剪切应力对毛细血管内皮细胞代谢的影响

建立的平行平板流动腔装置适用于研究血管内皮细胞代谢对剪切流场的响应。将培养的人胚肾小球血管单层内皮细胞置于剪应力分别为５×１０－５Ｎ／ｃｍ２,１×１０－４Ｎ／ｃｍ２和１．５×１０－４Ｎ／ｃｍ２的定常层流中剪切２５小时,样品中的内皮素分泌量用放射免疫法测定。结果表明,剪应力水平对内皮细胞内皮素的代谢活动有显著影响。与静态培养对照,低水平的剪应力（５×１０－５Ｎ／ｃｍ２、１×１０－４Ｎ／ｃｍ２）促进内皮素的分泌,而较高水平的剪应力（１．５×１０－４Ｎ／ｃｍ２）抑制内皮素的分泌;剪应力对内皮素累积含量的影响比之分泌速率更大     

牛肺动脉单层内皮细胞长度与其血管紧张素II代谢的相关性

用离体培养的牛肺动脉内皮细胞灌流实验研究内皮细胞单层长度与其血管紧张素II代谢的相关性。牛肺动脉单层内皮细胞暴露于剪应力为0.64N/m2 的剪切流中24h后 ,两种不同长度(10cm和6cm)单层内皮细胞的血管紧张素II的平均分泌率有比较显著差异 ,10cm处理的血管紧张素II平均分泌率 (8.61±0.28pg/cm2.h)比6cm处理 (6.14±0.12pg/cm2.h)高40 % ,10cm处理的最小分泌率 (7.55pg/cm2.h)较6cm处理 (5.75pg/cm2.h)高31 % ,10cm处理的最小分泌率出现的剪切时间点比6cm处理要早6个小时。表明牛肺动脉内皮细胞单层长度与其血管紧张素II代谢 (分泌率 )间有密切的相关关系 ,进而从细胞代谢角度间接证实血管内皮细胞膜张应力存在累积效应。     

剪切应力对毛细血管内皮细胞代谢的影响

建立的平行平板流协腔装置适用于研究血管内皮细胞代谢对剪切流场的响应。将培养的人胚肾小球血管单层内皮细胞置于剪应力分别为５＊１０＾－５Ｎ／ｃｍ＾２,１＊１０＾－４Ｎ／ｃｍ＾２和１．５＊１０＾－４Ｎ／ｃｍ＾２的定常层流中剪切２５小时。     

血管壁剪切应力系统及用于内皮细胞与白细胞粘附的研究

本文建立了对血管壁上的内皮细胞施加剪切应力的系统。通过对系统中的血管内流场分析表明,该系统中的血管段的中间部分作为研究剪切应力对内皮细胞作用,以及研究内皮细胞与其它细胞粘附力的场所是比较理想的。采用该系统对在体内皮细胞研究发现：当对内皮细胞施加２８ｄｙｎ／ｃｍ２的剪切应力时,内皮细胞并未出现暴发性释放前列环素的现象,前列环素释放水平略有升高（０．４１±０．０５ｎｇ／ｃｍ２ｍｉｎ）以后呈线性下降并稳定在一定水平上（０．１７±０．０４ｎｇ／ｃｍ２ｍｉｎ）。在２８ｄｙｎ／ｃｍ２剪切应力下,受机械损伤的内皮细胞和动脉粥样硬化的内皮细胞仍能与较多白细胞粘附,而在正常内皮细胞和去掉内皮细胞的动脉壁几乎无白细胞粘附,说明在内皮细胞受损或动脉粥样硬化时,内皮细胞与白细胞的粘附增强     

急性缺氧对冠状动脉内皮细胞释放ET-1、NO、PGI2的影响

冠状动脉内皮细胞能分泌多种血管收缩和舒张物质。其中缩血管物质以内皮素的缩血管作用最强 ,且作用持久。内皮源性舒张因子主要为ＮＯ和花生四烯酸代谢产物ＰＧＩ2 。三种内皮源性血管活性因子均通过旁分泌方式作用于血管内皮下的平滑肌细胞 ,调节局部血管紧张度和血流量。在体研究表明缺氧可引起内皮素 1、ＮＯ、ＰＧＩ2 合成分泌改变。但在体研究时血浆及组织匀浆中ＥＴ 1、ＮＯ、ＰＧＩ2 的浓度改变难以反映其在血管内皮细胞形成和释放的状况。国外有关缺氧对离体冠状动脉内皮细胞合成和释放ＥＴ 1、ＮＯ、ＰＧＩ2 影响的研究也才刚起…     

剪切流场中内皮细胞膜张应力累加效应的实验研究 －离体血管段长度与其蛋白质代谢的相关性

用离体血管灌流试验验证冯元桢等关于血管内皮细胞的膜张应力逆血流方向累加的理论分析．用0.12N/m     

白细胞介素—8扩血管效应与内皮舒张因子的关系

为探讨内皮舒张因子在白细胞介素-8(IL-8)扩血管效应中的作用,本实验在大鼠离体主动脉条上,观察IL-8对血管反应性及血管组织cGMP含量的影响。实验发现,IL-8显著地扩张离体血管,其作用在去内皮后明显减弱。IL-8还能显著地提高离体血管组织cGMP含量,一氧化氮合成抑制剂L-NNA可阻断这一作用,一氧化氮前体L-精氨酸可逆转L-NNA的效应。结果表明IL-8可以通过促进血管内皮细胞释放一氧化氮而扩张血管。     

恒河猴睾丸内激素调控的离体研究 I.恒河猴睾丸Leydig细胞的离体培养及激素分泌的研究

为了研究恒河猴睾丸的激素分泌及其调节作用,作者发展了一种简便分离睾丸Ｌｅｙｄｉｇ细胞和用无血清培养的实验模型,研究了人绒毛膜促性腺激素（ｈＣＧ）、恒河猴绒毛膜促性腺激素（ｍＣＧ）、环磷酸腺苷（ｃＡＭＰ）、Ｆｏｒｓｋｏｌｉｎ和雌二醇（Ｅ２）对睾丸Ｌｅｙｄｉｇ细胞激素分泌的影响。结果显示离体的Ｌｅｙｄｉｇ细胞对低剂量同源的ｍＣＧ和异源的ｈＣＧ产生睾酮都有依赖反应,而高剂量的前者对Ｌｅｙｄｉｇ细胞分泌睾酮有明显的抑制作用,后者不出现反向调节;ｃＡＭＰ和Ｆｏｒｓｋｏｌｉｎ对Ｌｅｙｄｉｇ细胞分泌睾酮有刺激作用;雌二醇具有明显抑制ｈＣＧ和ｍＣＧ促睾丸分泌的作用     

土层厚度对旱地小麦花后根系衰老的影响

随土层加深,根系活力、根系ＳＯＤ、ＣＡＴ活性及可溶性蛋白质含量增加,ＭＤＡ含量减少,土层愈薄,其根系衰老愈快。从整个土层看,处理３（１６０ｃｍ）与对照（２００ｃｍ）无显著差异,处理１（８０ｃｍ）和处理２（１２０ｃｍ）与对照差异显著,由此可以认为延迟旱地小麦衰老的土层厚度应在１６０ｃｍ左右,并可据此制定管理方案,以获得高产高效。     

剪切应力对血管内皮细胞NO合成酶活性的影响及其机理探讨

目的：观测流体剪切应力对血管内皮细胞ＮＯ合成酶（ｎｉｔｒｉｃ ｏｘｉｄｅｓｙｎｔｈａｓｅ,ＮＯＳ）活性的影响并探讨其发生机制。方法：采用Ｇｒｉｅｓｓ 方法测定不同流体剪切应力作用下血管内皮细胞中ＮＯＳ活性的变化;并观测多种ＮＯＳ干预物质对这种变化的影响。结果：剪切应力显著提高血管内皮细胞中ＮＯＳ活性;地塞米松（ｄｅｘａｍｅｔｈａｓｏｎｅ）实验表明,剪切应力这种作用主要是通过对结构型ＮＯＳ活性的增强实现的,且具有明显的剂量和时间依赖性;放线菌酮（ｃｙｃｌｏｈｅｘｉｍｉｄｅ）非特异性地抑制细胞中ＮＯＳ酶蛋白合成,但ｃｙｃｌｏｈｅｘｉｍｉｄｅ 处理组中受剪切应力作用细胞ＮＯＳ活性仍显著高于其对照细胞,仅升高幅度明显降低。Ａ２３１８７ 处理后细胞中ＮＯＳ活性升高约达２ 倍,其中剪切应力作用细胞的ＮＯＳ活性显著高于其对照,但这种变化程度亦较Ａ２３１８７ 未处理组明显减小。结论：剪切应力显著提高血管内皮细胞ｅＮＯＳ活性：ｅＮＯＳ酶蛋白合成增加和细胞内Ｃａ２＋ 浓度的升高在剪切应力对血管内皮细胞ＮＯＳ活性的调节机制中具有重要意义     

血管内皮细胞内皮素代谢的数值分析

将培养的人胚肾小球血管单层内皮细胞置于剪应力分别为6．5dyn/cm^2和13．0dyn/cm^2的定常层流中剪切10小时,样品中的内皮素（ET）分泌量用放射免疫法测定。结果表明,剪切应力和剪切作用时间对内皮细胞内皮素的代谢均有显著的影响,其影响不是简单的线性增减,而呈较复杂的非线性特征,内皮素累积含量和分泌速率,随剪切时间的变化的关系可用Logistic方程来描述,获得了反映内皮素（ET）分泌规律的大量特征方程和特征数,为了解体内发生于血管壁的病理生理过程中内皮素分泌规律提供了实验数据。     

Shear stress-dependent expression of apoptosis-regulating genes in endothelial cells

Laminar shear stress exerts potent anti-apoptotic effects. Therefore, we analyzed the influence of laminar shear stress on the expression of apoptosis-regulating genes in human umbilical vein endothelial cells (HUVEC). Application of high levels of laminar shear stress (15 and 30 dyn/cm(2)) decreased the susceptibility of HUVEC to undergo apoptosis, whereas low shear stress (1 dyn/cm(2)) had no effect. These diminished signs of apoptosis were accompanied by a decreased mRNA expression of apoptosis-inducing Fas receptor. Furthermore, mRNA and protein expression of anti-apoptotic, soluble Fas isoform FasExo6Del and anti-apoptotic Bcl-x(L) were induced. Surprisingly, high shear stress also elevated mRNA and protein expression of pro-apoptotic Bak. The shear stress-induced up-regulation of Bcl-x(L) and Bak mRNA can be abrogated by inhibition of the endothelial NO synthase. We propose that altered expression of Bcl-x(L) and the Fas system is involved in the protective effect of laminar shear stress against apoptosis in human endothelial cells.     

Role of xanthine oxidoreductase and NAD(P)H oxidase in endothelial superoxide production in response to oscillatory shear stress

Oscillatory shear stress occurs at sites of the circulation that are vulnerable to atherosclerosis. Because oxidative stress contributes to atherosclerosis, we sought to determine whether oscillatory shear stress increases endothelial production of reactive oxygen species and to define the enzymes responsible for this phenomenon. Bovine aortic endothelial cells were exposed to static, laminar (15 dyn/cm2), and oscillatory shear stress (+/-15 dyn/cm2). Oscillatory shear increased superoxide (O2.-) production by more than threefold over static and laminar conditions as detected using electron spin resonance (ESR). This increase in O2*- was inhibited by oxypurinol and culture of endothelial cells with tungsten but not by inhibitors of other enzymatic sources. Oxypurinol also prevented H2O2 production in response to oscillatory shear stress as measured by dichlorofluorescin diacetate and Amplex Red fluorescence. Xanthine-dependent O2*- production was increased in homogenates of endothelial cells exposed to oscillatory shear stress. This was associated with decreased xanthine dehydrogenase (XDH) protein levels and enzymatic activity resulting in an elevated ratio of xanthine oxidase (XO) to XDH. We also studied endothelial cells lacking the p47phox subunit of the NAD(P)H oxidase. These cells exhibited dramatically depressed O2*- production and had minimal XO protein and activity. Transfection of these cells with p47phox restored XO protein levels. Finally, in bovine aortic endothelial cells, prolonged inhibition of the NAD(P)H oxidase with apocynin decreased XO protein levels and prevented endothelial cell stimulation of O2*- production in response to oscillatory shear stress. These data suggest that the NAD(P)H oxidase maintains endothelial cell XO levels and that XO is responsible for increased reactive oxygen species production in response to oscillatory shear stress.     

Shear stress enhances human endothelial cell wound closure in vitro

Repair of the endothelium occurs in the presence of continued blood flow, yet the mechanisms by which shear forces affect endothelial wound closure remain elusive. Therefore, we tested the hypothesis that shear stress enhances endothelial cell wound closure. Human umbilical vein endothelial cells (HUVEC) or human coronary artery endothelial cells (HCAEC) were cultured on type I collagen-coated coverslips. Cell monolayers were sheared for 18 h in a parallel-plate flow chamber at 12 dyn/cm(2) to attain cellular alignment and then wounded by scraping with a metal spatula. Subsequently, the monolayers were exposed to a laminar shear stress of 3, 12, or 20 dyn/cm(2) under shear-wound-shear (S-W-sH) or shear-wound-static (S-W-sT) conditions for 6 h. Wound closure was measured as a percentage of original wound width. Cell area, centroid-to-centroid distance, and cell velocity were also measured. HUVEC wounds in the S-W-sH group exposed to 3, 12, or 20 dyn/cm(2) closed to 21, 39, or 50%, respectively, compared with only 59% in the S-W-sT cells. Similarly, HCAEC wounds closed to 29, 49, or 33% (S-W-sH) compared with 58% in the S-W-sT cells. Cell spreading and migration, but not proliferation, were the major mechanisms accounting for the increases in wound closure rate. These results suggest that physiological levels of shear stress enhance endothelial repair.     

Plaque-prone hemodynamics impair endothelial function in pig carotid arteries

Hemodynamic forces play an active role in vascular pathologies, particularly in relation to the localization of atherosclerotic lesions. It has been established that low shear stress combined with cyclic reversal of flow direction (oscillatory shear stress) affects the endothelial cells and may lead to an initiation of plaque development. The aim of the study was to analyze the effect of hemodynamic conditions in arterial segments perfused in vitro in the absence of other stimuli. Left common porcine carotid segments were mounted into an ex vivo arterial support system and perfused for 3 days under unidirectional high and low shear stress (6 +/- 3 and 0.3 +/- 0.1 dyn/cm(2)) and oscillatory shear stress (0.3 +/- 3 dyn/cm(2)). Bradykinin-induced vasorelaxation was drastically decreased in arteries exposed to oscillatory shear stress compared with unidirectional shear stress. Impaired nitric oxide-mediated vasodilation was correlated to changes in both endothelial nitric oxide synthase (eNOS) gene expression and activation in response to bradykinin treatment. This study determined the flow-mediated effects on native tissue perfused with physiologically relevant flows and supports the hypothesis that oscillatory shear stress is a determinant factor in early stages of atherosclerosis. Indeed, oscillatory shear stress induces an endothelial dysfunction, whereas unidirectional shear stress preserves the function of endothelial cells. Endothelial dysfunction is directly mediated by a downregulation of eNOS gene expression and activation; consequently, a decrease of nitric oxide production and/or bioavailability occurs.     

Acute laminar shear stress reversibly increases human glomerular endothelial cell permeability via activation of endothelial nitric oxide synthase

Laminar shear stress is a key determinant of systemic vascular behavior, including through activation of endothelial nitric oxide synthase (eNOS), but little is known of its role in the glomerulus. We confirmed eNOS expression by glomerular endothelial cells (GEnC) in tissue sections and examined effects of acute exposure (up to 24 h) to physiologically relevant levels of laminar shear stress (10-20 dyn/cm(2)) in conditionally immortalized human GEnC. Laminar shear stress caused an orientation of GEnC and stress fibers parallel to the direction of flow and induced Akt and eNOS phosphorylation along with NO production. Inhibition of the phophatidylinositol (PI)3-kinase/Akt pathway attenuated laminar shear stress-induced eNOS phosphorylation and NO production. Laminar shear stress of 10 dyn/cm(2) had a dramatic effect on GEnC permeability, reversibly decreasing the electrical resistance across GEnC monolayers. Finally, the laminar shear stress-induced reduction in electrical resistance was attenuated by the NOS inhibitors l-N(G)-monomethyl arginine (l-NMMA) and l-N(G)-nitroarginine methyl ester (l-NAME) and also by inhibition of the PI3-kinase/Akt pathway. Hence we have shown for GEnC in vitro that acute permeability responses to laminar shear stress are dependent on NO, produced via activation of the PI3-kinase/Akt pathway and increased eNOS phosphorylation. These results suggest the importance of laminar shear stress and NO in regulating the contribution of GEnC to the permeability properties of the glomerular capillary wall.     

KLF2-dependent, shear stress-induced expression of CD59: a novel cytoprotective mechanism against complement-mediated injury in the vasculature

Complement activation may predispose to vascular injury and atherogenesis. The atheroprotective actions of unidirectional laminar shear stress led us to explore its influence on endothelial cell expression of complement inhibitory proteins CD59 and decay-accelerating factor. Human umbilical vein and aortic endothelial cells were exposed to laminar shear stress (12 dynes/cm(2)) or disturbed flow (+/- 5 dynes/cm(2) at 1Hz) in a parallel plate flow chamber. Laminar shear induced a flow rate-dependent increase in steady-state CD59 mRNA, reaching 4-fold at 12 dynes/cm(2). Following 24-48 h of laminar shear stress, cell surface expression of CD59 was up-regulated by 100%, whereas decay-accelerating factor expression was unchanged. The increase in CD59 following laminar shear was functionally significant, reducing C9 deposition and complement-mediated lysis of flow-conditioned endothelial cells by 50%. Although CD59 induction was independent of PI3-K, ERK1/2 and nitric oxide, an RNA interference approach demonstrated dependence upon an ERK5/KLF2 signaling pathway. In contrast to laminar shear stress, disturbed flow failed to induce endothelial cell CD59 protein expression. Likewise, CD59 expression on vascular endothelium was significantly higher in atheroresistant regions of the murine aorta exposed to unidirectional laminar shear stress, when compared with atheroprone areas exposed to disturbed flow. We propose that up-regulation of CD59 via ERK5/KLF2 activation leads to endothelial resistance to complement-mediated injury and protects from atherogenesis in regions of laminar shear stress.