$338 Journal of Biomechanics 2006, Vol. 39 (Suppl 1)

6503 Th, 08:45-09:00 (P39) G-Protein coupled membrane receptors may serve as mechanosensors for fluid shear stress in neutrophils A. Makino, G.W. Schmid-Sch6nbein. Department of Bioengineering, University of California, San Diego, La Jell& CA, USA

Neutrophils exposed to physiological levels of fluid shear stress (1 to 5 dyn/cm 2) exhibit a response that is unique to this cell type. Upon exposure to fluid shear, migrating neutrophils retract their pseudopods and they may detach from the substrate. This response is different than that observed in endothelial cells, which elongate or smooth muscle cells that also elongate - but at right angle - to the fluid shear direction. Thus the sensory mechanism appears to be cell type specific. We hypothesize that specific receptors in the cell membrane are activated/deactivated by fluid shear. We found that fluid shear decreases the constitutive activity of G-protein coupled receptor (GPCR). Inhibition of GPCR constitutive activity by inverse agonists abolished shear-induced cell area reduction. We found further that the formyl peptide receptor (FPR) on neutrophils, with high constitutive activity, is directly involved in the disassembly of F-actin in pseudopods and their retraction. Transfection of the FPR cDNA into a neutrophil cell line (undifferentiated HL60) without shear stress response and without FPR expression, leads to formation of pseudopods and retraction upon application of fluid shear. Suppression of FPR expression by siRNA in differentiated HL60 cells, that expressed other GPCRs, leads to a suppression of the fluid shear response. These results suggest that besides the phospholipid membrane and the glycocalyx, specific membrane receptors serve as mechanosensors for fluid shear stress. Ligand binding receptors may not only respond to biochemical agonist/antagonist binding but also to purely mechanical shear stress. Supported by NIH Grant HL 43026.

4304 Th, 09:00-09:15 (P39) The role of endothelial P2X4 receptors in flow-dependent control of vascular tone and remodeling

J. Ando 1 , K. Yamamoto 1 , T. Sokabe 1 , A. Kamiya 2. 1Department efBiemedical Engineering, Graduate School of Medicine, University of Tokyo, Tokyo, Japan, 2Interdisciplinary Science Center, Nihon University, Tokyo, Japan

Vascular endothelial cells are in direct contact with blood flow and exposed to shear stress, the frictional force exerted by flowing blood. A number of recent studies have revealed that vascular endothelial cells recognize changes in shear stress and transmit signals to the interior of the cell, which leads to cell responses that involve changes in cell morphology, cell function, and gene expression. These endothelial cell responses to shear stress are thought to play important roles in blood-flow-dependent phenomena, such as vascular tone control, angiogenesis, and vascular remodeling, and Ca2÷signaling plays an important role in shear-stress signal transduction. Our previous studies demonstrated that a shear stress-dependent Ca 2÷ influx occurs in endothelial cells when exposed to flow, and an ATP-operated cation channel P2X4 is the major contributor to flow-induced Ca 2÷ influx. However, the physiological and pathological significance of this shear-sensing mechanism via P2X4 is not fully understood. To gain insight into its significance, we generated a P2X4- deficient ( - / - ) mouse. P2X4-deficient mice did not exhibit normal endothelial cell responses to flow, such as Ca 2÷ influx and subsequent production of nitric oxide (NO), a potent vasodilator. The dilation of vessels induced by acute increase in blood flow in situ was markedly suppressed in P2X4-deficient mice. P2X4-deficient mice had higher blood pressure values and excreted smaller amounts of NO products in their urine than wild-type mice. No adaptive vascular remodeling, i.e., decrease in vessel size in response to chronic decrease in blood flow, was observed in the P2X4-deficient mice. We therefore conclude that endothelial P2X4 channels are critical to the blood-flow-sensitive mechanism that regulates vascular tone and remodeling.

Oral Presentations

shifts the balance between cell-cell and cell-matrix interactions to favor cell- matrix interactions and force transmission. Human umbilical vein endothelial cells were suspended at 1 106cells/ml within type-I rat-tail collagen gels. Gels formed with initially 1.0mg/ml of collagen are hereafter referred to compliant gels (E = 124±8 Pa) and gels with initially with 3.0 mg/ml collagen referred to as stiff gels (501 ±48 Pa). Relative to those in compliant gels, endothelial cells in stiff gels stained more intensely for the 151 integrin subunit (~J,21~1 is the integrin primarily responsible for cell binding to type-I collagen, P <0.05) but less intensely for PECAM (a homotypic cell-cell adhesion molecule prevalent in endothelial cells, P <0.05). In further supports of stiff gels stimulating cell-matrix interactions, endothelial cells isolated from stiff gels adhered to tissue more rapidly than did those from compliant gels with 8-fold more cells adhering within 30 minutes (P < 0.0001 ). Stiff gels also favored the formation of focal adhesion as indicated by increased immunostaining for vinculin (P <0.05). Extrapolating these in vitro data to an in vivo setting would predict that the microvasculature in stiff tissue would be more permeable, as is commonly observed in tumors, suggesting that the mechanical environment of a tumor may potentially contribute to abnormalities in its microvasculature.

References [1] Sieminski AL, Hebbel RP, Gooch KJ. Exp Cell Res. 2004 Ju115; 297(2): 574-84.

7387 Th, 09:30-09:45 (P39) Shear stress-induced gene response in human microvascular lymphat ic endothelial cells C. Yong 1 , E.A. Bridenbaugh 2, D.C. Zawieja 2, M.A. Swartz 1. 1Institute ef Bioengineering, Ecole Polytechnique F~d~rale de Lausanne, Switzerland, 2Department of Medical Physiology, College of Medicine, Texas A&M University System Health Science Center, College Station, TX, USA

The lymphatic system serves, among other things, to drain excess fluid from the interstitial space and return it to the blood. Under normal functioning conditions, increased load leads to increased draining (and presumably higher shear stress on the lymphatic vessels), and their functional capacity is quite high. However, fluid stagnation and thus loss of shear stress may result from pathological conditions like edema. Lymphatic biology remains poorly under- stood, and it is unknown whether lymphatic capillaries have any active control over their function in relationship to their functional load. This is in contrast to blood endothelium, whose sensitivity and responsiveness to even subtle changes in shear stress has been well documented. Although microvascular blood and lymphatic endothelial cells (BECs and LECs) share many common properties and features, they exhibit molecular heterogeneity obviated by distinct structure-function relationships. Additionally, the normal levels of shear experienced by the two cell types differ by an order of magnitude. As a first investigation into the molecular response of lymphatics to shear, we compared gene expression profiles in human dermal microvascular LECs exposed to low and high shear stress with those under static conditions using cDNA microarrays and confirmed with QRT-PCR. Specifically, 2-D shear condi- tions of 2 dyn/cm 2 (normal) and 20 dyn/cm 2 (elevated) were directly compared to static conditions. We find that key changes in gene expression patterns related to immune response, morphogenesis, cytoskeletal reorganization, cell- cell and cell-matrix interactions result from increases in shear stress. These studies provide a basis for analysis of the molecular mechanisms responsible for lymphatic vascular function modulation by fluid mechanical stimuli.

Track 16

Reproductive Biomechanics

6174 Th, 09:15-09:30 (P39) Matrix stiffness regulates the architecture of microvascular networks: role of cell-cell and cell-substrate interactions F.J. Byfield 1 , K.J. Gooch 2. 1Institute for Medicine and Engineering, Penn, Philadelphia, PA, USA, 2Department of Biomedical Engineering, Ohio State University, Columbus, OH, USA

Mechanical factors, especially pressure and shear stress, are widely acknowl- edged to play major roles in regulating the structure of the microvasculature. In addition to these well-established factors that act, at least initially, on the luminal surface, a growing body of evidence suggests that the mechanical environment presented on the abluminal surface also regulates microvascular structure. We recently presented evidence from an in vitro model that increas- ing matrix stiffness significantly disrupted the formation of interconnecting muti- cellular networks [1]. This inhibition was not due to an inability of the cells to elongate within the stiffer matrix but appeared to secondary to the formation of cell-cell junctions. Here we explore the hypothesis that increased stiffness

16.1. Non-Pregnant Uterine Peristalsis 7827 Mo, 08:15-08:45 (P6) Transport phenomena in human female genital tract

L. Wildt 1 , D. Hadziomerovic 1 , H.W. Ott 1 , D.W. Heute 2, I. Virgolini 2. 1 Department of Gynecologic Endocrinology and Reproductive Medicine; 2Department of Nuclear Medicine, Medical University of Innsbruck, Innsbruck, Austria

Transport of spermatozoa from the vagina to the site of fertilization in the pars ampullaris of the Fallopian tube represents one of the critical steps in the process of reproduction. Mechanical patency, as well as functional integrity, of the uterus and the oviducts are required for its successful completion. In this presentation physiological and pathophysiological data on normal and disturbed uterine transport mechanisms in human female will be presented and discussed.