0, 12, 24, and 48 hours, respectively. Nitric oxide (NO) levels and endothlin-1 (ET-1) levels were determined by spectrophotometer and immunehistory. RESULTS: The NO levels of HUVECs in the PAPP-A groups were significantly lower than those in the control group (P<0.05). The ET-1 levels of HUVECs in the PAPP-A groups were significantly higher than those in the control group (P<0.05). The changes were all dose-dependent. CONCLUSION PAPP-A may affect the function of vascular endothelial cells by reducing the secretion of NO and increasing the level of ET-1.
Cell Line ; Endothelial Cells ; cytology ; metabolism ; physiology ; Endothelin-1 ; biosynthesis ; Female ; Humans ; Nitric Oxide ; biosynthesis ; Pregnancy-Associated Plasma Protein-A ; pharmacology ; Umbilical Veins ; cytology ; metabolism

OBJECTIVETo investigate the expressions of VEGF in prostate cancer (PCa) and benign prostatic hyperplasia (BPH), their clinical significance and their relationship with that of ET-1.

METHODSA total of 44 specimens of PCa and 36 of BPH tissues were examined by the immunohistochemical Elivision plus method for the expressions of VEGF and ET-1. The intensity of staining for VEGF and ET-1 was assessed by light microscopy on a scale from "-" to "+ + +".

RESULTSThe rates of positive expression of VEGF were 69.4% in BPH and 80.9% in PCa, positive staining mostly in the cytoplasm of glandular epithelia and cancer cells, and strongly positive in all the stroma vascular endothelial cells. The staining intensity of VEGF was significantly higher in the PCa than in the BPH group (P < 0.05) , in the bone metastasis (BM) than in the non-BM group (P < 0.01), and in the lowly than in the highly and moderately differentiated PCa tissues (P < 0.01). The expression of VEGF was positively correlated with that of ET-1 ( r(s) = 0.780, P < 0.01).

CONCLUSIONVEGF is involved in the development, progression and metastasis of PCa. VEGF and ET-1 may play a joint role in its development and progression.

Aged ; Aged, 80 and over ; Endothelin-1 ; biosynthesis ; Humans ; Male ; Middle Aged ; Neoplasm Metastasis ; Neoplasm Staging ; Prostatic Hyperplasia ; metabolism ; pathology ; Prostatic Neoplasms ; metabolism ; pathology ; Vascular Endothelial Growth Factor Receptor-1 ; biosynthesis

OBJECTIVETo investigate the mechanism of rosiglitazone (RSG, the activator of peroxisome proliferators activated receptor lambda) for inhibiting endothelin-1 (ET-1)-induced neonatal rat cardiac myocyte hypertrophy and the role of protein kinase C (PKC) and c-fos.

METHODSIn vitro cultured neonatal rat cardiac myocytes were treated with ET-1, phorbol ester (PMA, the PKC activator), ET-1+RSG, ET-1+chelerythrine (che, the PKC inhibitor), PMA+RSG, or without treatment (control), respectively. The effects of RSG on the protein content, (3)H-leucine incorporation, PKC activity and C-fos protein expression were observed in the cardiac myocytes stimulated with ET-1 or PMA.

RESULTSAfter two days of culture, the intracellular protein content in ET-1 group and PMA group were increased by 15% (339-/+15 microg/ml) and 13% (329-/+14 microg/ml) as compared with the control cells (290-/+13 microg/ml), respectively (P<0.01). Compared with the ET-1 group, cells treated with ET-1+10(-8) mol/L RSG, ET-1+10(-7) mol/L RSG, and ET-1+che showed decreased intracellular protein content by 10% (303-/+14 microg/ml, P<0.05), 12% (292-/+11 microg/ml, P<0.05), and 13% (291-/+12 microg/ml, P<0.01), respectively. The intracellular protein content in PMA+10(-7) mol/LRSG group was decreased by 10% (P<0.05) in comparison with the PMA group. RSG inhibited protein synthesis enhancement and increased (3)H-leucine incorporation induced by ET-1 and PMA, and antagonized the effects of ET-1 and PMA in promoting PKC activity and c-fos protein expression in the myocytes.

CONCLUSIONThe inhibitory effect of RSG on ET-1- or PMA-induced myocyte hypertrophy is associated with PKC-c-fos pathway.

Animals ; Animals, Newborn ; Blotting, Western ; Cell Enlargement ; drug effects ; Cells, Cultured ; Dose-Response Relationship, Drug ; Endothelin-1 ; pharmacology ; Hypoglycemic Agents ; pharmacology ; Myocytes, Cardiac ; cytology ; drug effects ; metabolism ; Protein Kinase C ; metabolism ; Proto-Oncogene Proteins c-fos ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Tetradecanoylphorbol Acetate ; pharmacology ; Thiazolidinediones ; pharmacology

No abstract available.
Blotting, Western ; Cells, Cultured ; Connective Tissue Cells/drug effects/metabolism ; Endothelin-1/*biosynthesis ; Fibroblasts/*drug effects/metabolism ; Humans ; Transforming Growth Factor beta1/*pharmacology

OBJECTIVETo preliminarily investigate the influence of hypoxia on human umbilical vein endothelial cells (HUVECs), and the effect of Ginkgo biloba extract 50 (GBE50) on it.

METHODSFlow cytometry, TUNEL, RT-PCR, Western blot, etc. were applied, to study the effect of hypoxia and GBE50 on endothelial cells.

RESULTSAfter being interfered by hypoxia for 24 h, the levels of reactive oxygen species (ROS) in HUVECs and the apoptotic rate either in the early or in the late stage significantly increased, and accompanied with the increased levels of endothelin-1 mRNA (ET-1) and endothelial oxide synthase (eNOS) protein. However, when HUVECs were pretreated with GBE50 (25 [microg/ml) 4 h before hypoxia, the apoptotic rate in the early or late stage and expression of ET-1 mRNA significantly decreased (P < 0.05), and the heightened ROS level and eNOS expression partially decreased (P > 0.05).

CONCLUSIONHypoxia can induce endothelial dysfunction, which could be partially or significantly reversed by GBE50, it shows a certain protective effect on hypoxia induced endothelial dysfunction.

Blotting, Western ; Cell Hypoxia ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Endothelial Cells ; drug effects ; metabolism ; Endothelin-1 ; biosynthesis ; genetics ; Endothelium, Vascular ; drug effects ; metabolism ; physiopathology ; Free Radical Scavengers ; pharmacology ; Ginkgo biloba ; chemistry ; Humans ; Plant Leaves ; chemistry ; RNA, Messenger ; biosynthesis ; genetics ; Reactive Oxygen Species ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate the different expressions of endothelin-1 ET-1) in prostate cancer (PCa) and benign prostatic hyperplasia (BPH) tissues and their clinical significance.

METHODSA total of 36 BPH and 44 PCa specimens were examined for the expression of ET-1 by immunohistochemical technique (Elivision plus method). The staining intensity for ET-1 was assessed by light microscopy on a scale from "-" to "+ + +".

RESULTSPositive immunoreactivity was found in BPH and PCa, with a positive rate of 100%. Positive staining was located mostly in the cytoplasm of glandular epithelia and smooth muscle cells of both BPH and PCa and was noted in all stroma vascular endothelial cells. These were no significant differences in the intensity of positive staining for ET-1 between the groups of BPH and PCa (P > 0.05), bone metastasis (BM) and non-BM (P > 0.05), and highly and moderately differentiated PCa (P > 0.05), but the staining intensity for ET-1 was significantly higher in the poorly than in the highly and moderately differentiated PCa (P < 0.01).

CONCLUSIONET-1 has a high expression and the localization is the same in both BPH and PCa. It is involved in the development and progression of BPH and PCa.

Aged ; Aged, 80 and over ; Endothelin-1 ; biosynthesis ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Male ; Middle Aged ; Prostate ; metabolism ; pathology ; Prostatic Hyperplasia ; metabolism ; pathology ; Prostatic Neoplasms ; metabolism ; pathology

AIMTo investigate the effects of omapatrilat (OMA) on endothelin-1-induced proliferation of cardiac fibroblasts (CFs).

METHODSIsolated and cultured CFs from neonatal Sprague-Dawley rats (SD) were randomly divided into 7 groups: (1) Control, (2) ET-1, (3) OMA, (4) ET-1 + OMA 10(-9) mol/L, (5) ET-1 + OMA 10(-8) mol/L, (6) ET-1 + OMA 10(-7) mol/L and (7) ET-1 + OMA 10(-6) mol/L. CFs were counted by MTT assay. Cell cycle distribution was determined with a flow cytometer (FCM). [3H]-Proline incorporation was evaluated by scintillation counting. Nitric oxide (NO) was measured by colorimetry.

RESULTS10(-7) mol/L ET-1 significantly increased A490 value and [3H]-Pro incorporation and decreased NO secretion compared with the control group (P < 0.01). 10(-9)-10(-6) mol/L OMA inhibited the effects of ET-1 on CFs in a concentration-dependent manner (P < 0.01 vs. ET-1). In the ET-1 group, the percentage of cells in the S phase was higher than control, which was inhibited by l0(-6) mol/L OMA (P < 0.01 vs. ET-1 and control).

CONCLUSIONOMA can restrain the proliferation and collagen synthesis of cardiac fibroblasts induced by endothelin-1, and this effect may be partially mediated by NO.

Animals ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen ; biosynthesis ; Endothelin-1 ; pharmacology ; Fibroblasts ; cytology ; drug effects ; Myocytes, Cardiac ; cytology ; drug effects ; Nitric Oxide ; metabolism ; Pyridines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Thiazepines ; pharmacology

AIMTo investigate the mechanisms of 17beta-estradiol on the production of endothelin-1 in vascular smooth muscle cells.

METHODSAfter incubation VSMC with various concentrations of 17beta-estradiol (10(-9) - 10(-7) mol/L) or plus L-NAME(10(- 6) mol/L) for different times, the concentration of endothelin-1 was measured. At the same time, the activity of endothelin converting enzyme-1 was analyzed, and the expression of preproET-1mRNA was measured by RT-PCR.

RESULTSIn basal conditions, 17beta-estradiol could inhibit the production of endothelin-1 in VSMC, and the action of 17beta-estradiol had nothing to do with the activity of endothelin converting enzyme-1. L-NAME inhibited the effect of 17-estradiol on the production of endothelin-1 in VSMC. RT-PCR results showed that 17-estradiol inhibited the preproET-1 mRNA expression, and whereas L-NAME reversed this action of 17beta-estradiol.

CONCLUSIONIn basal conditions, 17beta-estradiol decreases the preproET-1 mRNA expression through NO-pathway to inhibit the production of endothelin-1 in cultured VSMC.

Animals ; Cells, Cultured ; Endothelin-1 ; biosynthesis ; Estradiol ; pharmacology ; Muscle, Smooth, Vascular ; cytology ; drug effects ; metabolism ; Myocytes, Smooth Muscle ; drug effects ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects

OBJECTIVETo observe the relationship between protein sythesis and cardiomyocyte viability in neonatal rats.

METHODSThe protein sythesis in neonatal rat cardiomyocytes was measured according to Brandford's method, the absorbance at 490 nm (A(490 nm)) of the cells was measured with MTT assay and the cell viability evaluated by the ratio of A(490 nm) to the total cell number.

RESULTSET-1 increased cardiomyocyte protein synthesis dose-dependently, and this effect was attenuated by the application of lacidipine and tetramethylpyrazines Higher doses of ET-1 resulted in lower A(490 nm)/total cell number ratio, which was further lowered by larcidipine and tetramethylpyrazine.

CONCLUSIONThe status of protein synthesis is not associated with the viability of neonatal rat cardiomyocytes.

Animals ; Animals, Newborn ; Calcium Channel Blockers ; pharmacology ; Cell Survival ; drug effects ; Cells, Cultured ; Dihydropyridines ; pharmacology ; Dose-Response Relationship, Drug ; Endothelin-1 ; pharmacology ; Myocytes, Cardiac ; cytology ; drug effects ; metabolism ; Protein Biosynthesis ; Pyrazines ; pharmacology ; Rats ; Rats, Sprague-Dawley