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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 2009 Feb 17;106(10):3782–3787. doi: 10.1073/pnas.0804037106

Aldosterone activates endothelial exocytosis

Youngtae Jeong a,b,c,1, Damian F Chaupin a, Kenji Matsushita a,1, Munekazu Yamakuchi a, Scott J Cameron b, Craig N Morrell b,d, Charles J Lowenstein a,b,2
PMCID: PMC2656157  PMID: 19223584

Abstract

Although elevated levels of aldosterone are associated with vascular inflammation, the proinflammatory pathways of aldosterone are not completely defined. We now show that aldosterone triggers endothelial cell exocytosis, the first step in leukocyte trafficking. Exogenous aldosterone stimulates endothelial exocytosis of Weibel-Palade bodies, externalizing P-selectin and releasing von Willebrand factor. Spironolactone, a nonselective mineralocorticoid receptor (MR) blocker, antagonizes aldosterone-induced endothelial exocytosis. Knockdown of the MR also decreases exocytosis, suggesting that the MR mediates exocytosis. Aldosterone triggers exocytosis within minutes, and this effect is not inhibited by actinomycin D, suggesting a nongenomic effect of aldosterone. Aldosterone treatment of endothelial cells increases leukocyte adherence to endothelial cells in culture. Taken together, our data suggest that aldosterone activates vascular inflammation in part through nongenomic, MR-mediated pathways. Aldosterone antagonism may decrease vascular inflammation and cardiac fibrosis in part by blocking endothelial exocytosis.

Keywords: inflammation, mineralocorticoid, P-selectin, vascular, nitric oxide

Aldosterone regulates Na+ and K+ homeostasis, in part by modulating Na+ resorption in the kidney. The mineralocorticoid receptor (MR) in epithelial cells lining the distal nephron mediates aldosterone regulation of electrolytes. However, aldosterone may have additional physiological effects, such as increasing fibrosis, modulating oxidative stress, and triggering vascular inflammation (1–7).

Hyperaldosteronism is associated with vascular inflammation (1, 2, 8–14). Aldosterone infusion induces leukocyte infiltration into arteries of rats and increases the expression of proinflammatory markers (15, 16). Mineralocorticoid receptor (MR) antagonists decrease monocyte infiltration into the vascular wall in hypertensive rats, in rats treated with aldosterone, and in hypertensive mice (17–21). Clinical studies also show that aldosterone increases markers of vascular inflammation. Patients with primary hyperaldosteronism have increased inflammatory biomarkers (22). Infusion of aldosterone increases IL-6 and IL-12 levels in humans (23, 24). Conversely, MR antagonists decrease markers of oxidative stress and acute-phase reactants in humans (25). Thus animal and clinical data suggest that aldosterone promotes vascular inflammation.

The mechanisms by which aldosterone affects vascular inflammation are not fully elucidated. Aldosterone interacting with the MR activates the transcription of a set of proinflammatory genes (26, 27). For example, aldosterone increases vascular smooth muscle cell expression of IL-6, PAI-1, and CTLA-4. Furthermore, aldosterone increases oxidant stress, which in turn promotes vascular inflammation. In particular, aldosterone increases expression of the NADPH oxidase subunits p22phox and gp91phox in the aorta, leading to an increase in reactive oxygen species (28, 29). Aldosterone also decreases endothelial expression of glucose-6-phosphate dehydrogenase (G6PD), leading to increased oxidant stress and decreased nitric oxide (NO) levels (30). Finally, aldosterone stimulates endothelial transcription of the proinflammatory intercellular adhesion molecule-1 (ICAM-1) (31).

In addition to its transcriptional effects, aldosterone can also promote inflammation through nontranscriptional pathways (5, 32–35). Aldosterone rapidly activates mitogen-activated protein kinase (MAPK) signaling in fibroblasts (36, 37). Furthermore, aldosterone rapidly activates calcium influx and protein kinase C (PKC), cAMP levels and protein kinase A (PKA), and protein kinase D (PKD) signaling pathways (32, 34, 38–40).

We hypothesized that aldosterone promotes vascular inflammation in part by triggering endothelial exocytosis. A variety of vascular agonists can bind to endothelial cell receptors and activate the secretion of endothelial granules, also called Weibel-Palade bodies (WPBs) (41, 42). WPB contain a variety of proinflammatory mediators, such as P-selectin, and prothrombotic signals, such as von Willebrand factor (vWF). Exocytosis leads to the translocation of P-selectin from the interior of the granule to the exterior surface of the endothelial cell. P-selectin then interacts with its receptor on the surface of leukocytes, mediating leukocyte rolling, the first step in leukocyte trafficking and vascular inflammation. Because hypertension is associated with vascular inflammation, it is possible that hypertensive messengers like aldosterone also stimulate vascular inflammation. In particular, because aldosterone leads to leukocyte infiltration, we hypothesized that aldosterone regulates one of the steps in leukocyte trafficking, such as endothelial exocytosis.

Results

Aldosterone Activates Weibel-Palade Body Exocytosis.

To explore the effect of aldosterone on endothelial exocytosis, we added increasing amounts of aldosterone to human aortic endothelial cells (HAEC) and measured the release of vWF with an ELISA. Aldosterone increases vWF release at a dose of 1.0 nM (Fig. 1A). Aldosterone increases vWF release by 31% of the level of increase by thrombin, a potent stimulator of endothelial exocytosis. Aldosterone also induces the release of interleukin-8, another component of endothelial WPBs (Fig. 1B). WPBs contain vWF, P-selectin, and IL-8. In contrast, tissue plasminogen activator (TPA) is stored in endothelial cells inside vesicles distinct from WPBs (43). We found that aldosterone does not stimulate TPA release from endothelial cells, emphasizing the distinct compartmentalization of TPA from vWF (Fig. 1C). Compared with other steroid hormones, aldosterone increases endothelial exocytosis by a level comparable to estrogen, but progesterone and hydrocortisone have no effect (Fig. 1D). These data suggest aldosterone stimulates endothelial release of vWF.

Fig. 1.

Fig. 1.

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Aldosterone induces endothelial exocytosis. (A) Dose-response study for vWF release: HAEC were incubated with aldosterone for 1 h. The amount of vWF released from cell into the media was measured by an ELISA (n = 3 ± SD; *, P < 0.05 vs. control). (B) Aldosterone activates interleukin-8 exocytosis. Endothelial cells were treated with TNF-a for 16 h to load IL-8 into granules. Endothelial cells were then stimulated with control (EtOH), thrombin, or aldosterone for 1 h. Cell media was analyzed for IL-8 with an ELISA as above (n = 3 ± SD; *, P < 0.05 vs. control). (C) Tissue plasminogen activator: aldosterone does not activate TPA release. Endothelial cells were treated with control, thrombin, or aldosterone for 1 h. Cell media was analyzed for TPA with an ELISA as described (n = 3 ± SD; P > 0.05 vs. control for all levels of aldosterone). (D) Specificity: aldosterone and estrogen activates vWF exocytosis. HAEC were treated with control, thrombin, aldosterone, estrogen, progesterone, or hydrocortisone for 1 h. Cell media was analyzed for vWF as described (n = 3 ± SD; *, P < 0.01 vs. control).

Aldosterone Rapidly Induces Endothelial Exocytosis.

Aldosterone can regulate transcription over several hours, and aldosterone can also have rapid nongenomic effects. We measured the kinetics of the release of vWF following aldosterone treatment, and we found that aldosterone stimulates release of vWF within 10 min (Fig. 2A). This effect is not transcriptional, as aldosterone stimulates vWF release even in cells treated with actinomycin D (Fig. 2B).

Fig. 2.

Fig. 2.

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Aldosterone rapidly induces endothelial exocytosis. (A) Time-course study: HAEC were treated with control, thrombin 1 U/ml, or aldosterone 10−9 M. Cell media was analyzed for vWF as described previously (n = 3 ± SD; *, P < 0.01 vs. control). (B) Aldosterone activation of exocytosis does not depend on new RNA transcription. HAEC were treated with actinomycin D 2.5 μM or DMSO for 1 h, and then stimulated with aldosterone 10−9 M or ethanol control for 1 h. Cell media was analyzed for vWF as described (n = 3 ± SD; *, P < 0.01 vs. control).

Mineralocorticoid Receptor Mediates Aldosterone Activation of Endothelial Exocytosis.

We next explored the role of the mineralocorticoid receptor (MR; also referred to as MLR, MCR, or NR3C2 gene product) in mediating aldosterone stimulation of endothelial exocytosis. Human aortic endothelial cells (HAEC) express MR by RT-PCR (Fig. 3A) and by immunoblotting (Fig. 3B). We then used pharmacological and genetic techniques to define the role of the MR in mediating the effects of aldosterone on endothelial exocytosis.

Fig. 3.

Fig. 3.

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Mineralocorticoid receptor mediates aldosterone activation of exocytosis: genetic data. (A) Expression of MR mRNA by RT-PCR of HAEC and HeLa cells. (B) Expression of MR protein by immunoblotting in HAEC, HUVEC, and HeLa cells (Top) and in HAEC, and mouse inner medullary collecting duct (mIMCD). Equal amounts 3.0 μg of cell lysates were loaded per lane. (C) Knockdown of mineralocorticoid receptor in endothelial cells. (D) Knockdown of mineralocorticoid receptor decreases effect of aldosterone on exocytosis. Endothelial cells were transfected with a control siRNA oligonucleotide or an oligonucleotide directed against MR; cells were treated or not with aldosterone 10−9 M or vehicle (EtOH) for 1 h and vWF release measured (n = 3 ± SD; *, P = 0.01 vs. control siRNA + aldo.).

First, we used siRNA to decrease MR expression in endothelial cells (Fig. 3C). Knockdown of the MR decreases aldosterone's effect on exocytosis (Fig. 3D). These genetic data suggest that MR mediates aldosterone induction of endothelial exocytosis.

Next, we pretreated endothelial cells with spironolactone, an inhibitor of the MR. Spironolactone inhibits aldosterone-triggered vWF release in a dose-dependent manner (Fig. 4A). Spironolactone does not inhibit thrombin activation of exocytosis (Fig. 4B). To confirm that the effects of spironolactone are specific for the MR, we also pretreated HAEC with tamoxifen, a selective estrogen receptor modulator, or mifepristone, a progesterone antagonist. These other inhibitors did not affect aldosterone stimulation of exocytosis (Fig. 4C). These inhibitors alone have no affect on exocytosis (Fig. 4D). Taken together, the pharmacological and genetic data suggest that the MR mediates aldosterone activation of vWF release.

Fig. 4.

Fig. 4.

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Mineralocorticoid receptor mediates aldosterone activation of exocytosis: pharmacological data. (A) Specificity: spironolactone inhibits aldosterone-induced vWF exocytosis. HAEC were pretreated with control or spironolactone for 1 h, and then treated with aldosterone 10−9 M for 1 h. Cell media was analyzed for vWF as described previously (n = 3 ± SD; *, P < 0.01 vs. media + aldosterone). (B) Spironolactone does not affect thrombin-induced exocytosis. HAEC were pretreated with EtOH (control vehicle) or spironolactone 10−7 M for 1 h, and then treated with thrombin 1 U/ml for 1 h. Cell media was analyzed for vWF as described (n = 3 ± SD; *, P < 0.01 vs. control). (C) Inhibitors of other steroid hormone receptors do not affect aldosterone-induced vWF exocytosis. HAEC were pretreated with estrogen receptor inhibitor tamoxifen 10−7 M, progesterone receptor inhibitor mifepristone 10−7 M, and spironolactone 10−7 M for 1 h, and then treated with aldosterone 10−9 M for 1 h. Cell media was analyzed for vWF as described (n = 3 ± SD; *, P < 0.01 vs. media + aldosterone). (D) Inhibitors of other steroid hormone receptors alone do not affect vWF exocytosis in the absence of aldosterone. HAEC were pretreated with estrogen receptor inhibitor tamoxifen 10−7 M, progesterone receptor inhibitor mifepristone 10−7 M, or aldosterone 10−9 M for 1 h. Cell media was analyzed for vWF as described (n = 3 ± SD; *, P < 0.01 vs. control).

Calcium Signal Transduction Mediates Aldosterone Activation of Endothelial Exocytosis.

We explored intracellular pathways that mediate aldosterone activation of exocytosis, focusing on calcium signaling. We added the calcium ionophore A23187 at varying concentrations to HAEC and measured the release of vWF. Calcium ionophore increases exocytosis in a dose-dependent manner [supporting information (SI) Fig. S1]. To define the pools of calcium that trigger exocytosis in response to aldosterone, we added aldosterone to endothelial cells incubated with normal media, calcium free media, or with 1,2-Bis[2-Aminophenoxy]ethane-N,N,N′,N′-tetraacetic acid (BAPTA) (which disperses intracellular calcium storage pools). BAPTA decreases endothelial release of vWF (Fig. S1), but removing extracellular calcium has no effect. The calmodulin antagonist trifluoperazine (TFP) inhibited aldosterone-triggered vWF release, suggesting that calmodulin may be involved in aldosterone signaling (Fig. S1). TFP alone did not affect exocytosis (Fig. S1). Taken together, these data suggest that intracellular calcium plays a critical role in aldosterone-triggered exocytosis.

Aldosterone Activates Leukocyte-Endothelial Interactions.

We then measured the effect of aldosterone on leukocyte interactions with endothelial cells ex vivo. We pretreated HAEC with vehicle or spironolactone for 1 h, and then added vehicle or aldosterone or thrombin or thrombin receptor-activating peptide (TRAP) for 1 h; BCEF-loaded HL-60 cells were added to the endothelial cells, incubated at 4 °C for 15 min, and then washed. The cocultured cells were then imaged with a digital camera. Few HL-60 cells adhere to control endothelial cells, but aldosterone increases the number of HL-60 cells adhering to the endothelial cells (Fig. 5 A and B). Pretreatment with spironolactone inhibits aldosterone-induced HL-60 adherence to HAEC (Fig. 5 A and B). To confirm that aldosterone triggers endothelial exocytosis of Weibel-Palade bodies and externalizes P-selectin, we pretreated endothelial cells with antibody to P-selectin before adding aldosterone. Antibody to P-selectin decreases HL-60 adherence to endothelial cells (Fig. 5 C and D). Taken together, these data show that aldosterone triggers leukocyte adherence to endothelial cells in a P-selectin-dependent manner.

Fig. 5.

Fig. 5.

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Aldosterone activates leukocyte-endothelial interactions ex vivo. (A) Images of leukocyte adherence to endothelial cells. HAEC were pretreated or not with spironolactone (10−7M) for 1 h, and then aldosterone (10−9M), thrombin 1 U/ml, or TRAP (10−6M) were added for 1 h. HL-60 promyelocytic leukemia cells were then loaded with BCEF-AM and cocultured with the HAEC. The cell mixtures were incubated at 4 °C for 15 min, washed twice with HBSS, and then imaged with a digital fluorescent camera. (B) Quantitation of above, measuring the effect of aldosterone (aldo.) and spironolactone (spiro.) on HL-60 adherence to HAEC (n = 4 wells with 3 different fields from each well ± SD. *, P < 0.001 compared with control). (C) Images of leukocyte adherence to endothelial cells. HAEC were treated with aldosterone (10−9 M) for 1 h with or without antibody to P-selectin (10−3 M) for 15 min, and then cocultured with HL-60 cells and imaged as described. (D) Quantitation of above, measuring the effect of antibody to P-selectin on aldosterone or calcium ionophore (A23187) stimulated HL-60 adherence to HAEC (n = 4 wells with 3 different fields from each well ± SD; *, P < 0.001 compared with control).

Discussion

Summary.

The major finding of our study is that aldosterone induces endothelial exocytosis, leading to leukocyte adherence to endothelial cells. Our data may partly explain why elevated aldosterone levels promote vascular inflammation.

Mineralocorticoid Receptor and Endothelial Exocytosis.

Two lines of evidence suggest that the MR mediates aldosterone's effect on exocytosis. First, spironolactone inhibits exocytosis after aldosterone treatment (Fig. 4). However, in theory, spironolactone might inhibit unidentified aldosterone receptors other than the MR. Second, direct siRNA targeting of MR also inhibits aldosterone activation of exocytosis (Fig. 3). Our findings support the studies of others showing that MR mediates vascular inflammation. For example, the MR antagonists decrease oxidative stress and vascular inflammation in animal models (17–21, 44).

Aldosterone Activates Endothelial Exocytosis Through a Nongenomic Effect.

We find that aldosterone activates exocytosis within 10 min of application (Fig. 2A). Actinomycin D fails to block the effects of aldosterone (Fig. 2B). This rapid effect suggests that aldosterone is signaling through previously synthesized messengers, rather than by regulating the transcription of proteins. Our data add to the growing literature describing rapid, nongenomic effects of aldosterone (2, 5, 32–35). For example, aldosterone activates signaling pathways such as PKA, PKC, PKD, MAPK, and the NADPH oxidase in vascular smooth muscle; and aldosterone activates phosphatidylinositol 3-kinase in endothelial cells (32, 34, 36–40, 45–47). However, nongenomic effects of aldosterone can be mediated through either the MR or other unidentified proteins. For instance, aldosterone increases calcium signaling in vascular smooth muscle cells in an MR-independent manner (48). In our studies, we used siRNA to show that the MR mediates aldosterone activation of endothelial exocytosis. Thus our studies show that aldosterone has a rapid and nongenomic effect on exocytosis, mediated in part by the MR.

Aldosterone and Thrombin Activate Secretion of Distinct Vesicles.

Endothelial granules are heterogeneous. Weibel-Palade bodies contain vWF, P-selectin, and other proteins. TPA appears to reside in vesicles distinct from Weibel-Palade bodies. Sucrose gradient centrifugation of human lung endothelial cells reveals TPA and vWF in different distributions (43). The small GTP binding protein Rab3D regulates vWF release but not TPA release from endothelial cells (49). Our findings support the idea that TPA and vWF are stored in separate vesicular compartments. We found that thrombin induces release of both TPA and vWF, but aldosterone selectively activates release of vWF but not TPA (Fig. 1C).

Proinflammatory Effects of Aldosterone in Humans.

Although aldosterone increases vascular inflammation in animals and in humans, the precise mechanisms are not completely understood. Clinical studies show that hyperaldosteronism or infusion of aldosterone is associated with elevated levels of inflammatory biomarkers (22–24). Conversely, aldosterone antagonists decrease inflammatory biomarkers in animals and humans (25, 50–52). For example, eplerenone decreases plasma levels of proinflammatory cytokines such as MCP-1 or IL-8 (53). Aldosterone may activate inflammation in vivo through a variety of mechanisms, inducing synthesis of new proinflammatory proteins, activating NADPH oxidase to produce superoxide, and decreasing glucose-6-phosphate dehydrogenase transcription. Our data suggest an additional new mechanism for the proinflammatory effects of aldosterone: aldosterone activates endothelial exocytosis, leading to increased leukocyte rolling along the vessel wall and vascular inflammation.

Experimental Procedures

Reagents.

Aldosterone was purchased from Sigma. Human alpha-thrombin was purchased from Enzyme Research Laboratories. Thrombin receptor activating peptide (TRAP) was purchased from Bachem. Beta-estradiol, progesterone, spironolactone, tamoxifen, mifepristone, hydrocortisone, trifluoperazine, calcium ionophore A223187, and BAPTA/AM were purchased from Sigma. Mouse anti-P-selectin (CD62P) monoclonal antibody was purchased from BD Bioscience. Rabbit polyclonal antibody to human MR was from Santa Cruz (sc-11412) and was used at 1:200. Mouse inner medullary collecting duct cells were from American Type Culture Collection (CRL-2123).

PCR.

PCR was performed on total RNA harvested from endothelial cells, using a hMR sense primer 5′-GAAGATGCATCAGTCTGCCA-3′ and a hMR antisense primer 5′-AGTCCAGCAGCTTGGTCAGT-3′. The PCR consisted of a 2-min denaturing step at 95 °C; followed by 36 cycles of denaturing at 95 °C and annealing at 58 °C and extending at 72 °C; and finishing with an extension at 72 °C for 5 min.

Cell Culture and Analysis of vWF Release.

Endothelial exocytosis was performed as described previously (54). HAEC were obtained from Lonza-Clonetics and grown in EGM-2 media supplemented with 2% serum and growth factors in a kit (Bullet Kit; Clonetics). Endothelial cells were purchased at passage 3. Exocytosis was measured in HAEC between passages 5 and 7. To measure the effect of VEGF on vWF release, HAEC were grown in EGM-2 media with growth factors, then washed and placed in EGM-2 media without growth factors and without serum, stimulated with various concentration of aldosterone. The amount of vWF released into the media was measured by an ELISA (American Diagnostica). Some HAEC were stimulated with thrombin, estrogen, progesterone, or hydrocortisone for up to 1 h at 37 °C. The supernatant from each well was collected separately at certain defined time points, and the concentration of vWF released into the media was measured by an ELISA.

siRNA Knockdown of MR in Rndothelial Cells.

Endothelial cells were electroporated with siRNA against MR (M-003425–01; Dharmacon) or a control siRNA (D-001206–14; Dharmacon) using a nucleofector (Amaxa). Electroporated cells were incubated for 48 h and then stimulated with aldosterone or vehicle.

Mineralocorticoid Receptor Expression, Inhibition, and Specificity.

HAEC were grown in EGM-2 media with growth factors, then washed and placed in EGM-2 media without growth factors or serum. The cells were pretreated with varying concentrations of the aldosterone antagonist spironolactone at 37 °C for 1 h and then aldosterone at 10−9 M was added for 1 h at 37 °C. The supernatant from each well was collected separately and the concentration of vWF released into the media was measured by an ELISA. Mineralocorticoid receptor expression was demonstrated using RT-PCR. RNA was isolated from HAEC and HELA cells and a 3′ gene-specific primer for the mineralocorticoid receptor gene was used for reverse transcription. The cDNA was then amplified using Taq polymerase and gene-specific forward and reverse primers. The samples both with and without primers were placed on a gel to show the MW of the specific isolated sequence present in both cell populations.

Calcium and Aldosterone-Mediated vWF Release.

HAEC were grown in EGM-2 media with growth factors, then washed and placed in EGM-2 media without growth factors or serum. The calcium ionophore A23187 was then added at varying concentrations to HAEC at 37° Celsius for 1 h. The amount of vWF released into the media was then quantified using ELISA. HAEC were pretreated with either the intracellular calcium chelator BAPTA/AM at a concentration of 20 μM and Dulbecco Modified Eagle's Medium (DMEM) or with calcium-free media and allowed to incubate for 30 min at 37 °C. Aldosterone 10−9 M was then added to the HAEC and placed in the incubator at 37 °C for 1 h. The amount of vWF released into the supernatant was then measured with ELISA.

The calmodulin antagonist trifluoperazine was used to determine the role of calmodulin in vWF release. HAEC were pretreated with trifluoperazine at varying concentrations at 37 °C for 30 min. Then aldosterone 10−9 M was added to the cells and placed in the incubator at 37 °C for 1 h. The amount of vWF release in the supernatant was measured by ELISA.mbf.

Aldosterone-Mediated Leukocyte and Endothelial Interactions.

The human promyelocytic cell line HL-60 was obtained from the American Type Culture Collection. HAEC were pretreated with or without spironolactone 10−7 M for 1 h at 37 °C and then aldosterone (10−9 M), thrombin 1 U/ml, or TRAP (10−6 M) were added for 1 h. HL-60 cells were then loaded with BCEF-AM and cocultured with the HAEC. The cell mixtures were incubated at 4 °C for 15 min, washed twice with HBSS, and then imaged with a digital fluorescent camera. HAEC were also treated with aldosterone (10−9 M) or A223187 at 1 μM for 1 h with or without antibody to P-selectin (10−3M) for 15 min, cocultured with HL-60 cells, and then imaged with the digital fluorescent camera as described.

Statistical Analyses.

Data are expressed as the mean ± SD. Statistical comparisons were made by analysis of variance followed by a Bonferroni's t test for multiple comparison. A P value <0.05 was considered significant.

Supplementary Material

Supporting Information
supp_106_10_3782__index.html (576B, html)

Acknowledgments.

Supported by grants from the National Institutes of Health (NIH Grants P01 HL56091, R01 HL074061, R01 HL78635, and P01 HL65608), the American Heart Association (AHA Grant EIG 0140210N), the Ciccarone Center, the John and Cora H. Davis Foundation, and the Clarence P. Doodeman Professorship in Cardiology (to C.J.L.), and by Grants RR07002 and HL074945 from NIH (to C.N.M.) and Grant 0815093E from the AHA (to Y.J.).

Footnotes

The authors declare no conflict of interest.

This article is a PNAS Direct Submission.

This article contains supporting information online at www.pnas.org/cgi/content/full/0804037106/DCSupplemental.

References

  • 1.Cohn JN, Colucci W. Cardiovascular effects of aldosterone and post-acute myocardial infarction pathophysiology. Am J Cardiol. 2006;97:4F–12F. doi: 10.1016/j.amjcard.2006.03.004. [DOI] [PubMed] [Google Scholar]
  • 2.Brown NJ. Aldosterone and vascular inflammation. Hypertension. 2008;51:161–167. doi: 10.1161/HYPERTENSIONAHA.107.095489. [DOI] [PubMed] [Google Scholar]
  • 3.Ferrario CM, Strawn WB. Role of the renin-angiotensin-aldosterone system and proinflammatory mediators in cardiovascular disease. Am J Cardiol. 2006;98:121–128. doi: 10.1016/j.amjcard.2006.01.059. [DOI] [PubMed] [Google Scholar]
  • 4.Fiebeler A, Muller DN, Shagdarsuren E, Luft FC. Aldosterone, mineralocorticoid receptors, and vascular inflammation. Curr Opin Nephrol Hypertens. 2007;16:134–142. doi: 10.1097/MNH.0b013e32801245bb. [DOI] [PubMed] [Google Scholar]
  • 5.Funder JW. Minireview: Aldosterone and the cardiovascular system: Genomic and nongenomic effects. Endocrinology. 2006;147:5564–5567. doi: 10.1210/en.2006-0826. [DOI] [PubMed] [Google Scholar]
  • 6.Funder JW. The role of aldosterone and mineralocorticoid receptors in cardiovascular disease. Am J Cardiovasc Drugs. 2007;7:151–157. doi: 10.2165/00129784-200707030-00001. [DOI] [PubMed] [Google Scholar]
  • 7.Young MJ. Mechanisms of mineralocorticoid receptor-mediated cardiac fibrosis and vascular inflammation. Curr Opin Nephrol Hypertens. 2008;17:174–180. doi: 10.1097/MNH.0b013e3282f56854. [DOI] [PubMed] [Google Scholar]
  • 8.Weber KT, et al. Aldosteronism in heart failure: A proinflammatory/fibrogenic cardiac phenotype. Search for biomarkers and potential drug targets. Curr Drug Targets. 2003;4:505–516. doi: 10.2174/1389450033490948. [DOI] [PubMed] [Google Scholar]
  • 9.Funder JW. Aldosterone, mineralocorticoid receptors and vascular inflammation. Mol Cell Endocrinol. 2004;217:263–269. doi: 10.1016/j.mce.2003.10.054. [DOI] [PubMed] [Google Scholar]
  • 10.Fiebeler A, Luft FC. The mineralocorticoid receptor and oxidative stress. Heart Fail Rev. 2005;10:47–52. doi: 10.1007/s10741-005-2348-y. [DOI] [PubMed] [Google Scholar]
  • 11.Joffe HV, Adler GK. Effect of aldosterone and mineralocorticoid receptor blockade on vascular inflammation. Heart Fail Rev. 2005;10:31–37. doi: 10.1007/s10741-005-2346-0. [DOI] [PubMed] [Google Scholar]
  • 12.Duprez DA. Role of the renin-angiotensin-aldosterone system in vascular remodeling and inflammation: A clinical review. J Hypertens. 2006;24:983–991. doi: 10.1097/01.hjh.0000226182.60321.69. [DOI] [PubMed] [Google Scholar]
  • 13.Savoia C, Schiffrin EL. Vascular inflammation in hypertension and diabetes: Molecular mechanisms and therapeutic interventions. Clin Sci (Lond) 2007;112:375–384. doi: 10.1042/CS20060247. [DOI] [PubMed] [Google Scholar]
  • 14.Young MJ, Lam EY, Rickard AJ. Mineralocorticoid receptor activation and cardiac fibrosis. Clin Sci (Lond) 2007;112:467–475. doi: 10.1042/CS20060275. [DOI] [PubMed] [Google Scholar]
  • 15.Blasi ER, et al. Aldosterone/salt induces renal inflammation and fibrosis in hypertensive rats. Kidney Int. 2003;63:1791–1800. doi: 10.1046/j.1523-1755.2003.00929.x. [DOI] [PubMed] [Google Scholar]
  • 16.Neves MF, et al. Role of aldosterone in angiotensin II-induced cardiac and aortic inflammation, fibrosis, and hypertrophy. Can J Physiol Pharmacol. 2005;83:999–1006. doi: 10.1139/y05-068. [DOI] [PubMed] [Google Scholar]
  • 17.Benetos A, Lacolley P, Safar ME. Prevention of aortic fibrosis by spironolactone in spontaneously hypertensive rats. Arterioscler Thromb Vasc Biol. 1997;17:1152–1156. doi: 10.1161/01.atv.17.6.1152. [DOI] [PubMed] [Google Scholar]
  • 18.Rocha R, et al. Aldosterone induces a vascular inflammatory phenotype in the rat heart. Am J Physiol Heart Circ Physiol. 2002;283:H1802–H1810. doi: 10.1152/ajpheart.01096.2001. [DOI] [PubMed] [Google Scholar]
  • 19.Lacolley P, et al. Prevention of aortic and cardiac fibrosis by spironolactone in old normotensive rats. J Am Coll Cardiol. 2001;37:662–667. doi: 10.1016/s0735-1097(00)01129-3. [DOI] [PubMed] [Google Scholar]
  • 20.Oestreicher EM, et al. Aldosterone and not plasminogen activator inhibitor-1 is a critical mediator of early angiotensin II/NG-nitro-L-arginine methyl ester-induced myocardial injury. Circulation. 2003;108:2517–2523. doi: 10.1161/01.CIR.0000097000.51723.6F. [DOI] [PubMed] [Google Scholar]
  • 21.Kuster GM, et al. Mineralocorticoid receptor inhibition ameliorates the transition to myocardial failure and decreases oxidative stress and inflammation in mice with chronic pressure overload. Circulation. 2005;111:420–427. doi: 10.1161/01.CIR.0000153800.09920.40. [DOI] [PubMed] [Google Scholar]
  • 22.Irita J, et al. Plasma osteopontin levels are higher in patients with primary aldosteronism than in patients with essential hypertension. Am J Hypertens. 2006;19:293–297. doi: 10.1016/j.amjhyper.2005.08.019. [DOI] [PubMed] [Google Scholar]
  • 23.Clapp BR, et al. Inflammation and endothelial function: Direct vascular effects of human C-reactive protein on nitric oxide bioavailability. Circulation. 2005;111:1530–1536. doi: 10.1161/01.CIR.0000159336.31613.31. [DOI] [PubMed] [Google Scholar]
  • 24.Luther JM, et al. Angiotensin II induces interleukin-6 in humans through a mineralocorticoid receptor-dependent mechanism. Hypertension. 2006;48:1050–1057. doi: 10.1161/01.HYP.0000248135.97380.76. [DOI] [PubMed] [Google Scholar]
  • 25.Sawathiparnich P, Kumar S, Vaughan DE, Brown NJ. Spironolactone abolishes the relationship between aldosterone and plasminogen activator inhibitor-1 in humans. J Clin Endocrinol Metab. 2002;87:448–452. doi: 10.1210/jcem.87.2.7980. [DOI] [PubMed] [Google Scholar]
  • 26.Jaffe IZ, Mendelsohn ME. Angiotensin II and aldosterone regulate gene transcription via functional mineralocortocoid receptors in human coronary artery smooth muscle cells. Circ Res. 2005;96:643–650. doi: 10.1161/01.RES.0000159937.05502.d1. [DOI] [PubMed] [Google Scholar]
  • 27.Fejes-Toth G, Naray-Fejes-Toth A. Early aldosterone-regulated genes in cardiomyocytes: Clues to cardiac remodeling? Endocrinology. 2007;148:1502–1510. doi: 10.1210/en.2006-1438. [DOI] [PubMed] [Google Scholar]
  • 28.Calo LA, et al. Effect of aldosterone and glycyrrhetinic acid on the protein expression of PAI-1 and p22(phox) in human mononuclear leukocytes. J Clin Endocrinol Metab. 2004;89:1973–1976. doi: 10.1210/jc.2003-031545. [DOI] [PubMed] [Google Scholar]
  • 29.Hirono Y, et al. Angiotensin II receptor type 1-mediated vascular oxidative stress and proinflammatory gene expression in aldosterone-induced hypertension: The possible role of local renin-angiotensin system. Endocrinology. 2007;148:1688–1696. doi: 10.1210/en.2006-1157. [DOI] [PubMed] [Google Scholar]
  • 30.Leopold JA, et al. Aldosterone impairs vascular reactivity by decreasing glucose-6-phosphate dehydrogenase activity. Nat Med. 2007;13:189–197. doi: 10.1038/nm1545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Caprio M, et al. Functional mineralocorticoid receptors in human vascular endothelial cells regulate intercellular adhesion molecule-1 expression and promote leukocyte adhesion. Circ Res. 2008;102:1359–1367. doi: 10.1161/CIRCRESAHA.108.174235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32.Grossmann C, Gekle M. Nongenotropic aldosterone effects and the EGFR: Interaction and biological relevance. Steroids. 2008;73:973–978. doi: 10.1016/j.steroids.2007.12.008. [DOI] [PubMed] [Google Scholar]
  • 33.Brown NJ. Aldosterone and end-organ damage. Curr Opin Nephrol Hypertens. 2005;14:235–241. doi: 10.1097/01.mnh.0000165889.60254.98. [DOI] [PubMed] [Google Scholar]
  • 34.Harvey BJ, et al. Rapid responses to aldosterone in the kidney and colon. J Steroid Biochem Mol Biol. 2008;108:310–317. doi: 10.1016/j.jsbmb.2007.09.005. [DOI] [PubMed] [Google Scholar]
  • 35.Wehling M. Specific, nongenomic actions of steroid hormones. Annu Rev Physiol. 1997;59:365–393. doi: 10.1146/annurev.physiol.59.1.365. [DOI] [PubMed] [Google Scholar]
  • 36.Haseroth K, et al. Rapid nongenomic effects of aldosterone in mineralocorticoid-receptor-knockout mice. Biochem Biophys Res Commun. 1999;266:257–261. doi: 10.1006/bbrc.1999.1771. [DOI] [PubMed] [Google Scholar]
  • 37.Min LJ, et al. Aldosterone and angiotensin II synergistically induce mitogenic response in vascular smooth muscle cells. Circ Res. 2005;97:434–442. doi: 10.1161/01.RES.0000180753.63183.95. [DOI] [PubMed] [Google Scholar]
  • 38.Mihailidou AS, Mardini M, Funder JW. Rapid, nongenomic effects of aldosterone in the heart mediated by epsilon protein kinase C. Endocrinology. 2004;145:773–780. doi: 10.1210/en.2003-1137. [DOI] [PubMed] [Google Scholar]
  • 39.Alzamora R, Brown LR, Harvey BJ. Direct binding and activation of protein kinase C isoforms by aldosterone and 17beta-estradiol. Mol Endocrinol. 2007;21:2637–2650. doi: 10.1210/me.2006-0559. [DOI] [PubMed] [Google Scholar]
  • 40.Alzamora R, Harvey BJ. Direct binding and activation of protein kinase C isoforms by steroid hormones. Steroids. 2008;73:885–888. doi: 10.1016/j.steroids.2008.01.001. [DOI] [PubMed] [Google Scholar]
  • 41.Lowenstein CJ, Morrell CN, Yamakuchi M. Regulation of Weibel-Palade body exocytosis. Trends Cardiovasc Med. 2005;15:302–308. doi: 10.1016/j.tcm.2005.09.005. [DOI] [PubMed] [Google Scholar]
  • 42.Rondaij MG, et al. Dynamics and plasticity of Weibel-Palade bodies in endothelial cells. Arterioscler Thromb Vasc Biol. 2006;26:1002–1007. doi: 10.1161/01.ATV.0000209501.56852.6c. [DOI] [PubMed] [Google Scholar]
  • 43.Emeis JJ, et al. An endothelial storage granule for tissue-type plasminogen activator. J Cell Biol. 1997;139:245–256. doi: 10.1083/jcb.139.1.245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 44.Sun Y, et al. Aldosterone-induced inflammation in the rat heart: Role of oxidative stress. Am J Pathol. 2002;161:1773–1781. doi: 10.1016/S0002-9440(10)64454-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 45.Christ M, Meyer C, Sippel K, Wehling M. Rapid aldosterone signaling in vascular smooth muscle cells: Involvement of phospholipase C, diacylglycerol and protein kinase C alpha. Biochem Biophys Res Commun. 1995;213:123–129. doi: 10.1006/bbrc.1995.2106. [DOI] [PubMed] [Google Scholar]
  • 46.Liu SL, et al. Aldosterone regulates vascular reactivity: Short-term effects mediated by phosphatidylinositol 3-kinase-dependent nitric oxide synthase activation. Circulation. 2003;108:2400–2406. doi: 10.1161/01.CIR.0000093188.53554.44. [DOI] [PubMed] [Google Scholar]
  • 47.Callera GE, et al. Aldosterone activates vascular p38MAP kinase and NADPH oxidase via c-Src. Hypertension. 2005;45:773–779. doi: 10.1161/01.HYP.0000154365.30593.d3. [DOI] [PubMed] [Google Scholar]
  • 48.Christ M, et al. Rapid effects of aldosterone on sodium transport in vascular smooth muscle cells. Hypertension. 1995;25:117–123. doi: 10.1161/01.hyp.25.1.117. [DOI] [PubMed] [Google Scholar]
  • 49.Knop M, Aareskjold E, Bode G, Gerke V. Rab3D and annexin A2 play a role in regulated secretion of vWF, but not tPA, from endothelial cells. EMBO J. 2004;23:2982–2992. doi: 10.1038/sj.emboj.7600319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 50.Takebayashi K, Matsumoto S, Aso Y, Inukai T. Aldosterone blockade attenuates urinary monocyte chemoattractant protein-1 and oxidative stress in patients with type 2 diabetes complicated by diabetic nephropathy. J Clin Endocrinol Metab. 2006;91:2214–2217. doi: 10.1210/jc.2005-1718. [DOI] [PubMed] [Google Scholar]
  • 51.Brown NJ, et al. Effect of activation and inhibition of the renin-angiotensin system on plasma PAI-1. Hypertension. 1998;32:965–971. doi: 10.1161/01.hyp.32.6.965. [DOI] [PubMed] [Google Scholar]
  • 52.Godfrey V, et al. Effect of spironolactone on C-reactive protein levels in patients with heart disease. Int J Cardiol. 2007;117:282–284. doi: 10.1016/j.ijcard.2006.05.069. [DOI] [PubMed] [Google Scholar]
  • 53.Savoia C, Touyz RM, Amiri F, Schiffrin EL. Selective mineralocorticoid receptor blocker eplerenone reduces resistance artery stiffness in hypertensive patients. Hypertension. 2008;51:432–439. doi: 10.1161/HYPERTENSIONAHA.107.103267. [DOI] [PubMed] [Google Scholar]
  • 54.Matsushita K, et al. Nitric oxide regulates exocytosis by S-nitrosylation of N-ethylmaleimide-sensitive factor. Cell. 2003;115:139–150. doi: 10.1016/s0092-8674(03)00803-1. [DOI] [PMC free article] [PubMed] [Google Scholar]

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