Insulin mediated upregulation of the renin angiotensin system in human subcutaneous adipocytes is reduced by Rosiglitazone

Background: Obesity associated hypertension is likely to be due to multiple mechanisms. Identification of the renin-angiotensin system (RAS) within adipose tissue does, however, suggest a potential causal role for it in obesity-associated hypertension. Obese patients are often hyperinsulinaemic, but mechanisms underlying insulin upregulation of the RAS in adipose tissue are unclear. TNFα, an inducer of angiotensinogen in hepatocytes, is elevated in hyperinsulinaemic, obese individuals, and may provide a link in mediating insulin upregulation of the RAS in adipose tissue. Further, thiazolidinediones lower blood pressure in vivo and downregulation of the RAS in adipose tissue may contribute to this effect. We therefore examined the effect of rosiglitazone (RSG), on the insulin mediated upregulation of the RAS. Methods and Results: Sera were obtained from the arterial circulation and from venous blood draining subcutaneous abdominal adipose tissue. Isolated human abdominal subcutaneous adipocytes (n=12) were treated with insulin (1-1000nM) and insulin in combination with RSG (10nM), and RSG (10nM) alone to determine angiotensinogen expression, angiotensin II, bradykinin and TNFα secretion. Subcutaneous adipocytes were also treated with TNFα (10-100ng/mL) to examine the direct effect on angiotensinogen expression and angiotensin II secretion. The findings showed that the arterio-venous difference in angiotensin II levels was significant (↑23%; p<0.001). Insulin increased TNFα secretion in a concentration-dependent manner (p<0.01) whilst RSG (10nM) significantly reduced the insulin mediated rise in TNFα (p<0.001), as well as AGT and angiotensin II. TNFα also increased angiotensinogen and angiotensin II in isolated adipocytes. Conclusions: Our in vivo data suggest that human subcutaneous adipose tissue is a significant source of angiotensin II. This study also demonstrates a potential TNFα mediated mechanism through which insulin may stimulate the RAS and may contribute to explain obesity associated hypertension. RSG downregulates the RAS in subcutaneous adipose tissue and this effect may contribute to the long-term effect of RSG on blood pressure

Vascular smooth muscle Sirtuin-1 protects against aortic dissection during Angiotensin II-induced hypertension

BACKGROUND: Sirtuin-1 (SirT1), a nicotinamide adenine dinucleotide(+)-dependent deacetylase, is a key enzyme in the cellular response to metabolic, inflammatory, and oxidative stresses; however, the role of endogenous SirT1 in the vasculature has not been fully elucidated. Our goal was to evaluate the role of vascular smooth muscle SirT1 in the physiological response of the aortic wall to angiotensin II, a potent hypertrophic, oxidant, and inflammatory stimulus. METHODS AND RESULTS: Mice lacking SirT1 in vascular smooth muscle (ie, smooth muscle SirT1 knockout) had drastically high mortality (70%) caused by aortic dissection after angiotensin II infusion (1 mg/kg per day) but not after an equipotent dose of norepinephrine, despite comparable blood pressure increases. Smooth muscle SirT1 knockout mice did not show any abnormal aortic morphology or blood pressure compared with wild-type littermates. Nonetheless, in response to angiotensin II, aortas from smooth muscle SirT1 knockout mice had severely disorganized elastic lamellae with frequent elastin breaks, increased oxidant production, and aortic stiffness compared with angiotensin II-treated wild-type mice. Matrix metalloproteinase expression and activity were increased in the aortas of angiotensin II-treated smooth muscle SirT1 knockout mice and were prevented in mice overexpressing SirT1 in vascular smooth muscle or with use of the oxidant scavenger tempol. CONCLUSIONS: Endogenous SirT1 in aortic smooth muscle is required to maintain the structural integrity of the aortic wall in response to oxidant and inflammatory stimuli, at least in part, by suppressing oxidant-induced matrix metalloproteinase activity. SirT1 activators could potentially be a novel therapeutic approach to prevent aortic dissection and rupture in patients at risk, such as those with hypertension or genetic disorders, such as Marfan's syndrome.R01 HL098028 - NHLBI NIH HHS; HL098028 - NHLBI NIH HHS; HL105287 - NHLBI NIH HHS; T32 HL07224 - NHLBI NIH HH

In Vitro Effects of DuP 753, a Nonpeptide Angiotensin II Receptor Antagonist, on Human Platelets and Rat Vascular Smooth Muscle Cells

These experiments were designed to assess the ability of the new nonpeptide angiotensin II antagonist DuP 753 to inhibit the binding and, particularly, to antagonize the cellular response to angiotensin II in human platelets and primary cultures of rat aortic smooth muscle cells (SMC). The binding of 125I-angiotensin II was competitively inhibited by DuP 753 with a 50% binding inhibition (IC50) of 5 to 6 × 10—8 mol/L in platelets and 1 × 10—8 mol/L in vascular SMC as compared to an IC50 of 5 to 7.5 × 10×9 mol/L with nonlabeled angiotensin II. In vascular SMC, DuP 753 completely abolished the effects of angiotensin II on 45CaCl2 efflux and 45CaCl2 uptake. Moreover, in these latter cells, DuP 753 prevented the angiotensin II but not the vasopressin induced increase in cytosolic calcium. These results demonstrate that DuP 753 competes with angiotensin II binding to its receptor in both animal and human cells and selectively blocks the cellular response to angiotensin II. Am J Hypertens 1991;4:438-44

ACE-versus chymase-dependent angiotensin II generation in human coronary arteries: a matter of efficiency?

OBJECTIVE: The objective of this study was to investigate ACE- and chymase-dependent angiotensin I-to-II conversion in human coronary arteries (HCAs). METHODS AND RESULTS: HCA rings were mounted in organ baths, and concentration-response curves to angiotensin II, angiotensin I, and the chymase-specific substrate Pro(11)-D-Ala(12)-angiotensin I (PA-angiotensin I) were constructed. All angiotensins displayed similar efficacy. For a given vasoconstriction, bath (but not interstitial) angiotensin II during angiotensin I and PA-angiotensin I was lower than during angiotensin II, indicating that interstitial (and not bath) angiotensin II determines vasoconstriction. PA-angiotensin I increased interstitial angiotensin II less efficiently than angiotensin I. Separate inhibition of ACE (with captopril) and chymase (with C41 or chymostatin) shifted the angiotensin I concentration-response curve approximately 5-fold to the right, whereas a 10-fold shift occurred during combined ACE and chymase inhibition. Chymostatin, but not captopril and/or C41, reduced bath angiotensin II and abolished PA-Ang I-induced vasoconstriction. Perfused HCA segments, exposed luminally or adventitially to angiotensin I, released angiotensin II into the luminal and adventitial fluid, respectively, and this release was blocked by chymostatin. CONCLUSIONS: Both ACE and chymase contribute to the generation of functionally active angiotensin II in HCAs. However, because angiotensin II loss in the organ bath is chymase-dependent, ACE-mediated conversion occurs more efficiently (ie, closer to AT(1) receptors) than chymase-mediated conversion

Atypical angiotensin II receptors coupled to phosphoinositide turnover/calcium signalling in catfish hepatocytes

AbstractIn catfish (Ictalurus punctatus) hepatocytes angiotensin II induced an immediate increase in cytosolic Ca2+ concentration. Other angiotensin analogues also induced this effect including: human angiotensin II, fish angiotensin II, human angiotensin III, human angiotensin I, fish angiotensin I and saralasin. CGP 42112A induced a very small effect at the highest concentration tested and angiotensin IV was without effect. Angiotensin II also increased the resynthesis of phosphatidylinositol and the production of IP3. These physiological effects were not blocked by losartan (AT1-selective antagonist) or PD 123177 (AT2-selective antagonist).[125I]Angiotensin II bound to liver plasma membranes in a saturable fashion with high affinity (KD 2.7 nM) and a Bmax of 185 fmol/mg of protein. Binding competition experiments showed the following order of potency: human angiotensin II=fish angiotensin II>human angiotensin III≥human angiotensin I=fish angiotensin I. These sites were insensitive to losartan or PD 123177.The data indicate that the angiotensin II receptors expressed in catfish hepatocytes are coupled to the phosphoinositide turnover/calcium mobilization signal transduction pathway and are atypical receptors, i.e., pharmacologically distinct from mammalian AT1 and AT2 receptors

Angiotensin II diminishes the effect of SGK1 on the WNK4-mediated inhibition of ROMK1 channels

ROMK1 channels are located in the apical membrane of the connecting tubule and cortical collecting duct and mediate the potassium secretion during normal dietary intake. We used a perforated whole-cell patch clamp to explore the effect of angiotensin II on these channels in HEK293 cells transfected with green fluorescent protein (GFP)-ROMK1. Angiotensin II inhibited ROMK1 channels in a dose-dependent manner, an effect abolished by losartan or by inhibition of protein kinase C. Furthermore, angiotensin II stimulated a protein kinase C-sensitive phosphorylation of tyrosine 416 within c-Src. Inhibition of protein tyrosine kinase attenuated the effect of angiotensin II. Western blot studies suggested that angiotensin II inhibited ROMK1 channels by enhancing its tyrosine phosphorylation, a notion supported by angiotensin II's failure to inhibit potassium channels in cells transfected with the ROMK1 tyrosine mutant (R1Y337A). However, angiotensin II restored the with-no-lysine kinase-4 (WNK4)-induced inhibition of R1Y337A in the presence of serum–glucocorticoids-induced kinase 1 (SGK1), which reversed the inhibitory effect of WNK4 on ROMK1. Moreover, protein tyrosine kinase inhibition abolished the angiotensin II-induced restoration of WNK4-mediated inhibition of ROMK1. Angiotensin II inhibited ROMK channels in the cortical collecting duct of rats on a low sodium diet, an effect blocked by protein tyrosine kinase inhibition. Thus, angiotensin II inhibits ROMK channels by two mechanisms: increasing tyrosine phosphorylation of the channel and synergizing the WNK4-induced inhibition. Hence, angiotensin II may have an important role in suppressing potassium secretion during volume depletion

Exogenous angiotensin II does not facilitate norepinephrine release in the heart

Studies on the effect of angiotensin II on norepinephrine release from sympathetic nerve terminals through stimulation of presynaptic angiotensin II type 1 receptors are equivocal. Furthermore, evidence that angiotensin II activates the cardiac sympathetic nervous system in vivo is scarce or indirect. In the intact porcine heart, we investigated whether angiotensin II increases norepinephrine concentrations in the myocardial interstitial fluid (NE(MIF)) under basal conditions and during sympathetic activation and whether it enhances exocytotic and nonexocytotic ischemia-induced norepinephrine release. In 27 anesthetized pigs, NE(MIF) was measured in the left ventricular myocardium using the microdialysis technique. Local infusion of angiotensin II into the left anterior descending coronary artery (LAD) at consecutive rates of 0.05, 0.5, and 5 ng/kg per minute did not affect NE(MIF), LAD flow, left ventricular dP/dt(max), and arterial pressure despite large increments in coronary arterial and venous angiotensin II concentrations. In the presence of neuronal reuptake inhibition and alpha-adrenergic receptor blockade, left stellate ganglion stimulation increased NE(MIF) from 2.7+/-0.3 to 7.3+/-1.2 before, and from 2.3+/-0.4 to 6.9+/-1.3 nmol/L during, infusion of 0.5 ng/kg per minute angiotensin II. Sixty minutes of 70% LAD flow reduction caused a progressive increase in NE(MIF) from 0.9+/-0.1 to 16+/-6 nmol/L, which was not enhanced by concomitant infusion of 0.5 ng/kg per minute angiotensin II. In conclusion, we did not observe any facilitation of cardiac norepinephrine release by angiotensin II under basal conditions and during either physiological (ganglion stimulation) or pathophysiological (acute ischemia) sympathetic activation. Hence, angiotensin II is not a local mediator of cardiac sympathetic activity in the in vivo porcine heart

Studies into the Regulation of FTO by Angiotensin II in Cardiomyocyte Hypertrophy

Leptin has been shown to upregulate the fat mass and obesity-associated protein (FTO) in the cardiomyocyte, an effect that has been linked to cardiomyocyte hypertrophy. In this current study, we sought to determine if other cardiomyocyte hypertrophic agonists could upregulate FTO. Angiotensin II displayed the ability to significantly upregulate mRNA expression and protein expression of FTO. Thus, in this study, we explored the cellular mechanisms behind angiotensin II-induced FTO upregulation. Angiotensin II-induced FTO upregulation was attenuated by 10 µM valsartan, an AT-1 receptor antagonist, 5 µM EMD87580, a sodium-hydrogen exchanger isoform 1 (NHE-1) inhibitor, and 2 nM FK506, a calcineurin inhibitor. This data suggests that angiotensin II-induced FTO upregulation is AT-1 receptor mediated and has a signal transduction pathway involving the NHE-1/calcineurin system. All the above agents also abolished angiotensin II-induced hypertrophy. In addition, our studies show that a super-ovine leptin receptor antagonist (LRA; 100 nM) and JAK2 inhibitor, 50 µM AG490, also attenuate angiotensin II-induced FTO upregulation. These results implicate the leptin-JAK2-STAT3 signalling system in angiotensin II-induced FTO upregulation. The above agents also abolished angiotensin II-induced hypertrophy. The data suggests that NHE-1/calcineurin as well as leptin-mediated JAK2/STAT3 signalling play a critical role in angiotensin II-induced FTO upregulation

Transgenic amplification of glucocorticoid action in adipose tissue causes high blood pressure in mice

Obesity is closely associated with the metabolic syndrome, a combination of disorders including insulin resistance, diabetes, dyslipidemia, and hypertension. A role for local glucocorticoid reamplification in obesity and the metabolic syndrome has been suggested. The enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) regenerates active cortisol from inactive 11-keto forms, and aP2-HSD1 mice with relative transgenic overexpression of this enzyme in fat cells develop visceral obesity with insulin resistance and dyslipidemia. Here we report that aP2-HSD1 mice also have high arterial blood pressure (BP). The mice have increased sensitivity to dietary salt and increased plasma levels of angiotensinogen, angiotensin II, and aldosterone. This hypertension is abolished by selective angiotensin II receptor AT-1 antagonist at a low dose that does not affect BP in non-Tg littermates. These findings suggest that activation of the circulating renin-angiotensin system (RAS) develops in aP2-HSD1 mice. The long-term hypertension is further reflected by an appreciable hypertrophy and hyperplasia of the distal tubule epithelium of the nephron, resembling salt-sensitive or angiotensin II–mediated hypertension. Taken together, our findings suggest that overexpression of 11β-HSD1 in fat is sufficient to cause salt-sensitive hypertension mediated by an activated RAS. The potential role of adipose 11β-HSD1 in mediating critical features of the metabolic syndrome extends beyond obesity and metabolic complications to include the most central cardiovascular feature of this disorder

Cyclooxygenase-1 Is Involved in Endothelial Dysfunction of Mesenteric Small Arteries From Angiotensin II–Infused Mice

Angiotensin II induces endothelial dysfunction by reducing NO availability and increasing reactive oxygen species. We assessed whether cyclooxygenase (COX)-1 or COX-2 participate in the angiotensin II–induced endothelial dysfunction in murine mesenteric small arteries and examined the role of reduced nicotinamide-adenine dinucleotide phosphate–dependent reactive oxygen species production. Mice received angiotensin II (600 ng/kg per minute, SC), saline (controls), angiotensin II + apocynin (reduced nicotinamide-adenine dinucleotide phosphate oxidase inhibitor, 2.5 mg/day), or apocynin alone for 2 weeks. Endothelial function of mesenteric arteries was assessed by pressurized myograph. In controls, acetylcholine-induced relaxation was inhibited by N G -monomethyl- l -arginine and unaffected by DFU (COX-2 inhibitor), SC-560 (COX-1 inhibitor), or ascorbic acid. In angiotensin II–infused animals, the attenuated response to acetylcholine was less sensitive to N G -monomethyl- l -arginine, unaffected by DFU, and enhanced by SC-560 and, similarly, by SQ-29548, a thromboxane–prostanoid receptor antagonist. Moreover, response to acetylcholine was unchanged by ozagrel, a thromboxane synthase inhibitor, and normalized by ascorbic acid. Apocynin prevented the angiotensin II–induced vascular dysfunctions. In angiotensin II–infused mice, RT-PCR analysis showed a significant COX-2 downregulation, whereas COX-1 expression was upregulated. These changes were unaffected by apocynin. Modulation of COX isoform by angiotensin II was also documented by immunohistochemistry. In small mesenteric vessels, the reduced NO availability and oxidant excess, which characterize endothelial dysfunction secondary to angiotensin II, are associated with a reduced COX-2 and an increased COX-1 function and expression. Angiotensin II causes an oxidative stress–independent COX-1 overexpression, whereas angiotensin II–mediated oxidant excess production stimulates COX-1 activity to produce a contracting prostanoid endowed with agonist activity on thromboxane–prostanoid receptors