卡格列净改善自发性高血压大鼠主动脉重构的作用及其机制

目的：探索卡格列净通过调控血管平滑肌细胞（VSMCs）增殖迁移及肾素-血管紧张素-醛固酮系统（RAAS）/转化生长因子β1(TGF-β1)信号通路,改善自发性高血压大鼠（SHR）胸主动脉重构。方法 ：将大鼠分为正常血压大鼠对照组（WKY组）、SHR组和SHR+卡格列净组。尾袖血压计监测大鼠尾动脉血压,H-E染色评估胸主动脉血管壁中膜厚度及中膜厚度/管腔直径比值,Masson染色检测血管壁纤维化程度。体外分离培养WKY和SHR胸主动脉VSMCs,细胞分为WKY组、WKY+卡格列净组、SHR组和SHR+卡格列净组。CCK-8检测细胞增殖,划痕实验检测细胞迁移,RT-qPCR和免疫印迹检测α-平滑肌肌动蛋白（α-SMA）、分泌型磷酸蛋白1(SPP1)、 I型胶原蛋白（Col1a）、Ⅲ型胶原蛋白（Col3a）、血管紧张素原（AGT）、血管紧张素Ⅱ受体1 (AGTR1)、TGF-β1mRNA与蛋白表达。结果 ：给予卡格列净灌胃8周后,与SHR组相比,SHR+卡格列净组大鼠尾动脉血压、胸主动脉中膜厚度、中膜厚度/管腔直径比值、胶原蛋白沉积程度降低。SHR组VSMCs增殖、迁移能力较WKY组明显增强...     

短链酰基辅酶A脱氢酶在高血压血管重构中的作用

目的:研究短链酰基辅酶A脱氢酶(short-chain acyl-CoA dehydrogenase,SCAD)在高血压血管重构中的变化,探讨SCAD与高血压血管重构之间的关系。方法:采用游泳耐力训练方式训练16周龄的自发性高血压大鼠(spontaneously hypertensive rats,SHR)和健康Wistar大鼠8周,分别以24周龄的SHR和Wistar大鼠作为实验对照,定期测量鼠尾收缩压,测定各组大鼠胸主动脉血管腔内径和血管壁中层厚度、SCAD的mRNA和蛋白表达水平、SCAD酶活性的变化、ATP和ROS水平及血清和胸主动脉游离脂肪酸含量。结果:与Wistar组比较,SHR组大鼠血压升高,血管腔内径减小,血管壁中层厚度增大,血管壁中层厚度与血管腔内径比值增大;与SHR组相比,SHR游泳组血压下降,血管腔内径增大,血管壁中层厚度减小,血管壁中层厚度与血管腔内径比值减小(P0.05)。与Wistar组比较,SHR组大鼠主动脉SCAD的mRNA和蛋白表达水平显著下调,主动脉中SCAD酶活性下降,ATP含量降低,血清和主动脉游离脂肪酸含量明显增加,ROS含量增加;分别与Wistar组和SHR组比较,Wistar游泳组和SHR游泳组主动脉SCAD的mRNA和蛋白表达水平均明显上调,主动脉中SCAD的酶活性增高,ATP含量增加,血清和主动脉的游离脂肪酸含量明显减少,ROS含量减少。结论:主动脉SCAD的表达下调可能与高血压血管重构密切相关。游泳运动可能通过上调SCAD表达,从而逆转高血压血管重构。     

在高血压动脉重建中microRNA-21对血管平滑肌细胞细胞外基质表达的调控作用

目的探讨在高血压动脉重建中microRNA-21(miR-21)对血管平滑肌细胞(vascular smooth muscle cells,VSMCs)细胞外基质(extracellular matrix,ECM)的调控作用及其机制。方法建立腹主动脉缩窄型大鼠高血压模型,大鼠分为假手术对照组、高血压2周组和高血压4周组;对体外培养的大鼠主动脉VSMCs施加频率为1.25 Hz周期性张应变,加载幅度分别为0%(静态对照组)、5%(正常张应变组)、15%(模拟高血压状态的高张应变组),加载持续时间均为12 h。采用Western blotting和Real time RT-PCR技术,分别检测动脉和细胞样品ECM以及miR-21的表达。用miR-21特异干扰片段抑制培养的VSMCs miR-21表达,然后检测VSMCs的ECM、miR-21和Smad 7表达变化。结果与假手术对照组相比,高血压2周组胸主动脉ECM和miR-21的表达显著上升;高血压4周组胸主动脉的I型胶原、III型胶原和miR-21表达显著上升。与静态对照组和5%张应变组相比,15%张应变组VSMCs的I型胶原表达无显著变化,而III型胶原表达显著升高,Smad 7表达显著下降,周期性张应变增强VSMCs的miR-21表达。干扰miR-21降低周期性张应变状态下VSMCs的miR-21表达以及III型胶原蛋白水平表达,上调VSMCs的Smad 7表达。结论高血压血管重建导致大鼠胸主动脉ECM和miR-21高表达。周期性高张应变可诱导VSMCs的miR-21高表达,再通过其调节Smad 7蛋白,进而调控VSMCs的ECM,尤其是III型胶原的表达,参与高血压血管重建。     

四逆汤对兔球囊损伤后血管平滑肌细胞增殖和凋亡的影响

目的: 观察四逆汤(SND)对兔腹主动脉球囊拉伤后血管平滑肌细胞(VSMCs)增殖和凋亡的影响,探讨VSMCs增殖和凋亡在动脉损伤后再狭窄(RS)中的作用及SND预防PCI术后RS的可行性。方法: 建立兔腹主动脉球囊拉伤模型,用SND进行干预,电镜观察中膜和内膜增殖和凋亡的VSMCs超微结构特征。用α-actin标记VSMCs并检测其增殖细胞核抗原(PCNA)及细胞周期素E(cyclin E)的表达;Tunel法检测凋亡细胞,计算增殖和凋亡指数。84 d亚组行腹主动脉造影观察损伤段血管的狭窄程度。结果: SND治疗组中膜内膜VSMCs增殖程度明显轻于对照组,凋亡指数明显高于对照组(P<0.05),而且细胞凋亡持续的时间较长,到术后14 d才达高峰,以后才逐渐下降。术后84 d亚组血管造影显示治疗组损伤段腹主动脉狭窄程度明显小于对照组(P<0.05)。结论: 四逆汤能有效抑制VSMCs增殖,诱导其凋亡,减轻血管损伤后的狭窄程度。     

CaN-NFAT信号通路参与苯肾上腺素介导的血管平滑肌细胞增殖

目的 研究钙调神经磷酸酶-活化T细胞核因子(CaN-NFAT)信号通路对苯肾上腺素诱导的血管平滑肌细胞(VSMCs)增殖的调节作用。方法：组织贴块法原代培养SD大鼠血管平滑肌细胞,MTT法和细胞计数法测定VSMCs增殖,间接免疫荧光法测定NFATc1细胞定位,Western blot测定CaN蛋白表达,定磷法检测CaN活性。结果：苯肾上腺素(PE,α1-受体激动剂)促进VSMCs增殖,哌唑嗪(prazosin,α1-受体抑制剂)和环胞霉素A(CsA,CaN抑制剂)降低PE诱发的VSMCs增殖,白屈菜红碱(chelerythrine,蛋白激酶C抑制剂)预处理VSMCs后,PE诱发的VSMCs吸光度和细胞数被抑制, 并且这种抑制作用可以被CsA进一步加强。CsA抑制PE诱发的CaN表达与活性。PE促进NFATc1从胞质易位入核,CsA抑制NFATc1核转位。结论：CaN-NFATc1信号通路参与调节苯肾上腺素诱导的VSMCs增生肥大。     

内皮素和血管紧张素Ⅱ对大鼠血管平滑肌细胞分裂的促进作用

血管平滑肌细胞(SMC)的增生、肥厚是高血压的一个重要致病因素,但其机制尚不清楚。本工作选用自发性高血压大鼠(SHR)和其对照大鼠(WKY)培养的SMC,以细胞计数和[~3H]-Thymidine参入为指标,观察到内皮素(ET)和血管紧张素Ⅱ(ANGⅡ)以剂量依赖方式刺激SMC的增殖和细胞内DNA合成,但其效应在SHR SMC更明显;心钠素(ANF)可显著抑制上述效应。SHR SMC对ET和ANGⅡ促分裂作用的反应性增强以及ANF对其抑制效应提示,ET和ANGⅡ的促血管壁增生作用在高血压的发生发展中起着重要作用,ANF可能具有机体内源性抗高血压因子的作用,抑制和缓解高血压的发生发展。     

尼古丁对大鼠血管平滑肌细胞增殖和凋亡的影响

目的 探讨尼古丁对大鼠胸主动脉血管平滑肌细胞(VSMCs)增殖和凋亡的影响.方法用终浓度为100 μmol/L的尼古丁作用于体外培养的大鼠VSMCs 24h,不加尼古丁的细胞作为对照组,应用荧光显微镜和扫描电子显微镜观察各组细胞凋亡的形态,BrdU-ELISA和流式细胞术检测各组细胞增殖和凋亡率,进而确定尼古丁对VSM...     

血管紧张素Ⅱ对血管外膜成纤维细胞Ⅰ型胶原合成及其mRNA表达的影响

目的:观察血管紧张素Ⅱ对血管外膜成纤维细胞Ⅰ型胶原合成及其mRNA表达的影响.方法:血管紧张素Ⅱ作用于培养的大鼠血管外膜成纤维细胞,采取酶联免疫吸附法(ELISA)、竞争性逆转录聚合酶链反应(RT-PCR)法分别检测Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达.结果:在血管紧张素Ⅱ作用下,血管外膜成纤维细胞Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达均增加,且有一定的剂量依赖性.结论:血管紧张素Ⅱ可刺激血管外膜成纤维细胞Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达,提示血管紧张素Ⅱ是调节血管外膜成纤维细胞胶原代谢的重要因子,血管外膜成纤维细胞可能参与高血压血管重塑过程.     

血管紧张素Ⅱ对血管外膜成纤维细胞Ⅰ型胶原合成及其mRNA表达的影响

目的:观察血管紧张素Ⅱ对血管外膜成纤维细胞Ⅰ型胶原合成及其mRNA表达的影响。方法:血管紧张素Ⅱ作用于培养的大鼠血管外膜成纤维细胞,采取酶联免疫吸附法(ELISA)、竞争性逆转录聚合酶链反应(RT-PCR)法分别检测Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达。结果:在血管紧张素Ⅱ作用下,血管外膜成纤维细胞Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达均增加,且有一定的剂量依赖性。结论:血管紧张素Ⅱ可刺激血管外膜成纤维细胞Ⅰ型胶原蛋白合成及Ⅰ型胶原mRNA的表达,提示血管紧张素Ⅱ是调节血管外膜成纤维细胞胶原代谢的重要因子,血管外膜成纤维细胞可能参与高血压血管重塑过程。     

大鼠血管平滑肌增殖器官模型的建立及其机制探讨

为更好地研究血管平滑肌细胞增生性疾病的机理及防治,建立一种体外VSMC增殖器官模型,并对其机制作初步探讨.HE染色和免疫组化染色显示,大鼠主动脉段在体外拉伤内皮并经20%血清培养后,会出现中膜VSMCs的增殖.体外培养5 d后血管壁VSMC就出现不同程度增生,其中13 d的血管有明显斑块形成;免疫组织化学染色见有标记的增生VSMCs; RT-PCR检测显示,Hrg-1和SM22αmRNA的表达随培养天数增多而减少,至13 d检测不出.而在同样体外培养10 d情况下,与对照组相比,内皮损伤组培养上清中ET-1明显增多,Hrg-1、SM22a mRNA表达下调,Brdu标记增殖细胞增多,当加入内皮素受体阻断剂BQ123后标记细胞明显减少,其中血清培养组与无血清培养组相比,以上检测变化有显著性差异.结果表明,大鼠主动脉经体外培养能诱导平滑肌细胞异常增生并且表型从收缩型向合成型转化, ET-1和血清作用是该模型平滑肌增殖的主要因素.本模型为研究血管平滑肌增殖性疾病的机理和防治提供了一个较好的实验平台.     

Ultraviolet light-irradiated collagen III modulates expression of cytoskeletal and surface adhesion molecules in rat aortic smooth muscle cells in vitro

Systemic and pulmonary hypertension is characterised by structural reconstruction of the vascular wall which includes hypertrophy and hyperplasia of vascular smooth muscle cells (VSMCs) and fibroproduction. We hypothesise that these changes are stimulated by non-enzymatic modification of collagen molecules in the injured vascular wall by radicals. We exposed collagen III to ultraviolet (UV) light irradiation which, as indicated by fluorescence and electrophoretic analyses, resulted in its fragmentation. Both irradiated and control unmodified collagen were adsorbed on culture dishes and seeded with VSMCs derived from the rat thoracic aorta. During the first week after seeding, the cells on the modified collagen attained significantly higher population density (by 15-83%), higher mitotic index (by 31-135%) and higher BrdU labelling index (by 32%). However, these cells were less resistant to spontaneous and trypsin-mediated detachment from the growth support. As revealed using enzyme-linked immunosorbent assay in 3-day-old cultures, the cells growing on the irradiated collagen exhibited a lower concentration of beta-1 integrins (-10%, measured per milligram of protein), vinculin (-18%), talin (-6%) and vimentin (-15%). Immunofluorescence staining showed that these molecules were distributed more diffusely and less organised into focal adhesion plaques or cytoskeletal fibres. The concentration of two adhesion molecules of immunoglobulin type, ICAM-1 and VCAM-1, was increased by 11% and 16%, respectively. The concentration of alpha-v integrins and alpha-actin was unchanged; the latter, however, formed fewer distinct microfilament bundles in cells on the modified collagen. Our results suggest that the VSMCs growing on UV-modified collagen are more prone to escape the growth control mediated by cell-extracellular matrix contact and can bind the cells of the immune system.     

抗高血压因子对大鼠血管平滑肌钙内流的影响

实验观察了从原发性高血压病患者红细胞中提取的抗高血压因子(AHF)对自发性高血压大鼠(SHR)、肾性高血压大鼠(RHR)和正常血压的WKY大鼠和Wistar大鼠主动脉和肠系膜动咏平滑肌Ca~(2+)内流的影响。结果表明,SHR和RHR的肠系膜动脉Ca~(2+)内流显著高于主动脉;AHF可显著抑制SHR和RHR主动脉和肠系膜动脉Ca~(2+)内流,抑制作用呈剂量依赖性,且对肠系腆动脉Ca~(2+)内流抑制作用更明显;AHF也可抑制正常动物血管平滑肌Ca~(2+)内流。本工作提示,AHF的降压机制可能与其抑制血管平滑肌特别是小动脉血管平滑肌Ca~(2+)内流有关。     

Vascular damages in rats immunized by alpha1-adrenoceptor peptides

Autoantibodies against the α1-adrenoceptor which had agonist activity as norepinephrine might play roles in the progression of hypertension, but whether the autoantibodies could induce vascular remodeling as norepinephrine is not clear. In this paper, the models with antibodies against the α1-adrenoceptor were made by immunizing Wistar rats with the synthesized the second extracellular loop of α1-adrenoceptor peptides. The homo-age male Wistar rats received BSA in the same immunizing manner and male spontaneous hypertensive rats （SHR） were used as control. All the rats were raised for one year. The blood pressure and morphological changes of arteries were measured. In the end, despite the systolic blood pressure of immunized rats had no difference with normal control, the media thickness of aortas and ratio of media to lumen in the third-order arteries of mesenteric vasculature were increased in immunized rats. The observation with electron microscope showed that the mitochondria of vascular smooth muscle cells （VSMCs） had notable hyperplasia, and the interstitial collagen fibril was increased too. The effects of purified antibodies against α1-adrenoceptor on the proliferation of cultured VSMCs, and the expressions of c-jun, c-fos and α1-adrenoceptor were detected. The results showed that the antibodies could promote the proliferation of cultured VSMCs, and enhance the expression of c-jun both in vitro and in vivo. So we concluded that antibodies against the α1-adrenoceptor could contribute to vascular damages in rats by stimulating the growth of VSMCs which might be caused by the increased c-jun expression, and might play particular roles in the pathological changes of hypertension.     

芝麻素对高脂、高糖饲养大鼠血脂血糖及血管重构的影响

目的：探讨芝麻素对高脂、高糖饲养大鼠血脂血糖及血管重构的影响。方法：大鼠高脂高糖饮食24周，于第9周开始连续口服芝麻素(120、60、30 mg·kg^-1·d^-1)16周。测血糖、血脂、血压、血清与血管总抗氧化能力和过氧化氢含量。主动脉HE染色和Masson染色分别观察主动脉组织学和胶原纤维变化。肠系膜动脉HE染色，病理图像分析系统测定血管中膜厚度(media thickness，M)、血管内径(luminal radius，L)和中膜厚度与内径比值（M/L）。免疫组化法观察主动脉诱导型一氧化氮合酶蛋白表达。结果：与模型组相比，芝麻素高中剂量组血糖、血脂和血压明显下降（P<0.01）；总抗氧化能力显著提高，过氧化氢含量明显降低（P<0.01或P<0.05）；主动脉iNOS蛋白表达和胶原纤维沉积明显减少；血管内膜和平滑肌细胞增殖减轻；系膜动脉血管M与M/L明显减少，L明显增加(P<0.01)。结论：芝麻素能纠正高脂、高糖诱导大鼠糖脂代谢异常,并抑制血压升高、改善血管重构。     

血压不稳定引起的大鼠血管重构

目的：观察去窦弓神经引起的血管重构。方法：10周龄SD大鼠行去窦弓神经术或假手术,术后4、8、16和32周采用组织病理学和计算机图像分析技术对大鼠各级动脉连续切片进行观察和比较。结果：与假手术组相比,去窦弓神经大鼠大动脉的重构主要以血管中层平滑肌细胞肥大和基质改变为主;中动脉的重构在血管中层平滑肌细胞肥大的同时可能有轻度的增生;小动脉主要以血管平滑肌细胞增生为主。结论：大鼠去窦弓神经后各级动脉均可发生重构。     

Age-related changes in adrenaline content of rat splanchnic blood vessels

The influence of ageing on the adrenaline content of the superior mesenteric artery and vein, renal artery and vein and portal vein was studied in 3-month- (young), 12-month- (adult) and 24-month-old (old) male Wistar rats using radioenzymatic assay for the measurement of catecholamine levels. Adrenaline concentrations were unchanged in the vascular wall of the blood vessels examined in adult rats, but were significantly decreased in the vascular wall of the superior mesenteric, renal and portal veins of old rats. In contrast, no age-dependent changes of adrenaline levels were found in the vascular wall of the superior mesenteric or renal arteries. The possibility that the loss of adrenaline concentrations in the venous vascular wall may be in some way related to the cardiovascular impairment occurring with age is discussed.     

Vascular area with marked resistance elevation in one-clip, two-kidney renovascular hypertensive rats

Blood flow in the terminal aorta or superior mesenteric artery was recorded in conscious one-clip, two-kidney renovascular hypertensive rats and normal control rats with a chronically implanted electromagnetic flow probe. An indwelling catheter for direct measurement of arterial pressure was inserted into the terminal aorta in rats with a flow probe around the superior mesenteric artery. In the hindquarter area supplied by the terminal aorta, the elevation of vascular resistance in hypertensive rats in relation to normal rats was close to the average over the whole body. However, it was about 40% more intense than the average in the superior mesenteric area. Quantitatively, the hindquarter and superior mesenteric beds in hypertensive rats contributed about a quarter and a third of the decrease in total vascular conductance, respectively. Plots of superior mesenteric flow per body weight against arterial pressure at rest for 51 measurements in 9 hypertensive rats revealed an inverse relation between these two variables. These findings indicate that elevation of vascular resistance in the superior mesenteric area and probably the splanchnic area in general play an important role in one-clip, two-kidney renovascular hypertension.     

桡动脉与足背动脉的解剖学特点及其临床意义

目的研究桡动脉和足背动脉的直径及管壁厚度,为桡动脉和足背动脉置管以及经桡动脉冠状动脉造影等提供解剖学依据。方法用体视显微镜测量31具尸体62侧桡动脉远侧段和60侧足背动脉的内径(ID)及外径(ED)。结果①桡动脉平桡骨茎突上5cm处断面(SARA)的外径、内径和管壁厚度(WT)分别为(2.33±0.49)mm、(1.31±0.45)mm和(0.51±0.13)mm,与平桡骨茎突处断面(SBRA)比较,SARA小于SBRA(P<0.05)。②足背动脉中点断面(SMDPA)的外径、内径和管壁厚度分别为(2.47±0.49)mm、(1.28±0.50)mm和(0.60±0.16)mm。与近端和远端比较,3组均数差异均有统计学意义。③桡动脉远侧段与足背动脉的内、外径及管壁厚度无侧别差异,但有年龄和性别差异,50岁以上组大于中、青年组,男性管壁厚度大于女性。④SARA和SMDPA的内、外径和管壁厚度有正相关关系。结论:本研究结果对桡动脉和足背动脉穿刺置管术的开展与应用等具有参考价值。     

Innervation and functional changes in mesenteric perivascular calcitonin gene-related peptide- and neuropeptide Y-containing nerves following topical phenol treatment

We have previously shown that age-related reduction of innervation and function in mesenteric perivascular calcitonin gene-related peptide-containing vasodilator nerves takes place in spontaneously hypertensive rats. The present study was performed to investigate innervation and functional changes in perivascular calcitonin gene-related peptide- and adrenergic neuropeptide Y-containing nerves after topical treatment with phenol, which damages nerve fibers, around the rat superior mesenteric artery. Under pentobarbital-Na anesthesia, 8-week-old Wistar rats underwent in vivo topical application of phenol (10% phenol in 90% ethanol) or saline (sham rats) to the superior mesenteric artery proximal to the bifurcation of the abdominal aorta. After the treatment, the animals were subjected to immunohistochemistry of the 3rd branch of small arteries proximal to the intestine and to vascular responsiveness testing on day 3 through day 14. The innervation levels of calcitonin gene-related peptide-like immunoreactivity containing fibers and neuropeptide Y-like immunoreactivity containing fibers were markedly reduced on day 3 to day 14 and on day 5 to day 14 after the treatment, compared with those in sham-operated rats, respectively. In perfused mesenteric vascular beds isolated from phenol-treated rats, adrenergic nerve-mediated vasoconstriction and calcitonin gene-related peptide nerve-mediated vasodilation in response to periarterial nerve stimulation (2-12 Hz) were significantly decreased on day 3 and day 7. Neurogenic release of norepinephrine in phenol-treated rats on day 7 was significantly smaller that that in sham-operated rats. Nerve growth factor content in the mesenteric arteries of phenol-treated rats was significantly lower than that in sham-operated rats. Administration of nerve growth factor using osmotic mini-pumps for 7 days after the phenol treatment resulted in greater density of calcitonin gene-related peptide- and neuropeptide Y-like immunoreactivity fibers than in phenol-treated rats and restored decreased vascular responses to periarterial nerve stimulation. These results suggest that topical phenol-treatment of the mesenteric artery effectively induces functional denervation of perivascular nerves, which can be prevented or reversed by nerve growth factor treatment.     

nor-NOHA逆转自发性高血压大鼠血管重构

目的 探讨精氨酸酶抑制剂nor-NOHA对自发性高血压大鼠（SHR）血管重构的影响。方法WKY大鼠作为对照组,SHR随机分为模型组与受试组,受试组每天腹腔注射nor-NOHA(40 mg/kg),定期测量大鼠收缩压和体质量,10周后,采用酶谱法检测心、肾及胸主动脉组织中活性MMP-2的含量,并对胸主动脉进行Tricrome- Masson,Picric-Sirius red 及Orcein组织特染观察中膜厚度（MT）、管腔内径（LD）、径腔比（M/L）、Ⅰ型和Ⅲ型胶原及弹力纤维面积百分比值等。 结果 受试组收缩压显著下降、左心室MMP-2较SHR组含量显著降低（P＜0.05),胸主动脉的MT、M/L、Ⅰ型和Ⅲ型胶原蛋白均下降（P＜0.05)。结论 nor-NOHA可能通过抑制MMP-2活性及降低Ⅰ型和Ⅲ型胶原的合成而缓解SHR血管重构。