降钙素基因相关肽对血管升压素缩血管效应的影响

目的 :探讨降钙素基因相关肽 （CGRP）对血管升压素 （AVP）缩血管效应的影响。方法 :培养大鼠主动脉血管平滑肌细胞 （VSMC） ,给予 AVP和 CGRP干预 ,检测 VSMC的细胞内游离钙离子浓度 （[Ca2 + ] i）。结果 :AVP增加VSMC的 [Ca2 + ] i,而 CGRP明显抑制 AVP的这一作用。结论 :CGRP可能通过抑制 VSMC的 [Ca2 + ] i的升高而发挥其对 AVP所致缩血管效应的抑制作用     

槲皮素对家兔主动脉血管平滑肌细胞胞内游离钙浓度的影响

目的：探讨槲皮素（Ｑｕｅ）对智力这平滑肌细胞胞内游离钙浓度（〖Ｃａ＾２＋）〗ｉ的影响。方法：采用新一代钙荧光探剂Ｆｌｕｏ－３／ＡＭ检测Ｑｕｅ对培养的兔主动脉血管平滑肌细胞（ＡＳＭＣ）〖Ｃａ＾２＋〗ｉ在高Ｋ＾＋、去甲肾上腺素（ＮＥ）、血管紧张素Ⅱ（ＡｎｇⅡ）刺作用下的改变,并与Ｖｅｒａｐａｍｉｌ进行对照研究。结果：Ｑｕｅ（１０＾－６、１０＾－４ｍｏｌ／ｌ）呈剂量依赖性抑制高Ｋ＾＋去极化引起的〖Ｃａ＾     

红细胞抗高血压因子对自发性高血压大鼠血管平滑肌细胞游离钙浓度的影响

观察从人红细胞中提取的抗高血压因子（ＡＨＦ）对自发性高血压大鼠（ＳＨＲ）和正常血压ＷＫＹ大鼠培养的主动脉平滑肌细胞胞内游离Ｃａ２＋浓度（［Ｃａ２＋］ｉ）的影响。结果ＳＨＲ和ＷＫＹ大鼠静息［Ｃａ２＋］ｉ无显著差别。ＫＣｌ（６０ｍｍｏｌ／Ｌ）对ＳＨＲ的激活显著大于ＷＫＹ大鼠，ＡＨＦ（１０－４ｇ／ｍｌ）可明显抑制由高钾诱导的［Ｃａ２＋］ｉ升高，对ＳＨＲ的抑制程度明显高于ＷＫＹ大鼠；去甲肾上腺素（ＮＥ，０．１ｍｍｏｌ／Ｌ）对两种大鼠的激活程度无显著差异。ＡＨＦ（１０－４ｇ／ｍＬ）也可明显抑制由ＮＥ（０．１ｍｍｏｌ／Ｌ）所诱导的ＶＳＭＣ［Ｃａ２＋］ｉ升高，对两种大鼠的抑制程度无显著差异。结论ＡＨＦ的降压作用可能与抑制细胞内［Ｃａ２＋］ｉ升高有关。     

A dihydropyridine calcium channel blocker with phosphodiesterase inhibitory activity: Effects on cultured vascular smooth muscle and cultured heart cells

To define the cellular mechanism of action of a dihydropyridine Ca channel antagonist in an experimental model system devoid of neural influences and reflex effects, we studied the actions of RS93522 on cultured vascular smooth muscle cells and on cultured chick embryo ventricular cells. 45Ca uptake by monolayer cultures of vascular smooth muscle cells was inhibited in a concentration-dependent manner. The IC50 for this effect was 10 nM, similar to that for nifedipine (7 nM) in the same system. 10(-6) M RS93522 inhibited 45Ca uptake more fully than 10(-6) M nifedipine (P less than 0.05). Using an optical-video system, the effect of RS93522 on amplitude of contraction of spontaneously beating cultured ventricular cells was studied. Amplitude of contraction was inhibited with IC50 = 7.9 x 10(-8)M. 45Ca uptake in myocytes was depressed by 15% at 5 min. RS93522 had the additional property of inhibiting phosphodiesterase activity in myocardial homogenates with IC50 = 1.6 x 10(-5)M; the potency and efficacy of phosphodiesterase inhibition was similar to that for milrinone in the same system. As expected of Ca channel antagonists, it has a negative inotropic effect on cultured myocardial cells. The compound also has phosphodiesterase inhibitory activity that possibly may potentiate vasodilatation and ameliorate, in part, negative inotropic effects. Thus, RS93522 has two distinct pharmacodynamic effects in myocytes and is a potent calcium channel blocker.     

TRPV1通道蛋白在呼吸系统的研究进展

瞬时感受器电位离子通道蛋白(transient receptor potential ion channel protein,TRP)是细胞膜或细胞器膜上的一类非选择性Ca2+通道蛋白,通过影响细胞内Ca2+浓度来发挥众多的生理、病理功能.在TRP家族中,瞬时感受器电位香草酸受体1(transient receptor potential vanilloid 1,TRPV1)是目前分布最广、研究最多、最为关注的TRP通道蛋白,多数研究发现TRPV1在炎症的产生和痛觉的传递中发挥重要作用.本文就TRPV1在呼吸系统中的表达、功能调节和临床应用等方面作一综述.     

血管平滑肌细胞凋亡与动脉粥样硬化研究进展

血管平滑肌细胞是构成血管壁的重要组成成分,其凋亡参与了动脉粥样硬化及再狭窄的发生发展,现就最近研究对血管平滑肌细胞凋亡诱导因素、相关基因、及其在动脉粥样硬化发生发展作用作一综述。     

激动网织红核因子相关因子2对大鼠氧化应激所致血管平滑肌细胞损伤的影响

目的探讨激动网织红核因子相关因子2（Nrf2）对氧化应激诱导的大鼠血管平滑肌细胞（VSMCs）损伤的影响。方法原代培养大鼠VSMCs,随机分为4组：对照组、氧化损伤组、Nrf2激动剂组、Nrf2干扰慢病毒组。Western免疫印迹法检NtJNrf2蛋白表达变化,MTT法检测各组细胞活力,Hoechst33342法及AnnexinV／FITC法检测各组细胞凋亡情况。结果荧光显微镜显示,Nrf2干扰慢病毒成功感染VSMCs;Western免疫印迹检测显示,Nrf2干扰慢病毒感染细胞后,Nrf2蛋白表达与对照组比较显著降低（P〈0．05）。与氧化损伤组比较,Nrf2激动剂组VSMCs活力显著增加（P〈0．05）,细胞凋亡显著降低（P〈0．05）;而Nrf2干扰慢病毒组VSMCs活力显著减弱（P〈0．05）,细胞凋亡明显增加（P〈0．05）。结论激动Nrf2能减轻氧化应激引起的VSMCs损伤,这可为VSMCs损伤的防治提出新的研究方向。     

胰岛素对SHR大鼠血管平滑肌细胞体外增殖相关基因表达的影响

目的　探讨胰岛素对体外培养的血管平滑肌细胞 (vascularsmoothmusclecell,VSMC)增殖相关基因表达的影响。方法　用贴块法分离培养VSMC ,应用含 4 4 8个与细胞增殖相关基因靶基因芯片分析大鼠血管平滑肌细胞在胰岛素干预后基因表达的差异 ,RT PCR检测胰岛素作用后VSMC中bFGF、TGFβ、PDGF、MatrixGla和OPN各目的基因mRNA相对表达水平 ,采用免疫组织化学检测胰岛素作用后VSMC肌动蛋白 (α SMactin)。结果　胰岛素组VSMC的3 H TdR掺入值 (cpm)比对照组高 5 4 .6 % (P <0 0 1) ;基因芯片分析发现与细胞增殖相关等 36个基因的mRNA胰岛素干预前后培养的细胞表达差异 (8 0 4 % ) ;RT PCR检测MatrixGla和OPN在培养的VSMC中mRNA的表达量 ,胰岛素组明显高于对照组 (P <0 0 5 ) ,同时bFGF、TGFβ、PDGF的表达也是胰岛素组明显高于对照组 (P <0 0 5 ) ;α SMactin免疫组化结果显示对照组的α SMactin比胰岛素组染色深。结论　提示胰岛素介导大鼠血管平滑肌细胞增殖是多基因效应 ,胰岛素促进了VSMC的增殖与VSMC表型转换有关。     

血管内皮细胞对平滑肌细胞表型转化的影响

目的分析共培养时大鼠血管内皮细胞(VECs)对大鼠血管平滑肌细胞(VSMCs)表型转化的影响。方法 VSMCs直接种植于培养板表面,VECs则种植于与VSMCs相对的浮胶底面。免疫荧光方法观察和鉴定VECs和VSMCs,RT-PCR和共聚焦显微镜分析表型相关基因的表达。结果原代培养的大鼠VECs呈铺路石样形态,vWF染色阳性。原代培养的大鼠VSMCs免疫荧光染色α-SMA呈阳性。RT-PCR检测结果表明,48h共培养组VSMCs的合成表型相关基因CRBP-1、Smemb的表达水平显著高于单独培养组,分别为1.4倍、1.5倍;72h达到峰值,分别为1.7倍、2.1倍,96h开始下降;共培养组中收缩表型标记物Smoothelin-B和SM-MHC的基因表达水平在48h、72h显著低于单独培养组,96hSmoothelin-B却高于单独培养组。单独培养组上述各基因的变化趋势不变或保持稳定。免疫荧光结果显示SM-MHC蛋白表达在共培养组中96h后从下降转为升高(P<0.05)。结论在共培养体系中,血管内皮细胞对血管平滑肌细胞表型转化的作用表现为先促进向合成型转化,96h后促进向收缩型转化。     

miR-155通过调控CaSR表达影响VSMC生长的机制研究

目的 miR-155可以通过干扰血管平滑肌细胞(VMSC)生物活性来抑制动脉粥样硬化斑块的形成,但具体机制不十分清楚.本研究主要观察miR-155对VSMC中钙敏感受体(CaSR)表达的影响,同时探讨CaSR在miR-155调控VSMC生物活性过程中的作用.方法 培养VSMC,将miR-155 mimic、Ad-CaS...     

去除Mfn2基因PKA磷酸化位点对血管平滑肌细胞增殖的影响

目的:研究大鼠线粒体融合素基因2(Mfn2)在去除蛋白激酶A(PKA)磷酸化位点后对大鼠血管平滑肌细胞(VSMC)增殖的影响及其相关的信号通路.方法:构建携带去除PKA磷酸化位点的Mfn2重组腺病毒[Adv-Mfn2-PKA(△)]和携带Mfn2的重组腺病毒(Adv-Mfn2)并感染VSMC.Western blot分析法Mfn2-PKA(△)和Mfn2蛋白的表达;激光共聚焦显微镜观察其亚细胞定位;荧光显微镜观察细胞形态变化;四甲基偶氮唑盐(MTT)法比较其对细胞增殖的影响;Western blot法分析磷酸化ERK1/2(p-ERK1/2)蛋白表达变化.结果:激光共聚焦显微镜示Mfn2-PKA(△)与Mfn2蛋白都主要分布于线粒体外膜;荧光显微镜示Mfn2组细胞数较少,而Mfn2-PKA(△)组与对照组相似;MTT示,Mfn2-PKA(△)抑制VSMC增殖作用较Mfn2显著减弱(P＜0.01),与对照组无显著差异;Western blot结果显示,Mfn2-PKA(△)较Mfn2组p-ERK1/2表达显著升高(P＜0.01),与对照组无明显差异.结论:Mfn2-PKA(△)与Mfn2蛋白一样都定位于线粒体外膜,但抑制VSMC增殖的作用消失,对ERK1/2信号通路无抑制作用.表明PKA磷酸化位点是调控Mfn2抗VSMC增殖的重要功能位点.     

内质网应激与自噬的交互作用对血管钙化的影响

目的证实内质网应激(ERS)和自噬的交互作用对血管钙化(VC)的影响。方法维生素D3肌注和尼古丁灌胃制备大鼠在体血管钙化模型,取主动脉行茜素红染色和钙含量检测,Western blot检测相关蛋白的表达水平。结果与对照组相比,钙化组大鼠主动脉管壁钙沉积显著增加,血管平滑肌细胞(VSMC)收缩表型标志蛋白SM-22α和Calponin表达显著降低,而成骨细胞样表型标志蛋白骨形态发生蛋白2(BMP-2)和Runt相关转录因子2(RUNX2)表达显著升高,ERS标志蛋白葡萄糖调节蛋白(GRP78)和C/EBP同源蛋白(CHOP)以及自噬标志蛋白轻链3(LC3Ⅱ)和Beclin-1表达显著升高。钙化大鼠应用ERS激动剂衣霉素[10μg/(kg·d)]可进一步增加血管壁钙沉积及BMP-2和RUNX2表达水平,而SM-22α和Calponin表达进一步减少,GRP78和CHOP以及LC3Ⅱ和Beclin-1表达水平进一步增加。钙化大鼠应用自噬抑制剂3-甲基腺嘌呤[10 mg/(kg·d)]可降低LC3Ⅱ和Beclin-1水平,同时GRP78和CHOP表达升高,增加血管壁钙沉积及BMP-2和RUNX2表达水平,降低SM-22α和Calponin表达。结论内质网应激与自噬的交互作用影响血管钙化的发展。     

Biology of the Vessel Wall and Atherosclerosis

In humans, atherosclerotic plaques develop within pre-existing diffuse intimal thickenings. The vast majority of cells in these intimal thickenings are smooth muscle, and it is their nodular proliferation, synthesis of extracellular matrix and accumulation of intra- and extracellular lipid which results in the development of the lesion. Two of the crucial initiating or progression events in atherogenesis are the invasion of monocyte/macrophages into the vessel wall as a result of hyperlipidemia, and endothelial denudation or dysfunction with resultant platelet aggregation and release. Products from these cells interact with smooth muscle, modifying its phenotype and influencing its behaviour and response.     

Leptin-enhanced neointimal hyperplasia is reduced by mTOR and PI3K inhibitors

Despite the use of the sirolimus (rapamycin) drug-eluting coronary stent, diabetics are at increased risk of developing in-stent restenosis for unclear reasons. Hyperleptinemia, which often coexists with diabetes and metabolic syndrome, is an independent risk factor for progression of coronary artery disease. It has not been determined whether elevated circulating leptin decreases the efficacy of the sirolimus drug-eluting stent in inhibiting neointimal hyperplasia, the process underlying restenosis after stenting. Here we show that leptin activates the mammalian target of rapamycin (mTOR) signaling pathway in primary murine vascular smooth muscle cells (VSMC) and stimulates VSMC proliferation in a PI3K-dependent fashion. Exogenous leptin, administered at levels comparable to those found in obese humans, promotes neointimal VSMC hyperplasia in a murine femoral artery wire injury model. Leptin significantly increases the dose of the mTOR inhibitor sirolimus that is required for effective inhibition of neointimal formation. Combination therapy with LY294002, a PI3K inhibitor, and sirolimus effectively inhibits leptin-enhanced neointimal hyperplasia. These data show that, in the setting of hyperleptinemia, higher doses of an mTOR inhibitor, or combination therapy with mTOR and PI3K inhibitors, inhibits neointimal hyperplasia after arterial injury. These studies may explain the higher rates of restenosis observed in diabetics treated with a sirolimus-eluting coronary stent and suggest a potential novel therapeutic approach for inhibiting in-stent restenosis in such patients.     

多巴胺受体与动脉粥样硬化

内皮细胞损伤,血液循环中的单核细胞迁移到内皮下分化为巨噬细胞,巨噬细胞及血管平滑肌细胞吞噬化学修饰的脂蛋白转变为泡沫细胞,巨噬细胞和平滑肌细胞的迁移、增生及炎症因子的释放等过程被认为是动脉粥样硬化的主要病理生理过程。多巴胺受体可以通过影响平滑肌细胞、内皮细胞的功能等直接或间接参与动脉粥样硬化的上述病理生理过程,对动脉粥样硬化具有保护作用。现探讨多巴胺受体在动脉粥样硬化这一病理生理过程中的作用。     

Evidence for the expression of transient receptor potential proteins in guinea pig airway smooth muscle cells

OBJECTIVE: The present study investigates the expression of transient receptor potential (TRPC) proteins in airway smooth muscle (ASM) cells in order to determine whether these proteins may be candidate molecular counterparts of plasma membrane Ca2+-permeable channels involved in the contraction of ASM. METHODS: Expression of TRPC mRNA was detected using specific primers and RT-PCR. Expression of the TRPC1, TRPC3 and TRPC6 proteins was detected using antibodies in immunoprecipitation and Western blot. RESULTS: Guinea pig ASM cells exhibited thapsigargin- and acetylcholine-initiated Ca2+ inflow but none by 1-oleoyl-2-acetyl-sn-glycerol. mRNA encoding each of the TRPC1 to TRPC6 proteins was detected in ASM cells. mRNA encoding TRPC1, TRPC3, TRPC4 and TRPC6 was detected in ASM cells at a concentration approximately equivalent to that in guinea pig brain. mRNA encoding TRPC2 and TRPC5 was more abundant in ASM cells than in brain. The TRPC1 protein, but not the TRPC3 or TRPC6 proteins, was detected in extracts of ASM cells, while all three proteins were detected in brain. CONCLUSION: The results provide evidence for a low level of expression of the TRPC1 to TRPC6 proteins in ASM cells. These proteins may function as store-operated Ca2+ and/or second messenger-activated non-selective cation channels in ASM cells.