芍药苷对血管紧张素Ⅱ诱导的大鼠动脉平滑肌细胞增殖的影响

目的 观察芍药苷对血管紧张素Ⅱ刺激下离体大鼠胸主动脉平滑肌细胞(VSMC)增殖的影响和基质金属蛋白酶-2活性的影响.方法 体外培养大鼠胸主动脉VSMC,使用血管紧张素Ⅱ(AngⅡ)、芍药苷干预血管平滑肌细胞,采用MMT法检测细胞增殖,硝酸还原酶及化学比色法测定细胞上清液中一氧化氮(NO)、一氧化氮合酶(NOS)、诱导型一氧化氮合酶(iNOS)水平,应用酶谱法测基质金属蛋白酶-2活性的变化.结果 AngⅡ能明显促进平滑肌增殖、提高基质金属蛋白酶-2活性,芍药苷在125～1000 μg/mL能显著提高NO、NOS、iNOS水平,抑制平滑肌增殖,并呈剂量及时间依赖性;能显著抑制AngⅡ诱导的血管平滑肌基质金属蛋自酶-2活性的升高.结论 芍药苷可能通过升高NO和NOS水平抑制AngⅡ诱导的平滑肌胞增殖.     

通脉Ⅰ号抑制自发性高血压大鼠血管平滑肌细胞增殖的机理探讨

目的 探讨通脉Ⅰ号抑制自发性高血压大鼠(SHR)血管平滑肌细胞(VSMC)增殖的可能机理。方法细胞计数、逆转录-聚合酶链反应(RT-PCR)及放射免疫等细胞、分子生物学方法。结果通脉Ⅰ号显著地抑制CSMC的增殖；15mg／L的浓度可极显著地抑制VSMC上AngⅡ受体AT1 mRNA的表达，并可降低SHR-VSMC内AngⅡ含量。结论 通脉Ⅰ号对SHR的VSMC增殖的抑制作用机理之一系降低了AT1受体的mRNA表达，拮抗AngⅡ受体。     

通脉I号抑制自发性高血压大鼠血管平滑肌细胞增殖的机理探讨

目的 探讨通脉I号抑制自发性高血压大鼠（SHR）血管平滑肌细胞（VSMC）增殖的可能机理。方法 细胞计数、逆转录－聚合酶链反应（RT－PCR）及放射免疫等细胞、分子生物学方法。结果 通脉I号显著地抑制VSMC的增殖；15mg/L的浓度可极显著地抑制VSMC上AngⅡ受体AT1 mRNA的表达，并可降低SHR－VSMC内AngⅡ含量。结论 通脉I号对SHR的VSMC增殖的抑制作用机理之一系降低了AT1受体的mRNA表达，拮抗AngⅡ受体。     

清达颗粒对血管紧张素Ⅱ诱导大鼠血管平滑肌细胞增殖和迁移的影响

目的探讨清达颗粒(QDG)对血管紧张素Ⅱ(AngⅡ)诱导大鼠血管平滑肌细胞(VSMCs)增殖和迁移的影响。方法采用组织贴块法培养大鼠胸主动脉平滑肌细胞,用表型标志蛋白α-SM-actin对VSMCs进行免疫荧光染色鉴定,取3～6代的VSMCs用于实验。将VSMCs分为对照组、AngⅡ组、AngⅡ+QDG 0.125 mg/m L组、AngⅡ+QDG 0.25 mg/m L组、AngⅡ+QDG 0.5 mg/m L组,共5组。以10~(-6) mol/L AngⅡ作为诱导剂,干预建立VSMCs增殖、迁移的模型。采用CCK-8法检测VSMCs的细胞活力,计算AngⅡ对VSMCs的增殖率以及QDG对AngⅡ诱导VSMCs增殖的抑制率;采用划痕实验检测VSMCs划痕损伤修复情况;transwell法检测VSMCs迁移情况。结果与对照组比较,AngⅡ能显著促进VSMCs的增殖和迁移;不同浓度QDG可在一定程度上抑制AngⅡ诱导的VSMCs增殖和迁移,且随着浓度的升高,抑制作用越明显。结论 QDG具有抑制AngⅡ诱导的VSMCs增殖与迁移的作用。     

玄参对心室重构大鼠血管紧张素Ⅱ及其1型受体基因表达的影响

目的 探讨玄参对心室重构大鼠血管紧张素Ⅱ(AngⅡ)及其1型受体基因表达(AT1AmRNA)的影响.方法 采用甲状腺素致大鼠心室重构及腹主动脉不完全结扎致大鼠心室重构两种动物模型,观察玄参对心脏指数(heart weight in-dex,HWI)、心肌血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)、醛固酮(aldosterone,ALD)及血管紧张素Ⅱ受体1型基因(AT1A mR-NA)表达水平的影响.结果 与模型组相比,玄参可显著降低动物HWI,降低心肌AngⅡ,ALD含量,降低AT1A>mRNA的过量表达.结论 玄参对心室重构具有明显的改善作用,其机制可能与抑制AngⅡ,ALD生成和AT1AmBNA表达有关.     

降压宝蓝片含药血清对血管紧张素Ⅱ诱导的血管平滑肌细胞增殖、迁移及MMP-9表达的影响

目的:研究降压宝蓝片含药血清对血管紧张素Ⅱ诱导的血管平滑肌细胞增殖、迁移MMP-9表达的影响。方法:组织贴块法培养大鼠胸主动脉平滑肌细胞,分为空白对照组、模型对照组、降压宝蓝片含药血清低、中、高浓度组(体积分数为5%、10%、15%),用10-6mol/L AngⅡ诱导细胞,用噻唑蓝法检测细胞增殖,Transwell小室法检测细胞迁移,免疫组化法检测MMP-9表达。结果:与模型对照组比较,15%的降压宝蓝片含药血清可抑制Ang II诱导的平滑肌细胞增殖和迁移(P0.05或P0.01),10%、15%的降压宝蓝片含药血清显著降低细胞的MMP-9表达(P0.05或P0.01)。结论:降压宝蓝片含药血清能抑制血管紧张素Ⅱ诱导的血管平滑肌细胞增殖和迁移,降低MMP-9的表达。     

钩藤碱对AngⅡ诱导大鼠血管平滑肌细胞增殖的抑制作用

 目的研究钩藤碱(rhynchophylline,Rhy)对血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)诱导大鼠血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖的抑制作用及机制。方法采用细胞计数法和MTT比色法检测Rhy对AngⅡ诱导大鼠VSMCs增殖的影响;测定Rhy对AngⅡ作用的VSMCs上清液一氧化氮(nitric oxide,NO)含量和一氧化氮合酶(constitutive nitric oxide synthase,cNOS)活性的影响;实时定量聚合酶链式反应(RT-PCR)检测VSMCs中c-myc、NOS和高血压相关基因-1(hypertension related gene-1,rHRG-1)mRNA的表达。结果Rhy(3×10^-6～3×10^-7mol·L^-1)呈浓度依赖性地抑制AngⅡ诱导大鼠VSMCs的增殖,升高细胞上清液中NO的含量和NOS活性,降低c-myc mRNA,升高rHRG-1和NOS mRNA的表达。结论Rhy明显抑制AngⅡ诱导大鼠VSMCs的增殖,机制可能与增加VSMCs中NOS活性并促进NO的合成和释放以及下调c-myc、上调NOS和rHRG-1 mRNA的表达有关。     

通脉颗粒对自发性高血压大鼠血管重构的影响机理研究

高血压血管重构 是指高血压状态下血管结构和功能的改变.其形成机制为血液动力学的刺激、血管活性物质的作用及细胞外基质的调节等引起VSMC的增殖、凋亡与重塑[1].它既是高血压状态下机体的适应性行为 又是高血压病持续恶化的病理学基础.故对高血压病的治疗并不能仅仅局限于降压 还应着眼于逆转高血压血管重构(VR).ATⅡ是影响VR的肾素-血管紧张素-醛固酮(RAAS)系统中起主要作用的重要血管活性物质 通过增高蛋白激酶C(PKC)活性诱导多种生长因子(PDGF、TGF-β等)的基因表达 使上述生长因子合成增加 并通过此途径影响VSMC外基质的构成和VSMC的生长、增殖及表型转换 从而进一步影响血管的组织结构.[2]TMN是临床治疗高血压病的验方.本实验通过观察TMN对SHR胸主动脉血管重构的影响 探讨其作用的ATⅡ机理.     

从CaN信号通路探讨川芎嗪对AngⅡ诱导的大鼠VSMCs增殖的抑制作用及机制

目的从钙调神经磷酸酶(CaN)信号通路探讨川芎嗪对血管紧张素Ⅱ(AngⅡ)诱导的大鼠血管平滑肌细胞(VSMCs)增殖的抑制作用及机制。方法以AngⅡ诱导体外培养的大鼠主动脉平滑肌细胞建立细胞增殖模型,实验分为空白组、AngⅡ组、AngⅡ 川芎嗪(低、中、高)剂量组。采用定磷法分别测定各组细胞CaM和CaN活性,MTT法检测细胞增殖活度的变化,免疫组化定量技术观察细胞内c-myc、PCNA的表达水平。结果AngⅡ组VSMCs细胞增殖活度(吸光度值)、胞内CaM、CaN活性以及c-myc、PCNA表达量(光密度值)显著高于空白组(P<0.01);同时加入不同剂量(低、中、高剂量)川芎嗪温育,各组CaM、CaN活性以及c-myc、PCNA表达水平均显著低于AngⅡ组(P<0.05或P<0.01)。结论川芎嗪可通过降低细胞内CaM、CaN活性,下调胞内c-myc、PCNA表达水平,从而显著抑制AngⅡ诱导的VSMCs增殖。     

人参三七川芎提取物对增龄和高血压大鼠血管平滑肌细胞的影响

目的 观察和比较增龄与高血压对大鼠主动脉血管平滑肌细胞(vascular smooth muscle cells,VSMCs)的影响及人参、三七、川芎提取物的干预作用。方法 采用原代细胞培养的方法建立大鼠主动脉VSMCs模型,其中增龄实验分5组：青年对照组(Yon组)、老龄组(Old组)、老龄+普罗布考组(Old+Pro组)、老龄+中药低剂量组(Old+Low组)和老龄+中药高剂量组(Old+High组);高血压实验分5组：Wistar-Kyoto对照组(WKY组)、自发性高血压大鼠(spontaneously hypertensive rat,SHR)组、SHR+缬沙坦组(SHR+Val组)、SHR+中药低剂量组(SHR+Low组)和SHR+中药高剂量组(SHR +High组)。采用MTT法检测细胞增殖,免疫细胞化学法检测基质金属蛋白酶-9(MMP-9)表达,RT-PCR和Western blot分别检测过氧化物酶增殖物激活受体γ(peroxisome proliferator activated receptor-γ,PPAR-γ)mRNA和蛋白表达。结果 与Yon组比较,Old组VSMCs增殖水平和MMP-9表达升高,PPAR-γ mRNA和蛋白表达减少,差异均有统计学意义(P<0.05)。与Old组比较,Old+Pro组、Old+High组和 Old+Low组VSMCs增殖水平和MMP-9表达明显降低,PPAR-γ mRNA表达明显增加(均P<0.05),Old+Pro组和Old+Low组PPAR-γ蛋白表达明显增加(均P<0.05)。与WKY组比较,SHR组VSMCs增殖水平和MMP-9表达明显升高,PPAR-γ mRNA和蛋白表达明显减少(均P<0.05);与SHR组比较,SHR+Val组、SHR +High组和SHR+Low组VSMCs增殖水平和MMP-9表达明显降低,PPAR-γ mRNA和蛋白表达增加(均P<0.05)。结论 增龄和高血压均可造成大鼠主动脉VSMCs过度增殖及相关细胞活性因子的表达改变,中药人参、三七、川芎提取物能够降低其增殖水平,减少负性细胞因子的表达以降低增龄和高血压对VSMCs的损害,延缓血管细胞的老化。     

Effects of Tilianin on Proliferation,Migration and TGF‐β/Smad Signaling in Rat Vascular Smooth Muscle Cells Induced with Angiotensin II

Flavonoid Tilianin was isolated from Dracocephalum moldavica, and its pharmacological mechanism on proliferation, migration and the TGF‐β/Smad signaling pathway in rat vascular smooth muscle cells (VSMCs) induced with Angiotensin II (Ang II) was systematically evaluated. Primary rat VSMCs were stimulated with Ang II to induce proliferation. The cells were then treated with Tilianin for 24 or 48 h. MTT assay and Transwell assays were used to evaluate the effects of Tilianin on proliferation and migration. The expression of intracellular proliferating cell nuclear antigen (PCNA), intercellular adhesion molecule‐1 (ICAM‐1) and vascular cell adhesion molecule‐1 (VCAM‐1) were measured by immunohistochemistry as verification of effects on proliferation and migration. The expression of TGF‐β1, Smad2 and Smad3 mRNA was measured by qRT‐PCR, and the expression of TGF‐β1 and P‐Smad2/3 protein was measured by Western blotting. The results show that Tilianin can inhibit proliferation and expression of intracellular PCNA in VSMCs induced with Ang II, in a dose‐dependent manner. Tilianin also mediates a dose‐dependent inhibition of migration and the expression of intracellular ICAM‐1, VCAM‐1, MMP‐2 and MMP‐9. Furthermore, TGF‐β1, Smad2, Smad3, Smad2/3 and P‐Smad2/3 in Ang II‐induced VSMCs are suppressed by Tilianin. The inhibitory effects of Tilianin support its use in the suppression and treatment of atherosclerosis. Copyright © 2017 John Wiley & Sons, Ltd.     

辛通畅络中药复方对大鼠血管平滑肌细胞迁移的影响

[目的]观察辛通畅络中药复方对血管平滑肌细胞(VSMCs)迁移的影响。[方法]以含辛通畅络中药处方不同浓度(10%、20%、30%)的大鼠血清培养VSMCs,以正常大鼠空白血清为对照,采用Boyden小室法及划痕实验观察辛通畅络中药血清对VSMCs迁移的影响;采用Western blot检测MMP-2、MMP-9蛋白表达情况。[结果]辛通畅络中药复方含药血清可以抑制VSMCs的迁移及MMP-2、MMP-9蛋白的表达,并且这种抑制效应呈浓度依赖性。[结论]辛通畅络中药复方具有抑制VSMCs迁移的作用,其作用机制可能与影响MMP-2和MMP-9的表达有关。     

Crocetin inhibits cell cycle G1/S transition through suppressing cyclin D1 and elevating p27kip1 in vascular smooth muscle cells

Crocetin is a natural carotenoid compound isolated from Gardenia jasminoids Ellis. Our previous study shows that crocetin inhibits angiotensin II (Ang II)‐induced vascular smooth muscle cells (VSMCs) proliferation. To further explore the mechanism by which crocetin inhibits VSMCs proliferation, in the present study we examined the effect of crocetin on cell cycle progression and cell cycle regulatory proteins. Flow cytometry analysis showed that Ang II elicited significant increase in the percentage of VSMCs in the S phase, with a concomitant decline in the percentage of VSMCs in the G₀/G₁ phase. However, on pretreatment of VSMCs with crocetin, the percentage of VSMCs in the S phase decreased, while that in the G₀/G₁ phase increased significantly. In addition, Ang II‐induced increase of cell proliferation index was also decreased by crocetin. Western blotting analysis indicated that crocetin markedly inhibited the protein expression of cyclin D1 but not cyclin E. Crocetin also increased the level of cyclin‐dependent kinase inhibitor (CDKI) p27^kip1 but not CDKI p21^waf1/cip1. In conclusion, our present results suggest that the inhibition of cell cycle G₁/S transition in VSMCs by crocetin can be attributed, at least in part, to its suppression of cyclin D1 and elevation of CDKI p27^kip1. Copyright © 2009 John Wiley & Sons, Ltd.     

钩藤碱对自发性高血压大鼠的降压作用及其对血管的调节机制探讨

目的研究钩藤碱连续给药后对自发性高血压大鼠(SHR)收缩压(SBP)的影响及其对血管的调节机制。方法取雄性SHR,随机分为模型组(等容量蒸馏水),卡托普利阳性对照组(6.25 mg/kg),钩藤碱低、中、高剂量(1.25、2.50、5.00 mg/kg)组,另取SD雄性大鼠作为对照组(等容量蒸馏水),各给药组连续ig给药21 d,测定给药前和给药第7、14、21天的大鼠尾动脉SBP,末次给药后采血,检测血浆血管紧张素II(Ang II)、非对称二甲基精氨酸(ADMA)、血管紧张素II 1型受体(AT1R)和血清NO、一氧化氮合酶(NOS)水平。结果与模型组比较,钩藤碱各剂量组可显著降低SHR的SBP值;模型组大鼠血浆Ang II、ADMA、AT1R水平显著升高,且血清中NO和NOS水平显著下降,而钩藤碱能够逆转上述指标的改变(P<0.05、0.01)。结论钩藤碱对SHR有显著降低SBP作用,能降低血浆Ang II、ADMA水平,升高血清NO和NOS水平,有保护血管内皮功能的作用。     

Protein kinase C pathway is involved in the inhibition by crocetin of vascular smooth muscle cells proliferation

Crocetin is a natural carotenoid compound isolated from Gardenia jasminoids Ellis. Our previous study showed that crocetin inhibits angiotensin II (Ang II)-induced proliferation of vascular smooth muscle cells (VSMCs). The present study investigated the involvement of the protein kinase C (PKC) pathway in the growth-inhibitory action of crocetin in VSMCs. The findings showed that PKC activity in the membrane fraction of VSMCs increased following stimulation with Ang II, which was suppressed significantly by pretreating the cells with crocetin. Inhibition of PKC activity by crocetin appeared to be associated with growth inhibition in VSMCs, because chelerythrine chloride, a specific PKC inhibitor, likewise decreased cell proliferation. PKC-a, a conventional PKC isoform, was detected in bovine aorta VSMCs by RT-PCR and western blotting analysis. Crocetin inhibited Ang II-induced membrane translocation of PKC-a, and the inhibition of crocetin on PKC activity in membrane fraction coincided with its suppression on membrane translocation of PKC-a. In addition, Ang II-induced mRNA expressions of c-fos, c-jun and c-myc were also decreased by crocetin. Taken together, the data suggest that the inhibition by crocetin of PKC activity, at least in part due to inactivation of PKC-a, and the subsequent suppression of proto-oncogene expressions might mediate its inhibitory effect on VSMCs proliferation.     

Inhibitory effects of emodin on the proliferation of cultured rat vascular smooth muscle cell-induced by angiotensin II

Rhubarb, used as a traditional Chinese medicine for centuries, offers therapeutic potential for cardiovascular and other diseases. Emodin, extracted from the root extract of rhubarb has sparked increasing interest for therapeutic application. The main objective was to study the effect of emodin on cultured vascular smooth muscle cells (VSMCs) proliferation induced by angiotensin II (Ang II) and the expression of proto-oncogene c-myc. VSMCs were cultured by the explant method, then incubated for 24, 48 and 72 h with emodin (10-80 microm) and Ang II, or were left untreated (control). Cell proliferation was measured by MTT assay and immunohistochemical staining for proliferating cell nuclear antigen (PCNA), respectively. The expression of c-myc was measured by immunohistochemical staining and image analysis technique. Ang II increased the cell proliferation compared with the control group (p < 0.01). The expression of PCNA and c-myc was increased compared with the control group (p < 0.01). After pretreatment with emodin, the above indexes were obviously reduced compared with the Ang II group (p < 0.01). These findings suggested that emodin inhibited VSMCs proliferation induced by Ang II. Inhibition of the expression of c-myc might be correlated with the inhibitory effects.     

槲皮素对血管紧张素Ⅱ诱导大鼠胸主动脉平滑肌细胞IL-6的影响

目的:观察槲皮素对血管紧张素Ⅱ(AngⅡ)诱导大鼠胸主动脉平滑肌细胞产生IL-6的影响。方法:以不同浓度AngⅡ刺激培养的大鼠胸主动脉平滑肌细胞,并以不同剂量的槲皮素干预24 h,用ELISA法检测不同时间点上清液中IL-6含量,用半定量RT-PCR法检测槲皮素对IL-6 mRNA表达的影响。结果:AngⅡ刺激血管平滑肌细胞产生IL-6具有剂量和时间依赖性。槲皮素对不同浓度AngⅡ刺激血管平滑肌细胞产生IL-6有明显剂量依赖性抑制作用,1000μmol/L时作用最为明显;不同浓度槲皮素均可下调AngⅡ(10-7M)所诱导的血管平滑肌细胞IL-6mRNA的表达,浓度为1000μM时作用最明显。结论:槲皮素可在蛋白和转录水平下调AngⅡ诱导血管平滑肌细胞IL-6的产生,提示槲皮素的抗动脉粥样硬化作用可能与其抗炎作用有关。     

Antihypertensive Effects of Extract from Flower Buds of Coreopsis tinctoria on Spontaneously Hypertensive Rats

Objective To provide experimental evidence for the antihypertensive activity of the flavonoids in flower buds of Coreopsis tinctoria(CT-F)and to investigate the underlying mechanism.Methods The spontaneously hypertensive rats(SHRs)were divided into model,captopril(positive control),and CT-F groups,and the Wistar-Kyoto rats were set as control group,eight in each group.The blood pressure of SHRs,the activity of angiotensin II(Ang-II)in plasma,nitric oxide(NO),malondialdehyde(MDA)and thoracic aorta media thickness in SHRs were measured by tail-cuff method,radioimmunity method,nitrate reductase method,thibabituric acid(TBA)method,and the hematoxylin-eosin staining method.Q-PCR analysis was performed to determine the relative quantity of angiotensin-converting enzyme(ACE),ACEII,angiotensin type 1receptor(AT1R),and TGF-β1 mRNA in left ventricle.Results CT-F could lower the systolic blood pressure of SHRs dramatically(P0.01).The levels of MDA in serum and Ang II in plasma of SHRs treated with CT-F decreased markedly(P0.05,0.01),the level of NO in serum increased significantly(P0.01).In addition,thoracic aorta media thickness in SHRs treated with CT-F was thinner than that of the model group(P0.05).The mRNA expression of ACE,AT1R,and TGF-β1 in left ventricle was markedly decreased(P0.05),while that of ACE II was increased(P0.05).Conclusion CT-F is effective to lower the blood pressure of SHRs,and its antihypertensive effect is probably associated with lowering the oxidative stress by reducing MDA,ameliorating aorta remodeling,dilating vessel by increasing NO and decreasing Ang-II,and regulating the expression of rennin-angiotensin System-related various genes.     

蒙花苷与木犀草素联用对离体血管的舒张作用及其机制研究

为研究蒙花苷与木犀草素联用对SD大鼠离体血管环的舒张作用及其作用机制。采用离体血管环张力实验观察蒙花苷单用(7.5×10~(-6)g·mL~(-1))、木犀草素单用(7.5×10~(-6)g·mL~(-1))、蒙花苷与木犀草素联用(1∶4)对去甲肾上腺素引起的SD大鼠完整、去内皮、L-NAME及吲哚美辛预处理胸主动脉收缩的影响;Western blot检测胸主动脉p-Akt,p-eNOS等蛋白水平。实验结果显示,蒙花苷与木犀草素联用对完整、去内皮及L-NAME预处理的血管环舒张率高于两者单用;且两者联用能显著促进血管中p-Akt,p-eNOS蛋白表达。实验结果说明蒙花苷与木犀草素联用能有效舒张血管,其机制可能是通过激活PI3K/Akt/NO信号通路,增强eNOS活性,从而增加NO的合成与释放,达到舒张血管的作用。