Teruo Higashi

Benidipine stimulates nitric oxide synthase and improves coronary circulation in hypertensive rats.

We evaluated the effects of long-term treatment with benidipine, a long-acting calcium antagonist, on endothelial cell-type nitric oxide synthase (eNOS) activity and eNOS mRNA expression in the left ventricle (LV) and its relation to coronary flow reserve, and microvascular remodeling in renovascular hypertensive rats (RHR: 2K-1C Goldblatt). Benidipine (5 mg/kg/day) was given to RHR (B-RHR, n = 11) for 6 weeks. Vehicle-treated RHR (U-RHR, n = 11) and age-matched sham-operated rats (ShC, n = 11) served as control group. Coronary flow reserve was measured in conscious rats using colored microspheres. Fifty-micrometer slices of the LV were incubated with L-arginine to measure nitrite production using the Griess method and eNOS mRNA expression was determined by reverse transcription-polymerase chain reaction. An increased blood pressure in RHR was significantly decreased by benidipine. Nitrite production and eNOS mRNA expression in the LV of U-RHR was significantly lower than that of ShC. This suppression of nitrite production and eNOS mRNA expression was significantly reversed in B-RHR. U-RHR demonstrated a significant decrease in coronary flow reserve and capillary density, and a significant increase in wall-to-lumen ratio, perivascular fibrosis, myocardial fibrosis, and myocyte cross-sectional area. These impaired factors were improved significantly by benidipine. These findings suggest that benidipine therapy may increase nitrite production and eNOS mRNA expression not only by lessening the endothelial damage by the reduction of blood pressure levels, but also by the stimulation of NOS activity and eNOS mRNA, and this increased NOS activity and eNOS mRNA expression may play a role in the amelioration of coronary flow reserve and microvascular remodeling.

Effects of imidapril on NOS expression and myocardial remodelling in failing heart of Dahl salt-sensitive hypertensive rats

OBJECTIVES To elucidate the relationship between renin-angiotensin system and nitric oxide in hypertensive heart failure, we evaluated the effects of long-term treatment with imidapril, angiotensin-converting enzyme inhibitor, on endothelial-cell nitric oxide synthase (eNOS) and inducible NOS (iNOS) expression in the left ventricle (LV) and its relation to myocardial remodelling in failing heart of Dahl salt-sensitive hypertensive rats (DS) fed a high-salt diet. METHODS In DS rats fed an 8% NaCl diet after the age of 6 weeks, a stage of concentric left ventricular hypertrophy at 11 weeks (DSLVH) was followed by a distinct stage of fatal left ventricular failure with chamber dilatation at 18 weeks (DSCHF). Imidapril (DSCHF-I, n = 7, 1 mg/kg/day, subdepressor dose) or vehicle (DSCHF-V, n = 7) were given from DSLVH to DSCHF stage for 7 weeks, and age-matched (18 weeks) Dahl salt-resistant rats fed the same diet were served as control group (DR-C, n = 7). RESULTS Markedly increased left ventricular end-diastolic diameter and reduced fractional shortening in DSCHF-V was significantly ameliorated in DSCHF-I using transthoracic echocardiography. The level of eNOS mRNA and protein in the LV was significantly suppressed in DSCHF-V compared with DR-C, and significantly increased in DSCHF-I compared with DR-C and DSCHF-V. The iNOS mRNA and protein and the fibrosis factor expression of type I collagen mRNA were significantly increased in DSCHF-V compared with DR-C, and significantly decreased in DSCHF-I compared with DSCHF-V. DSCHF-V demonstrated a significant increase in wall-to-lumen ratio, perivascular fibrosis, and myocardial fibrosis. These changes in the microvasculature were improved significantly by imidapril. CONCLUSIONS Subdepressor dose of imidapril may ameliorate the endothelial damage not only by inhibiting production of angiotensin II but also by promoting eNOS and inhibiting iNOS mRNA and protein expression in the LV, and this increased eNOS mRNA and protein level may have a role in the improvement of congestive heart failure and myocardial remodelling.

Nicorandil enhances cardiac endothelial nitric oxide synthase expression via activation of adenosine triphosphate-sensitive K channel in rat

Summary: In the heart, nitric oxide activates an adenosine triphosphate (ATP)‐sensitive K (KATP) channel that is constructed of two subunits, i.e., an ATP‐binding cassette protein sulfonylurea receptor (SUR2) and a pore‐forming inward rectifier (Kir6.1 or 6.2). However, whether this KATP channel affects nitric oxide activation is unknown. Our aim was to assess whether pharmacologic activation of the KATP channel by nicorandil contributes to endothelial nitric oxide synthase (eNOS) levels. A total of 21 7‐week old male Sprague‐Dawley rats were used. Seven were treated by intraperitoneal injection of nicorandil at 3 mg/kg/d; seven were treated with intraperitoneal nicorandil at 3 mg/kg/d after glibenclamide at 12 mg/kg/d twice a day p.o.; and seven were left untreated (controls). At 24 h after treatment, blood pressure and heart rate were measured, and eNOS, SUR2, Kir6.1, and Kir6.2 mRNA levels and eNOS protein levels in the left ventricle were determined by reverse transcription polymerase chain reaction (RT‐PCR) and Western blot analysis. Nicorandil caused tachycardia without a change in blood pressure, whereas glibenclamide had no effect on the nicorandil‐induced change in heart rate or on blood pressure. RT‐PCR revealed that nicorandil increased the eNOS and SUR2 mRNA levels by 2.2‐ and 2.0‐fold, respectively, (p < 0.01 versus control), and that these increases were completely inhibited by glibenclamide. A significant correlation was observed between eNOS and SUR2 mRNA levels in all experimental rats (r = 0.760, p < 0.001). However, Kir6.1 or 6.2 mRNA level was constant. Western blot analysis revealed that nicorandil caused a 1.6‐fold increase in eNOS protein levels (p < 0.01 versus control). This increase was completely inhibited by glibenclamide. In conclusion, upregulation of eNOS mRNA and protein levels by nicorandil, and inhibition of this upregulation by glibenclamide, were demonstrated in normotensive conscious rat hearts. Nicorandil appears to enhance cardiac eNOS expression via activation of a KATP channel.

Effects of angiotensin II type 1 receptor antagonist on nitric oxide synthase expression and myocardial remodeling in Goldblatt hypertensive rats.

We evaluated the effects of long-term treatment with TCV-116, an angiotensin II type 1 receptor antagonist, on endothelial-cell nitric oxide synthase (eNOS) messenger RNA (mRNA) and protein expression in the left ventricle and its relation to myocardial remodeling in Goldblatt hypertensive rats. Two-kidney, one-clip Goldblatt hypertensive rats (RHR) were assigned either to a TCV-116 treatment group (RHR-TCV, n = 8, 3 mg/kg/day, subdepressor dose) or to a group without treatment (RHR-V, n = 7) after their kidneys had been clipped for 4 weeks. TCV-116 was administered to rats in the treatment group for 6 weeks, and age-matched sham-operated rats (ShC, n = 7) served as a control group. Blood pressure in RHR-V and RHR-TCV was similar and significantly higher than that in ShC. The eNOS mRNA and protein levels and NOS activity in the left ventricle was significantly decreased in RHR-V compared with ShC, and significantly increased in RHR-TCV compared with ShC and RHR-V. RHR-V demonstrated a significant increase in fibrosis factor (type I collagen) mRNA expression, perivascular fibrosis, and myocardial fibrosis. These parameters in the microvasculature were improved significantly by TCV-116. Subdepressor dose of TCV- 116 improved pathological myocardial changes in RHR, which may be due in part to an increased eNOS mRNA and protein expression and NOS activity in the left ventricle.

Effect of imidapril on myocardial remodeling in l-NAME–induced hypertensive rats is associated with gene expression of NOS and ACE mRNA☆

Chronically administered N(omega)nitro-L-arginine methyl ester (L-NAME) produces vascular structural changes and fibrosis of the left ventricle (LV). However, very few studies have evaluated whether the beneficial effects of angiotensin-converting enzyme (ACE) inhibitors on these myocardial remodelings are associated with local gene expression of nitric oxide synthase (NOS) and ACE mRNA in the LV. Effects of long term treatment with imidapril, an ACE inhibitor, on gene expression of endothelial-cell NOS (eNOS) and ACE mRNA in the LV and its relation to myocardial remodeling in L-NAME-induced hypertensive rats were evaluated. Fifteen male Sprague-Dawley rats were given L-NAME (60 mg/ kg/day) in drinking water for 6 weeks to induce hypertension, and then treated with imidapril (L-NAME-I, n = 8, 1 mg/kg/day, subdepressor dose), or a vehicle (L-NAME-V, n = 7) for 4 weeks. Age-matched rats (C, n = 7) served as a control group. Blood pressure in L-NAME-V and L-NAME-I was similar and significantly higher than that in C. The level of eNOS mRNA in the LV was significantly decreased in L-NAME-V compared with C, and was significantly increased in L-NAME-I compared with C and L-NAME-V. The ACE mRNA and type I collagen mRNA expression levels were significantly increased in L-NAME-V compared with C, and significantly suppressed in L-NAME-I compared with L-NAME-V. L-NAME-V demonstrated a significant increase in wall-to-lumen ratio, perivascular fibrosis, and myocardial fibrosis. These changes in the microvasculature were improved significantly by imidapril. Myocardial remodeling in L-NAME-induced hypertensive rats was significantly ameliorated by a subdepressor dose of imidapril, which may be due to an increase in local eNOS mRNA expression and a decrease in angiotensin II in the LV.

Effects of imidapril on endothelin-1 and ACE gene expression in failing hearts of salt-sensitive hypertensive rats.

The renin-angiotensin system and endothelin are important regulators of the cardiovascular system. Although increased production of endothelin-1 (ET-1) is reported in patients with heart failure, the detailed mechanism remains to be determined. To elucidate the relationship between the renin-angiotensin system and ET-1 in hypertensive heart failure, we evaluated the effects of long-term treatment with imidapril, an angiotensin converting enzyme (ACE) inhibitor, on preproET-1, endothelin A receptor (ETAR), and ACE mRNA expression in the left ventricle and evaluated these in relation to myocardial remodeling in the failing heart of Dahl salt-sensitive (DS) hypertensive rats fed a high salt diet. In DS rats fed an 8% NaCl diet after the age of 6 weeks, a stage of concentric left ventricular hypertrophy at 11 weeks (DSLVH) was followed by a distinct stage of left ventricular failure with chamber dilatation at 18 weeks (DSHF). Imidapril (DSHF-IM, n = 8, 1 mg/kg/day, subdepressor dose) or vehicle (DSHF-V, n = 8) was given from stage DSLVH to DSHF for 7 weeks, and age-matched (18 weeks) Dahl salt-resistant rats fed the same diet served as the control group (DR-C, n = 8). In both groups, blood pressure was similar and significantly higher than in DR-C. Markedly increased left ventricular end-diastolic diameter and reduced fractional shortening in DSHF-V was significantly ameliorated in DSHF-IM using transthoracic echocardiography. The preproET-1, ETAR, and ACE mRNA levels in the left ventricle were significantly increased in DSHF-V compared with DR-C, and significantly suppressed in DSHF-IM compared with DSHF-V. DSHF-V demonstrated a significant increase in the wall-to-lumen ratio and perivascular fibrosis in coronary arterioles, and myocardial fibrosis, with all these parameters being significantly improved by imidapril. In conclusion, myocardial remodeling and heart failure in DS rats fed a high salt diet were significantly ameliorated by a subdepressor dose of imidapril, which may be attributable to a decrease in ET-1 mRNA expression and angiotensin II in the left ventricle.