Effects of magnolol on vascular contraction in rat aortic rings

1. Magnolol (5,5′‐diallyl‐2,2′‐dihydroxybiphenyl) is a major phenolic compound purified from Magnolia officinalis. The aim of the present study was to elucidate the effects of magnolol on vascular contractions. 2. Rat aortic rings were mounted in organ baths. Magnolol was added cumulatively (0.3–30 μmol/L) to elicit relaxation in endothelium‐intact and ‐denuded rat aortic rings precontracted with U46619 (30 nmol/L, 20 min), NaF (8.0 mmol/L, 40 min), phenylephrine (1.0 or 0.1 μmol/L, 15 min) or phorbol‐12,13‐dibutyrate (PDBu, 0.3 or 0.1 μmol/L, 40 min). In separate experiments, cumulative concentration?response curves were obtained for NaF (2.0–12 mmol/L), U46619 (1.0 nmol/L?1.0 μmol/L) or PDBu (1.0 nmol/L?1.0 μmol/L) after pretreatment with either magnolol or vehicle for 30 min in endothelium‐denuded aortic rings. After completion of the functional study, we measured the amount of guanosine triphosphate (GTP) RhoA by using a G‐LISA RhoA Activation Assay, as well as the phosphorylation of 20 kDa myosin light chain (MLC₂₀), myosin phosphatase‐targeting subunit 1 (MYPT1) and protein kinase C‐potentiated inhibitory protein for heterotrimeric myosin light‐chain phosphatase of 17 kDa (CPI‐17) by immunobloting. 3. Magnolol (0.3–30 μmol/L) reduced vascular tension induced by the thromboxane A₂ agonist U46619 (30 nmol/L), sodium fluoride (NaF) (8.0 mmol/L) and the α₁‐adrenoceptor agonist phenylephrine (1.0 or 0.1 μmol/L) in both endothelium‐intact and ‐denuded rings. The magnitude of the relaxation effects of magnolol on the contraction induced by each of the drugs were similar. The magnitude of the effect of magnolol in endothelium‐intact and ‐denuded rings were similar. 4. Pretreatment of rat aortic rings with 1.0, 3.0 or 10 μmol/L magnolol for 30 min dose‐dependently inhibited the maximum response on the concentration–response curves to NaF and U46619, but not to phorbol‐12,13‐dibutyrate (PDBu). 5. Magnolol (3.0 or 10 μmol/L) decreased RhoA activation, as well as the phosphorylation of MLC₂₀, MYPT1_Thr855 and CPI‐17_Thr38 induced by either 8.0 mmol/L NaF or 30 nmol/L U46619. In contrast, magnolol did not affect PDBu (0.1 μmol/L)‐induced phosphorylation of CPI‐17_Thr38. 6. In conclusion, magnolol reduces vascular contraction by inhibiting the RhoA/Rho kinase pathway in endothelium‐denuded rat aorta.     

非肌肉肌球蛋白Ⅱ研究进展

非肌肉细胞中存在的肌球蛋白Ⅱ称为非肌肉肌球蛋白Ⅱ(non-muscle myosinⅡ,NMⅡ),与肌肉中肌球蛋白Ⅱ具有类似的化学结构,其活性受自身轻链和重链磷酸化水平调节。除了作为一种分子马达为细胞内各种分子运动提供动力外,NMⅡ还参与细胞迁移、黏附、胞质分裂等各种生理活动。     

温阳化饮益气活血法对慢性心功能不全大鼠心肌α-MHC、β-MHC基因表达的影响

目的：探讨温阳化饮益气活血法对慢性心功能不全大鼠心肌α-MHC、β-MHC基因表达的影响。方法：采用腹腔注射阿霉素法复制慢性心功能不全大鼠动物模型，利用RT—PCR技术检测心肌重构的相关调控因素α-MHC、β-MHC在慢性心功能不全发病中的作用，同时观察温阳化饮益气活血法对其干预作用。结果：慢性心功能不全大鼠模型组心肌组织α-MHC基因表达下调，β-MHC基因表达上调；温阳化饮益气活血法治疗组心肌组织α-MHC基因表达上调，β-MHC基因的表达下调。结论：温阳化饮益气活血法能上调慢性心功能不全大鼠α-MHC基因的表达，下调β-MHC基因的表达，为温阳化饮益气活血法的临床应用提供了实验依据，这可能是温阳化饮益气活血法防治慢性心功能不全的作用机制之一。     

褪黑素通过 ERK/MAPK 通路调控 AS 兔动脉内皮细胞 MLCK 表达

 目的 探讨褪黑素（ melatonin , MLT ）是否可通过信号调节激酶 / 丝裂原活化蛋白激酶（ ERK/MAPK ）通路调控动脉粥样硬化（ atherosclerosis , AS ）模型兔动脉内皮细胞肌球蛋白轻链激酶（ myosin light chain kinase , MLCK ）的表达。 方法 复制兔 AS 模型,分析血清中相关脂质成分的动态变化;γ -³²P-ATP 掺入法测定模型兔主动脉的 MLCK 活性;动脉经冰冻切片,免疫组化检测动脉内膜 MLCK 的表达,伊红（ HE ）染色观察动脉壁的形态结构; Western blot 检测 AS 兔动脉组织 MLCK 的表达和 ERK 磷酸化水平。 结果 AS 兔模型复制成功,实验兔在高胆固醇（ cholesterol , Cho ）饮食 4 , 8 , 12 周后,与正常对照相比,血清中相关生化指标有逐渐增加趋势（ <> P <0.05 ）;γ -³²P-ATP 掺入法显示动脉组织中 MLCK 的活性呈逐渐增强趋势（ <> P <0.05 ）, Western blot 和免疫组化则显示动脉内皮细胞 MLCK 的表达、 ERK 磷酸化水平亦呈逐渐增强趋势, HE 染色显示主动脉内膜逐渐增厚,细胞间隙逐渐增大;而加喂 MLT 后,动脉组织 MLCK 的表达、 ERK 磷酸化水平及 MLCK 的活性和主动脉内膜通透性均有所下降,而泡沫细胞形成量逐渐增多,至高 Cho 饮食 12 周时则形成了明显的 AS 斑块,加喂 MLT 后,主动脉内膜病变则有一定程度减轻。 结论 MLT 可能通过 ERK/MAPK 通路下调模型兔动脉内皮细胞 MLCK 活性而减轻 AS 斑块的病变程度。     

The mechanism of vasorelaxation induced by Schisandra chinensis extract in rat thoracic aorta

Aim of the study

Schisandra chinensis (SC) is a known medical herb for the treatment of cardiovascular symptoms associated with menopausal symptoms in Korea. However, the pharmacological action mechanisms involved have not been well studied. This study was aimed to investigate the vascular effects of SC in rat thoracic aorta.

Materials and methods

We isolated the hexane, chloroform, and methanol extracts from SC and evaluated their vasodilatory effects in the rat thoracic aorta.

Results

Hexane extracts of SC (SCHE, 5 × 10⁻⁵ to 10⁻³ g/L) caused a concentration-dependent relaxation in both endothelium-intact and -denuded aortas. The relaxant effect of SCHE on the endothelium-intact aorta was more prominent than on the endothelium-denuded aorta. The former was significantly attenuated by L-NAME, a nitric oxide synthase inhibitor, and ODQ, a soluble guanyl cyclase inhibitor, but not by tetraethylammonium, a nonselective blocker of K⁺ channels, and indomethacin, a cyclooxygenase inhibitor. Furthermore, SCHE caused nitrite production as well as eNOS activation in aortic segments, suggesting implication of NO signal pathway in SCHE-induced relaxation. In endothelium-denuded aorta, SCHE-induced vasorelaxation was also attenuated by calyculin A, an inhibitor of myosin light chain (MLC) phosphatase, but not by ML-9, a MLC kinase inhibitor, suggestive of implication of MLC phosphatase activation. Phenylephrine-enhanced MLC phosphorylation ratio was significantly attenuated by SCHE, which was recovered to the control level by pretreatment with calyculin A.

Conclusions

Taken collectively, these findings suggest that the vascular relaxation evoked by SCHE was mediated by not only endothelium dependent NO pathway but also direct effect on vascular smooth muscle cell via dephosphorylation of MLC.     

温阳化饮益气活血法对慢性心功能不全大鼠心肌α-MHC、β-MHC基因表达的影响

目的:探讨温阳化饮益气活血法对慢性心功能不全大鼠心肌α- MHC、β-MHC基因表达的影响。方法:采用腹腔注射阿霉素法复制慢性心功能不全大鼠动物模型,利用RT-PCR技术检测心肌重构的相关调控因素α- MHC、β-MHC在慢性心功能不全发病中的作用,同时观察温阳化饮益气活血法对其干预作用。结果:慢性心功能不全大鼠模型组心肌组织α-MHC基因表达下调,β-MHC基因表达上调;温阳化饮益气活血法治疗组心肌组织α-MHC基因表达上调,β-MHC基因的表达下调。结论:温阳化饮益气活血法能上调慢性心功能不全大鼠α-MHC基因的表达,下调β-MHC基因的表达,为温阳化饮益气活血法的临床应用提供了实验依据,这可能是温阳化饮益气活血法防治慢性心功能不全的作用机制之一。     

大鼠脑损伤后星形胶质细胞结蛋白和肌球蛋白表达的研究

对大鼠脑基底节穿刺损伤后星形胶质细胞结蛋白和肌球蛋白的表达进行了动态观察。结果显示结蛋白的表达显著增强，７ｄ时达高峰并维持这一高水平至损伤后３０ｄ，损伤后早期（２～４ｄ）其反应限于伤口周围，随后波及整个同侧半球，但强度随与伤口距离增加而逐渐减弱。肌球蛋白的阳性细胞主要见于伤口周围，部分出现在皮质分子层。结果表明结蛋白表达增强与反应性胶质化密切相关，而肌球蛋白的表达可能与增生星形胶质细胞的迁移、收缩有关。文中还对脑损伤后星形胶质细胞骨架蛋白表达改变的意义进行了讨论。     

人血小板肌球蛋白轻链激酶的研究

报道了人血小板匀浆上清液肌球蛋白轻链酶微量快速测定方法，对该酶的性质进行了初步研究，并建立了较为合适的测定酶活性的体系。该酶活性依赖于Ｃａ＾２＋和ＣａＭ，其半激活浓度分别为０．３５ｍｍｏｌ／Ｌ和１０μｍｏｌ／Ｌ，ＴＦＰ能抑制该酶活性，其半抑制浓度为１５０μｍｏｌ／Ｌ。     

心肌急性损伤肌球蛋白轻链激酶酶促反应动力学研究

以内毒素休克狗作为急性心肌损害的动物模型，从心肌中提纯肌球蛋白轻链（ＭＬＣ）和肌球蛋白轻链激酶（ＭＬＣＫ），根据酶动力学分析法研究急性损害对ＭＬＣＫ活性的影响。结果发现在此种状态下狗心肌ＭＬＣＫ活性显著降低，推测ＭＬＣＫ活性降低导致可磷酸化轻链（ＰＬＣ）磷酸化的磷酸酶相对占优势，磷酸化型Ｐ－ＬＣ向去磷酸化型转变。     

Chronic ouabain treatment enhances cardiac myosin ATPase activity in rats

1. Chronic ouabain administration increases blood pressure and produces a positive inotropic effect. However, the temporal changes capable of affecting both arterial and ventricular pressures and myosin ATPase activity during the induced hypertension have not been determined. 2. The aim of the present study was to investigate the time-course of the induction of hypertension to define when changes occur in Wistar rats treated with 25 mg/kg per day, s.c., ouabain for 3, 7, 15 or 30 days. 3. In anaesthetized rats, diastolic blood pressure increased after 7 days treatment with ouabain and after 15 and 30 days treatment, increases were observed in systolic blood pressure, left ventricular systolic pressure and myosin ATPase activity. After 15 days treatment, heart rate (HR) also increased, but after 30 days treatment HR returned to control levels. However, only after 30 days treatment did the left ventricular positive and negative first derivatives of intraventricular pressure (dP/dt(max) and dP/dt(min), respectively) increase. Increased arterial and left ventricular systolic pressures and myosin ATPase activity observed after 15 days treatment maintained similar levels as those after 30 days treatment. 4. The results suggest that changes in arterial and left ventricular pressures, HR and myosin ATPase activity induced by chronic ouabain treatment are time dependent, increasing after 15 days treatment. After 30 days treatment, the increase in systolic and diastolic arterial and ventricular pressures remained stable, as did inotropism. Normalization of HR after 30 days treatment suggests that during the period from Day 16 to Day 30 ouabain-induced hypertension is dependent, at least in part, on increased sympathetic activity.