Abstract-Mechanical forces are important modulators of cellular function in many tissues and are particularly important in the cardiovascular system. The endothelium, by virtue of its unique location in the vessel wall, responds rapidly and sensitively to the mechanical conditions created by blood flow and the cardiac cycle. In this study, we examine data which suggest that steady laminar shear stress stimulates cellular responses that are essential for endothelial cell function and are atheroprotective. We explore the ability of shear stress to modulate atherogenesis via its effects on endothelial-mediated alterations in coagulation, leukocyte and monocyte migration, smooth muscle growth, lipoprotein uptake and metabolism, and endothelial cell survival. We also propose a model of signal transduction for the endothelial cell response to shear stress including possible mechanotransducers (integrins, caveolae, ion channels, and G proteins N umerous studies suggest that normal functioning of the endothelium is critical in limiting the development of atherosclerosis, as illustrated by the correlation between risk factors for atherosclerosis (smoking, high cholesterol, high homocysteine, decreased estrogen, increasing age, and hypertension) and endothelial dysfunction.1 Endothelial cells play a critical role in vascular homeostasis by performing many functions. They sense and integrate hemodynamic and hormonal stimuli and effect alterations in vascular function through the secretion of various mediator proteins and molecules.2 As a result of these properties, endothelial cells modulate biological processes related to the blood vessel wall, including regulation of the permeability of plasma lipoproteins, adhesion of leukocytes, and release of prothrombotic and antithrombotic factors, growth factors, and vasoactive substances.3 Impairment of these endothelial cell-mediated processes has been postulated to play a central role in the pathogenesis of atherosclerosis. 1Just as other tissues have developed mechanisms to detect changes in the physiological conditions to which they are exposed, endothelial cells respond not only to humoral factors in the circulation but also to the mechanical conditions created by blood flow and the cardiac cycle. 4 As a result of their unique location, endothelial cells experience three primary mechanical forces: pressure, created by the hydrostatic forces of blood within the blood vessel; circumferential stretch or tension, created as a result of defined intercellular connections between the endothelial cells that exert longitudinal forces on the cell during vasomotion; and shear stress, the dragging frictional force created by blood flow. Of these forces, shear stress appears to be a particularly important hemodynamic force because it stimulates the release of vasoactive substances and changes gene expression, cell metabolism, and cell morphology. 4 The nature and magnitude of shear stress plays an important role in long-term maintenance of the structure and function of the blood vessel. T...

Fluid shear stress is one of the most important mechanical forces acting upon vascular endothelium, because of its location at the interface between the bloodstream and vascular wall. Recent evidence indicates that several intracellular signaling events are stimulated in endothelial cells in response to shear stress. Through these events, shear stress modulates endothelial cell function and vascular structure, but the molecular basis of shear stress mechanotransduction remains to be elucidated. In our research we have focused on three temporal signal responses to shear stress: (1) production of nitric oxide (NO) as an immediate response; (2) activation of extracellular-regulated kinases (ERK1/2; p44/p42 mitogen-activated protein (MAP) kinases) as a rapid response, and (3) tyrosine phosphorylation of focal adhesion kinase (FAK) as a sustained response. In terms of vessel biology, NO production, and ERK1/2 and FAK activation seem to be correlated with vascular homeostasis, gene expression and cytoskeletal rearrangement, respectively. In this review, we discuss the mechanisms that establish the temporal order of shear stress-stimulated responses based on a hierarchy for assembly of signal transduction molecules at the cell plasma membrane.

No abstract

Mechano-sensitive regulation of endothelial cells (EC)function by shear stress is critical for flow-induced vasodilation and gene expression. Previous studies by our laboratory demonstrated that shear stress activates the 44-and 42-kDa extracellular signal-regulated kinases (ERK1/2) in EC in a time-and force-dependent manner. ERK1/2 activation was inhibited by protein kinase C (PKC) down-regulation with phorbol 12,13-dibutyrate (1 M for 24 h) but not by calcium chelation with BAPTA-AM (acetoxymethyl ester of BAPTA) (75 M for 30 min), suggesting that a novel PKC isoform (␦, ⑀, , ) mediates shear stress-induced ERK1/2 activation. Western blotting with PKC isoform-specific antibodies demonstrated expression of PKC-␣, -⑀, and -isoforms in EC. PKC-⑀ was specifically inhibited by transfection with antisense PKC-⑀ phosphorothioate oligonucleotides (1,000 nM for 6 h). Antisense treatment decreased PKC-⑀ protein levels by 80 ؎ 13% after 72 h and completely inhibited shear stress-stimulated ERK1/2 activation. Scrambled PKC-⑀ oligonucleotides and antisense PKC-␣ and PKC-oligonucleotides had no effect on ERK1/2 activity. PKC-⑀ appeared specific for mechano-sensitive ERK1/2 activation, as antisense PKC-⑀ oligonucleotides did not inhibit ERK1/2 activation by EGF or bradykinin but did inhibit ERK1/2 activation upon EC adhesion to fibronectin. These results define a pathway for shear stress-mediated ERK1/2 activation and establish a new function for PKC-⑀ as part of a mechano-sensitive signal transduction pathway in EC.

Hemostatic mechanisms are an integral part of the human physiology. Traditionally divided into intrinsic and extrinsic arms, the coagulation cascade converges, through the interactions of many different factors, at a common element-thrombin. As a consequence, a number of different agents have been developed to supplement this common, critical step to aid surgical hemostasis. Intraoperative interventions most commonly include sutures and heat-generating cautery devices; however, these methods are sometimes insufficient or inappropriate for a specific procedure or anatomic location, leading to the development of other adjunctive therapies, including topical hemostats. Topical hemostatic agents generally act as active, passive, and combinations therapies, depending on their individual composition and mode of action. We provide a brief review of the normal coagulation cascade, including critical points, followed by a discussion of surgical strategies and adjuctive therapies used to achieve surgical hemostasis and concluding with a discussion of topical thrombins.

Fluid shear stress is an important regulator of endothelial cell (EC) function. To determine whether mechanosensitive ion channels participate in the EC response to shear stress, we characterized the role of ion transport in shear stress-mediated extracellular signalregulated kinase (ERK1/2) stimulation. Replacement of all extracellular Na ؉ with either N-methyl-D-glucamine or choline chloride increased the ERK1/2 stimulation in response to shear stress by 1.89 ؎ 0.1-fold. The Na ؉ effect was concentration-dependent (maximal effect, <12.5 mM) and was specific for shear stress-mediated ERK1/2 activation as epidermal growth factor-stimulated ERK1/2 activation was unaffected by removal of extracellular Na ؉ . Shear stress-mediated ERK1/2 activation was potentiated by the voltage-gated sodium channel antagonist, tetrodotoxin (100 nM), to a magnitude similar to that achieved with extracellular Na ؉ withdrawal. Transfection of Chinese hamster ovary cells with a rat brain type IIa voltage-gated sodium channel completely inhibited shear stress-mediated ERK1/2 activation in these cells. Inhibition was reversed by performing the experiment in sodium-free buffer or by including tetrodotoxin in the buffer. Western blotting of bovine and human EC lysates with SP19 antibody detected a 250-kDa protein consistent with the voltage-gated sodium channel. Degenerate polymerase chain reaction of cDNA from primary human EC yielded transcripts whose sequences were identical to the sodium channel SCN4a and SCN8a ␣ subunit genes. These results indicate that shear stress-mediated ERK1/2 activation is regulated by extracellular sodium and demonstrate that ion transport via Na ؉ channels modulates EC responses to shear stress.Mechanical stimuli are important modulators of cellular function in tissues, particularly in the cardiovascular system. A key physical force experienced by EC 1 by virtue of their unique location in the vascular wall is fluid shear stress created by the frictional force of blood flow (1). Changes in fluid shear stress have been shown to regulate EC function, including permeability of plasma lipoproteins, adhesion of leukocytes, and release of pro-and antithrombotic factors, growth factors, and vasoactive substances (reviewed in Refs. 1 and 2). These hemodynamically regulated events may contribute to the pathogenesis of vascular disease as atherosclerotic plaques are preferentially localized to areas of the vascular system that experience turbulent flow and low time-averaged shear stress (3, 4).Our laboratory has previously reported that ERK1/2 are activated by shear stress in EC (5). Whereas the mechanisms responsible for growth factor-mediated stimulation of ERK1/2 have been well characterized (6), the upstream signaling pathway that leads to activation of ERK1/2 by shear stress remains undefined. Of particular interest are the primary plasma membrane mechanisms by which the physical force of shear stress can be transduced into biochemical signals. Several candidate mechanotransducers have been proposed including G p...

Whether from surgical misadventure, inherent patient factors, or iatrogenic causes, postoperative bleeding can be a consequence of any surgical procedure. There are many methods and products available to assist in managing or preventing bleeding. For each method, there may be specific benefits and indications, but they may also carry some degree of risk. Topical thrombin is used extensively in many surgical specialties, especially in the cardiovascular and neurosurgical arenas where other hemostatic modalities may not be appropriate choices. As a class, topical thrombins are generally a safe and effective method for achieving intraoperative hemostasis; however, some members of the class carry associated risks with their use. For example, the United States Food and Drug Administration required the addition of a black-box warning to the prescribing information of bovine-derived thrombin, the oldest member of the class, due to concerns of immune-mediated coagulopathies developing after use. In addition, human thrombin derived from pooled plasma has its own, if theoretical, risk of transmitting infections due to viral or prion agents. We address the topical thrombin class and review each product in the context of the current literature.

Ca2+ plays a major role in vascular contraction, and a defect in intracellular Ca2+ regulation has been associated with increased vascular reactivity in hypertension. To test the hypothesis that the sarcoplasmic reticulum does not adequately buffer Ca2+ in deoxycorticosterone acetate (DOCA) hypertension, contractile experiments were performed with a specific inhibitor of the sarcoplasmic reticulum Ca2+ ATPase, cyclopiazonic acid (CPA). Contractile force in aortic strips from DOCA and control rats was measured, using standard muscle bath procedures, to evaluate (i) Ca2+ handling, assessing caffeine and serotonin (5HT) induced contractions in Ca(2+)-free buffer and (ii) relaxation rate after 5HT washout. Contractile responses elicited with 5HT (3 x 10(-6) mol/L) and caffeine (20 mmol/L) were greater in DOCA than in control arteries. CPA (1 x 10(-7) to 3 x 10(-5) mol/L) reduced phasic contractions to 5HT and caffeine in DOCA and control aorta, and no differences in the IC50 values were observed. Aortae from DOCA rats contracted when placed in normal buffer, subsequent to treatment with Ca(2+)-free buffer, but control aortae did not. CPA potentiated these responses in DOCA aorta and only caused a modest contraction in control aorta. CPA-induced contraction did not occur in Ca(2+)-free buffer, and it was inhibited by nifedipine (IC50 = 4 x 10(-9) mol/L). The relaxation rate, after 5HT washout (3 x 10(-6) mol/L), was increased in DOCA aorta (2.6 +/- 0.3 min) compared with control (1.7 +/- 0.2 min), and CPA (10(-5) mol/L) increased the relaxation rate in both groups. The results support the hypothesis of defective Ca2+ handling in DOCA hypertension. However, an increased Ca2+ influx, and not a decreased buffering ability of the sarcoplasmic reticulum, contributes to the enhanced vascular reactivity observed in DOCA hypertension.