It has been reported that a change in the cellular redox state may be involved in the regulation of vascular tone, but the underlying mechanism is not fully understood. The present study was designed to investigate the cellular effect of sulfhydryl modifying agents in the coronary artery of rabbit using the tension measurement and whole cell clamping method. The application of diamide, a sulfhydryl oxidizing agent, relaxed the endothelium denuded coronary arteries in a dose dependent manner. The fact that this diamide-induced relaxation was significantly attenuated by a pretreatment of 4-AP, and the coronary arteries precontracted with 100 mM K+ instead of histamine, suggests the involvement of 4-AP sensitive K+ channels in the diamide-induced relaxation of coronary arteries. Whole cell patch clamp studies revealed that the 4-AP sensitive IdK was significantly enhanced by the membrane permeant oxidizing agents, diamide and DTDP, and were reversed by subsequent exposure to the reducing agent, DTT. Neither the membrane impermeant oxidizing or reducing agents, GSSG or GSH, had any effect on the activity of IdK, indicating that intracellular sulfhydryl modification is critical for modulating IdK activity. The Diamide failed to significantly alter the voltage dependence of the activation and inactivation parameters, and did not change the inactivation process, suggesting that diamide increases the number of functional channels without altering their gating properties. Since IdK has been believed to play an important role in regulating membrane potential and arterial tone, our results about the effect of sulfhydryl modifying agents on coronary arterial tone and IdK activity should help understand the pathophysiology of the diseases, where oxidative damage has been implicated.
Animal ; Arteries/physiology ; Arteries/drug effects ; Arteries/cytology ; Coronary Vessels/physiology ; Coronary Vessels/drug effects* ; Coronary Vessels/cytology ; Female ; Male ; Oxidants/pharmacology* ; Potassium Channels/physiology ; Rabbits ; Reducing Agents/pharmacology* ; Sulfhydryl Compounds/metabolism*

To elucidate the Ca2+ release mechanisms in the rabbit coronary artery, arterial preparations were permeabilized with beta-escin and changes in tension were measured under varying experimental conditions. Additionally, we investigated properties and distribution of two kinds of Ca2+ release mechanisms, Ca2+-induced Ca2+ release (CICR) and IP3-induced Ca2+ release (IICR). The results obtained were summarized as follows; 1. When a rabbit coronary artery was incubated in a relaxing solution containing 30 microM beta-escin for 40 min. sensitivity to externally added Ca2+ was much higher in beta-escin permeabilized muscle than in intact preparations. The contractile effect of IP3 in beta-escin permeabilized muscle was also demonstrated; 2. Caffeine and IP3 contracted coronary arteries were permeabilized with beta-escin, but the amplitude of contraction was much larger in the presence of caffeine than of IP3. 3. Intracellular heparin completely inhibited the contractions induced by IP3, but not those by caffeine. On the other hand, procaine inhibited the responses to caffeine, but not those to IP3. Ryanodine inhibited both the caffeine- and IP3-induced contractions. 4. The amplitude of contractile responses was much larger to the maximal stimulation of CICR by applying caffeine than to the maximal stimulation of IICR by applying IP3. After the maximal CICR stimulation by caffeine, the activation of IICR by IP3 without the reloading of Ca2+ could no longer evoke contraction. On the other hand, after the maximal IICR activation, the activation of CICR could still evoke contraction although the amplitude of the contraction was smaller when compared with the case without the initial IICR stimulation. 5. Acetylcholine contracted coronary artery smooth muscles were permeabilized with beta-escin. However, in the absence of added guanosine triphosphate (GTP), the responses were very small. Acetylcholine-induced contraction was inhibited by heparin, but not by procaine. From the above results, it may be concluded that there are two kinds of mechanisms of Ca2+ release, CICR and IICR, in the rabbit coronary artery smooth muscle cell. Also, whereas the CICR mechanism distributes on the membrane of the whole smooth muscle Ca2+ store, the IICR mechanism distributes only on a part of it.
Animal ; Arteries/metabolism ; Calcium/*metabolism ; Capillary Permeability/drug effects ; Coronary Vessels/drug effects/*metabolism ; Escin/pharmacology ; In Vitro ; Intracellular Membranes/*metabolism ; Rabbits ; Support, Non-U.S. Gov't ; Tissue Distribution

Cardiovascular diseases, like coronary heart disease and myocardial infarction, are the most common cause of death worldwide. Chinese medicines have demonstrated rich cardioprotective activities for clinical applications. Salvia miltiorrhiza, a very important component of traditional Chinese medicine, can promote blood circulation and relieve blood stasis. Salvia miltiorrhiza is widely used in treatment of cardiovascular and cerebrovascular disease such as coronary heart disease and cerebral infarction ( CI). Tanshinone II(A), the major lipophilic components extracted from the root of S. miltiorrhiza, possesses anti-atherosclerosis, anti-cardiac hypertrophy, anti-oxidant, anti-arrhythmia and so on. This paper discusses current research status of tanshinone II(A) in cardioprotective effects.
Animals ; Cardiovascular Diseases ; drug therapy ; genetics ; metabolism ; physiopathology ; Coronary Vessels ; drug effects ; physiopathology ; Diterpenes, Abietane ; therapeutic use ; Humans

OBJECTIVETo observe the effects of rapamycin on the expressions of Rho-kinase and p27 mRNA during vascular intimal proliferation in a porcine model of coronary stenosis induced by interleukin-1beta (IL-1beta).

METHODSThe proximal segments of LAD and LCX were wrapped with cotton mesh that had absorbed sepharose bead solution with or without IL-1beta. Selective coronary angiography was performed two weeks later and the animals were killed for collecting the samples for histopathology and RT-PCR analyzing of Rho-kinase and p27 mRNA.

RESULTSThe expressions of Rho-kinase and p27 mRNA could be visualized in normal coronary wall. The expression of Rho-kinase mRNA was significantly enhanced and the expression of p27 mRNA was significantly decreased during the process of intimal proliferation induced by IL-1beta. Rapamycin significantly inhibited the intimal proliferation, reduced the infiltration of inflammatory cells, reduced the expression of Rho-kinase mRNA and increased the expression of p27 mRNA.

CONCLUSIONSThe expression of Rho-kinase mRNA is upregulated and p27 mRNA downregulated in coronary artery stenosis induced by IL-1beta and these effects could be abolished by cotreatment with rapamycin.

Animals ; Coronary Angiography ; Coronary Vessels ; drug effects ; metabolism ; pathology ; Disease Models, Animal ; Interleukin-1beta ; pharmacology ; Male ; RNA, Messenger ; metabolism ; Sirolimus ; pharmacology ; Swine ; Tunica Intima ; drug effects ; metabolism ; pathology ; rho-Associated Kinases ; metabolism

OBJECTIVEThe present study was to explore signaling mechanisms underlying nicotine-induced inhibition of large-conductance calcium-activated potassium channels (BK(Ca)).

METHODS8 week male Wistar rats were divided randomly into saline group and nicotine group and received respectively injection with saline or nicotine (Sigma, Shanghai, China) at 2 mg/(kg x d) for 21 days. Coronary vascular smooth muscle cells were dissociated enzymatically. Dissociated smooth muscle cells were interfered with CPT-cAMP (100 micromol/L) or forskolin (10 micromol/L). The signal channel open dwell-time (To), close dwell-time (Tc) and open probability (Po) were recorded.

RESULTSCPT-cAMP or forskolin significantly prolonged To, shorten Tc and increased Po in saline group (P < 0.01). But in nicotine group To, Tc and Po did not been changed.

CONCLUSIONThis phenomenon may serve as a physiological mechanism that nicotine inhibits BK(Ca) channel activity to increase via cAMP/PKA-dependent pathway.

Animals ; Arteries ; cytology ; drug effects ; metabolism ; Coronary Vessels ; cytology ; drug effects ; metabolism ; Large-Conductance Calcium-Activated Potassium Channels ; metabolism ; Male ; Myocytes, Smooth Muscle ; drug effects ; metabolism ; Nicotine ; pharmacology ; Patch-Clamp Techniques ; Rats ; Rats, Wistar ; Signal Transduction

The relationship between the level of testosterone and the incidence of coronary heart disease is still controversial in the view of the results of clinical and epidemiologic studies. This uncertainty might be partly due to relatively small number of experimental studies undertaken to investigate the cellular mechanism underlying the vascular responses to testosterone. To further investigate the cellular mechanisms of testosterone with respect to vascular response, we investigated the effect of testosterone on contractility and intracellular Ca2+ regulation in a rabbit coronary artery and evaluated the underlying mechanism of testosterone-induced changes of coronary vascular tone by using various pharmacological blockers. Testosterone was found to relax rabbit coronary arteries in a dose-dependent manner, and no significant difference was found in the relaxation response to testosterone with or without endothelium. Similar results were obtained in male and non-pregnant female rabbit coronary arteries. The relaxation response of rabbit coronary arteries to testosterone was greater for PGF2alpha-contracted rings than for KCl contracted rings, which suggest the involvement of K+ channels. Furthermore, the relaxation response to testosterone was significantly reduced by 4-aminopyridine, a sensitive blocker of voltage dependent K+ channels, but not by low doses of tetraethylammonium or iberiotoxin, a Ca2+ activated K+ channel blocker. Testosterone simultaneously reduced the intracellular Ca2+ concentration ([Ca2+]i) and tension, and 4-AP effectively antagonized the testosterone-induced change of [Ca2+]i and tension. Therefore, it may be concluded that the stimulation of voltage dependent K channels is responsible, at least in part, for the testosterone-induced relaxation of rabbit coronary arteries.
Androgens/*pharmacology ; Animals ; Arteries/drug effects ; Calcium/*metabolism ; Coronary Vessels/*drug effects ; Female ; Intracellular Membranes/*metabolism ; Male ; Osmolar Concentration ; Potassium Channels, Voltage-Gated/drug effects/*metabolism ; Rabbits ; Support, Non-U.S. Gov't ; Testosterone/*pharmacology ; *Vasodilation

Isoproterenol (ISO), a beta agonist, causes hyperpolarization of coronary smooth muscle cells via an increase in K+ conductance. This hyperpolarization may cause the coronary vasorelaxation by decreasing the cytoplasmic Ca2+ concentration. It is well known that the activation of beta adrenoreceptors stimulates the adenylate cyclase activity, and the resulting K+ channel phosphorylation by cAMP-dependent protein kinase may be responsible for ISO-induced increase in K+ channel activity. However, it is not clear whether the increase in K+ channel activity by ISO is exclusively due to the activation of adenylate cyclase or not. In this research, the effect of ISO on the isometric tension and the mechanism of ISO-induced K+ channel activation were investigated in various patch clamp conditions. The summarized results are as follows. ISO- and pinacidil induced vasorelaxation was significantly inhibited by the application of TEA or by increasing the external K+ concentration. In the whole cell clamp mode, application of ISO increased K+ outward current, and this effect was completely eliminated by propranolol. In the cell-attached patch, application of ISO or forskolin increased Ca(2+)-activated K+ channel activity. Application of ISO to the bath in the outside-out patches or GTP in the inside-out patches stimulated Ca(2+)-activated K+ channels. From the above results, both A-kinase dependent channel phosphorylation and direct GTP-binding protein mediated effect might be responsible for the the activation of Ca(2+)-activated K+ channel by ISO in rabbit coronary smooth muscle cells. And this K+ channel activation also contributes to the ISO-induced vasorelaxation.
Animal ; Calcium/*metabolism ; Coronary Vessels/*drug effects/physiology ; Cyclic AMP-Dependent Protein Kinases/physiology ; Female ; GTP-Binding Proteins/physiology ; Isoproterenol/*pharmacology ; Male ; Muscle, Smooth, Vascular/*drug effects/physiology ; Potassium Channels/*drug effects ; Rabbits ; Support, Non-U.S. Gov't ; Vasodilation/drug effects

The aim of the present study was to explore the effect of nitric oxide (NO) on iron-induced toxicity in rat hearts. Langendorff perfused rat heart and enzymatically isolated cardiomyocytes were used. It was shown that lipophilic Fe-HQ reduced the contractile amplitude, velocity and end-diastolic cell length in the cardiomyocyte, while the left ventricular developed pressure (LVDP), +/-dp/dt(max), heart rate and coronary flow showed biphasic alterations, which increased in the first 2 min and then was followed by a decline in isolated perfused rat heart; the contents of lactate dehydrogenase (LDH) and creatine kinase (CK) in the coronary effluent and the malondialdehyde (MDA) in the myocardium were increased. L-arginine (L-Arg), an NO precursor, reduced the contractile amplitude and end-diastolic cell length in the cardiomyocyte; but reversibly increased LVDP, +/-dp/dt(max), and coronary flow in isolated perfused rat heart. Pretreatment with L-Arg aggravated the Fe-HQ-induced decrease in contractile amplitude, velocity and end-diastolic cell length in the cardiomyocyte; LVDP, +/-dp/dt(max), heart rate and coronary flow were significantly reduced in the perfused heart, and the levels of LDH and CK increased in the coronary effluent. In contrast, the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) blocked the Fe-HQ induced change in contractile amplitude, velocity and end-diastolic cell length in the cardio- myocyte; it inhibited the decrease in LVDP, LVEDP and +/-dp/dt(max), and reduced the LDH and CK. Removing endothelial cells in coronary vessels attenuated the increase in LVDP and +/-dp/dt(max) at the beginning of Fe-HQ perfusion. It is suggested that L-Arg aggravates the iron-induced cardiac dysfunction, NO can mediate the iron-induced toxicity in heart, and endothelial cells in coronary vessels play an important role in the early stage of the effect of iron.
Animals ; Arginine ; pharmacology ; Coronary Vessels ; cytology ; Creatine Kinase ; metabolism ; Endothelial Cells ; drug effects ; Heart ; drug effects ; Iron ; toxicity ; L-Lactate Dehydrogenase ; metabolism ; Malondialdehyde ; metabolism ; Myocardium ; metabolism ; Myocytes, Cardiac ; cytology ; NG-Nitroarginine Methyl Ester ; pharmacology ; Nitric Oxide ; metabolism ; Rats

OBJECTIVETo explore the protein expression of heme oxygenase-1 (HO-1) and platelet endothelial cell adhesion molecules-1 (PECAM-1) in human coronary artery endothelial cells induced with Zinc Oxide Nanoparticle (ZnO-NPs).

METHODSMTT assay was used to determine the cell viability of ZnO-NPs. Levels of HO-1 and PECAM-1 protein in culture supernatants were measured using ELISA after human coronary artery endothelial cells were treated with different concentrations (0, 10, 20, 40µg/ml) of ZnO-NPs for 24 h.

RESULTSThe cell viability of human coronary artery endothelial cells in each group was 89.76%, 83.61%, 63.10%, 53.20%, 48.11%, 42.35%, 38.06%, 25.44% respectively when treated with different concentrations of ZnO-NPs (12.5, 25, 50, 70, 80, 90, 100, 200µg/ml). Protein levels of HO-1 (ng/L) in each group were 0.041±0.011, 0.512±0.076, 0.906±0.059, 1.062±0.089 respectively after the stimulation of different concentrations of ZnO-NPs (0, 10, 20, 40µg/ml). Comparisons in each group were statistically significant (P < 0.05). Protein levels of PECAM-1 (µg/L) in each group were 7.966 ± 0.046, 7.993 ± 0.036, 8.629 ± 0.052, 8.811 ± 0.039 respectively after the stimulation of different concentrations of ZnO-NPs (0, 10, 20, 40 µg/ml). Compared with the control group, protein levels of PECAM-1 increased (P < 0.05) when the concentration of ZnO-NPs was 20µg/ml or 40 µg/ml.

CONCLUSIONZnO-NPs stimulation could inhibit the viability of human coronary artery endothelial cells and upregulate the protein expression of HO-1 and PECAM-1.

Blood Platelets ; Cell Survival ; Coronary Vessels ; Endothelial Cells ; drug effects ; Heme Oxygenase-1 ; metabolism ; Humans ; Nanoparticles ; toxicity ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Zinc Oxide ; toxicity

OBJECTIVEPeriplocin is an active digitalis-like component from Cortex Periplocae, which has been widely used in the treatment of heart diseases in China for many years. According to the recommendations on the cardiovascular effect of periplocin from in vivo experiments, subsequent in vitro experiments are greatly needed for the global assessment of periplocin. The objective of this study is to investigate the cell proliferation effect and the mechanism of periplocin on endothelial cells.

METHODSThe proliferative activity of periplocin (0.4, 2, 10, 50, 250 micromol/L; 6, 12, 24, 48, 72 h) was investigated by a comparison with the well-reported cardiac glycoside, ouabain, on mouse cardiac microvascular endothelial cells (CMEC). 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide (MTT), lactate dehydrogenase (LDH) and 5-bromo-2-deoxyuridine (BrdU) assays were used to evaluate cell proliferation and viability. Subsequently, cDNA microarray experiments were performed on periplocin- (50 micromol/L) and ouabain- (50 micromol/L) treated cells, and data was analyzed by ArrayTrack software.

RESULTSPeriplocin could increase cell viability to a level lower than ouabain in the MTT analysis, but decrease LDH release simultaneously. The BrdU incorporation assay showed an increase in cell proliferation with 2-50 micromol/L periplocin. Genes related to protein serine/threonine kinase were the most significantly enriched in the 160 genes identified in periplocin versus the control. In the 165 genes regulated by periplocin versus ouabain, GTP-binding was the most altered term.

CONCLUSIONSThe results demonstrated the proliferation action of periplocin on CMEC. Meanwhile, its lower cytotoxicity compared to ouabain provides a new insight into the treatment of heart failure.

Animals ; Animals, Newborn ; Cardiac Glycosides ; pharmacology ; Cardiotonic Agents ; pharmacology ; Cell Proliferation ; drug effects ; Cell Survival ; drug effects ; genetics ; Cells, Cultured ; Coronary Vessels ; drug effects ; metabolism ; physiology ; Drug Evaluation, Preclinical ; Endothelial Cells ; drug effects ; metabolism ; physiology ; Gene Expression Profiling ; Gene Expression Regulation ; drug effects ; Mice ; Microvessels ; drug effects ; metabolism ; physiology ; Models, Biological ; Myocardium ; metabolism ; Oligonucleotide Array Sequence Analysis ; Ouabain ; pharmacology ; Saponins ; pharmacology