Keisuke Kohno

Single LDL Apheresis Improves Endothelium-Dependent Vasodilatation in Hypercholesterolemic Humans

BACKGROUND Although long-term lipid-lowering therapy improves endothelium-dependent vasodilatation in humans, it remains unknown whether the short-term removal of LDL per se ameliorates endothelial dysfunction. METHODS AND RESULTS To examine the effects of a single session of LDL apheresis on endothelial function in patients with hypercholesterolemia, we measured forearm blood flow (FBF) by strain-gauge plethysmography before and after single LDL apheresis while infusing acetylcholine (ACh; 4 to 24 micrograms/min) and sodium nitroprusside (SNP; 0.2 to 1.2 micrograms/min). The single session of LDL apheresis reduced total LDL (from 142.2 +/- 15.0 to 32.6 +/- 5.0 mg/mL, P < .0005) and oxidized LDL (from 111.6 +/- 22.8 to 30.0 +/- 5.4 ng/mL, P < .005). Although ACh and SNP increased FBF dose-dependently before and after LDL apheresis, the endothelium-dependent vasodilatation responses to ACh were significantly augmented (P < .01) after the single session of LDL apheresis without changes in the endothelium-independent vasodilatation responses to SNP. The plasma levels of total and oxidized LDL correlated with the degree of ACh-induced vasodilatation. Furthermore, the local production of nitrate/nitrite, metabolites of NO, during ACh infusion was significantly (P < .05) augmented by LDL apheresis, and there was a significant correlation between the degree of ACh-induced vasodilatation and the production in nitrate/nitrite (r = .99, P < .0005). CONCLUSIONS We demonstrated that even a single session of LDL apheresis with the reduction of total LDL and oxidized LDL improved endothelial function. Our results suggest that total LDL and/or oxidized LDL may directly impair endothelial function in the human forearm vessel.

Asymmetrical Dimethylarginine, an Endogenous Nitric Oxide Synthase Inhibitor, in Experimental Hypertension

NG,NG-dimethyl-L-arginine (ADMA) is an endogenously synthesized nitric oxide (NO) synthase inhibitor which has potent pressor/vasoconstrictor effects. Dimethylargininase metabolizes ADMA to L-citrulline and plays a key role in determining the in vivo levels of ADMA. To investigate the role of ADMA in the pathogenesis of hypertension, we measured 24-hour urinary excretion of ADMA (UADMA) and nitrate/nitrite (NOx) in Dahl salt-sensitive hypertensive rats and spontaneously hypertensive rats (SHR). In Dahl salt-resistant rats, high-salt diet (8% NaCl) did not increase blood pressure and increased urinary NOx (P < .01) without changes in UADMA compared with low-salt diet (0.3% NaCl). In contrast, in Dahl salt-sensitive rats, high-salt diet increased blood pressure (P < .01), did not change urinary NOx excretion, and increased UADMA (P < .01). There was a significant (r = .65, P < .01) correlation between UADMA and the level of blood pressure in Dahl salt-sensitive rats. Plasma levels of NOx and ADMA and renal dimethylargininase content were comparable among them. These results may suggest that in Dahl salt-resistant rats, blood pressure is kept constant during high-salt intake, possibly due to the compensatory increased production of NO, and that in Dahl salt-sensitive rats, high-salt intake increases the production of ADMA, attenuates the compensatory increases in NO, and increases blood pressure. These results also suggest that the systemic production of ADMA is not dependent on renal dimethylargininase. SHR had significantly greater urinary NOx excretion (P < .05) and smaller UADMA than Wistar-Kyoto rats (P < .05), and UADMA was inversely correlated with their mean arterial pressure (r =.64, P < .05). In conclusion. ADMA, independently of the renal dimethylargininase content, may play a role in the pathogenesis in Dahl salt-sensitive hypertensive rats but not in SHR.

Chronic l-Arginine Administration Attenuates Cardiac Hypertrophy in Spontaneously Hypertensive Rats

Nitric oxide inhibits proliferation and migration of vascular smooth muscle cells and contractility of cardiomyocytes in vitro. In spontaneously hypertensive rats (SHR), evidence suggests intrinsic abnormalities of the L-arginine-nitric oxide axis, such as low cGMP-dependent protein kinase in the heart and abnormal L-arginine metabolism. To investigate the in vivo effect of L-arginine on cardiac hypertrophy, 30 SHR and 30 Wistar-Kyoto rats (WKY) were randomly grouped to receive L-arginine (7.5 g/L in drinking water) or vehicle for 12 weeks. L-Arginine treatment did not affect body weight or arterial pressure in either strain. In vehicle-treated animals, the heart/body weight ratio was significantly higher in SHR than in WKY (P < .01). L-Arginine treatment decreased the heart/body weight ratio in SHR (P < .05) but did not affect it in WKY. Expression of skeletal alpha-actin mRNA, known to be expressed in the hypertrophied myocardium, was attenuated in L-arginine-treated SHR compared with vehicle-treated SHR. Cardiac cGMP content and nitrate/nitrite content were less in SHR than WKY. L-Arginine treatment increased these levels only in SHR, suggesting enhanced nitric oxide production. Thus, chronic L-arginine administration attenuated cardiac hypertrophy independently of blood pressure and increased myocardial content of cGMP and nitrate/nitrite. Our results suggest that abnormality of the cardiac L-arginine-nitric oxide axis may play an important role in the pathogenesis of cardiac hypertrophy in SHR.

Carbonic-adsorbent AST-120 reduces overload of indoxyl sulfate and the plasma level of TGF-β1 in patients with chronic renal failure

BackgroundWe previously reported a significant increase in plasma TGF-β1 in patients with chronic renal failure (CRF). Progression of CRF may be caused by persistent renal production of TGF-β1. In CRF rat models, an oral carbonic absorbent (AST-120) reduces the expression of the TGF-β1 gene in the kidney, and delays the progression of CRF, in part by alleviating the overload of indoxyl sulfate. The aim of this study was to evaluate the effect of AST-120 on plasma levels of indoxyl sulfate and TGF-β1 in CRF patients.MethodsTen CRF patients (aged 59.3 ± 9.5 years, 5 men, serum creatinine 4.37 ± 1.72 mg/dl) were enrolled in this study. All patients maintained a regular dietary therapy and the same medication throughout the study. AST-120 was added at a dose of 6 g/day. Parameters including the slope of the reciprocal of the serum creatinine – time plot, plasma indoxyl sulfate level, and plasma and urinary levels of TGF-β1 were compared before and after the treatment with AST-120. The mean observation periods before and after the treatment were 9.7 ± 2.8 and 6.5 ± 2.9 months, respectively.ResultsAdministration of AST-120 significantly reduced the plasma levels of indoxyl sulfate (1.42 ± 1.50 vs. 1.26 ± 1.40 mg/dl, P < 0.05) and TGF-β1 (17.9 ± 7.2 vs. 10.6 ± 4.7 ng/ml, P < 0.05) and improved the slope of the reciprocal of serum creatinine (−0.061 ± 0.041 vs. −0.032 ± 0.055 dl/mg/year, P < 0.05).ConclusionsThese results support the notion that indoxyl sulfate and TGF-β1 may be involved in the progression of CRF, and that the oral adsorbent AST-120 may suppress the progression, at least in part, by reducing overproduction of TGF-β1.

Erythropoietin impairs endothelium-dependent vasorelaxation through cyclooxygenase-dependent mechanisms in humans*

Hypertension is a major complication of recombinant human erythropoietin therapy in patients with end-stage renal disease. Although the mechanisms for this pressor effect remain unknown, in vitro and ex vivo experiments suggest that erythropoietin stimulates release of cyclooxygenase-dependent endothelium-derived contracting factors (EDCFs) and attenuates endothelium-dependent vasorelaxation. To investigate the effect of erythropoietin on human endothelial function, we measured forearm blood flow by strain gauge plethysmography before and after intraarterial incremental administration of erythropoietin (10 to 300 IU/min, total 3000 U), while infusing acetylcholine (4 to 24 microg/min) and sodium nitroprusside (0.2 to 1.2 microg/min) in healthy male volunteers (n = 12). To examine a possible role of EDCFs, we repeated the same protocol under the pretreatment with oral indomethacin (50 mg), a cyclooxygenase inhibitor (n = 8). Infusion of erythropoietin into the brachial artery increased the erythropoietin blood concentration of venous effluents significantly (P < .0001) without changes in blood pressure and basal forearm blood flow. Acetylcholine and sodium nitroprusside increased forearm blood flow dose dependently before and after erythropoietin administration. However, acetylcholine-induced vasodilation (endothelium dependent) was significantly attenuated (P < .001) after erythropoietin administration, whereas vasodilation to sodium nitroprusside (endothelium independent) was unchanged. After indomethacin pretreatment, erythropoietin no longer attenuated endothelium-dependent vasorelaxation. Our results indicate that erythropoietin may impair endothelial function acutely, probably through cyclooxygenase-dependent EDCFs in humans. Long-term effects of erythropoietin on endothelial function in uremic patients remain to be elucidated.

Renal depressor mechanisms of physical training in patients with essential hypertension

To clarify characteristics of the patients in whom exercise training lowers blood pressure and to elucidate the mechanisms by which exercise training lowers blood pressure, we evaluated 24-h blood pressure, glomerular filtration rate (GFR), renal blood flow (RBF), filtration fraction (FF), plasma renin activity (PRA), plasma aldosterone concentration (PAC), plasma norepinephrine concentration (PNE), and incremental area of insulin/glucose (sigmaI/sigmaG) during 75 g oral glucose tolerance test, and assessed arterial baroreceptor function (BSI) before and after a 3-week exercise training program (four 6-min sessions daily at 75% VO2 max). Patients were classified as responders (n = 15) if they showed statistically significant reduction in the multiple comparison of 24-h mean arterial pressure (MAP), or as nonresponders (n = 15) if they did not. Although there were no significant differences between responders and nonresponders in age, weight, MAP, GFR, RBF, RPF, FF, PNE, sigmaI/sigmaG, or BSI before exercise, renal vascular resistance (RVR; P < .05), PRA (P < .05), and PAC (P < .05) were significantly higher in responders than in nonresponders. The fractional excretion of sodium (FENa) (P < .05) were significantly lower in responders than in nonresponders. After exercise training, FF (P < .01), RVR (P < .05), PNE (P < .05) PRA (P < .01), and sigmaI/sigmaG (P < .05) decreased significantly only in responders. The decrease in MAP significantly correlated with the reductions in FF (r = 0.46, P < .05), PNE (r = 0.52, P < .01) and RVR (r = 0.40, P < .05). Thus, in patients who have higher RVR and PRA, exercise training lowered blood pressure in parallel to a reduction in RVR associated with decreases in sympathetic tone and improvement of insulin resistance. Our results suggest that exercise-induced changes in renal hemodynamics may contribute to the reduction in blood pressure in these patients.

Renal Denervation Prevents Intraglomerular Platelet Aggregation and Glomerular Injury Induced by Chronic Inhibition of Nitric Oxide Synthesis

Nitric oxide (NO) inhibits platelet adhesion and aggregation in vitro. In vivo, chronic inhibition of NO synthesis induces nephrosclerosis and hypertension. Although the pathophysiological mechanism of this glomerular injury has not been clarified, sympathetic nerve activation, a potent procoagulant stimulus elicited by NO inhibition, may play a role. To investigate the role of renal sympathetic nerves in the development of renal injury induced by NG-nitro-L-arginine methyl ester (L-NAME), a specific NO synthesis inhibitor, we examined renal histological changes in four groups of Sprague-Dawley rats: (1) sham operated, vehicle treated; (2) sham operated, L-NAME treated; (3) denervated, vehicle treated, and (4) denervated, L-NAME treated. Following renal denervation or sham operation, L-NAME was administered orally for 4 weeks. Chronic NO inhibition induced platelet aggregation and erythrocyte stasis in the glomerular capillary lumen accompanied by electron-microscopic glomerular injury. Renal denervation abrogated platelet aggregation and glomerular injury in L-NAME-treated animals. Thus, chronic NO synthesis inhibition induced intraglomerular platelet aggregation and glomerular injury, which was attenuated by renal nerve denervation. These results suggest that intrinsic NO may have an antithrombotic effect in the glomeruli and may play a protective role in the progression of glomerular injury possibly mediated by renal sympathetic nerves.

Effects of switching from calcium carbonate to lanthanum carbonate on bone mineral metabolism in hemodialysis patients.

Phosphate binders are useful for the treatment of hyperphosphatemia in hemodialysis (HD) patients. This study was performed to examine the effects of switching from calcium carbonate (CC) to lanthanum carbonate (LC) on bone mineral metabolism and inflammatory markers in HD patients. We conducted 29 stable HD patients receiving CC, which was replaced by LC and followed‐up for 12 weeks. Patients underwent determinants of blood chemistries such as serum calcium (Ca), phosphorus, parathyroid hormone (PTH) and vitamin D status, and interleukin‐6 (IL‐6) mRNA levels in whole blood cells were evaluated by real‐time PCR just before and after the treatment with LC. Corrected levels were significantly reduced, but serum phosphorus levels (P levels) were unchanged after LC treatment. Switching to LC increased whole‐PTH, osteocalcin, 1,25(OH)2D3 levels and 1,25(OH)2D3/25(OH)D3 ratio. 1,25(OH)2D3/25(OH)D3 ratio was negatively correlated with HD duration. Furthermore, whole blood cell IL‐6 mRNA levels were significantly reduced by LC treatment. We provided that the switching from CC to LC improved Ca overload and ameliorated vitamin D and inflammatory status in HD patients. These observations suggest that LC may play a protective role for the progression of atherosclerosis and vascular calcification in these patients.