Michiaki Abe

Possible Role of P-450 Metabolite of Arachidonic Acid in Vasodilator Mechanism of Angiotensin II Type 2 Receptor in the Isolated Microperfused Rabbit Afferent Arteriole

Although angiotensin II type 2 (AT2) receptor has recently been cloned, its functional role is not well understood. We tested the hypothesis that selective activation of AT2 receptor causes vasodilation in the preglomerular afferent arteriole (Af-Art), a vascular segment that accounts for most of the preglomerular resistance. We microperfused rabbit Af-Arts at 60 mmHg in vitro, and examined the effect of angiotensin II (Ang II; 10(-11)-10(-8) M) on the luminal diameter in the presence or absence of the Ang II type 1 receptor antagonist CV11974 (CV; 10(-8) M). Ang II was added to both the bath and lumen of preconstricted Af-Arts. Ang II further constricted Af-Arts without CV (by 74+/-7% over the preconstricted level at 10(-8) M; P < 0.01, n = 7). In contrast, in the presence of CV, Ang II caused dose-dependent dilation; Ang II at 10(-8) M increased the diameter by 29+/-2% (n = 7, P < 0.01). This dilation was completely abolished by pretreatment with an AT2 receptor antagonist PD123319 (10(-7) M, n = 6), suggesting that activation of AT2 receptor causes vasodilation in Af-Arts. The dilation was unaffected by inhibiting either nitric oxide synthase (n = 7) or cyclooxygenase (n = 7), however, it was abolished by either disrupting the endothelium (n = 10) or inhibiting the cytochrome P-450 pathway, particularly the synthesis of epoxyeicosatrienoic acids (EETs, n = 7). These results suggest that in the Af-Art activation of the AT2 receptor may cause endothelium-dependent vasodilation via a cytochrome P-450 pathway, possibly by EETs.

Strain vessel hypothesis : a viewpoint for linkage of albuminuria and cerebro-cardiovascular risk

Albuminuria is closely associated with stroke and cardiovascular diseases (CVDs) as well as the salt sensitivity of blood pressure (BP). Although albuminuria may reflect generalized endothelial dysfunction, there may be more specific hemodynamic mechanisms underlying these associations. Cerebral hemorrhage and infarction occur most frequently in the area of small perforating arteries that are exposed to high pressure and that have to maintain strong vascular tone in order to provide large pressure gradients from the parent vessels to the capillaries. Analogous to the perforating arteries are the glomerular afferent arterioles of the juxtamedullary nephrons. Hypertensive vascular damage occurs first and more severely in the juxtamedullary glomeruli. Therefore, albuminuria may be an early sign of vascular damages imposed on ‘strain vessels’ such as perforating arteries and juxtamedullary afferent arterioles. Coronary circulation also occurs under unique hemodynamic conditions, in which the entire epicardial segments are exposed to very high pressure with little flow during systolic phases. From the evolutionary point of view, we speculate that such circulatory systems in the vital organs are mandatory for survival under the danger of hypoperfusion due to difficult access to salt and water as well as high risks of wound injuries. In addition, albuminuria would indicate an impairment of renal medullary circulation, downstream from the juxtamedullary glomeruli, and therefore an impaired pressure natriuresis, which would lead to salt sensitivity of BP. Our ‘strain vessel hypothesis’ may explain why hypertension and diabetes, unforeseen in the concept of evolution, preferentially affect vital organs such as the brain, heart and kidney.

Rare variant discovery by deep whole-genome sequencing of 1,070 Japanese individuals

The Tohoku Medical Megabank Organization reports the whole-genome sequences of 1,070 healthy Japanese individuals and construction of a Japanese population reference panel (1KJPN). Here we identify through this high-coverage sequencing (32.4 × on average), 21.2 million, including 12 million novel, single-nucleotide variants (SNVs) at an estimated false discovery rate of <1.0%. This detailed analysis detected signatures for purifying selection on regulatory elements as well as coding regions. We also catalogue structural variants, including 3.4 million insertions and deletions, and 25,923 genic copy-number variants. The 1KJPN was effective for imputing genotypes of the Japanese population genome wide. These data demonstrate the value of high-coverage sequencing for constructing population-specific variant panels, which covers 99.0% SNVs of minor allele frequency ≥0.1%, and its value for identifying causal rare variants of complex human disease phenotypes in genetic association studies.

SLCO4C1 Transporter Eliminates Uremic Toxins and Attenuates Hypertension and Renal Inflammation

Hypertension in patients with chronic kidney disease (CKD) strongly associates with cardiovascular events. Among patients with CKD, reducing the accumulation of uremic toxins may protect against the development of hypertension and progression of renal damage, but there are no established therapies to accomplish this. Here, overexpression of human kidney-specific organic anion transporter SLCO4C1 in rat kidney reduced hypertension, cardiomegaly, and inflammation in the setting of renal failure. In addition, SLCO4C1 overexpression decreased plasma levels of the uremic toxins guanidino succinate, asymmetric dimethylarginine, and the newly identified trans-aconitate. We found that xenobiotic responsive element core motifs regulate SLCO4C1 transcription, and various statins, which act as inducers of nuclear aryl hydrocarbon receptors, upregulate SLCO4C1 transcription. Pravastatin, which is cardioprotective, increased the clearance of asymmetric dimethylarginine and trans-aconitate in renal failure. These data suggest that drugs that upregulate SLCO4C1 may have therapeutic potential for patients with CKD.

Plasticity of Renal Erythropoietin-Producing Cells Governs Fibrosis

CKD progresses with fibrosis and erythropoietin (Epo)-dependent anemia, leading to increased cardiovascular complications, but the mechanisms linking Epo-dependent anemia and fibrosis remain unclear. Here, we show that the cellular phenotype of renal Epo-producing cells (REPs) alternates between a physiologic Epo-producing state and a pathologic fibrogenic state in response to microenvironmental signals. In a novel mouse model, unilateral ureteral obstruction-induced inflammatory milieu activated NFκB and Smad signaling pathways in REPs, rapidly repressed the Epo-producing potential of REPs, and led to myofibroblast transformation of these cells. Moreover, we developed a unique Cre-based cell-fate tracing method that marked current and/or previous Epo-producing cells and revealed that the majority of myofibroblasts are derived from REPs. Genetic induction of NFκB activity selectively in REPs resulted in myofibroblastic transformation, indicating that NFκB signaling elicits a phenotypic switch. Reversing the unilateral ureteral obstruction-induced inflammatory microenvironment restored the Epo-producing potential and the physiologic phenotype of REPs. This phenotypic reversion was accelerated by anti-inflammatory therapy. These findings demonstrate that REPs possess cellular plasticity, and suggest that the phenotypic transition of REPs to myofibroblasts, modulated by inflammatory molecules, underlies the connection between anemia and renal fibrosis in CKD.

Enhanced Superoxide Production in Renal Outer Medulla of Dahl Salt-Sensitive Rats Reduces Nitric Oxide Tubular-Vascular Cross-Talk

Studies were conducted to determine whether the diffusion of NO from the renal medullary thick ascending limb (mTAL) to the contractile pericytes of surrounding vasa recta was reduced and, conversely, whether diffusion of oxygen free radicals was enhanced in the salt-sensitive Dahl S rat (SS/Mcwi). Angiotensin II ([Ang II] 1 &mgr;mol/L)–stimulated NO and superoxide (O2·−) production were imaged by fluorescence microscopy in thin tissue strips from the inner stripe of the outer medulla. In prehypertensive SS/Mcwi rats and a genetically designed salt-resistant control strain (consomic SS-13BN), Ang II failed to increase either NO or O2·− in pericytes of isolated vasa recta. Ang II stimulation resulted in production of NO in epithelial cells of the mTAL that diffused to vasa recta pericytes of SS-13BN rats but not in SS/Mcwi rats except when tissues were preincubated with the superoxide scavenger TIRON (1 mmol/L). Ang II resulted in a greater increase of O2·− in the mTAL of SS/Mcwi compared with SS.13BN mTAL. The O2·− diffused to adjoining pericytes in tissue strips only in SS/Mcwi rats but not in control SS-13BN rats. Diffusion of Ang II-stimulated O2·− from mTAL to vasa recta pericytes was absent when tissue strips from SS/Mcwi rats were treated with the NO donor DETA-NONOate (20 &mgr;mol/L). We conclude that the SS/Mcwi rat exhibits increased production of O2·− in mTAL that diffuses to surrounding vasa recta and attenuates NO cross-talk. Diffusion of O2·− from mTAL to surrounding tissue could contribute to reduced bioavailability of NO, reductions of medullary blood flow, and interstitial fibrosis in the outer medulla of SS/Mcwi rats.

Localization of Aldosterone-Producing Adrenocortical Adenomas: Significance of Adrenal Venous Sampling

Accurate localization of aldosterone-producing adenoma (APA) is essential for the treatment of primary aldosteronism (PA). In order to confirm the clinical usefulness of adrenal venous sampling (AVS), we retrospectively studied 87 cases of PA in whom AVS was conducted. We collected right and left adrenal venous effluents simultaneously before and after adrenocorticotropic hormone (ACTH) stimulation for measurements of aldosterone concentration (A) and cortisol concentration (C). Based on AVS results, we judged 66 cases as having unilateral aldosterone hypersecretion and the remaining 21 cases as having no apparent laterality. Of the above 66 subjects, 61 underwent laparoscopic removal of the adrenal gland through a retroperitoneal approach. The presence of APA was histopathologically confirmed, and blood pressure decreased significantly with normalization of plasma aldosterone concentration (PAC) in all cases. The receiver operator characteristics (ROC) curve analysis between the operated and no–apparent-laterality groups revealed that the ratio of A/C on the higher side to A/C on the lower side (A/C ratio) after ACTH stimulation is a useful index, with a cutoff value of 2.6, a sensitivity of 0.98 and a specificity of 1.0. The ROC curve analysis between the APA side and contralateral side within the operated patients revealed that the cutoff value of A was 1,340 ng/dL, with a sensitivity of 0.92 and a specificity of 1.00. Our results indicate the usefulness of simultaneous AVS and ACTH stimulation for localizing APA.

Reactive oxygen species as important determinants of medullary flow, sodium excretion, and hypertension.

The physiological evidence linking the production of superoxide, hydrogen peroxide, and nitric oxide in the renal medullary thick ascending limb of Henle (mTAL) to regulation of medullary blood flow, sodium homeostasis, and long-term control of blood pressure is summarized in this review. Data obtained largely from rats indicate that experimentally induced elevations of either superoxide or hydrogen peroxide in the renal medulla result in reduction of medullary blood flow, enhanced Na(+) reabsorption, and hypertension. A shift in the redox balance between nitric oxide and reactive oxygen species (ROS) is found to occur naturally in the Dahl salt-sensitive (SS) rat model, where selective reduction of ROS production in the renal medulla reduces salt-induced hypertension. Excess medullary production of ROS in SS rats emanates from the medullary thick ascending limbs of Henle [from both the mitochondria and membrane NAD(P)H oxidases] in response to increased delivery and reabsorption of excess sodium and water. There is evidence that ROS and perhaps other mediators such as ATP diffuse from the mTAL to surrounding vasa recta capillaries, resulting in medullary ischemia, which thereby contributes to hypertension.

Methylglyoxal augments intracellular oxidative stress in human aortic endothelial cells

Abstract Methylglyoxal (MGO) is a non-enzymatic metabolite in the glycolytic pathway and its concentration in blood and tissues is elevated in diabetes and renal failure. MGO induces tissue injuries via ROS; however, the mechanism remains to be clarified. The present study examined the harmful actions of MGO. Human aortic endothelial cells were assessed under real-time fluorescent microscopy with continuous superfusion. Increases in intracellular ROS were measured with fluorescent indicator, 5-(and-6)-chloromethyl-2’,7’-dichlorodihydrofluorescein diacetate acetyl ester (DCFH-DA). The addition of MGO rapidly increased the ROS in a dose-dependent manner. The increment of DCF was entirely abolished by pre-treatment with superoxide anion scavenger and membrane-permeable catalase, indicating that MGO induces superoxide production. The increment was completely inhibited by 2-thenoyltrifluoroacetone or carbonyl cyanide 3-chlorophenylhydrazone and partially inhibited by N-methyl-L-arginine. These data suggest that MGO stimulates superoxide production from mitochondria and partially stimulates nitric oxide synthase in human endothelial cells.

Luminal Alkalinization Attenuates Proteinuria-Induced Oxidative Damage in Proximal Tubular Cells

A highly acidic environment surrounds proximal tubular cells as a result of their reabsorption of HCO(3)(-). It is unclear whether this luminal acidity affects proteinuria-induced progression of tubular cell damage. Here, we investigated the contribution of luminal acidity to superoxide (O(2)(·-)) production induced by oleic acid-bound albumin (OA-Alb) in proximal tubular cells. Acidic media significantly enhanced OA-Alb-induced O(2)(·-) production in the HK-2 proximal tubular cell line. Simultaneous treatment with both OA-Alb and acidic media led to phosphorylation of the intracellular pH sensor Pyk2. Highly phosphorylated Pyk2 associated with activation of Rac1, an essential subcomponent of NAD(P)H oxidase. Furthermore, knockdown of Pyk2 with siRNA attenuated the O(2)(·-) production induced by cotreatment with OA-Alb and acid. To assess whether luminal alkalinization abrogates proteinuria-induced tubular damage, we studied a mouse model of protein-overload nephropathy. NaHCO(3) feeding selectively alkalinized the urine and dramatically attenuated the accumulation of O(2)(·-)-induced DNA damage and proximal tubular injury. Overall, these observations suggest that luminal acidity aggravates proteinuria-induced tubular damage and that modulation of this acidic environment may hold potential as a therapeutic target for proteinuric kidney disease.