Experiments were performed in male Sprague-Dawley rats (150-200 g). A silver clip (0.2 mm internal diameter) was placed on the left renal artery of rats. After operation the rats were divided into 4 groups sham group, 2K1C (two-kidney one clip) group, 2K1C+Arg (2K1C and L-Arg 150 mg/kg x d(-1) by drinking) group, and 2K1C+NAME (2K1C and L-NAME 10 mg/kg x d(-1) i.p.) group. The animals were studied at two time points (4 and 7 weeks after operation) corresponding to phases I and II of Goldblatt hypertension. The animals were deeply anesthetized with pentobarbital and perfused by the cardiac route with saline (100 ml) and freshly prepared 4% paraformaldehyde in phosphate buffer (300 ml). The caudal medulla was removed, then placed in 25% sucrose in PB for 12 h in a 4 degrees C fridge. The 40 microm coronal slices of brainstems were cut with cryostat, collected in PBS for nNOS study by immunohistochemistry. The results showed that (1) only a few neuronal nitric oxide synthase (nNOS) positive neurones were found in caudal medulla, including the depressor area of the ventral surface of medulla oblongata (VSMd) and the caudal pressor area (CPA) of the sham operated animals. The number of nNOS positive neurons in caudal medulla was significantly increased in 2K1C Goldblatt hypertension rats at 4 and 7 weeks. (2) The number of nNOS positive neurons in VSMd and CPA were 2K1C+Arg > 2K1C >2K1C +NAME > sham. (3) L-Arg enhanced the expression of nNOS whereas L-NAME inhibited the expression of nNOS in caudal medulla. (4) The character of nNOS expression was similar in Goldblatt hypertension rats at 4 weeks with that of the rats at 7 weeks.
Animals ; Hypertension, Renovascular ; enzymology ; physiopathology ; Male ; Medulla Oblongata ; enzymology ; metabolism ; Nitric Oxide Synthase ; biosynthesis ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the changes in pulmonary artery protein kinase C (PKC) activity in rats with chronic inflammatory pulmonary hypertension (PHT).

METHODSChronic inflammatory PHT was induced in rats with monocrotaline. The PKC activities in the rat pulmonary arteries were measured by radioactive assay during the development of PHT.

RESULTSWith the development of chronic inflammatory PHT, the total and cytosolic fractions of PKC activity in PHT rat pulmonary arteries increased initially with subsequent decrease (Plt;0.05), but the membranous fraction of PKC activity and the membrane-to-cytosol PKC activity ratio increased continuously (P<0.05).

CONCLUSIONThe up-regulation of PKC activity and the translocation of PKC might be associated with the development of chronic inflammatory PHT in rats.

Animals ; Chronic Disease ; Hypertension, Pulmonary ; chemically induced ; enzymology ; Inflammation ; chemically induced ; enzymology ; Male ; Monocrotaline ; Protein Kinase C ; metabolism ; Pulmonary Artery ; enzymology ; Rats ; Rats, Wistar

OBJECTIVETo evaluate the expression of nitric oxide synthase III (NOS III) mRNA in the heart, aorta, kidney and liver of spontaneously hypertensive rats (SHR).

METHODSTwo hundred and ninety-four total RNA samples were obtained from the tissues of ventricle, aortic smooth muscle, kidney and liver of SHR and normotensive rats (Wistar-Kyoto rats, WKY). RNA array was used to determine the mRNA levels of NOS III of the two groups.

RESULTSCompared with WKY, the systolic blood pressure increased significantly in SHR at 6-week-old, 8-week-old, 10-week-old and 12-week-old [(158.50 +/-7.69 vs 108.67 +/-5.89) mmHg, (174.33 +/-4.46 vs 128.50 +/-4.97) mmHg, (198.00 +/-13.45 vs 142.00 +/-3.58) mmHg, (216.67 +/-8.91 vs 141.17 +/-4.92) mmHg, P<0.01], and the ventricle/body weight ratio was significant higher at 10-week-old and 12-week-old [(4.08 +/-0.17 vs 3.59 +/-0.11, 4.05 +/-0.18 vs 3.40 +/-0.19)mg/g, P<0.01]. In the heart tissue and the kidney, the mRNA levels of NOS III were significantly increased at 6-week-old, 8-week-old, 10-week-old and 12-week-old (1.12 +/-0.18 vs 0.90 +/- 0.15, 1.46 +/- 0.34 vs 1.06 +/-0.18, 1.66 +/- 0.31 vs 1.21 +/- 0.30, 1.98 +/- 0.40 vs 1.31 +/-0.38, P <0.05) and at 4-week-old, 6-week-old, 8-week-old, 10-week-old and 12-week-old (1.10 +/- 0.21 vs 0.81 +/-0.11, 1.28 +/-0.18 vs 0.95 +/-0.13,1.31 +/-0.23 vs 0.99 +/-0.23, 1.70 +/-0.30 vs 1.08 +/-0.25, 1.83 +/-0.33 vs 1.15 +/-0.20, P<0.05 or P<0.01), respectively. There was no significant difference of the NOS III expression in the liver and no significant signals were detected in the aortic smooth muscle.

CONCLUSIONThe results provide the evidence of the increased expression of NOS III in different tissues in SHR and suggests the progressive nature of essential hypertension.

Animals ; Hypertension ; enzymology ; genetics ; Kidney ; enzymology ; Liver ; enzymology ; Male ; Myocardium ; enzymology ; Nitric Oxide Synthase ; biosynthesis ; genetics ; Nitric Oxide Synthase Type III ; Oligonucleotide Array Sequence Analysis ; methods ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY

OBJECTIVETo observe the effects of up- or down-regulation of haemoxygenase 1 (HO-1) gene expression on intestinal mucosa injury induced by intra-abdominal hypertension (IAH).

METHODS(1) Reproduction of rat model of up- or down-regulation of HO-1 gene expression. Twenty-four healthy adult Wistar rats were divided into Co-PP (HO-1 specific revulsive) 2.5 mg, Co-PP 5.0 mg, Sn-PP (HO-1 specific inhibitor) 2.5 mg, and control groups according to the random number table, with six rats in each group. Rats in groups Co-PP 2.5 mg and Sn-PP 2.5 mg were respectively given Co-PP 2.5 mg/kg and Sn-PP 2.5 mg/kg by intraperitoneal injection, once every 12 hours for 3 days. The rats in group Co-PP 5.0 mg were intraperitoneally injected with Co-PP 5.0 mg/kg, once a day for 3 days. The rats in control group were treated with equal volume of normal saline by intraperitoneal injection. All rats were sacrificed on post injection day (PID) 4, and intestinal mucosa tissues were collected for determination of HO-1 mRNA expression. Optimal dose of Co-PP was chosen for the following experiment. (2) The influence of up- or down-regulation of HO-1 gene expression on intestinal mucosa injury under IAH condition. Another 24 healthy adult Wistar rats were divided into control, IAH, Co-PP+IAH, and Sn-PP+IAH groups according to the random number table, with six rats in each group. The rats in groups Co-PP+IAH and Sn-PP+IAH were intraperitoneally injected with 2.5 mg/kg Co-PP and 2.5 mg/kg Sn-PP, once every 12 hours for 3 days. Equal volume of normal saline was intraperitoneally injected into the rats in control group, once every 12 hours for 3 days. Then, nitrogen gas pneumoperitoneum was used to establish the model of IAH in rats of the latter three groups on PID 4, with IAP at 20 mm Hg (1 mm Hg = 0.133 kPa) , and it was maintained for 2 hours. Puncture and intubation were performed in rats of control group without inflating nitrogen gas. Jejunal segment in the length of 10-15 cm was harvested for collecting intestinal mucosa tissues to determine the HO-1 mRNA expression and diamine oxidase (DAO) content. Serum obtained from portal vein blood was collected to determine the D-lactate, TNF-α, and IL-6 contents. Another jejunal segment in the length of 1-2 cm was harvested for histopathological examination. Data were processed with one-way analysis of variance and t test.

RESULTS(1) The HO-1 mRNA expression in group Co-PP 2.5 mg was significantly higher than that in control and Co-PP 5.0 mg groups (with t values respectively 4.756, 3.175, P < 0.05 or P < 0.01). The HO-1 mRNA expression in group Sn-PP 2.5 mg was significantly lower than that in control group (t = 4.880, P < 0.01). The optimal dose of Co-PP for the following experiment was 2.5 mg/kg. (2) HO-1 mRNA expression in group Co-PP+IAH was 60 ± 5, and it was obviously higher than that of group IAH (49 ± 5, t = 3.811, P < 0.01) and control group (39 ± 4, t = 8.034, P < .001) . HO-1 mRNA expression was higher in group IAH than in control group (t = 3.826, P < 0.01). HO-1 mRNA expression in group Sn-PP+IAH was 29 ± 4, which was obviously lower than that of control group (t = 4.330, P < 0.01). The contents of DAO and D-lactate in group Co-PP+IAH were (0.52 ± 0.05) U/mL and (1.9 ± 0.6) mg/L, which were significantly lower than those in group IAH [(0.88 ± 0.06) U/mL and (4.3 ± 0.7) mg/L, with t values respectively 11.291, 6.376, P values all below 0.01], but still higher than those in control group [(0.34 ± 0.04) U/mL, (1.2 ± 0.5) mg/L, with t values respectively 6.886, 2.295, P < 0.05 or P < 0.01]. The contents of TNF-α and IL-6 were much lower in group Co-PP+IAH than in group IAH, but still higher than in control group (with t values from 3.781 to 18.557, P values all below 0.01). The contents of DAO, D-lactate, TNF-α, and IL-6 in group Sn-PP+IAH were all higher than those in the other 3 groups (with t values from 4.181 to 32.938, P values all below 0.01). Structure of epithelial cells from intestinal mucosa was intact and regularly arranged in rats of control group. Intestinal mucosal tissue was edematous, and the top of villi was anabrotic and necrotic in rats of group IAH. Compared with that of group IAH, the degree of intestinal mucosa injury was alleviated in rats of group Co-PP+IAH, while the pathology was aggravated in rats of group Sn-PP+IAH.

CONCLUSIONSUp-regulation of HO-1 gene expression can ameliorate intestinal mucosa injury caused by IAH, thus protecting intestinal mucosa tissues.

Animals ; Disease Models, Animal ; Gene Expression Regulation ; Heme Oxygenase (Decyclizing) ; metabolism ; Intestinal Mucosa ; enzymology ; pathology ; Intra-Abdominal Hypertension ; enzymology ; pathology ; Rats ; Rats, Wistar ; Up-Regulation

OBJECTIVETo study the effect of peroxisome proliferator activated receptor γ (PPAR-γ) agonist on the angiotensin converting enzyme 2 (ACE2) mRNA expression in monocyte-derived macrophages of essential hypertensive patients.

METHODSTotally 57 essential hypertensive patients were randomly divided into three groups: conventional treatment group (n=18), telmisartan group (n=19), and benazepril group (n=20); 20 patients with normal blood pressure were also selected as the control group. Monocyte-derived macrophages were isolated from blood samples of patients in all four groups. The expression of ACE2 mRNA in monocyte-derived macrophages was detected by RT-PCR before treatment and 4 and 12 weeks after treatment.

RESULTSFour and 12 weeks after treatment, the systolic pressure and diastolic pressure of telmisartan group and benazepril group were significantly lower than that of the conventional treatment group (all P<0.01), and the systolic pressure and diastolic pressure of telmisartan group were significantly lower than that of the benazepril group(both P<0.01) .The expression of ACE2 mRNA in monocyte-derived macrophages were significantly lower in essential hypertensive patients than that in control group (P<0.01). After having been treated for 4 weeks and 12 weeks, the expression of ACE2 mRNA in monocyte-derived macrophages of hypertensive patients in telmisartan and benazepril groups were significantly higher than that in conventional treatment group (all P<0.01), and the expression of ACE2 mRNA in telmisartan group was significantly higher than that in benazepril group (both P<0.01).

CONCLUSIONPPAR-γ agonist could increase the ACE2 mRNA expression in monocyte-derived macrophages of essential hypertensive patients.

Aged ; Benzazepines ; pharmacology ; Benzimidazoles ; pharmacology ; Benzoates ; pharmacology ; Female ; Humans ; Hypertension ; drug therapy ; enzymology ; Macrophages ; enzymology ; Male ; Middle Aged ; PPAR gamma ; agonists ; Peptidyl-Dipeptidase A ; genetics ; metabolism ; RNA, Messenger ; genetics

OBJECTIVETo investigate plasma gamma-glutamyl transpeptidase (γ-GGT) level as a cardiovascular risk factor in elderly patients with hypertension or hypertension with diabetes mellitus.

METHODSForty-nine elderly patients of hypertension and 42 elderly patients of hypertension with diabetes mellitus and 39 healthy elderly subjects were enrolled in the study. The height, weight and blood pressure of patients were measured, serum C reactive protein and other biochemical indicators were detected. The relation between plasma γ-GGT and cardiovascular risk factors in three groups were analyzed.

RESULTSThere was no significant difference in plasma γ-GGT levels among three groups. There was a positive correlation of plasma γ-GGT levels with systolic pressure, pulse pressure, hemoglobin A1c and CRP in control group. While in hypertension with diabetes mellitus group, plasma γ-GGT levels were correlated with systolic pressure, mean arterial pressure, fasting blood sugar and cystatin.

CONCLUSIONPlasma γ-GGT might be a risk factor for cardiovascular diseases, and may be used as a predictive indicator for kidney injury in early patients with hypertension with diabetes mellitus.

Aged ; Aged, 80 and over ; Cardiovascular Diseases ; etiology ; Case-Control Studies ; Diabetes Mellitus ; enzymology ; Female ; Humans ; Hypertension ; complications ; enzymology ; Male ; Middle Aged ; Risk Factors ; gamma-Glutamyltransferase ; blood

The activation of protein kinase C (PKC) is not only a pivotal node during cardiac hypertrophy in chronic pressure-overloaded heart, but also involved in the regulation of cardiac contractility. The aim of this paper was to observe PKC modulation in cardiac contractility at different stages of cardiac hypertrophy in spontaneously hypertensive rat (SHR). The cardiomyocytes were isolated from 4- and 10-month-old normotensive Wistar-Kyoto (WKY) and SHR rat hearts. The shortening amplitude of unloading contraction in cardiomyocytes was observed by an Edge Detector system. The shortening amplitude in WKY rat cardiomyocytes increased gradually as the stimulating frequency increased from 1 to 3 Hz. The shortening amplitude was positively correlated with stimulating frequency. The shortening amplitude in 4-month-old SHR group was enhanced as compared with that in WKY group at the same stimulating frequency. When the stimulating frequency increased, the shortening amplitude did not increase in 4-month-old SHR group. There was no difference in shortening amplitude between 10-month-old SHR and WKY groups at 1-Hz stimulating frequency. But the shortening amplitude in 10-month-old SHR group decreased when the stimulating frequency increased to 3 Hz. The perfusion of 50, 100 or 200 nmol/L PMA (a non-specific agonist of PKCs) significantly reduced the shortening amplitude in WKY and SHR groups. The shortening amplitude was reduced to (69.8±1.9)%, (58.2±2.2)% and (22.7±2.5)% (all P<0.01), respectively, in WKY group, as compared with that in HEPES buffer perfusion (100%). It was reduced to (6.1±0.7)%, (2.4±0.2)% and (12.5±2.6)% (all P<0.01) in 4-month-old SHR group, and (65.7±1.6)%, (53.9±4.0)% and (16.3±2.0)% (all P<0.01) in 10-month-old SHR group. The decreases in shortening amplitude in 4-month-old SHR group were more significant than those in 10-month-old SHR and WKY groups. On the other hand, 200 nmol/L of staurosporine, an inhibitor of PKC, significantly increased the shortening amplitude of cardiomyocytes in WKY, 4-month-old SHR, and 10-month-old SHR groups by (63.63±4.53)%, (80.82±4.61)% and (80.97±4.59)%, respectively (all P<0.05). The results suggest that the PKC isoforms inducing negative inotropic effect may be activated at the early stage of cardiac hypertrophy in SHR rats, and are possibly involved in the modulation of cardiac contractility. The activated PKC isoforms return to their normal activity at the stable stage of cardiac hypertrophy in SHR rats.
Animals ; Cardiomegaly ; enzymology ; Heart ; physiopathology ; Hypertension ; physiopathology ; Myocardial Contraction ; Myocytes, Cardiac ; enzymology ; Protein Kinase C ; metabolism ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY

OBJECTIVETo investigate the expression of Rho-kinase and heme oxygenase and the interaction among NOS/NO, HO/CO and RhoA/Rho-kinase in the corpus cavernosum of spontaneous hypertensive rats (SHRs).

METHODSA series of electric stimuli were applied to the corpus cavernosum nerves of 7 SHRs, the changes of ICP/MAP observed continuously and the expressions of ROCK2, HO-2 and eNOS analyzed by Western blot and immunohistochemistry. Another 7 WKY rats were taken as controls.

RESULTSCompared with the controls, ICP/MAP did not rise obviously (P < 0.05), HO-2 dropped sharply (P = 0.006) and the expression of the ROCK2 protein was elevated markedly (P = 0.017) in the SHRs. Immunohistochemistry showed HO-2 to be distributed in the smooth muscle cells and nervous cells of the corpus cavernosum, and eNOS mainly in the vascular endothelial cells. The expressions of HO-2 and eNOS were obviously reduced in the SHRs.

CONCLUSIONNOS/NO, HO/CO and RhoA/Rho-kinase are related with erectile dysfunction in SHRs and may interact on one another.

Animals ; Blotting, Western ; Erectile Dysfunction ; enzymology ; physiopathology ; Heme Oxygenase (Decyclizing) ; metabolism ; Hypertension ; enzymology ; physiopathology ; Immunohistochemistry ; Male ; Penis ; enzymology ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY ; rho-Associated Kinases ; metabolism

OBJECTIVETo observe myocardial cathepsin (Cat) S expression and activity in hypertensive heart failure rats.

METHODSThe expression and activity of Cat S were determined in the left ventricular (LV) myocardium (LVM) of Dahl salt-sensitive rats fed either a high-salt (HS, 8%) or low-salt (LS, 0, 3%, controls) diet starting at age 7 weeks for 12 weeks (hypertrophy model, H-LVH) or 19 weeks (heart failure model, H-HF). Age-matched rats served as controls and human normal, hypertensive and heart failure myocardial specimen were also examined for changes on the expression and activity of Cat S.

RESULTSReverse transcription and real-time polymerase chain reaction analysis revealed significantly upregulated Cat S mRNA in rats with H-HF than in rats with H-LVH or in control rats and Cat S mRNA expression is negatively correlated with LVEF (r = -0.88, P < 0.05). In situ and immunohistochemistry examinations showed that Cat S was localized predominantly in cardiac myocytes (CMCs) and coronary vascular smooth muscle cells (SMC). Elastic lamina fragmentations and Cat S-dependent elastolytic activity were significantly increased in H-HF-rats. The expression of interleukin-1 beta was also increased in the LVM of H-HF rats, and this cytokine was found to increase the Cat S protein expression in culture neonatal CMCs. Similar results were revealed in human myocardial specimens.

CONCLUSIONElastolytic Cat S might play an important role in the pathogenesis of myocardial remodeling and heart failure and Cat S might serve as a novel therapeutic target in preventing or reversing hypertension induced LV remodeling and heart failure.

Adult ; Aged ; Animals ; Case-Control Studies ; Cathepsins ; metabolism ; Disease Models, Animal ; Enzyme Activation ; Heart Failure ; enzymology ; etiology ; physiopathology ; Humans ; Hypertension ; complications ; enzymology ; Male ; Middle Aged ; Myocardium ; enzymology ; Rats ; Rats, Inbred Dahl ; Ventricular Function, Left ; Ventricular Remodeling

OBJECTIVES: The present study was aimed at exploring whether the pathogenesis of hypertension is related with an altered expression of nitric oxide synthase (NOS) isozymes, i.e., bNOS, iNOS and ecNOS. METHOD: By Western blot analysis, the expression of NOS isozymes were determined in the kidney isolated from spontaneously hypertensive rats (SHR) and their normotensive control, Wistar-Kyoto rats (WKY). The NOx (nitrite/nitrate) contents were also determined in the kidney and plasma. RESULTS: The plasma NOx was significantly increased in SHR compared with that in WKY. The basal level of NOx was higher in the medulla and cortex of the kidney in SHR compared with that in WKY rat. bNOS proteins were expressed higher in the outer medulla and cortex, and iNOS proteins were higher in the inner medulla, outer medulla and cortex in SHR. ecNOS expression did not significantly differ between the SHR and WKY. CONCLUSIONS: These results indicate that the NO generation may not be impaired, but rather increased. It is likely that the increased expression of NOS isozymes is a counter-reactive phenomenon secondary to the increased blood pressure in this model of hypertension.
Animal ; Hypertension/physiopathology ; Hypertension/enzymology* ; Isoenzymes/metabolism ; Kidney/enzymology* ; Male ; Nitrates/metabolism ; Nitrates/blood ; Nitric Oxide/biosynthesis ; Nitric-Oxide Synthase/metabolism* ; Nitrites/metabolism ; Nitrites/blood ; Rats ; Rats, Inbred SHR ; Rats, Inbred WKY