Kengo Azushima

Activation of angiotensin II type 1 receptor-associated protein exerts an inhibitory effect on vascular hypertrophy and oxidative stress in angiotensin II-mediated hypertension

AIMS Activation of tissue angiotensin II (Ang II) type 1 receptor (AT1R) plays an important role in the development of vascular remodelling. We have shown that the AT1R-associated protein (ATRAP/Agtrap), a specific binding protein of AT1R, functions as an endogenous inhibitor to prevent pathological activation of the tissue renin-angiotensin system. In this study, we investigated the effects of ATRAP on Ang II-induced vascular remodelling. METHODS AND RESULTS Transgenic (Tg) mice with a pattern of aortic vascular-dominant overexpression of ATRAP were obtained, and Ang II or vehicle was continuously infused into Tg and wild-type (Wt) mice via an osmotic minipump for 14 days. Although blood pressure of Ang II-infused Tg mice was comparable with that of Ang II-infused Wt mice, the Ang II-mediated development of aortic vascular hypertrophy was partially inhibited in Tg mice compared with Wt mice. In addition, Ang II-mediated up-regulation of vascular Nox4 and p22(phox), NADPH oxidase components, and 4-HNE, a marker of reactive oxygen species (ROS) generation, was significantly suppressed in Tg mice, with a concomitant inhibition of activation of aortic vascular p38MAPK and JNK by Ang II. This protection afforded by vascular ATRAP against Ang II-induced activation of NADPH oxidase is supported by in vitro experimental data using adenoviral transfer of recombinant ATRAP. CONCLUSION These results indicate that activation of aortic vascular ATRAP partially inhibits the Nox4/p22(phox)-ROS-p38MAPK/JNK pathway and pathological aortic hypertrophy provoked by Ang II-mediated hypertension, thereby suggesting ATRAP as a novel receptor-binding modulator of vascular pathophysiology.

Enhanced Angiotensin Receptor-Associated Protein in Renal Tubule Suppresses Angiotensin-Dependent Hypertension

We have previously shown that angiotensin II type 1 receptor-associated protein (ATRAP/Agtrap) interacts with the angiotensin II type 1 receptor and promotes constitutive internalization of the receptor so as to inhibit the pathological activation of its downstream signaling but preserve baseline physiological signaling activity. The present study was designed to investigate the role of renal ATRAP in angiotensin II–dependent hypertension. We generated transgenic mice dominantly expressing ATRAP in the renal tubules, including renal distal tubules. The renal ATRAP transgenic mice exhibited no significant change in blood pressure at baseline on normal salt diet. However, in the renal ATRAP transgenic mice compared with wild-type mice, the following took place: (1) the development of high blood pressure in response to angiotensin II infusion was significantly suppressed based on radiotelemetry, (2) the extent of daily positive sodium balance was significantly reduced during angiotensin II infusion in metabolic cage analysis, and (3) the renal Na+-Cl− cotransporter activation and &agr;-subunit of the epithelial sodium channel induction by angiotensin II infusion were inhibited. Furthermore, adenoviral overexpression of ATRAP suppressed the angiotensin II–mediated increase in the expression of &agr;-subunit of the epithelial sodium channel in mouse distal convoluted tubule cells. These results indicate that renal tubule–dominant ATRAP activation provokes no evident effects on blood pressure at baseline but exerts an inhibitory effect on the pathological elevation of blood pressure in response to angiotensin II stimulation, thereby suggesting that ATRAP is a potential target of interest in blood pressure modulation under pathological conditions.

The physiology and pathophysiology of a novel angiotensin receptor-binding protein ATRAP/Agtrap.

The Ang II type 1 receptor (AT1R)-associated protein (ATRAP/Agtrap) is a molecule specifically interacting with the carboxyl- terminal domain of AT1R. The results of in vitro studies showed that ATRAP suppresses Ang II-mediated pathological responses in cardiovascular cells by promoting AT1R internalization. With respect to the tissue distribution and regulation of ATRAP expression in vivo, ATRAP is broadly expressed in many tissues as is AT1R. Accumulating evidence indicates that a tissue-specific regulatory balancing of ATRAP and AT1R expression may be involved in the modulation of AT1R signaling at local tissue sites and also in the pathophysiology of hypertension and its associated end-organ injury. Furthermore, the activation of ATRAP in transgenic-models inhibited inflammatory vascular remodeling and cardiac hypertrophy in response to Ang II stimulation. These results suggest the clinical potential benefit of an ATRAP activation strategy in the treatment of hypertension and related organ injury.

Effects of aliskiren-based therapy on ambulatory blood pressure profile, central hemodynamics, and arterial stiffness in nondiabetic mild to moderate hypertensive patients.

J Clin Hypertens (Greenwich). 2012;00:000–000. ©2012 Wiley Periodicals, Inc.

Bofu-Tsu-Shosan, an Oriental Herbal Medicine, Exerts a Combinatorial Favorable Metabolic Modulation Including Antihypertensive Effect on a Mouse Model of Human Metabolic Disorders with Visceral Obesity

Accumulating evidence indicates that metabolic dysfunction with visceral obesity is a major medical problem associated with the development of hypertension, type 2 diabetes (T2DM) and dyslipidemia, and ultimately severe cardiovascular and renal disease. Therefore, an effective anti-obesity treatment with a concomitant improvement in metabolic profile is important for the treatment of metabolic dysfunction with visceral obesity. Bofu-tsu-shosan (BOF) is one of oriental herbal medicine and is clinically available to treat obesity in Japan. Although BOF is a candidate as a novel therapeutic strategy to improve metabolic dysfunction with obesity, the mechanism of its beneficial effect is not fully elucidated. Here, we investigated mechanism of therapeutic effects of BOF on KKAy mice, a model of human metabolic disorders with obesity. Chronic treatment of KKAy mice with BOF persistently decreased food intake, body weight gain, low-density lipoprotein cholesterol and systolic blood pressure. In addition, both tissue weight and cell size of white adipose tissue (WAT) were decreased, with concomitant increases in the expression of adiponectin and peroxisome proliferator-activated receptors genes in WAT as well as the circulating adiponectin level by BOF treatment. Furthermore, gene expression of uncoupling protein-1, a thermogenesis factor, in brown adipose tissue and rectal temperature were both elevated by BOF. Intriguingly, plasma acylated-ghrelin, an active form of orexigenic hormone, and short-term food intake were significantly decreased by single bolus administration of BOF. These results indicate that BOF exerts a combinatorial favorable metabolic modulation including antihypertensive effect, at least partially, via its beneficial effect on adipose tissue function and its appetite-inhibitory property through suppression on the ghrelin system.

The angiotensin II type 1 receptor blocker olmesartan preferentially improves nocturnal hypertension and proteinuria in chronic kidney disease

Accumulated evidence suggests that an altered ambulatory blood pressure (BP) profile, particularly elevated nighttime BP, reflects target organ injury and is a better predictor of further cardiorenal risk than the clinic BP or daytime BP in hypertensive patients complicated by chronic kidney disease (CKD). In this study, we examined the beneficial effects of olmesartan, an angiotensin II type 1 receptor blocker (ARB), on ambulatory BP profiles and renal function in hypertensive CKD patients. Forty-six patients were randomly assigned to the olmesartan add-on group (n=23) or the non-ARB group (n=23). At baseline and after the 16-week treatment period, ambulatory BP monitoring was performed and renal function parameter measurements were collected. Although the baseline clinic BP levels and the after-treatment/baseline (A/B) ratios of clinic BP levels were similar in the olmesartan add-on and non-ARB groups, the A/B ratios of ambulatory 24-h and nighttime BP levels in the olmesartan add-on group were significantly lower. Furthermore, the A/B ratios of urinary protein, albumin and type IV collagen excretion in the olmesartan add-on group were significantly lower than those in the non-ARB group (urinary protein excretion, 0.72±0.41 vs. 1.45±1.48, P=0.030; urinary albumin excretion, 0.73±0.37 vs. 1.50±1.37, P=0.005; urinary type IV collagen excretion, 0.87±0.42 vs. 1.48±0.87, P=0.014) despite comparable A/B ratios for the estimated glomerular filtration rate in the two groups. These results indicate that in hypertensive patients with CKD, olmesartan add-on therapy improves the ambulatory BP profile via a preferential reduction in nighttime BP with concomitant renal injury inhibition.

Deletion of the angiotensin II type 1 receptor–associated protein enhances renal sodium reabsorption and exacerbates angiotensin II–mediated hypertension

Angiotensin II type 1 receptor (AT1R)–associated protein (ATRAP) promotes AT1R internalization along with suppression of pathological activation of tissue AT1R signaling. However, the functional significance of ATRAP in renal sodium handling and blood pressure regulation under pathological stimuli is not fully resolved. Here we show the blood pressure of mice with a gene-targeted disruption of ATRAP was comparable to that of wild-type mice at baseline. However, in ATRAP-knockout mice, angiotensin II–induced hypertension was exacerbated and the extent of positive sodium balance was increased by angiotensin II. Renal expression of the sodium-proton antiporter 3, a major sodium transporter in the proximal tubules, urinary pH, renal angiotensinogen production, and angiotensin II content was unaffected. Stimulation of the renal expression and activity of the epithelial sodium channel (ENaC), a major sodium transporter in the distal tubules, was significantly enhanced by chronic angiotensin II infusion. The circulating and urinary aldosterone levels were comparable. The blood pressure response and renal ENaC expression by aldosterone were not affected. Thus, ATRAP deficiency exacerbated angiotensin II–mediated hypertension by pathological activation of renal tubular AT1R by angiotensin II. This directly stimulates ENaC in the distal tubules and enhances sodium retention in an aldosterone-independent manner.

Effects of the oriental herbal medicine Bofu-tsusho-san in obesity hypertension: A multicenter, randomized, parallel-group controlled trial (ATH-D-14-01021.R2)

OBJECTIVE There is no clinical evidence that supports the benefit of integrative medicine, defined as combination therapy of oriental and western medicine, on obesity-related hypertension. This study evaluates the efficacy of Bofu-tsusho-san (BOF), an oriental herbal medicine, on the ambulatory blood pressure (BP) profile in hypertensive patients with obesity. METHODS The study design was a multicenter, randomized, open-label, parallel-group controlled trial in 107 hypertensive patients with obesity. Participants were randomly assigned to receive either the conventional control therapy or BOF add-on therapy. In both groups antihypertensive therapy was aimed at achieving the target clinic BP. The primary outcome was change in the ambulatory BP profile from baseline to 24 weeks after randomization. RESULTS Daytime systolic BP variability, an important parameter of ambulatory BP profile, was decreased in the BOF group, and the difference in the changes in daytime systolic BP variability was significant between the BOF and control group (Control vs BOF; the change from baseline in daytime systolic BP variability, 1.0±3.3 vs -1.0±3.3%; p=0.006). CONCLUSION The BOF add-on therapy effectively improved the ambulatory BP variability. This is the first report suggesting that an integrative medicine approach may exert favorable effects on obesity-related hypertension compared with conventional pharmaceutical treatment. CLINICAL TRIAL REGISTRATION UMIN000003878.

Angiotensin receptor-binding protein ATRAP/Agtrap inhibits metabolic dysfunction with visceral obesity.

Background Metabolic disorders with visceral obesity have become a major medical problem associated with the development of hypertension, type 2 diabetes, and dyslipidemia and, ultimately, life‐threatening cardiovascular and renal diseases. Adipose tissue dysfunction has been proposed as the cause of visceral obesity‐related metabolic disorders, moving the tissue toward a proinflammatory phenotype. Methods and Results Here we first report that adipose tissues from patients and mice with metabolic disorders exhibit decreased expression of ATRAP/Agtrap, which is a specific binding modulator of the angiotensin II type 1 receptor, despite its abundant expression in adipose tissues from normal human and control mice. Subsequently, to examine a functional role of ATRAP in the pathophysiology of metabolic disorders, we produced homozygous ATRAP deficient (Agtrap−/−) mice, which exhibited largely normal physiological phenotype at baseline. Under dietary high fat loading, Agtrap−/− mice displayed systemic metabolic dysfunction, characterized by an increased accumulation of pad fat, hypertension, dyslipidemia, and insulin resistance, along with adipose tissue inflammation. Conversely, subcutaneous transplantation of donor fat pads overexpressing ATRAP derived from Agtrap transgenic mice to Agtrap−/− recipient mice improved the systemic metabolic dysfunction. Conclusions These results demonstrate that Agtrap−/− mice are an effective model of metabolic disorders with visceral obesity and constitute evidence that ATRAP plays a protective role against insulin resistance, suggesting a new therapeutic target in metabolic disorders. Identification of ATRAP as a novel receptor binding modulator of adipose tissue inflammation not only has cardiovascular significance but may have generalized implication in the regulation of tissue function.

Involvement of Runx3 in the basal transcriptional activation of the mouse angiotensin II type 1 receptor-associated protein gene.

We previously cloned a molecule that interacts with angiotensin II type 1 (AT1) receptor to exert an inhibitory function on AT1 receptor signaling that we named ATRAP/Agtrap (for AT1 receptor-associated protein). In the present study we examined the regulation of basal ATRAP gene expression using renal distal convoluted tubule cells. We found that serum starvation upregulated basal expression of ATRAP gene, a response that required de novo mRNA and protein synthesis. Luciferase assay revealed that the proximal promoter region directs transcription and that a putative binding site of runt-related transcription factors (RBE) is important for transcriptional activation. The results of RBE-decoy transfection and endogenous knockdown by small interference RNA showed that the runt-related transcription factor Runx3 is involved in ATRAP gene expression. Chromatin immunoprecipitation assay also supported the binding of Runx3 to the ATRAP promoter in renal distal convoluted tubule cells. Immunohistochemistry demonstrated the expression of Runx3 and ATRAP proteins in the distal convoluted and connecting tubules of the kidney in consecutive sections. Furthermore, the Runx3 immunostaining was decreased together with a concomitant suppression of ATRAP expression in the affected kidney after 7 days of unilateral ureteral obstruction. These findings indicate that Runx3 plays a role in ATRAP gene expression in renal distal tubular cells both in vitro and in vivo.