Hidehiko Ono

Relations of plasma high-sensitivity C-reactive protein to traditional cardiovascular risk factors

Variations of circulating C-reactive protein (CRP) levels are supposed to reflect chronic inflammatory process of the cardiovascular system. In particular, it has been reported that high-sensitivity CRP (hsCRP) is a promising marker of coronary heart disease. In the present study, we assessed the relationship between hsCRP and classic cardiovascular risk factors, such as age, blood pressure, smoking habit and serum lipids. Plasma hsCRP was measured by ELISA in 908 subjects, aged 30-79 years, who entered our health-check program. Plasma hsCRP level was 0.54+/-0.02 mg/l in 566 subjects without any disease currently treated. The level was significantly higher in patients treated for hypertension (0.74+/-0.06 mg/l, P=0.002), diabetes mellitus (0.77+/-0.09 mg/l, P=0.016) or coronary artery disease (0.99+/-0.16 mg/l, P=0.008) than in subjects without diseases. In a simple regression analyses of the 566 subjects without diseases, plasma hsCRP positively correlated with male gender, smoking, body mass index, systolic blood pressure, white blood cell count, blood hemoglobin, fasting blood glucose, serum gamma-GTP, uric acid and triglycerides, and inversely correlated with serum albumin and HDL-cholesterol. In multiple regression analysis, white blood cell count (r=0.276, P<0.001), body mass index (r=0.246, P<0.001), age (r=0.122, P=0.001) and smoking (r=0.112, P=0.009) showed independent correlations with plasma hsCRP. It is suggested that variation of circulating hsCRP, even within normal range, is involved in the interrelation of cardiovascular risk factors, such as age, smoking, obesity, high blood pressure and dyslipidemia, which are supposed to promote atherosclerosis and ultimately provoke cardiovascular diseases, such as coronary artery disease.

Receptor-Mediated Intrarenal Angiotensin II Augmentation in Angiotensin II–Infused Rats

Chronic low-dose angiotensin II (Ang II) infusion for 13 days mimics two-kidney, one clip Goldblatt hypertension and increase intrarenal Ang II levels. We performed studies to determine the time course for the enhancement of intrarenal Ang II levels and whether the increased intrarenal Ang II is a tissue-specific event and requires a receptor-mediated step. Male Sprague-Dawley rats were uninephrectomized, and either vehicle or Ang II (40 ng/min) was infused via a subcutaneous osmotic minipump. Plasma and renal Ang II levels were measured 3, 7, 10, and 13 days after minipump implantation. Compared with controls (126 +/- 2 mm Hg), systolic pressure in Ang II-infused rats exhibited a detectable increase by day 6 (146 +/- 2 mm Hg) and continued to increase to 189 +/- 5 mm Hg by day 12. Plasma Ang II levels were elevated by day 3, whereas intrarenal Ang II levels were not significantly elevated until 10 days of Ang II infusion. Renal injury characterized by focal and segmental glomerulosclerosis was evident after 13 days of Ang II infusion. Losartan (30 mg/kg per day) prevented the development of hypertension in the Ang II-infused rats for the duration of the infusion period (125 +/- 1 mm Hg) and reduced the degree of glomerular injury. Plasma renin activity was suppressed in the Ang II-infused group but was elevated markedly in both losartan-treated groups. Plasma Ang II levels were elevated in the Ang II-infused rats and were even higher during losartan treatment. Intrarenal Ang II levels were enhanced significantly (354 +/- 60 versus 164 +/- 23 fmol/g) in the Ang II-infused rats. However, losartan treatment prevented the augmentation of intrarenal Ang II caused by Ang II infusion. Heart and adrenal Ang II levels were not significantly increased in the Ang II-infused rats but were significantly elevated during losartan treatment. These results suggest that the tissue-specific elevations of intrarenal Ang II levels caused by chronic Ang II infusion are mediated by angiotensin type 1 receptor activation, which leads to either receptor-mediated internalization of Ang II, enhancement of intrarenal Ang II formation, or both.

Nitric Oxide Synthase Inhibition in Spontaneously Hypertensive Rats: Systemic, Renal, and Glomerular Hemodynamics

To investigate the prolonged effects of nitric oxide inhibition on systemic, renal, and glomerular hemodynamics, the effects of the nitric oxide synthase inhibitor N omega-nitro-L-arginine methyl ester (L-NAME) on cardiac index, renal micropuncture results, urinary excretion, and histology were obtained in 20-week-old male spontaneously hypertensive rats (SHR) that were divided into two groups: untreated and L-NAME-treated (50 mg/L), each followed for 3 weeks. Cardiac index and effective renal plasma flow decreased (P < .01) in L-NAME-treated SHR, exhibiting a positive correlation (r = .816; P < .0001). Single-nephron plasma flow (123 +/- 8 versus 80 +/- 12 nL/min per gram; P < .01) and ultrafiltration coefficient (P < .05) were also reduced in L-NAME-treated SHR versus controls. Most notably, the L-NAME-treated SHR had increased afferent (4.4 +/- 0.3 versus 9.5 +/- 1.3 U; P < .01) and efferent (1.4 +/- 0.1 versus 2.7 +/- 0.3 U; P < .01) glomerular arteriolar resistances versus controls. These functional changes were associated with significantly altered afferent arteriolar (P < .001) and glomerular (P < .005) histological injury scores accompanied by marked proteinuria (P < .001). Because of the intense afferent glomerular artery constriction and lesser increase in efferent glomerular arteriolar resistance associated with reduced single-nephron plasma flow, glomerular capillary pressure did not increase in the L-NAME-treated SHR.(ABSTRACT TRUNCATED AT 250 WORDS)

Fasudil, a Rho-kinase inhibitor, attenuates glomerulosclerosis in Dahl salt-sensitive rats.

Objective The present study was designed to clarify whether the Rho–Rho-kinase pathway is involved in the process of hypertensive glomerulosclerosis and to assess the therapeutic effect of fasudil, a specific Rho-kinase inhibitor. Method and results Dahl salt-sensitive rats (DS) and Dahl salt-resistant rats (DR) were fed a high-salt diet at 6 weeks of age. Fasudil (30 mg/kg per day) was administered for 7 weeks to DS starting at the age of 11 weeks. After 7 weeks, untreated DS were characterized by decreased kidney function, increased proteinuria, abnormal morphological findings, increased adrenomedullin and atrial natriuretic peptide (ANP) levels, and increased renal messenger RNA expression of RhoB, Rho-kinaseα, Rho-kinaseβ, collagen I and collagen III, and transforming growth factor-beta (TGF-β) in the renal cortex compared with DR. Chronic fasudil treatment significantly improved renal function (serum creatinine, –26%; blood urea nitrogen, –41%; creatinine clearance, +42%), proteinuria (–24%) and histological findings (glomerular injury score, –49%; afferent arteriolar injury score, –17%) without changing blood pressure compared with untreated DS. Interestingly, long-term fasudil treatment decreased the plasma adrenomedullin (–25%) and ANP (–49%), but did not change the plasma renin or aldosterone. Furthermore, fasudil significantly decreased the messenger RNA expression of TGF-β (–20%), collagen I (–23%), and collagen III (–24%) in the renal cortex. However, there were still significant differences in the aforementioned parameters between DR and fasudil-treated DS. Conclusion These results suggest that the Rho–Rho-kinase pathway may be partly responsible for the pathogenesis of hypertensive glomerulosclerosis independently of blood pressure in DS, and that chronic inhibition of the Rho–Rho-kinase pathway may be a new strategy for treating hypertensive nephrosclerosis.

Glomerular Dynamics and Morphology of Aged Spontaneously Hypertensive Rats: Effects of Angiotensin-Converting Enzyme Inhibition

Relationships between glomerular dynamics and renal injury, micropuncture and histological studies were assessed in 73 week-old normotensive Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats divided into untreated and angiotension-converting enzyme inhibitor-treated (quinapril; 3 mg/kg/day; for 3 weeks) groups. Urinary protein excretion (UPE) and histologic arteriolar (AIS) and glomerular (GIS) injury scores were determined. Mean arterial pressure (MAP) of untreated SHR was increased compared with WKY (200 +/- 6 vs 119 +/- 4 mm Hg; P < 0.01), effective renal plasma flow (ERPF) was reduced (1.47 +/- 0.21 vs 3.06 +/- 0.26 ml/min/per g; P > 0.01), and filtration fraction (FF) and total renal vascular resistance (RVR) of SHR were increased (P < 0.01). Single-nephron plasma flow (SNPF) of untreated SHR was decreased (174 +/- 17 vs 80 +/- 9 ml/min; P < 0.01), and single-nephron filtration fraction and afferent arteriolar resistance (RA) were increased (19.4 +/- 1.8 vs 30.0 +/- 2.5% and 1.90 +/- 0.25 vs 9.05 +/- 1.35 U, respectively; both P < 0.01). Despite reduced SNPF, glomerular capillary pressure (PG) increased (49.7 +/- 0.7 vs 53.8 +/- 1.3 mm Hg; P < 0.05), the result of efferent arteriolar constriction (1.15 +/- 0.18 vs 2.84 +/- 0.36 U; P < 0.01). Untreated SHR had higher UPE (13.9 +/- 1.5 vs 42.8 +/- 3.2; mg/100 g per day; P < 0.01) and GIS and AIS scores than WKY (4.3 +/- 1.1 vs 64.3 +/- 8.4 and 16.6 +/- 3.1 vs 96.3 +/- 14.4; both P < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

ACE Inhibition Prevents and Reverses L-NAMEExacerbated Nephrosclerosis in Spontaneously Hypertensive Rats

Chronic nitric oxide inhibition exacerbates hypertension and nephrosclerosis in spontaneously hypertensive rats (SHRs). In this study, we determined whether angiotensin-converting enzyme (ACE) inhibition could prevent or reverse the systemic, renal, and glomerular hemodynamic alterations and the pathological changes of nephrosclerosis. Four groups of 20-week-old SHRs were studied: group 1, untreated controls; group 2, treated with N omega-nitro-L-arginine methyl ester (L-NAME, 50 mg/L for 3 weeks); group 3, L-NAME cotreated with quinapril (3 mg.kg-1.d-1 for 3 weeks); and group 4, L-NAME for 3 weeks followed by quinapril for 3 weeks (same doses). The results of this study demonstrated that both cotreatment (group 3) and posttreatment (group 4) with quinapril reduced mean arterial pressure (186 +/- 9 and 192 +/- 9 mm Hg, respectively, compared with group 2 SHRs, 221 +/- 5 mm Hg) and total peripheral resistance index associated with significant reductions in afferent and efferent arteriolar resistances; nephrosclerosis pathological scores; and urinary protein excretion (all at least P < .01). ACE inhibition also significantly increased stroke index, single-nephron glomerular filtration rate, and ultrafiltration coefficient compared with the L-NAME SHRs. Most notable were the findings that cotreatment with quinapril completely prevented the renal glomerular hemodynamic alterations with reduced glomerular capillary hydrostatic pressure and efferent arteriolar resistance compared with both the untreated and the L-NAME-treated SHRs (all at least P < .01). Posttreatment with quinapril also reversed the glomerular injury (subcapsular, -83%; juxtamedullary, -56%) and arteriolar (-87%) injury scores obtained from renal biopsy specimens (P < .005 and P < .0001, respectively). These changes were associated with decreased periarteriolar fibronectin and increased afferent arteriolar alpha-smooth muscle actin deposition (immunohistochemistry). These data, therefore, demonstrate that ACE inhibition not only prevents but also reverses L-NAME-exacerbated severe nephrosclerosis in SHRs, as indicated by improved systemic, renal, and glomerular hemodynamic changes, proteinuria, and histological alterations.

N- and L-type calcium channel antagonist improves glomerular dynamics, reverses severe nephrosclerosis, and inhibits apoptosis and proliferation in an L-NAME/SHR model

Objective To determine the responses of the new dihydropyridine N- and L-type calcium antagonist, cilnidipine, on systemic and renal hemodynamics, glomerular dynamics, renal function, and histopathology in an Nω-nitro-l-arginine methylester spontaneously hypertensive rat (l-NAME/SHR) model of nephrosclerosis. Methods Five groups of 20-week-old male SHR were studied using renal micropuncture techniques and histopathological analyses: group 1, control; group 2, cilnidipine (10 mg/kg per day) by gavage, for 3 weeks; group 3, l-NAME (50 mg/l) in drinking water, for 3 weeks; group 4, combination of l-NAME and cilnidipine, for 3 weeks; group 5, l-NAME for 3 weeks, followed by cilnidipine for a subsequent 3 weeks. Results Cilnidipine significantly reduced mean arterial pressure, total peripheral resistance and renal vascular resistance, while increasing effective renal blood flow and glomerular filtration rate (P < 0.01) in l-NAME/SHR. These hemodynamic changes were associated with significantly increased single nephron glomerular filtration rate (SNGFR) and plasma flow (SNPF) and decreased afferent glomerular arteriolar resistances when cilnidipine was used alone, and with increased SNGFR and SNPF, but decreased glomerular capillary pressure, afferent and efferent arteriolar resistances, urinary protein excretion, serum creatinine and uric acid concentrations (at least P < 0.05) in l-NAME-exacerbated SHR nephrosclerosis. In addition, glomerular and arteriolar injuries were markedly reversed (both P < 0.01), and glomerular apoptosis and cellular proliferation were inhibited and associated with glomerular tuft enlargement and an increase in cell number. Conclusion Cilnidipine not only prevented, but reversed, the severe renal hemodynamic and glomerular dynamic changes, including apoptosis and glomerular cellular proliferation, in l-NAME/SHR-exacerbated nephrosclerosis. This dual-channel calcium antagonist thus exerted renoprotective pathophysiological effects in the l-NAME/SHR.

Long-Term Administration of Rho-Kinase Inhibitor Ameliorates Renal Damage in Malignant Hypertensive Rats

We have shown recently that fasudil, a Rho-kinase inhibitor, has renoprotective effects in salt-sensitive hypertensive rats. We hypothesized that activation of Rho-kinase is involved in the pathogenesis of glomerulosclerosis in malignant hypertensive rats. To test this hypothesis, we studied the following 4 groups: control Wistar–Kyoto rats, untreated deoxycorticosterone-acetate salt spontaneously hypertensive rats (DOCA-SHR), low-dose fasudil-treated DOCA-SHR, and high-dose fasudil-treated DOCA-SHR. After 3 weeks of treatment, the effects of fasudil were examined. DOCA-SHR was characterized by increased blood pressure (BP); increased kidney weight; decreased renal function; increased proteinuria; abnormal histological findings; increased monocyte/macrophage infiltration; increased urinary 8-isoprostran levels; increased gene expression of collagen I, collagen III, transforming growth factor-β, and reduced nicotinamide-adenine dinucleotide phosphate oxidase subunits (p40phox, p47phox, and p67phox); and decreased gene expression of endothelial NO synthase (eNOS) in the renal cortex as compared with Wistar–Kyoto rats. Long-term high-dose fasudil treatment significantly improved renal function and histological findings without changing BP, as compared with untreated DOCA-SHR. Interestingly, long-term fasudil treatment significantly decreased monocyte/macrophage infiltration and urinary 8-isoprostran excretion, in association with decreased mRNA levels of transforming growth factor-β, collagen I, collagen III, and NADPH oxidase subunits (p40phox, p47phox, and p67phox), and increased mRNA levels of eNOS in the renal cortex. Long-term low-dose fasudil treatment tended to improve these variables slightly but did not affect most of them significantly. Our results suggest that long-term fasudil treatment provides renoprotective effects independent of BP-lowering activity. These renoprotective effects are associated with inhibition of extracellular matrix gene expression, monocyte/macrophage infiltration, oxidative stress, and upregulation of eNOS gene expression.

Renoprotective Effects of Felodipine and/or Enalapril in Spontaneously Hypertensive Rats With and Without L-NAME

To determine the renoprotective effects of a calcium antagonist (felodipine) and an angiotensin-converting enzyme (ACE) inhibitor (enalapril), alone or in combination, 10 groups of 19-week-old spontaneously hypertensive rats (SHR) (with or without N(G)-nitro-L-arginine methyl ester [L-NAME]) were studied using renal micropuncture techniques. Group 1 (control), group 2 (felodipine, 30 mg x kg(-1) x d[-1]), group 3 (enalapril, 30 mg x kg(-1) x d[-1]), and group 4 (felodipine plus enalapril, 15 mg x kg(-1) x d(-1) each agent) were studied after 3 weeks of treatment without L-NAME. L-NAME (50 mg/L) cotreatment was administered in drinking water to groups 6 through 10 using the same doses of each agent as in groups 1 through 4: group 5 (only L-NAME), group 6 (felodipine), group 7 (enalapril), and group 8 (felodipine plus enalapril). Groups 9 and 10 received L-NAME initially for 3 weeks followed by felodipine or felodipine plus enalapril, respectively, for the subsequent 3 weeks. All three treatments resulted in reductions in mean arterial pressure and total peripheral vascular resistance (P<.001) that were associated with important structural and functional renal microcirculatory improvements. Thus, the pathological nephrosclerosis (subcapsular and juxtamedullary) glomerular and arteriolar injury scores were improved (P<.05 at least) in association with normalization of afferent and efferent arteriolar resistances, and single-nephron glomerular filtration rate, plasma flow, and blood flow were significantly improved, as well as the ultrafiltration coefficient (compared with group 5, L-NAME). Thus, the calcium antagonist felodipine, alone or in combination with an ACE inhibitor, not only prevented but also reversed L-NAME-exacerbated hypertensive nephrosclerosis in SHR.

Hydrochlorothiazide exacerbates nitric oxide-blockade nephrosclerosis with glomerular hypertension in spontaneously hypertensive rats.

OBJECTIVE To determine whether a diuretic can also reverse the clinical, systemic, renal and glomerular haemodynamic and pathological changes caused by nephrosclerosis. METHODS Three groups of 20-week-old spontaneously hypertensive rats (SHR) were investigated: control male SHR; a similar group, administered 50 mg/l NG-nitro-L-arginine methyl ester (L-NAME) for 3 weeks; and SHR treated similarly with L-NAME but also with 80 mg/kg per day hydrochlorothiazide (HCTZ) by gavage for 3 weeks. RESULTS The mean arterial pressure, cardiac output, effective renal plasma flow and glomerular filtration rate decreased as urinary volume increased in the SHR treated with HCTZ and L-NAME. A micropuncture study demonstrated increased glomerular capillary pressure (PG, 56 +/- 1 versus 68 +/- 3 mmHg) associated with increased efferent (2.1 +/- 0.2 versus 2.9 +/- 0.3 u) but no change in afferent arteriolar resistances compared with the SHR group treated with L-NAME only. In addition, HCTZ administration increased the juxtamedullary glomerular injury score (47 +/- 13 versus 114 +/- 29) associated with elevated urinary protein excretion (35 +/- 1 versus 53 +/- 13 mg/100 g body weight per 24 h) The afferent arteriolar injury score was not changed. The PG elevation was related not only to severe glomerulosclerosis but also to increased fibronectin and alpha-smooth muscle actin deposition. CONCLUSION HCTZ administration exacerbated the changes in renal and micropuncture dynamics, proteinuria and histopathological nephrosclerosis produced by L-NAME in SHR.