No abstract available.
Adrenal Cortex Neoplasms/diagnosis/*metabolism/surgery ; Adrenalectomy ; Adrenocortical Adenoma/diagnosis/*metabolism/surgery ; Adult ; Aldosterone/*biosynthesis ; Cushing Syndrome/diagnosis ; Female ; Humans ; Hydrocortisone/*biosynthesis ; Hyperaldosteronism/metabolism

OBJECTIVEIt has been known that the intrahepatic renin-angiotensin-aldosterone system (RAAS) plays a key role in the fibrogenesis in livers. Aldosterone (Aldo), the principal effector molecule of the RAAS, exerts local effects on cell growth and fibrogenesis. However, the signal transduction mechanisms underlying the effects of Aldo on hepatic fibrogenesis remain to be fully elucidated. The present study aims to investigate the signal transduction mechanism underlying the effects of Aldo on the signal passageway of active protein-1 (AP-1).

METHODSIn vitro, HSCs-T6 cell line was treated with Aldo for 10 min, 30 min, 60 min, 120 min and 180 min, and protein expression of Phospho-P42/44 was detected by Western blot. In addition, HSCs-T6 cell line was preincubated for 60 min or not with U0126 (an inhibitor of the MAPK/ERK kinase), and also with antioxidant-N-acetylcysteine (NAC) prior to exposure to Aldo for the indicated times. Protein expression of Phospho-P42/44 was measured by Western blot. DNA biding activity of AP-1 was analyzed by electrophoretic gel mobility shift assay (EMSA). By means of RT-PCR, expression of alpha1(1) procollagen mRNA was detected.

RESULTSAldo stimulated HSC via extracellular signal-regulated kinase (ERK1/2) pathway. Time course experiments showed that Aldo induced Phospho-P42/44 expression, which was abrogated by U0126, reaching a maximum at 10 minutes, and then declined progressively. NAC inhibited the Phospho-P42/44 expression. EMSA showed that stimulation of HSC by Aldo markedly increased AP-1 DNA binding activity. U0126 markedly reduced AP-1 DNA binding activity induced by Aldo; NAC partly decreased AP-1 activity induced by Aldo. Aldo up-regulated expression of alpha1(1) procollagen mRNA, which was attenuated by U0126 and NAC.

CONCLUSIONStimulation of HSC by Aldo results in activation of AP-1 via ERK1/2 pathway, leading to up-regulation of AP-1 target gene alpha1(1) procollagen mRNA expression.

Aldosterone ; pharmacology ; Cell Line ; Collagen Type I ; biosynthesis ; genetics ; Hepatocytes ; cytology ; metabolism ; Humans ; Mitogen-Activated Protein Kinase 3 ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Transcription Factor AP-1 ; metabolism

OBJECTIVEIt is known that intrahepatic renin-angiotensin-aldosterone system (RAAS) plays a key role in liver fibrogenesis. Aldosterone (Aldo), the principal effector molecule of the RAAS, exerts local effects on cell growth and fibrogenesis. However, the signal transduction mechanisms underlying the effects of Aldo on hepatic fibrogenesis remain to be fully elucidated. The present study aims to investigate the signal transduction mechanism underlying the effects of Aldo on extracellular signal-regulated kinase 1/2 (ERK1/2), early growth response-1 (EGR-1) and on the platelet-derived growth factor-B (PDGF-B).

METHODSIn vitro, hepatic stellate cell (HSC)-T6 cell line was treated with Aldo for 10 min, 0.5 h, 1 h, 2 h and 3 h. Protein expression of phospho-p42/44 was detected by Western blot. In addition, HSC-T6 were preincubated for 1 h or not at all with U0126 (an inhibitor of the MAPK/ERK kinase), and antioxidant-N-acetylcysteine (NAC) prior to exposure to Aldo for the indicated times. Protein expressions of phospho-p42/44 and PDGF-B were measured by Western blot. DNA biding activity of EGR-1 was analyzed by electrophoretic gel mobility shift assay (EMSA). By means of immunohistochemistry, expression of PDGF-B was detected.

RESULTSAldo induced phospho-p42/44 expression could be abrogated by U0126; NAC did not inhibit phospho-p42/44 expression. Gel shift study showed that stimulation of HSC by Aldo markedly increased the EGR-1 DNA binding activity, which was abrogated by U0126, reaching a maximum at 60 minutes, and then declined progressively. NAC did not reduce the EGR-1 activity. Aldo increased the PDGF-B protein level in HSC, which was not attenuated by NAC and U0126.

CONCLUSIONSStimulation of HSC by Aldo results in activation of EGR-1 via ERK1/2 pathway, leading to up-regulation of PDGF-B expression.

Aldosterone ; pharmacology ; Cell Line ; Early Growth Response Protein 1 ; biosynthesis ; genetics ; Hepatocytes ; cytology ; metabolism ; Humans ; Mitogen-Activated Protein Kinase 3 ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-sis ; biosynthesis ; genetics ; Signal Transduction

OBJECTIVETo compare the mRNA expression of renin-angiotensin-aldosterone system in human subcutaneous and visceral adipose tissues.

METHODSTotal RNA was extracted from 12 human subcutaneous adipose tissues, 12 perirenal adipose tissue and 9 periadrenal adipose tissues. The expressions of angiotensinogen ( AGT) , renin, angiotensin converting enzyme ( ACE) , angiotensin converting enzyme 2 (ACE2), angiotensin I1 receptor type 1 (AT1), angiotensin II receptor type 2 (AT2 ), CYP11 B2, and their internal reference glyceraldehyde phosphate (GAPDH) were studied by reverse transcription-polymerase chain reaction. The ratios of each target genes were used to evaluate the expression levels of AGT, renin, ACE, ACE2, AT1, AT2, and CYP11B2 in different adipose tissues.

RESULTSThe mRNA expressions of AGT, ACE, ACE2, AT1, and AT2 were detected in human subcutaneous, perirenal, and periadrenal adipose tissues. However, CYPI B2 mRNA expression was not found in these three adipose tissues. The mRNA expressions of renin was only detected in perirenal and periadrenal adipose tissues, which was significantly higher in perirenal adipose tissues than in periadrenal adipose tissues ( P < 0. 05 ). The mRNA expressions of ACE and ACE2 in perirenal adipose tissues were significantly higher than that in subcutaneous adipose tissues ( P < 0. 05). The mRNA expressions of ACE were significantly higher than that of ACE2 in subcutaneous, perirenal, and periadrenal adipose tissues (P <0. 05). The mRNA expressions of AT1 were significantly lower than that of AT2 in periadrenal adipose tissues (P < 0. 05).

CONCLUSIONLocal renin-angiotensin system exists in the adipose tissues; however, aldosterone is not synthesized in the adipose tissues.

Adipose Tissue ; metabolism ; Adult ; Aged ; Aldosterone ; physiology ; Angiotensinogen ; biosynthesis ; Cytochrome P-450 CYP11B2 ; biosynthesis ; Female ; Humans ; Male ; Middle Aged ; Peptidyl-Dipeptidase A ; biosynthesis ; RNA, Messenger ; biosynthesis ; Receptor, Angiotensin, Type 1 ; biosynthesis ; Receptor, Angiotensin, Type 2 ; biosynthesis ; Renin ; biosynthesis ; Renin-Angiotensin System ; physiology ; Reverse Transcriptase Polymerase Chain Reaction

The aim of this study was to examine the effects of endoplasmic reticulum (ER) stress on aldosterone (Aldo)-induced apoptosis of endothelial cells. Glucose-regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP, a hallmark of ER-associated apoptosis) were used to evaluate ER stress. Western blotting and real-time PCR were used to analyze indicators of ER molecule. Apoptosis was detected by annexin V/propidium iodide staining and flow cytometry. Human umbilical vein endothelial cells (HUVECs) were stimulated with different concentrations of Aldo for different durations. Aldo promoted apoptosis of HUVECs and induced ER stress, as evidenced by increased expression of GRP78 and CHOP. siRNA knockdown of CHOP attenuated Aldo-mediated apoptosis. These results indicate that ER stress may be involved in Aldo-induced apoptosis of HUVECs.
Aldosterone ; pharmacology ; Apoptosis ; drug effects ; Endoplasmic Reticulum Stress ; drug effects ; Gene Expression Regulation ; drug effects ; Heat-Shock Proteins ; biosynthesis ; Human Umbilical Vein Endothelial Cells ; cytology ; metabolism ; Humans ; Transcription Factor CHOP ; biosynthesis

Aldosterone ; pharmacology ; Animals ; Cells, Cultured ; Collagen Type I ; biosynthesis ; Collagen Type III ; biosynthesis ; Dose-Response Relationship, Drug ; Liver ; cytology ; drug effects ; metabolism ; Liver Cirrhosis ; etiology ; RNA, Messenger ; analysis ; Rats ; Tissue Inhibitor of Metalloproteinase-1 ; genetics

Emerging evidence indicates that aldosterone and mineralocorticoid receptors (MRs) are associated with the pathogenesis of erectile dysfunction. However, the molecular mechanisms remain largely unknown. In this study, freshly isolated penile corpus cavernosum tissue from rats was treated with aldosterone, with or without MRs inhibitors. Nuclear factor (NF)-kappa B (NF-κB) activity was evaluated by real-time quantitative PCR, luciferase assay, and immunoblot. The results demonstrated that mRNA levels of the NF-κB target genes, including inhibitor of NF-κB alpha (IκB-α), NF-κB1, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6), were higher after aldosterone treatment. Accordingly, phosphorylation of p65/RelA, IκB-α, and inhibitor of NF-κB kinase-β was markedly increased by aldosterone. Furthermore, knockdown of MRs prevented activation of the NF-κB canonical pathway by aldosterone. Consistent with this finding, ectopic overexpression of MRs enhanced the transcriptional activation of NF-κB by aldosterone. More importantly, the MRs antagonist, spironolactone blocked aldosterone-mediated activation of the canonical NF-κB pathway. In conclusion, aldosterone has an inflammatory effect in the corpus cavernosum penis, inducing NF-κB activation via an MRs-dependent pathway, which may be prevented by selective MRs antagonists. These data reveal the possible role of aldosterone in erectile dysfunction as well as its potential as a novel pharmacologic target for treatment.
Aldosterone/pharmacology* ; Animals ; Cytokines/biosynthesis* ; Gene Knockdown Techniques ; I-kappa B Kinase/antagonists & inhibitors* ; Interleukin-6/genetics* ; Male ; Mineralocorticoid Receptor Antagonists/pharmacology* ; NF-kappa B/genetics* ; Penis/metabolism* ; Protein Serine-Threonine Kinases/antagonists & inhibitors* ; RNA, Messenger/biosynthesis* ; Rats ; Rats, Inbred WKY ; Receptors, Mineralocorticoid/genetics* ; Signal Transduction/drug effects* ; Spironolactone/pharmacology* ; Transcriptional Activation ; Tumor Necrosis Factor-alpha/biosynthesis* ; NF-kappaB-Inducing Kinase

Aldosterone has been shown to stimulate renal TGF-beta1 expression. However, the mechanisms for aldosterone-induced TGF-beta1 expression have not been clearly determined in mesangial cells. We examined the role of extracellular-signal regulated kinase 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK) and activator protein- 1 (AP-1) in the aldosterone-induced TGF-beta1 expression in rat mesangial cells. TGF-beta1 protein in the conditioned medium released from rat mesangial cells was measured by sandwich ELISA, TGF-beta1 mRNA expression was analyzed by Northern blotting, AP-1 DNA binding activity was measured by EMSA and the ERK1/2, JNK activity was analyzed by western blotting. Aldosterone significantly stimulated TGF-beta1 protein production and TGF-beta1 mRNA expression in mesangial cells in a dose-dependent manner. Aldosterone significantly increased AP-1 DNA binding activity in mesangial cells. Pre-treatment of cells with AP-1 inhibitor, curcumin, blocked aldosterone-induced AP-1 DNA binding activity as well as aldosterone-induced TGF-beta1 production. Aldosterone increased phosphorylation of ERK1/2 and JNK in mesangial cells. Pre-treatment of cells with ERK1/2 inhibitor, PD98059, or JNK inhibitor, SP600125 significantly inhibited aldosterone-induced ERK1/2 and JNK activity and subsequently TGF-beta1 production, respectively. We conclude that aldosteroneinduced TGF-beta1 expression in mesangial cells is regulated by the ERK1/ 2, JNK and AP-1 intracellular signaling pathways.
Aldosterone/*pharmacology ; Animals ; Culture Media, Conditioned/pharmacology ; DNA/metabolism ; Extracellular Signal-Regulated MAP Kinases/metabolism ; *Gene Expression Regulation, Enzymologic ; Humans ; *MAP Kinase Signaling System ; Mesangial Cells/*metabolism ; Models, Biological ; Phosphorylation ; Protein Binding ; Rats ; Transcription Factor AP-1/*metabolism ; Transforming Growth Factor beta1/*biosynthesis