Hypertension and Hypercholesterolemia: Biochemical and Pharmacological Studies in the Rabbit

1. Four groups of rabbits were set up to investigate the separate and combined effects of hypertension and hypercholesterolemia on a number of cardiovascular parameters, including in vivo vascular reactivity, phosphoinositide metabolism in aortic tissue, and platelet [Ca2+]i concentrations. The four groups were: a control group, a hypertensive group (perinephritis hypertension), a hypercholesterolemic group (0. 3% cholesterol diet), and a hypertensive-hypercholesterolemic group. 2. As the basic pharmacology of one of the agonists used in this work, endothelin, was poorly understood at the time these studies were planned, preliminary investigations into its effects in normotensive-normocholesterolemic animals were undertaken before embarking on the main project. In addition, some phosphoinositide studies were done using rat aorta. Endothelin-1 caused a dose-related increase in phosphoinositide hydrolysis in both rat and rabbit aorta. In rat aorta, endothelin-1-induced phosphoinositide hydrolysis increased with stimulation time for the first 30 minutes, and thereafter plateaued. The endothelin-stimulated effects were attenuated with endothelium removal, or with extracellular Ca2+ depletion. The endothelin-l-induced phosphoinositide hydrolysis was greater in rat aorta than in rabbit aorta. In vivo endothelin-1 caused a short-lived depressor response followed by a long-lasting pressor response in the rabbit. The pressor response was dose-related, and could be attenuated by the calcium antagonist nifedipine. 3. The imposition of perinephritis hypertension caused an increase of about 40 mmHg in mean arterial pressure in the operated rabbits, which stabilized after 6-7 weeks, while feeding a 0. 3% cholesterol diet induced a continual rise in plasma cholesterol levels in the rabbits, which reached 30 mmol/L after 4 months. In contrast to 0.3% cholesterol diet, perinephritis hypertension was a significant risk factor for cardiovascular deaths in the course of a 4-month study. There were significantly greater numbers of cardiovascular deaths in both the hypertensive and the hypertensive-hypercholesterolemic groups versus the control group (p= 0.038 and 0.009, respectively), but no significant difference between the hypercholesterolemic and the control group, or between the hypertensive-hypercholesterolemic and the hypertensive group. However, it is noteworthy that when the hypertensive-hypercholesterolemic group was compared to the hypertensive group, 0.3% cholesterol diet tended to augment the cardiovascular deaths in these hypertensive animals. 4. The imposition of perinephritis hypertension enhanced the pressor responses to angiotensin II, and endothelin-1, as well as the depressor responses to acetylcholine, isoproterenol, and nitroprusside at 2-3, 6-7, and 13-16 weeks of study. The differences tended to increase with the duration of hypertension for the pressor responses, but not for the depressor responses. The patterns of changes in the duration of hypertension differed from one pressor agonist to another, suggesting that structural changes in the vessel wall were not the sole explanation for enhanced vascular reactivity. Similar conclusions held for the depressor agonists. In contrast, the imposition of 0. 3% cholesterol diet had no effect on the in vivo vascular reactivity at any time point of the study, whether given to normotensive or hypertensive animals. 5. Neither perinephritis hypertension nor 0. 3% cholesterol diet caused any changes in the basal platelet [Ca2+] at the 17th week of study. 6. At the 18th week of study, perinephritis hypertension tended to enhance the noradrenaline-stimulated, but not the endothelin-stimulated phosphoinositide hydrolysis in rabbit aorta, but the difference was significant only at 10e-4 M noradrenaline for the hypertensive versus the control group. In contrast, 0.3% cholesterol feeding tended to decrease both noradrenaline- and endothelin-stimulated, phosphoinositide hydrolysis. The difference was significant at 10e-4 M noradrenaline for the hypercholesterolemic versus the control group, as well as for the hypertensive-hypercholesterolemic versus the hypertensive group, and at 10e-6 & 10e-5 M endothelin-1 for the hypertensive-hypercholesterolemic versus the hypertensive group. There was no difference in the basal [ 3H]-inositol phosphates formation between any experimental group and the control group, suggesting that neither perinephritis hypertension nor 0.3% cholesterol diet for 18 weeks altered the basal phosphoinositide metabolism in rabbit aorta. 7. Overall, in our study no significant additive effects of the two disease states were observed in any of the parameters examined. However, the number of biochemical responses, vessels and animals examined were limited, and further studies might identify sites of interaction between the two parameters

Inhibitory effects of armepavine against hepatic fibrosis in rats

Activation of hepatic stellate cells (HSCs) plays a crucial role in liver fibrogenesis. armepavine (Arm, C19H23O3N), an active compound from Nelumbo nucifera, has been shown to exert immunosuppressive effects on T lymphocytes and on lupus nephritic mice. The aim of this study was to investigate whether Arm could exert anti-hepatic fibrogenic effects in vitro and in vivo. A cell line of rat HSCs (HSC-T6) was stimulated with tumor necrosis factor-α (TNF-α) or lipopolysaccharide (LPS) to evaluate the inhibitory effects of Arm. An in vivo therapeutic study was conducted in bile duct-ligated (BDL) rats. BDL rats were given Arm (3 or 10 mg/kg) by gavage twice daily for 3 weeks starting from the onset of BDL. Liver sections were taken for fibrosis scoring, immuno-fluorescence staining and quantitative real-time mRNA measurements. In vitro, Arm (1-10 μM) concentration-dependently attenuated TNF-α- and LPS-stimulated α-SMA protein expression and AP-1 activation by HSC-T6 cells without adverse cytotoxicity. Arm also suppressed TNF-α-induced collagen collagen deposition, NFκB activation and MAPK (p38, ERK1/2, and JNK) phosphorylations. In vivo, Arm treatment significantly reduced plasma AST and ALT levels, hepatic α-SMA expression and collagen contents, and fibrosis scores of BDL rats as compared with vehicle treatment. Moreover, Arm attenuated the mRNA expression levels of col 1α2, TGF-β1, TIMP-1, ICAM-1, iNOS, and IL-6 genes, but up-regulated metallothionein genes. Our study results showed that Arm exerted both in vitro and in vivo antifibrotic effects in rats, possibly through anti-NF-κB activation pathways

Utilization patterns of Chinese medicine and Western medicine under the National Health Insurance Program in Taiwan, a population-based study from 1997 to 2003

<p>Abstract</p> <p>Background</p> <p>In 1995, Taiwan has launched a national health-care system (the National Health Insurance Program, NHI) covering the use of both Western medicine (WM) and Chinese medicine (CM). This population-based study was conducted to understand the role of CM in this dual medical system by determining the utilization patterns of CM and WM and to analyze the demographic characteristics and primary indications influencing the choice of the medical services for the development of strategies to enhance the appropriate use and reduce unnecessary use of CM.</p> <p>Methods</p> <p>This study used the NHI sample files from 1997 to 2003 consisting of comprehensive utilization and enrolment information for a random sample of 200,432 NHI beneficiaries of the total enrolees from 1995 to 2000. A total of 136,720 subjects with valid and complete enrolment and utilization data were included in this study. The logistic regression method was employed to estimate the odds ratios (ORs) for utilization of CM and WM. The usage, frequency of services, and primary indications for CM and WM were evaluated. A significance level of α = 0.05 was selected.</p> <p>Results</p> <p>Compared with WM, the odds of CM increased from 1997 to 2003. The odds of using CM (OR = 1.48; 95% CI: 1.45–1.50; p < 0.001) and WM (OR = 1.74; 95% CI: 1.72–1.77; p < 0.001) were higher in females and that of CM increased with age to a peak in the 45–54-year-group (OR = 1.75; 95% CI: 1.68–1.82; p < 0.001) and WM (OR = 1.09; 95% CI: 1.05–1.13; p < 0.001) in the elderly subjects (≥ 65 years). The odds of CM and WM were similar in all income groups. However, those of CM were higher in Central (OR = 1.65; 95% CI: 1.56–1.74; p < 0.001) and Southern Taiwan (OR = 1.18; 95% CI: 1.12–1.25; p < 0.001) and lower in the remote areas (OR = 0.57; 95% CI: 0.52–0.63; p < 0.001). Most of the patients had one ambulatory visit of both medical services annually. However, the utilization of WM predominated over CM. Over 90% of CM service was provided by clinics, whereas over 60% of WM service by hospitals. Diseases of the respiratory system was the most frequent primary indication in CM and WM. Herbal medication was the most commonly used form of CM (68.4–72.7%).</p> <p>Conclusion</p> <p>In recent years, there is an increasing trend in the utilization of CM in Taiwan. This increasing trend may be due to the covering of CM in the national health insurance system.</p

Investigation of Mitomycin-C-Treated Fibroblasts in 3-D Collagen Gel and Conditioned Medium for Keratinocyte Proliferation

Fibroblasts produce a spectrum of necessary growth factors essential for growth and proliferation of a variety of cell types. In this study, the paracrine effect of mitomycin-C- treated fibroblasts with various densities in collagen gel for keratinocyte proliferation was investigated from which an optimum cell density and optimum conditioned medium would be determined to expand keratinocyte without further differentiation for skin equivalent tissue engineering. The optimum cell density in collagen feeder gel for optimum collected medium preparation will be determined by checking the level of keratinocyte growth factor and granulocyte macrophage colony-stimulating factor in conventional medium. The results showed that the cell density of 1 x 10(5) cells /gel in the feeder gel is better to produce optimum collected medium. The conditioned medium is prepared by mixing together the optimum collected medium and molecular cellular and developmental biology (MCDB) 153 medium in different ratios for keratinocyte growth. The keratinocyte viability will be measured by 3- (4,5-dimethyl-thiazol-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay to determine the optimum conditioned medium. From the study, 67% conditioned medium was supposed as the better medium for keratinocyte proliferation. In this experiment, the optimum cell density in feeder gel to coculture with keratinocytes is also determined as 1 x 10(5) cells/gel. Keratin 10 (K10) and Terminal Deoxynucleotidyl Transferase Mediated dUTP Nick End Labeling stain will be used to check the cell differentiation and apoptosis, respectively. The results suggest that keratinocytes should not be cultured in postconfluent conditions due to undesired apoptosis and differentiation. The result of cell viability from passages to passages shows that the optimum feeder gel plays a more important role to the keratinocyte proliferation than that of optimum conditioned medium. Keratinocytes cultured with optimum feeder gel in 67% conditioned medium could effectively promote proliferation, inhibit apoptosis, and prevent differentiation. The combination of conditioned media and feeder gel to culture keratinocytes without external supplements can provide an inexpensive way for keratinocyte proliferation and construct an environment for real-time communication between the two cells. The results conclude that keratinocyte cultivation in feeder gel with modified medium should be feasible in the production of high quality keratinocytes for skin equivalents preparation

Hemodynamic-Effects of 8-Day Dl-028 and Octreotide Administration in Rats with Portal-Hypertension

Background: DL-028 (chemical name: 3-[[4-(2-methoxyphenyl) piperazin-1-yl ]methyl]-2,3-dihydroimidazo[1,2-c]quinazolin-5 (6H)-one (27b)) is a synthetic alpha(1)-adrenoceptor antagonist. The present study was undertaken to investigate the hemodynamic effects of chronic DL-028 administration, alone or in combination with octreotide, in rats with portal hypertension. Methods: Portal hypertension was induced by partial portal-vein ligation. Portal-hypertensive rats were allocated to one of the four groups: vehicle group (saline, 0.5 ml/12 h), octreotide group (30 mu g/kg/12 h), DL-028 group (0.4 mg/kg/12 h), and octreotide (30 mg/kg/12 h) plus DL-028 (0.4 mg/kg/12 h) group, with eight rats in each group . DL -028 or saline was administered by gavage and octreotide by subcutaneous injection. Drugs were given immediately after ligation and for 8 consecutive days thereafter. Systemic and splanchnic hemodynamic variables were measured thereafter. Results: Portal-vein-ligated rats showed a typical hyperdynamic state as compared with sham-operated rats. The portal venous pressure, portal tributary blood flow, and cardiac index were significantly reduced by treatment with octreotide, DL-028, or octreotide plus DL-028 in portal-hypertensive rats. Hyperdynamic variables of systemic, renal, hepatocollateral, and portal territory vascular resistances and renal and hepatic arterial blood flow were ameliorated by treatment with octreotide or octreotide plus DL-028 in portal-hypertensive rats. Octreotide plus DL-028 treatment exerted better hemodynamic effects on the cardiac index but worse effects on systemic and hepatocollateral vascular resistance than octreotide treatment alone. Conclusion: Although either DL-028 or octreotide ameliorated portal hypertension and splanchnic hyperemia in portal-hypertensive rats, octreotide treatment exerted more beneficial hemodynamic effects than DL-028 treatment. The combination of octreotide and DL-028 conferred no better hemodynamic benefits than octreotide alone, except on the cardiac index

Portal Hypotensive Effects of Dl-028 and Prazosin on Portal Hypertensive Rat

The portal hypotensive effects of prazosin and DL-028 ( chemical name: 3- [[4-(2-methoxyphenyl)piperazin-1-yl]methyl] 2,3-dihydroimidazo[1,2-c] quinazolin-5(6H)-one(27b)), a synthetic alpha(1)-adrenoceptor antagonist, were assessed in portal hypertensive rats. Portal hypertension was induced by partial portal vein ligation in Sprague-Dawley rats. Two weeks after ligation, when the hyperdynamic state was stabilized the rats were anesthetized after an overnight fast and cannulated for measuring mean arterial pressure ( MAP), portal venous pressure (PVP), cardiac index (CI) and heart rate (HR). Both DL 028 and prazosin (1, 3.3 and 10 mu g/kg) induced dose-dependent decreases of PVP and MBP after intravenous infusion , with effects lasting for longer than 30 min. The maximum percentage reduction of PVP after DL-028 was 10, 10 and 15%, respectively, for the dosages given (1, 3.3 and 10 mu g/kg), and 5, 12 and 25%, respectively, after prazosin. CI was not changed by either drug. HR was not changed by either drug except DL- 028 at 10.0 mu g/kg with a bradycardiac effect. Our results showed that both DL-028 and prazosin reduced PVP in portal hypertensive rats

Inhibitory Effect of Tanshinone IIA on Rat Hepatic Stellate Cells

<div><p>Background</p><p>Anti-inflammation via inhibition of NF-κB pathways in hepatic stellate cells (HSCs) is one therapeutic approach to hepatic fibrosis. Tanshinone IIA (C₁₉H₁₈O₃, Tan IIA) is a lipophilic diterpene isolated from <i>Salvia miltiorrhiza</i> Bunge, with reported anti-inflammatory activity. We tested whether Tan IIA could inhibit HSC activation.</p><p>Materials and Methods</p><p>The cell line of rat hepatic stellate cells (HSC-T6) was stimulated with lipopolysaccharide (LPS) (100 ng/ml). Cytotoxicity was assessed by MTT assay. HSC-T6 cells were pretreated with Tan IIA (1, 3 and 10 µM), then induced by LPS (100 ng/ml). NF-κB activity was evaluated by the luciferase reporter gene assay. Western blotting analysis was performed to measure NF-κB-p65, and phosphorylations of MAPKs (ERK, JNK, p38). Cell chemotaxis was assessed by both wound-healing assay and trans-well invasion assay. Quantitative real-time PCR was used to detect gene expression in HSC-T6 cells.</p><p>Results</p><p>All concentrations of drugs showed no cytotoxicity against HSC-T6 cells<b>.</b> LPS stimulated NF-κB luciferase activities, nuclear translocation of NF-κB-p65, and phosphorylations of ERK, JNK and p38, all of which were suppressed by Tan IIA. In addition, Tan IIA significantly inhibited LPS-induced HSCs chemotaxis, in both wound-healing and trans-well invasion assays. Moreover, Tan IIA attenuated LPS-induced mRNA expressions of <i>CCL2, CCL3, CCL5, IL-1β, TNF-α, IL-6, ICAM-1, iNOS,</i> and <i>α-SMA</i> in HSC-T6 cells.</p><p>Conclusion</p><p>Our results demonstrated that Tan IIA decreased LPS-induced HSC activation.</p></div

Chemical structures and cytotoxicity of salvianolic acid B and tanshinone IIA.

<p>Chemical structures of Sal B (A) and Tan IIA (B). (C) MTT assay was performed to assess HSC-T6 cells viability with Tan IIA (1, 3, and 10 µM) and Sal B (200 µM) treatment for 24 hr. There was no cytotoxicity in all concentrations of drugs. n = 3.</p