肝素十二糖对血管平滑肌细胞增殖作用的研究

目的:研究肝素十二糖(dp12)对血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖作用的影响,并分析其分子水平的作用机制。方法:通过以10%新生牛血清诱导牛胸主动脉血管平滑肌细胞增殖建立模型,然后考察实验室精制的不同浓度的肝素寡糖对血管平滑肌细胞增殖作用的影响;四甲基偶氮唑盐法(MTT)检测dp12对VSMCs增殖的影响;流式细胞仪检测细胞周期分布;RT-PCR法检测ERK1/2转录变化情况;Western Blotting法检测ERK1/2和p-ERK1/2的表达。结果:MTT结果显示不同浓度的dp12可以明显抑制由10%新生牛血清诱导的血管平滑肌细胞的增殖;细胞周期实验揭示dp12抑制血清诱导的VSMCs从G1期进入S期,影响细胞周期;dp12通过下调ERK基因的转录进而下调ERK1/2的表达,此外dp12抑制ERK1/2的磷酸化进而影响细胞增殖。结论:dp12使VSMCs在G1期/S期阻滞,通过抑制ERK基因的转录和ERK蛋白的磷酸化抑制VSMCs增殖,可能是dp12抗VSMCs增殖的机制之一。     

羟基红花黄色素A通过影响PCNA表达和MEK-ERK1/2信号通路抑制大鼠血管平滑肌细胞增殖

目的探讨羟基红花黄色素A(HYSA)影响增殖细胞核抗原(PCNA)表达和MEK-ERK1/2信号通路抑制血管平滑肌细胞(VSMCs)增殖的分子机制。方法采用贴块法培养大鼠血管平滑肌细胞,用血小板源生长因子(PDGF)诱导其增殖,采用MTT法检测HYSA对平滑肌细胞增殖的抑制作用;采用Western blot和免疫组织化学方法检测HYSA对PCNA表达及对MEK-ERK1/2信号通路的影响。结果HYSA浓度依赖性地抑制PDGF诱导的VSMC增殖,降低PCNA的表达,阻断PDGF受体的激活和MEK/ERK信号通路的活化,但不影响JNK和p38MAPK的活性。结论 HYSA通过降低PCNA表达和阻断MEK-ERK1/2信号通路抑制大鼠血管平滑肌细胞增殖。     

油酰乙醇胺对血管紧张素Ⅱ诱导的血管平滑肌细胞增殖的影响

目的研究油酰乙醇胺(OEA)对血管紧张素Ⅱ(AngⅡ)刺激大鼠主动脉血管平滑肌细胞(VSMCs)增殖的抑制作用以及对p38信号通路的影响。方法分离大鼠胸主动脉,组织贴块法培养VSMCs,AngⅡ刺激VSMCs建立细胞增殖模型,不同浓度OEA(5,10,20μmol/L)作用后,用溴脱氧核苷尿嘧啶(BrdU)掺入的方法检测细胞增殖活性;流式细胞术检测细胞周期变化;Western-bolt法检测对P-p38和p38蛋白表达的影响。结果与AngⅡ组比较,随着OEA浓度升高,VSMCs的增殖受到抑制、G0/G1比例显著升高,G2/M比例显著降低,且P-p38和p38蛋白的表达量降低并呈浓度依赖关系。结论 OEA对VSMCs的增殖有抑制作用,其机制可能是抑制了p38MAPK信号通路。     

Apelin-13促大鼠血管平滑肌细胞增殖的PKC-ERK1/2信号通路研究

目的研究G蛋白偶联受体APJ(血管紧张素受体样受体或称血管紧张素受体AT1相关的受体蛋白,putativere-ceptor protein related to the angiotensin receptor AT1)的内源性配体apelin-13通过PKC-ERK1/2-Cyclins信号通路促进大鼠血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖的影响。方法培养SD大鼠胸主动脉VSMCs,Western blot检测p-ERK1/2、ERK1/2、细胞周期蛋白CyclinD1和CyclinE的表达,四噻唑蓝比色法观察PKC阻断剂GF109203X对apelin-13促大鼠VSMCs增殖的影响。结果Apelin-13剂量依赖性和时间依赖性地促进大鼠VSMCsp-ERK1/2表达增加,对ERK1/2表达没有明显影响,GF109203X可明显抑制apelin-13诱导的细胞增殖及p-ERK1/2、CyclinD1和CyclinE的表达。结论Apelin-13促进大鼠VSMCs增殖可能与ape-lin-APJ-PKC-ERK1/2-Cyclins信号通路有关。     

杜鹃素通过降低Cx43的表达抑制AngⅡ诱导的VSMCs增殖

目的研究Cx43在杜鹃素抑制血管紧张素Ⅱ(AngⅡ)诱导的血管平滑肌细胞(vascular smooth muscle cells,VSMCs)增殖中的作用。方法体外培养原代大鼠VSMCs细胞,随机分为对照组、AngⅡ刺激组(模型组)、AngⅡ+杜鹃素组(给药组)。CCK-8法检测细胞活力,Ed U法检测细胞增殖活性,流式细胞术观察细胞周期,real-time PCR和Western blot法检测细胞中Cx43的表达。用si RNA干扰Cx43的表达并测定细胞Ed U结合率和细胞周期。结果浓度为60μmol·L^-1的杜鹃素可使AngⅡ诱导的大鼠VSMCs的细胞增殖活性和Ed U结合率均明显降低(P<0.05),并阻止VSMCs由G0/G1期向S期的转化。Real-time PCR和Western blot结果显示,与模型组比较,杜鹃素能明显降低Cx43的m RNA和蛋白表达水平(P<0.01)。用si RNA干扰Cx43的表达后,杜鹃素对AngⅡ诱导的VSMCs增殖的抑制作用明显减弱。结论杜鹃素可明显抑制AngⅡ诱导的VSMCs增殖,抑制VSMCs有丝分裂,使其停滞于G_0/G_1期,其作用可能与杜鹃素抑制Cx43的表达有关。     

拉马克啉对内皮素-1诱导的大鼠血管平滑肌细胞增殖及蛋白激酶C表达的作用

目的：探讨拉马克啉（levcromakalim）对内皮素-1（ET-1）诱导的大鼠血管平滑肌细胞增殖及蛋白激酶C（protein kinase C，PKC）表达的影响。方法：体外培养Wistar大鼠主动脉血管平滑肌细胞（vascular smooth muscle cell，VSMCs），用ET-1诱导其增殖，用不同浓度拉马克啉共培养，MTT法评价VSMCs增殖情况，3H—TdR检测DNA合成，流式细胞术检测VSMCs凋亡，逆转录聚合酶链式反应（RT-PCR），Western blot检测PKCamRNA及蛋白表达。结果：拉马克啉对ET-1所致VSMCs增殖有显著抑制作用，随浓度增加，其MTT活性细胞含量和3H—TdR掺入量都明显减少（P〈0．05）；拉马克啉呈剂量依赖性地增加G0／G1期VSMCs（P〈0．05），促使VSMCs凋亡增多（P〈0．05）；拉马克啉抑制VSMCs内PKCamRNA及蛋白表达。结论：拉马克啉抑制ET-1诱导的VSMCs增殖作用，可能的机制是通过下调VSMCs内PKCa表达水平而影响vSMCs增殖和凋亡。     

Apelin促大鼠血管平滑肌细胞增殖的PI3K／Akt信号通路研究

目的：研究G蛋白偶联受体APJ的内源性配体多肽apelin-13是否通过P13K／Akt信号通路影响血管平滑肌细胞（vascular smooth muscle cells,VSMCs）的牛长增殖,发现apelin—APJ系统促血管平滑肌细胞增殖的新的信号转导机制;方法：培养大鼠胸主动脉血管平滑肌细胞,噻唑蓝比色法（MTT）观察VSMCs增殖;Westernblot检测P13K等信号蛋门表达。结果：Western blot检测结果显尔,apelin-13（0、0．5、1、2、4gmol／L）刺激大鼠血管平滑肌细胞P13K磷酸化、Akt磷酸化增加,以1p．M最为明显;lgMapelin-13分别刺激大鼠VSMCs0、5、15、30、45、60min,P13K磷酸化、Akt磷酸化表达在30min时最为明显,P13K阻断剂LY294002明显抑制apelin-13诱导的P13K磷酸化及Akt磷酸化表达,Akt阻断剂1701．1明显抑制apelin—13诱导的Akt磷酸化、ERK1／2磷酸化及cyclinDl表达,MTT法显示P13K抑制剂LY294002和Akt抑制剂1701—1能明显抑制apelin--13诱导的VSMCs增殖;结论：Apelin-13通过P13K／Akt信号转导通路促进大鼠血管平滑肌细胞增殖。     

氯沙坦抑制ROS/ERK1/2信号途径减轻AGEs诱导的大鼠脑血管平滑肌细胞增殖

目的探讨果糖制备的糖基化终末产物(AGEs)对大鼠脑基底动脉血管平滑肌细胞(BASMC)的增殖以及氯沙坦对增殖的影响和机制。方法培养大鼠脑基底动脉血管平滑肌细胞,用CCK8法测定不同浓度(1,5,10μmol/L)氯沙坦对果糖制备的AGEs(200mg/L)预处理48h诱导血管平滑肌细胞增殖的影响。用western blotting检测AGEs以及氯沙坦处理后对ERK蛋白表达影响。结果 AGEs刺激血管平滑肌细胞增殖(0.827±0.069比0.364±0.052,P<0.01)。给予氯沙坦(5,10μmol/L)后,细胞增殖明显下降(0.658±0.064和0.381±0.058比0.827±0.069,P<0.01)。AGEs还使平滑肌细胞内活性氧簇(ROS)生成增多,并显著增加ERK1/2蛋白表达,氯沙坦则明显抑制ROS和ERK1/2蛋白水平。结论果糖制备的AGEs诱导脑基底动脉血管平滑肌细胞增殖,氯沙坦可能通过阻断AT1受体而抑制ROS和ERK1/2表达,减少平滑肌细胞的异常增殖。     

粉防己碱对增生性瘢痕成纤维细胞增殖的影响

［摘要］目的观察粉防己碱（tetrandrine, Tet）对增生性瘢痕成纤维细胞增殖及MEK/ERK、MKP 1信号通路的影响。方法培养增生性瘢痕成纤维细胞，四甲基偶氮唑蓝（MTT）比色法、［3H］ TdR掺入率测定不同浓度Tet对细胞增殖的抑制作用；免疫沉淀法纯化蛋白并细胞外信号调节激酶（ERK）试剂盒测定ERK活性；Western blot测定p MEK1/2、p ERK1/2及蛋白激酶磷酸酶 1（MKP 1）表达。结果MTT实验、［3H］ TdR掺入率均显示Tet明显抑制细胞增殖，同时Tet明显抑制ERK活性，Western blot显示Tet明显抑制p MEK1/2及p ERK1/2表达，同时提高MKP 1表达。结论粉防己碱明显抑制增生性瘢痕成纤维细胞增殖，机制与下调ERK活性，抑制MEK/ERK通路及上调MKP 1表达有关。     

肝素寡糖通过抑制PKC-α影响血管平滑肌细胞增殖的作用(英文)

为考察肝素寡糖对血管平滑肌细胞增殖的抑制作用,并探索其相关细胞内信号转导机制,采用cell-based ELISA法、RT-PCR法、Western blotting法和免疫细胞化学法检测细胞内PKC-α蛋白水平和mRNA水平;采用RT-PCR法检测c-jun、c-myc和c-fos等3种原癌基因的mRNA水平。结果显示,肝素寡糖可以抑制新生牛血清诱导的PKC-α和原癌基因的表达,这可能是肝素寡糖抑制血管平滑肌细胞增殖的机制之一;流式细胞实验结果显示,肝素寡糖能将细胞阻滞于G0/G1期,从而阻断细胞周期进程。综上所述,肝素寡糖可能通过抑制PKC-α表达,进而抑制原癌基因的表达,阻断细胞G1/S期转换,从而抑制血管平滑肌细胞的增殖。     

High Glucose Induces Connective Tissue Growth Factor Expression and Extracellular Matrix Accumulation in Rat Aorta Vascular Smooth Muscle Cells Via Extracellular Signal-Regulated Kinase 1/2

Connective tissue growth factor (CTGF) is a potent pro-fibrotic factor, which is implicated in fibrosis through extracellular matrix (ECM) induction in diabetic cardiovascular complications. It is an important downstream mediator in the fibrotic action of transforming growth factor β (TGFβ) and is potentially induced by hyperglycemia in human vascular smooth muscle cells (VSMCs). Therefore, the goal of this study is to identify the signaling pathways of CTGF effects on ECM accumulation and cell proliferation in VSMCs under hyperglycemia. We found that high glucose stimulated the levels of CTGF mRNA and protein and followed by VSMC proliferation and ECM components accumulation such as collagen type 1, collagen type 3 and fibronectin. By depleting endogenous CTGF we showed that CTGF is indispensable for the cell proliferation and ECM components accumulation in high glucose-stimulated VSMCs. In addition, pretreatment with the MEK1/2 specific inhibitors, PD98059 or U0126 potently inhibited the CTGF production and ECM components accumulation in high glucose-stimulated VSMCs. Furthermore, knockdown with ERK1/2 MAPK siRNA resulted in significantly down regulated of CTGF production, ECM components accumulation and cell proliferation in high glucose-stimulated VSMCs. Finally, ERK1/2 signaling regulated Egr-1 protein expression and treatment with recombinant CTGF reversed the Egr-1 expression in high glucose-induced VSMCs. It is conceivable that ERK1/2 MAPK signaling pathway plays an important role in regulating CTGF expression and suggests that blockade of CTGF through ERK1/2 MAPK signaling may be beneficial for therapeutic target of diabetic cardiovascular complication such as atherosclerosis.     

离体和在体机械损伤致血管平滑肌增殖模型的建立

目的建立离体和在体机械损伤致血管平滑肌增殖模型。方法原代培养大鼠胸主动脉血管平滑肌细胞(VSMCs),体外建立机械划伤致VSMCs增殖模型,采用5-溴脱氧尿嘧啶(5-BrUd)掺入法,流式细胞术检测细胞增殖情况;整体动物上,采用新西兰兔左侧颈总动脉球囊损伤,术后7d取颈总动脉段,常规病理切片,HE染色,观测血管内膜厚度、内膜面积、内膜厚度/中膜厚度、内膜面积/中膜面积。结果机械划痕损伤12 h后划伤处两侧的细胞开始有部分增殖和迁移,24 h后细胞增殖较为明显。流式细胞术检测机械损伤后细胞增殖增多(P<0.05)。整体动物上,与假手术组比较,模型组术后7 d血管内膜厚度、内膜面积、内膜厚度/中膜厚度、内膜面积/中膜面积显著增加(P<0.01)。结论机械损伤可以导致离体和在体血管平滑肌细胞增殖。     

Corynoxeine isolated from the hook of Uncaria rhynchophylla inhibits rat aortic vascular smooth muscle cell proliferation through the blocking of extracellular signal regulated kinase 1/2 phosphorylation

The proliferation of vascular smooth muscle cells (VSMCs) induced by injury to the intima of arteries is an important etiologic factor in vascular proliferative disorders such as atherosclerosis and restenosis. Uncaria rhynchophylla is traditional Chinese herb that has been applied to the treatment of convulsive disorders, such as epilepsy, in China. In the present study, we examined whether corynoxeine exerts inhibitory effects on platelet-derived growth factor (PDGF)-BB-induced rat aortic VSMC proliferation and the possible mechanism of such effects. Pre-treatment of VSMCs with corynoxeine (5-50 microM) for 24 h resulted in significant decreases in cell number without any cytotoxicity; the inhibition percentages were 25.0+/-12.5, 63.0+/-27.5 and 88.0+/-12.5% at 5, 20 and 50 microM, respectively. Also, corynoxeine significantly inhibited the 50 ng/ml PDGF-BB-induced DNA synthesis of VSMCs in a concentration-dependent manner without any cytotoxicity; the inhibitions were 32.8+/-11.0, 51.8+/-8.0 and 76.9+/-7.4% at concentrations of 5, 20 and 50 microM, respectively. Pre-incubation of VSMCs with corynoxeine significantly inhibited PDGF-BB-induced extracellular signal-regulated kinase 1/2 (ERK1/2) activation, whereas corynoxeine had no effects on mitogen-activated protein kinase (MAPK/ERK)-activating kinase 1 and 2 (MEK1/2), Akt, or phospholipase C (PLC)gamma1 activation or on PDGF receptor beta (PDGF-Rbeta) phosphorylation. These results suggest that corynoxeine is a potent ERK1/2 inhibitor of key PDGF-BB-induced VSMC proliferation and may be useful in the prevention and treatment of vascular diseases and restenosis after angioplasty.     

Lercanidipine inhibits vascular smooth muscle cell proliferation and neointimal formation via reducing intracellular reactive oxygen species and inactivating Ras-ERK1/2 signaling

Lercanidipine, a calcium channel antagonist, is currently employed in the treatment of essential hypertension and angina pectoris. The purpose of this study was to elucidate the anti-proliferative effect of lercanidipine and to investigate the molecular role of this agent. Both in vitro studies and in a balloon injury rat carotid artery model were employed to study the effect of lercanidipine on smooth muscle cell proliferation. Lercanidipine-inhibited rat vascular smooth muscle cell (VSMC) proliferation and migration in a dose-dependent manner following stimulation of VSMC cultures with 10% fetal bovine serum (FBS) and 20 ng/ml platelet-derived growth factor (PDGF)-BB. FBS- and PDGF-BB-stimulated intracellular Ras, MEK1/2, ERK1/2, proliferative cell nuclear antigen (PCNA), and Akt activations were significantly inhibited by lercanidipine; however, lercanidipine did not affect FBS- and PDGF-BB-induced STAT3 phosphorylation. Lercanidipine also inhibited PDGF-receptor β chain phosphorylation and reactive oxygen species (ROS) production induced by PDGF-BB. Lercanidipine blocked the FBS-inducible progression through the G₀/G₁ to the S-phase of the cell cycle in synchronized cells. In vivo, 14 days after balloon injury, treatment with 3 and 10 mg/kg lercanidipine resulted in significant inhibition of the neointima/media ratio. Suppression of neointima formation by lercanidipine was dependent on its influence on ERK1/2 phosphorylation. These results demonstrate that lercanidipine can suppress the proliferation of VSMCs via inhibiting cellular ROS, Ras-MEK1/2-ERK1/2, and PI3K-Akt pathways, and suggesting that it may have therapeutic relevance in the prevention of human restenosis.     

Inhibition of reactive oxygen species/extracellular signal-regulated kinases pathway by pioglitazone attenuates advanced glycation end products-induced proliferation of vascular smooth muscle cells in rats

Advanced glycation end products (AGEs) have been shown to induce the proliferation of vascular smooth muscle cells (VSMCs) and contribute to atherogenesis and diabetes. In the present study, we investigated the effects of pioglitazone, a peroxisome proliferator activated receptor gamma (PPARγ) agonist, on AGE-induced rat VSMC growth and the underlying mechanism. In cultured rat VSMCs, AGE treatment induced VSMC proliferation in time- and dose-dependent manner, while down-regulated the expression of PPARγ. Pretreatment of pioglitazone not only prevented the down-regulation of PPARγ, but inhibited VSMC proliferation and prevented S-phase entry of cell via a G0-G1 block in the presence of AGEs. Western blotting analysis showed that AGE treatment potentiated to activate extracelluar signal-regulated kinases (ERK1/2) by the induction of intracellular reactive oxygen species (ROS) production, since ROS scavenger N-acetyl-L-cysteine pretreatment significantly inhibited AGE-induced ERK1/2 activation. Further, pretreatment with either N-acetyl-L-cysteine or the inhibitor of ERK1/2 activation suppressed AGE-induced proliferation of VSMCs, suggesting a role of ROS/ERK1/2 signaling. Notably, we demonstrated that pretreatment of pioglitazone significantly attenuated AGE-induced ROS and ERK1/2 activation. Collectively, these results suggest that pioglitazone inhibits AGE-induced VSMC proliferation via increasing PPARγ expression and inhibiting ROS/ERK1/2 signaling pathway.     

JM91, a newly synthesized indoledione derivative, inhibits rat aortic vascular smooth muscle cells proliferation and cell cycle progression through inhibition of ERK1/2 and Akt activations

The increased potential for growth of vascular smooth muscle cells (VSMCs) is a key abnormality in the development of atherosclerosis and postangioplasty restenosis. Platelet-derived growth factor (PDGF)-BB is a potent mitogen for VSMCs that plays an important role in the intimal accumulation of VSMCs. This study examined the effect of JM91, a newly synthesized indoledione derivative, on the proliferation of PDGF-BB-stimulated rat aortic VSMCs. The antiproliferative effect of JM91 on rat aortic VSMCs was examined by cell counting and [(3)H]thymidine incorporation assay. The pre-incubation of JM91 (0.5-3.0 microM) significantly inhibited the proliferation and DNA synthesis of 25 ng/mL PDGF-BB-stimulated rat aortic VSMCs in a concentration-dependent manner. JM91 inhibited the PDGF-BB-stimulated phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt kinase, while had no effect on PLCgamma1 and PDGF-Rbeta activation. In addition, treatment with JM91 (0.5-3.0 microM) induced cell-cycle arrest in the G(1) phase, which was associated with the down-regulation of cyclins and CDKs. These findings suggest that the inhibitory effects of JM91 against proliferation, DNA synthesis and cell cycle progression of PDGF-BB-stimulated rat aortic VSMCs are mediated by the suppression of the ERK1/2 and PI3K/Akt signaling pathways. Furthermore, JM91 may be a potential antiproliferative agent for the treatment of atherosclerosis and angioplasty restenosis.     

The effect of 2,3,4′,5-tetrahydroxystilbene-2-0-β-d glucoside on neointima formation in a rat artery balloon injury model and its possible mechanisms

2,3,4′,5-tetrahydroxystilbene-2-0-β-d glucoside (TSG) has been recognized to suppress the proliferation of vascular smooth muscle cells (VSMCs). The aim of the present study was to determine whether TSG inhibits neointimal hyperplasia in a rat carotid arterial balloon injury model. Balloon injury was induced in the left common carotid artery of rats. TSG (30, 60, 120 mg/kg/day) was treated from 3 days prior to, until 14 days after the induction of balloon injury. The ratio of intima-to-media was significantly reduced in the TSG-treated rats at 14 days after the induction of injury, which was associated with reduced expressions of proliferating cell nuclear antigen (PCNA), α-smooth muscle actin (α-SMA) and platelet-derived growth factor-BB (PDGF-BB), as markers of VSMCs proliferation and migration. Additionally, TSG significantly inhibited PDGF-BB induced cell migration in cultured VSMCs. Furthermore, we explored the underlying mechanisms for such effects of TSG. The result showed that TSG markedly reduced balloon injury-induced AKT, extracellular signal-regulated kinase (ERK1/2) and nuclear factor kappaB (NF-κB) activation as well as mRNA expressions of c-myc, c-fos and c-jun, which is important signal pathway for VSMCs proliferation. And in both vivo and vitro model, TSG markedly regulated matrix metalloproteinase-2, 9 expressions and collagen I, III expressions, which are key factors in extracellular matrix for VSMCs migration. These results suggest that the anti-proliferative and anti-migrative effects of TSG on VSMCs could help to explain the beneficial effects of TSG on neointima hyperplasia induced by balloon injury.     

Homocysteine regulates endothelin type B receptors in vascular smooth muscle cells

Vascular smooth muscle endothelin type B (ET_B) receptor is involved in the pathogenesis of cardiovascular diseases (CVDs). Hyperhomocysteinemia is an independent risk factor for CVDs. The present study was designed to examine the hypothesis that homocysteine (Hcy) up-regulates vascular smooth muscle ET_B receptors. In vitro experiments were performed in rat superior mesenteric artery (SMA) and vascular smooth muscle cells (VSMCs). The rat SMA or VSMCs were cultured in serum-free medium for 24 h in the presence and absence of Hcy with or without specific inhibitors for the ERK1/2 signaling pathway and NF-κB. In vivo, the rats received subcutaneous injections of Hcy in the presence or absence of specific inhibitors for the ERK1/2 signaling pathway (U0126) for 3 weeks. Levels of protein expression were determined using Western blot analysis. The contractile responses to sarafotoxin 6c (an ET_B receptor agonist) were studied using a sensitive myograph. The blood pressure of the rats was measured via a noninvasive tail-cuff plethysmography method. The results from in vitro experiments showed that Hcy concentration-dependently increased the ET_B receptor-mediated contractile responses, and up-regulated ET_B receptor expression, in rat SMA. Blockage of the ERK1/2 signaling pathway and NF-κB using the MEK1/2 inhibitor (PD98059 and U0126) or IκB kinase inhibitor (wedelolactone) significantly abolished Hcy-induced up-regulation of ET_B receptor. Finally, we used VSMCs as a cellular model to further validate our finding. In vivo study found that hyperhomocysteinemia up-regulated ET_B receptor expression, and elevated the blood pressure of rats via the ERK1/2 signaling pathway. In conclusion, Hcy up-regulated vascular smooth muscle ET_B receptor via activation of the ERK1/2 signaling pathway and NF-κB.     

Upregulation of cyclooxygenase-2 by motorcycle exhaust particulate-induced reactive oxygen species enhances rat vascular smooth muscle cell proliferation

Long-term exposure to particulate air pollution has been implicated as a risk factor for cardiovascular disease and mortality. Short-term exposure has also been suggested to contribute to complications of atherosclerosis. Aberrant regulation of smooth muscle cell proliferation is thought to associate with the pathophysiology of vascular disorders such as atherosclerosis. In this study, we investigate the influence of organic extracts of motorcycle exhaust particulates (MEPE) on rat vascular smooth muscle cell (VSMC) proliferation and related regulation signaling. Exposure of VSMCs to MEPE (10-100 microg/mL) enhanced serum-induced VSMC proliferation. The expression of proliferating cell nuclear antigen (PCNA) was also enhanced in the presence of MEPE. VSMCs treated with MEPE induced the increase in the extent of cyclooxygenase (COX)-2 mRNA and protein expression and prostaglandin E 2 production, whereas the level of COX-1 protein was unchanged. Moreover, MEPE increased the production of reactive oxygen species (ROS) in VSMCs in a dose-dependent manner. MEPE could also trigger time-dependently extracellular signal-regulated kinase (ERK)1/2 phosphorylation in VSMCs, which was attenuated by antioxidants N-acetylcysteine (NAC) and pyrrolidinedithiocarbamate (PDTC). The level of translocation of nuclear factor (NF)-kappaB-p65 in the nuclei of VSMCs was also increased under MEPE exposure. The potentiating effect of MEPE on serum-induced VSMC proliferation could be abolished by COX-2 selective inhibitor NS-398, specific ERK inhibitor PD98059, and antioxidants NAC and PDTC. Taken together, these findings suggest that MEPE may contribute to the enhancement of the pathogenesis of cardiovascular diseases by augmenting proliferation of VSMCs through a ROS-regulated ERK1/2-activated COX-2 signaling pathway.