Hormone aldosterone is promising target for stroke treatment
Posted on November 30, 2005 by Staff
A bi-polar hormone that can contribute to strokes and
minimize their damage is emerging as a therapeutic target in the battle
against these brain attacks, researchers say.
“It costs about $56 billion a year to look after stroke
patients, never mind the quality-of-life issues for these patients,” says
Dr. Anne M. Dorrance, Medical College of Georgia physiologist and senior
author of a review article on the cover of the November issue of Trends
in Endocrinology and Metabolism.
Despite better management of blood pressure – the
number-one risk factor for strokes – stroke incidence is not declining and
aging baby boomers likely will cause rates to spike, says Dr. Dorrance.
She is among an increasing number of scientists who think
the hormone, aldosterone, is part of the problem and blocking it may be part
of the solution. Scientific momentum surrounding the hormone secreted by the
adrenal gland prompted the journal to ask Dr. Dorrance to write the article,
“Aldosterone: Good Guy or Bad Guy in Cerebrovascular Disease.”
She calls aldosterone “a double-edged sword” that helps
maintain healthy blood pressure but also dangerously reshapes blood vessels
and makes the heart fibrotic. Its conflicting roles in the body are clearly
played out in the brain of hypertensives.
“What aldosterone does under normal circumstances is
regulate sodium balance in the body,” says Dr. Dorrance. When sodium levels
rise, aldosterone levels drop so the kidneys will eliminate more sodium to
help maintain a healthy blood pressure and vice versa. “Ten years ago, we
would have told you all aldosterone does is work on the kidneys,” she says.
In recent years, Dr. Dorrance and others have learned
aldosterone works in many ways – many of them bad – inside blood vessels
throughout the body and brain where there are many aldosterone receptors.
As with most things, too much aldosterone – resulting
from an adrenal gland tumor or abnormal production for unknown reasons – is
what causes problems, such as proliferation of the smooth muscle cells that
make up the elastic walls of blood vessels. Instead of the middle filling up
with plaque as it does in atherosclerosis, the middle, or lumen, of the
blood vessel shrinks because the walls thickens. To further set the stage
for stroke, the thicker walls are less stretchy, Dr. Dorrance says.
Conversely, after a stroke, emerging evidence suggests
aldosterone helps neurons survive. “This is still a very big ‘perhaps,’ but
perhaps at the time of a stroke, it may be beneficial, but only within the
brain,” Dr. Dorrance says of high aldosterone levels. “There is a huge
caveat because aldosterone does not get into the brain terribly well.”
Other good news emerging is that adolesterone blockers,
such as spironolactone and a newer, more specific blocker eplerenone, may
help turn the tide. Spironolactone was developed as an antihypertensive
years ago but didn’t work very well, Dr. Dorrance says. Then, a University
of Michigan study in the mid-1980s showed heart failure patients who got the
drug in addition to standard treatment, such as beta blockers and ACE
inhibitors, basically stopped dying, says Dr. Dorrance, who came from the
University of Michigan to MCG as a postdoctoral fellow in 2000.
Unfortunately spironolactone also blocks testosterone so
it increased men’s breast size. Last year, a more-specific second generation
of the drug, eplerenone, received Food and Drug Adminstration approval.
Still neither version has become a medicine cabinet
mainstay. However the potential for aldosterone blockers may still be
unfolding as researchers such as Dr. Dorrance discover what the hormone does
and what blocking it prevents.
In an animal model of stroke and hypertension, Dr.
Dorrance and MCG graduate student Christine Rigsby, a paper co-author, have
shown spironolactone won’t lower blood pressure but will reverse the
wall-thickening effect of aldosterone, at least in males.
Within six weeks, “the blood vessels look like a
normotensive animal, like the animal never had high blood pressure,” says
Dr. Dorrance. “But when you block aldosterone in the females, there is no
beneficial effect,” she says of the increasingly complex story. Her lab is
looking at whether blockers become protective after menopause.
Gender difference hold up in aldosterone’s effect on the
heart as well. One of the many findings of the Framingham Heart Study, a
long-term study of thousands of residents in a Boston suburb, was that in
women, higher levels of aldosterone correlate with worsening enlargement of
the pumping chamber of the heart. “That does not mean aldosterone doesn’t
impact a man’s heart in some way, but it’s definitely not as direct an
effect,” says Dr. Dorrance.
“We are literally at the tip of the iceberg,” she says.
Some scientists suspect the level of adolsterone receptors, not the hormone
level, may be the real culprit.
Either way, there is little doubt aldosterone’s obscurity
is over. “I think we are beginning an extremely complicated and terribly
exciting 10 to 15 years of research as we try to unravel what is happening
and why,” she says. “I think at the end, there will be better drugs and
better information so clinicians will know when patients would benefit from
these aldosterone blockers.”
Part of the excitement will come in identifying the
pathways of aldosterone’s many actions. Dr. Dorrance thinks a relationship
between aldosterone and epidermal growth factor, for example, causes the
dangerous thickening of blood vessel walls in the brain. “It’s not just a
blood pressure-dependent effect. This is something that is much more active
than that. I do get up in the middle of the night wondering why it happens.
It fascinates me.”
Dr. William E. Cannady, an MCG School of Medicine
graduate who worked in Dr. Dorrance’s lab, also is a paper co-author.
Extra-adrenal aldosterone biosynthesis
1University of Mississippi, Jackson, Mississippi, USA; 2G.V. Montgomery VA Medical Center, Jackson, Mississippi, USA.
Aldosterone (aldo) concentrations in the brain and heart are slightly lower, but parallel those in the plasma. Plasma, heart and urine aldosterone continue to be detected in very small amounts in adrenalectomized (ADX) rats, however the concentrations of aldosterone in the brain, though very low, are higher than plasma levels in ADX rats. StAR protein and all of the steroidogenic enzymes necessary for the synthesis of aldosterone are expressed in the central nervous system (CNS) of rats and humans. Brain tissue from ADX rats synthesizes aldo from endogenous substrate and converts 3H-deoxycorticosterone into 3H-aldosterone in vitro. Inappropriately high aldo levels are associated with increased sympathetic drive and hypertension of central origin. Aldo synthesized in the brain appears to play a role in the development of hypertension in the Dahl salt-sensitive rat. Aldo concentration in the hypothalamus of Dahl SS rats is higher than in Sprague-Dawley controls and inhibition of enzymes within the aldosterone biosynthetic pathway, including trilostane, a 3-hydroxysteroid dehydrogenase, and aldosterone synthase, decreases the BP in the Dahl SS rat. Rats in which the aldosterone synthase enzyme cDNA is over-expressed in neurons also have hypertension.
Though most of the aldo in the brain is sequestered from circulating aldo synthesized by the adrenal gland, a small proportion is locally synthesized in the normal rat. As the amount is exceedingly low and number of cells that produce aldo in the brain few, aldo produced in the brain, if relevant, would be expected to have autocrine or paracrine functions. Aldo synthesized in the CNS might have a role in those forms of hypertension and autonomic dysfunction in which circulating aldo is not elevated, but which respond nonetheless to mineralocorticoid receptor antagonists.
Differential effects of high and low steroidogenic factor-1 expression on CYP11B2 expression and aldosterone production in adrenocortical cells.
Ye P, Nakamura Y, Lalli E, Rainey WE.
Department of Physiology, Medical College of Georgia, Augusta, Georgia 30912, USA.
Steroidogenic factor-1 (SF-1/Ad4BP/NR5A1) plays a major role in regulating steroidogenic enzymes. We have previously shown that SF-1 inhibits aldosterone synthase (CYP11B2) reporter gene activity. Herein, we used the H295R/TR/SF-1 adrenal cells that increase SF-1 in a doxycycline-dependent fashion. Cells were incubated with or without doxycycline to induce SF-1 and then treated with angiotensin II (Ang II). Aldosterone was measured by immunoassay. SF-1 mRNA was silenced by small interfering RNA (siRNA) by Nucleofector technology. mRNA levels were measured by real-time RT-PCR. Ang II treatment without doxycycline increased aldosterone production by 11.3-fold and CYP11B2 mRNA by 116-fold. Doxycycline treatment increased SF-1 mRNA levels by 3.7-fold and inhibited Ang II-induced aldosterone by 84%. Doxycycline treatment inhibited Ang II-stimulated CYP11B2 mRNA levels by 86%. Doxycycline decreased basal CYP11B2 promoter activity by 68%. Doxycycline inhibited Ang II stimulation by 85%. Ang II increased CYP21 mRNA expression by 4.6-fold, whereas doxycycline inhibited induction by 69%. In contrast, doxycycline treatment increased CYP11B1 mRNA by 1.7-fold in basal cells and increased Ang II induction by 3.6-fold. SF-1-specific siRNA significantly reduced SF-1 mRNA expression as compared with cells treated with control siRNA. SF-1 siRNA reversed doxycycline stimulation of CYP B1 and its inhibition of CYP11B2. However, in H295R/TR/SF-1 cells without doxycycline treatment, both CYP11B1 and CYP11B2 mRNAs were significantly decreased, suggesting that both enzymes require a minimal level of SF-1 for basal expression. In summary, SF-1 overexpression dramatically inhibited CYP11B2 expression and decreased aldosterone production. The opposing effects of SF-1 on CYP11B1 and CYP11B2 suggest that the regulation of SF-1 activity may play a role that determines the relative ability to produce mineralocorticoid and glucocorticoid.
Central Infusion of Minocycline Inhibits Proinflammatory Cytokines in the PVN and Attenuates Ang II-Induced Hypertension
Carlos C Diez Freire; Univ of Florida, Gainesville, FL
Convincing evidence supports the view that the CNS plays a key role in angiotensin II (AngII) - dependant hypertension, although the precise signaling pathway in this peripheral/central communication remains elusive. Recent observations of an increased oxidative stress in the cardiovascular (CV) sensitive brain regions and emerging involvement of proinflammatory cytokines (PIC) in the vascular system by ang II has led us to propose the following hypothesis: Central effects of ang II leading to hypertension are, at least in part, mediated by activation of PIC in CV sensitive brain regions. Thus, our objectives in this study were to determine: i) if chronic ang II infusion increases PIC in the brain and ii) if central infusion of minocycline, an anti-inflammatory antibiotic, would decrease PIC and attenuate hypertension. We decided to focus on the PVN since it is the key nucleus that perceives signals from the systemic circulation and relays to the NTS and RVLM for the regulation of sympathetic activity. Male Sprague Dawly rats were implanted with ICV cannulae to infuse minocycline (5ug/kg/hr) and fitted with telemetry probes to measure changes in blood pressure (BP) by SQ infusion of ang II (150ng/kg/min). Basal MAP of SD rats was 94 5 mmHg (n 6). Chronic ang II infusion for 4 weeks resulted in a 29 5 mmHg increase in MAP. In addition, HW/BW ratio was increased by 38% in ang II treated rats. The hypertensive state was accompanied by a 5.6 fold increase in TNF- and 3.8 fold increase in IL-6 in the PVN. Also, cardiac hypertrophy was associated with significant increases in the expression of hypertrophy markers (ANP, 7.6 fold; TNF- , 3.7 fold; IL-6, 8.5 fold). ICV infusion of minocycline resulted in normalization of high BP and cardiac hypertrophy and significant reductions in PIC in the PVN (TNF- 1.05 fold and IL-6, 1.3 fold) and hypertrophy gene markers in the heart (ANP, 0.98; TNF- , 1.2; IL-6,1.3 folds). These observations indicate that central infusion of an anti-inflammatory antibiotic protects rats from the development of ang II - dependant hypertension at least, in part, by inhibiting PIC expression in the PVN. They suggest that PVN cytokines are key contributors in the genesis of ang II - dependant hypertension.
MMP activity is triggered in the vessel wall by multiple stimuli. Alterations in flow (1) and increases in transmural pressure (7) stimulate MMP-2 and -9 activity in blood vessels. In addition, vasoactive compounds, particularly the components of the renin-angiotensin-aldosterone system, are linked to increased MMP activity. Angiotensin II stimulates MMP-2 and -9 expression in the rat thoracic aorta, independently of pressure (45), and MMP-2 gene expression is increased in aldosterone-induced hypertensive rats (46). We have shown that the renin-angiotensin-aldosterone system is an important factor involved in the MCA remodeling observed in SHRSP (14, 42). It is possible that one of the mechanisms underlying aldosterone-dependent remodeling in the SHRSP is an upregulation of MMP activity.
Thus we hypothesized that inhibition of MMP activity with doxycycline (DOX) would attenuate vascular remodeling.
These results suggest that MMPs are involved in hypertensive vascular remodeling in both the peripheral and cerebral vasculature and that DOX reduced brain damage after cerebral ischemia.
Another important effect is protection of the blood-brain-barrier (BBB). MMPs have been implicated in BBB disruption following transient ischemia (31, 43) and hemorrhagic transformation after reperfusion (44). The potential that BBB breakdown is reduced in SHRSP + DOX warrants further investigation.
Effect of steroids on CSF matrix metalloproteinases in multiple sclerosis
Relation to blood-brain barrier injury
G. A. Rosenberg, MD,
J. E. Dencoff, BS,
N. Correa Jr., BS,
M. Reiners, MD and
C. C. Ford, MD, PhD
+ Author Affiliations
From the Departments of Neurology and Physiology, University of New Mexico School of Medicine, and the Neurology Service, Veterans Health Administration Medical Center, Albuquerque, NM.
Supported by a grant to the University of New Mexico for the Clinical Research Center (NIH: NCRR, 5M01 RR00997-18). Other support came from NIH (RO1 NS21169-07) and the VA Research Service to GR.
Received August 15, 1995. Accepted in final form October 12, 1995.
Address correspondence and reprint requests to Dr. Gary A. Rosenberg, Department of Neurology, University of New Mexico, Albuquerque, NM 87131.
Contrast-enhanced MRI in patients with MS shows that increased permeability of the blood-brain barrier (BBB) commonly occurs. The changes in capillary permeability often precede T2-weighted MRI evidence of tissue damage. In animal studies, intracerebral injection of the matrix metalloproteinase (MMP) 72-kDa type IV collagenase (gelatinase A) opens the BBB by disrupting the basal lamina around capillaries. Steroids affect production of endogenous MMPs and tissue inhibitors to metalloproteinases (TIMPs). To determine the role of MMP activity in BBB damage during acute exacerbations of MS, we measured MMPs in the CSF of patients with MS. Patients (n = 7) given steroids to treat an acute episode of MS had CSF sampled before and after 3 days of methylprednisolone (1 g/day). Patients had a graded neurologic examination and gadolinium-enhanced MRI before treatment. CSF studies included total protein, cell count, and a demyelinating profile. We measured levels of MMPs, urokinase-type plasminogen activator (uPA), and TIMPs by zymography, reverse zymography, and Western blots. The MMP, 92-kDa type IV collagenase (gelatinase B), fell from 216 +/- 70 before steroids to 54 +/- 26 relative lysis zone units (p < 0.046) after treatment. Similarly, uPA dropped from 3880 +/- 800 to 2655 +/- 353 (p < 0.03). Four patients with gadolinium enhancement on MRI had the most pronounced drop in gelatinase B and uPA. Western immunoblots showed an increase in a complex of gelatinase B and TIMPs after treatment, suggesting an increase in a TIMP (p < 0.05). Reverse zymography of CSF samples showed that steroids increased a TIMP with a molecular weight similar to that of mouse TIMP-3 (p = 0.053). Our results suggest that increased gelatinase B is associated with an open BBB on MRI. Steroids may improve capillary function by reducing activity of gelatinase B and uPA and increasing levels of TIMPs.
AUG. 17, 2009 3 54Inexpensive hypertension drug could be multiple sclerosis treatment, study shows
BY BRUCE GOLDMAN
Steve Fisch Photography
Lawrence Steinman and his colleagues found that a drug used to treat high blood pressure may also be able to treat multiple sclerosis.
Turning serendipity into science, researchers at the Stanford University School of Medicine have found a link, in mice and in human brain tissue, between high blood pressure and multiple sclerosis. Their findings suggest that a safe, inexpensive drug already in wide use for high blood pressure may have therapeutic value in multiple sclerosis, as well.
While neurology professor Lawrence Steinman, MD, senior author of the new study, cautioned that extensive clinical trial work is needed to determine if the drug, known as lisinopril, can do in humans what it does in mice, he is excited that “we were able to show that all the targets for lisinopril are there and ready for therapeutic manipulation in the multiple-sclerosis lesions of human patients. Without that, this would be just another intriguing paper about what’s possible in the mouse.”
The paper was published online Aug. 17 by the Proceedings of the National Academy of Sciences.
The genesis for the paper can be traced to about seven years ago, when Steinman learned he had high blood pressure. His doctor put him on lisinopril, which is used by millions of people all over the world and has an excellent safety profile. Chagrined, Steinman went home and, researcher that he is, immediately did a Google search on the drug. (Steinman is a renowned multiple sclerosis investigator whose earlier work on the inflammatory features of the disease spurred development of a blockbuster class of anti-inflammatory multiple-sclerosis therapeutics. The drug natalizumab, marketed under the trade name Tysabri, is one.)
» Study yields 'gold mine' of potential drug targets for treating multiple sclerosis
» Researchers show how protein protects nerves from multiple sclerosis
» Study identifies molecule linked to multiple sclerosis relapses
Long ago, a glitch crept into Steinman’s home computer: No matter what keywords he types into the search field, the computer automatically inserts the additional term, “multiple sclerosis.” Thus, to his surprise, a list of medical literature popped up offering tantalizing, if vague, hints of a possible connection between multiple sclerosis and a fast-acting hormone, angiotensin, whose receptors abound on blood-vessel walls throughout the body.
In response to, say, a change in posture, angiotensin immediately causes blood vessels to constrict. “That raises your blood pressure so when you stand up to get out of a chair, you don’t fall down and faint,” said Steinman, who is also the George A. Zimmerman Professor in the medical school. But angiotensin overactivity causes chronic hypertension. Lisinopril controls blood pressure by blocking an enzyme that converts angiotensin’s precursor into the active hormone. The drug also appears to have certain anti-inflammatory properties.
Multiple sclerosis is a chronic and occasionally lethal autoimmune disease in which the body’s immune system mounts recurring assaults on the myelin sheathing of nerve cells in the brain. This causes nerves to malfunction and can lead to blindness and paralysis. Both multiple sclerosis and atherosclerosis involve inflammatory processes.
Eventually, Steinman and his colleagues decided to test the angiotensin/multiple-sclerosis relationship using modern scientific techniques. First, they examined the multiple-sclerosis lesions of brain samples from autopsied patients. In those lesions, well-established molecular-detection methods turned up significantly elevated levels of both the angiotensin receptor and the angiotensin-producing enzyme blocked by lisinopril.
Next, the investigators turned to an equally well-established animal model: a laboratory-bred strain of mouse that, after being immunized with a particular chemical, develops brain lesions very similar to those observed in multiple sclerosis. When, before immunization with the disease-triggering chemical, mice got lisinopril dosages equivalent to those prescribed for humans with high blood pressure, they didn’t develop the paralysis characteristic of disease progression. Strikingly, if it was given after the mice developed full-blown symptoms, lisinopril reversed their paralysis.
The team also found that lisonopril administration reduced numerous molecular measures of inflammation that accompany multiple sclerosis in humans and its analog in the animal model. But, importantly, the drug didn’t inhibit the mice’s overall immune competence.
An additional observation was that lisinopril administration triggered proliferation of an important class of immune cells, called regulatory T cells, that prevent autoimmune diseases by dialing down the activity of other immune cells erroneously targeting cells and tissues that should be left alone. It’s likely, Steinman said, that this proliferation was a key component in the protection provided by the drug, as an infusion of regulatory T cells from mice that had been given lisinopril was sufficient to prevent or reverse the disease process in mice that had been given none.
Steinman’s results have major public-health implications, said Marc Feldmann, an Imperial College London immunologist who is familiar with the study but did not participate in it. He noted that the current therapies for multiple sclerosis (including Tysabri) are pricey monoclonal antibodies, costing tens of thousands of dollars annually for each patient treated. “If multiple-sclerosis patients can be treated with lisinopril at something like 1 percent of the price of treatment with Tysabri, then far more patients will receive adequate therapy, at a substantially lower cost to those paying for it,” Feldmann said.
First authorship of the paper is shared by two postdoctoral researchers in Steinman’s lab: Michael Platten, MD, and Sawsan Youssef, PhD. (Platten is now a neurology professor at University Hospital of Heidelberg, in Germany.) Other Stanford co-authors include May Han, MD, acting assistant professor of neurology and neurological sciences, and Raymond Sobel, MD, professor of pathology.
The study was financed by the National Institutes of Health, the National MS Society, the Phil N. Allen Trust, the German Research Foundation, the Helmholtz Association, the U.S. Public Health Service and the Biomedical Sciences Exchange Program.
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Angiotensin I in the DD Genotype Are Blunted by Low Sodium Intake
Frank G. H. van der Kleij*†,
Paul E. de Jong*†,
Rob H. Henning*‡,
Dick de Zeeuw*†‡ and
+ Author Affiliations
*Groningen University Institute for Drug Exploration, †Department of Internal Medicine, Division of Nephrology, and ‡Department of Clinical Pharmacology, University Hospital Groningen and State University Groningen, Groningen, The Netherlands.
Correspondence to Dr. Gerjan Navis, Department of Internal Medicine, Division of Nephrology, University Hospital Groningen, Hanzeplein 1 P.O. Box 30.001, 9700 RB Groningen, The Netherlands. Phone: +31-50-3612621; Fax: +31-50-3619310; E-mail: g.j.navis@ int.azg.nl
Received for publication August 9, 2001.
Accepted for publication November 27, 2001.
ABSTRACT. Angiotensin-converting enzyme (ACE) activity is increased in the DD genotype, but the functional significance for renal function is unknown. Blunted responses of BP and proteinuria to ACE inhibition among DD renal patients during periods of high sodium intake were reported. It was therefore hypothesized that sodium status affects the phenotype in the ACE I/D polymorphism. The effects of angiotensin I (AngI) and AngII among 27 healthy subjects, with both low (50 mmol sodium/d) and liberal (200 mmol sodium/d) sodium intakes, were studied. Baseline mean arterial pressure (MAP) values, renal hemodynamic parameters, and renin-angiotensin system parameters were similar for all genotypes with either sodium intake level. With liberal sodium intake, the increases in MAP, renal vascular resistance, and aldosterone levels during AngI infusion (8 ng/kg per min) were significantly higher for the DD genotype, compared with the ID and II genotypes (all parameters presented as percent changes ± 95% confidence intervals), with mean MAP increases of 22 ± 2% (DD genotype), 13 ± 5% (ID genotype), and 12 ± 6% (II genotype) (P < 0.05), mean increases in renal vascular resistance of 100.1 ± 19.7% (DD genotype), 73.0 ± 16.3% (ID genotype), and 63.2 ± 16.9% (II genotype) (P < 0.05), and increases in aldosterone levels of 650 ± 189% (DD genotype), 343 ± 71% (ID genotype), and 254 ± 99% (II genotype) (P < 0.05). Also, the decrease in GFR was more pronounced for the DD genotype, with mean decreases of 17.9 ± 4.7% (DD genotype), 8.8 ± 3.4% (ID genotype), and 6.4 ± 5.9% (II genotype) (P < 0.05). The effective renal plasma flow, plasma AngII concentration, and plasma renin activity values were similar for the genotypes. In contrast, with low sodium intake, the responses to AngI were similar for all genotypes. The responses to AngII were also similar for all genotypes, with either sodium intake level. In conclusion, the responses of MAP, renal hemodynamic parameters, and aldosterone concentrations to AngI are enhanced for the DD genotype with liberal but not low sodium intake. These results support the presence of gene-environment interactions between ACE genotypes and dietary sodium intake.
Synergistic induction of osteopontin by aldosterone and inflammatory cytokines in mesangial cells
Stefan Gauer1,*, Ingeborg A. Hauser1, Nicholas Obermüller1, Yvonne Holzmann1, Helmut Geiger1, Margarete Goppelt-Struebe2
Article first published online: 1 JUN 2007
Copyright © 2007 Wiley-Liss, Inc.
Journal of Cellular Biochemistry
Volume 103, Issue 2, pages 615–623, 1 February 2008
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Hypertensive nephrosclerosis is characterized by activation of the renin–angiotensin–aldosterone system in combination with an inflammatory response characterized by an infiltration of T-cells and mononuclear cells, which release proinflammatory cytokines like IL-1β/TNFα. In various models of experimental hypertensive disease the chemokine osteopontin (OPN) enhances further leukocyte infiltration. Therefore, we investigated the induction of OPN expression in renal mesangial cells (MCs) by aldosterone and the inflammatory cytokines IL-1β/TNFα. Incubation with aldosterone resulted in a time- and concentration-dependent increase in OPN mRNA and protein. OPN mRNA expression followed a biphasic time course with an early increase between 4 and 8 h and the second phase starting at 14 h. The early phase was independent of protein synthesis, indicating a direct effect of aldosterone. Aldosterone-mediated induction of OPN was prevented by spironolactone, indicative of a receptor-mediated aldosterone effect. The mineralocorticoid receptor (MR) was identified in MCs by RT-PCR and immunoprecipitation, and shown to interact with a putative aldosterone-response element of the OPN promoter. The proinflammatory cytokines IL-1β and TNFα only marginally affected OPN expression in MCs. However, coincubation of aldosterone and the cytokines synergistically increased OPN mRNA and protein levels. Since the synergistic effect on OPN mRNA was inhibited by diphenyleneiodonium, we assume an involvement of reactive oxygen species (ROS). We conclude that the chemokine OPN is a target gene of aldosterone in renal MCs, which is activated via the MR, and that proinflammatory cytokines enhance aldosterone-dependent OPN expression. In vivo, this may result in further leukocyte infiltration aggravating hypertensive nephrosclerosis. J. Cell. Biochem. 103: 615–623, 2008. © 2007 Wiley-Liss, Inc.
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