Cyclic strain-mediated regulation of vascular endothelial cell migration and tube formation

Hemodynamic forces exerted by blood flow (cyclic strain, shear stress) affect the initiation and progression of angiogenesis; however, the precise signaling mechanism(s) involved are unknown. In this study, we examine the role of cyclic strain in regulating bovine aortic endothelial cell (BAEC) migration and tube formation, indices of angiogenesis. Considering their well-documented mechanosensitivity, functional inter-dependence, and involvement in angiogenesis, we hypothesized roles for matrix metalloproteinases (MMP-2/9), RGD-dependent integrins, and urokinase plasminogen activator (uPA) in this process. BAECs were exposed to equibiaxial cyclic strain (5% strain, 1Hz for 24h) before their migration and tube formation was assessed by transwell migration and collagen gel tube formation assays, respectively. In response to strain, both migration and tube formation were increased by 1.83+/-0.1- and 1.84+/-0.1-fold, respectively. Pertussis toxin, a Gi-protein inhibitor, decreased strain-induced migration by 45.7+/-32% and tube formation by 69.8+/-13%, whilst protein tyrosine kinase (PTK) inhibition with genistein had no effect. siRNA-directed attenuation of endothelial MMP-9 (but not MMP-2) expression/activity decreased strain-induced migration and tube formation by 98.6+/-41% and 40.7+/-31%, respectively. Finally, integrin blockade with cRGD peptide and siRNA-directed attenuation of uPA expression reduced strain-induced tube formation by 85.7+/-15% and 84.7+/-31%, respectively, whilst having no effect on migration. CONCLUSIONS: Cyclic strain promotes BAEC migration and tube formation in a Gi-protein-dependent PTK-independent manner. Moreover, we demonstrate for the first time a putative role for MMP-9 in both strain-induced events, whilst RGD-dependent integrins and uPA appear only to be involved in strain-induced tube formation.     

IgE-dependent regulation of human basophil adherence to vascular endothelium

Mechanisms favoring the recruitment of circulating human basophils to extravascular sites of allergic inflammation are unknown. The basophil secretagogues anti-IgE, and pollen allergens rye grass I and ragweed Ag E (Lol p I and Amb a I) were tested for their ability to promote basophil adherence to umbilical vein endothelial cells. Co-incubation of endothelial cells and basophils with anti-IgE resulted in time and dose-dependent increases in basophil adhesion. These effects were due to activation of the basophil, required both magnesium and calcium, occurred before or in the absence of histamine release, and were seen at concentrations of stimulus below the usual range of secretagogue activity. In contrast, anti-IgE or Ag stimulation of neutrophils, or basophils from donors non-responsive to anti-IgE or Ag with respect to histamine release, had no effect on cell adherence. mAb 60.3, recognizing the CD18 leukocyte adhesion molecule, inhibited anti-IgE-induced enhancement of basophil-endothelial cell binding. Exposure of basophils to low concentrations of Ag in vivo may selectively initiate basophil infiltration into tissue sites of allergic inflammation by enhancing their adherence to endothelium.     

Cyclic changes in the matrix metalloproteinase system in the ovary and uterus

With each estrous or menstrual cycle, extensive alterations occur in the extracellular matrix and connective tissue of the ovary and uterus. In the ovary, these changes occur during follicular development, breakdown of the follicular wall and extrusion of the oocyte, as well as during the formation and regression of the corpus luteum. In the uterus, the endometrium undergoes dramatic connective tissue turnover associated with tissue breakdown and subsequent regrowth during each menstrual cycle. These changes in the ovarian and uterine extracellular architecture are regulated, in part, by the matrix metalloproteinase (MMP) system. This system is comprised of both a proteolytic component, the MMPs, and associated inhibitors, and it is involved in connective tissue remodeling processes throughout the body. The current review highlights the key features of the MMP system and focuses on the changes in the MMPs and the tissue inhibitors of metalloproteinases during the dynamic remodeling that takes place in the ovary and uterus during the estrous and menstrual cycles.     

The cell surface: the stage for matrix metalloproteinase regulation of migration

Matrix metalloproteinases are important for the turnover of extracellular matrix in tissue. Recent studies have expanded their roles well beyond extracellular matrix degradation - they also cleave many growth factors, cytokines and cell adhesion molecules in the extracellular milieu, modulating their functions irreversibly. In particular, some matrix metalloproteinases that associate with the cell surface have arisen as intriguing regulators of cellular functions, including migration.     

Effect of contact time and force on monocyte adhesion to vascular endothelium

In this study we examined whether monocytic cell attachment to vascular endothelium was affected by elevating shear stress at a constant shear rate. Contact time, which is inversely related to the shear rate, was fixed and viscosity elevated with dextran to increase the shear stress (and hence the net force on the cell) independently of shear rate. At a fixed contact time, tethering frequencies increased, rolling velocities decreased, and median arrest durations increased with increasing shear stress. Rolling and short arrests (< 0.2 s) were well fit by a single exponential consistent with adhesion via the formation of a single additional bond. The cell dissociation constant, k(off), increased when the shear stress was elevated at constant shear rate. Firmly adherent cells arresting for at least 0.2 s were well fit by a stochastic model involving dissociation from multiple bonds. Therefore, at a fixed contact time and increasing shear stress, bonds formed more frequently for rolling cells resulting in more short arrests, and more bonds formed for firmly arresting cells resulting in longer arrest durations. Possible mechanisms for this increased adhesion include greater monocyte deformation and/or more frequent penetration of microvilli through steric and charge barriers.     

Low matrix metalloproteinase levels precede vascular lesion formation in the JCR:LA-cp rat

Molecular and Cellular Biochemistry - Clinically significant occlusive vascular lesions contain more extracellular matrix (ECM) proteins and lipid deposition than healthy vascular tissue. The...     

Not a German past to be reckoned with: negotiating migrant subjectivities between Vergangenheitsbewältigung and the nationalization of history

In discussing the public commemoration of the Armenian genocide and the case of a Turkish refugee‐turned‐memory‐activist in Germany, this article focuses on migrants as they mediate political concerns by reference to the German discourse and practice of Vergangenheitsbewältigung (confronting the past). Such engagements shed light on the intersection of a performative and inclusionary dimension of citizenship inscribed in the figure of the repentant perpetrator and the continuous re‐ethnicization of subjects by reference to particular nationalized histories. The analysis thus reveals the ‘memorial’ contour of a normative German citizen‐subject, one that is self‐reflexive vis‐à‐vis his or her national past. The article charts how the politics of history and citizenship thus play out at the level of subjectivity, implicating registers of affect and aesthetics that are not captured by the more commonly employed framework of ‘memory’, which falls short of analytical import precisely because it lacks a theory of the subject.     

Cyclic phosphinamides and phosphonamides, novel series of potent matrix metalloproteinase inhibitors with antitumour activity

The design, synthesis, and structure–activity relationship (SAR) of a series of novel nonpeptidic cyclic phosphon- and phosphinamide-based hydroxamic acids as inhibitors of matrix metalloproteinases MMP-1, MMP-3, and MMP-9 are presented. Based on modelling studies and X-ray analysis, a model of the binding mode of these novel compounds in the MMP active site was obtained. This model provided a rational explanation for the observed SAR data, which included a systematic study of different S1′ directed substituents, zinc-complexing groups, chirality, and variation of the cyclic phosphon- and phosphinamide rings. The in vivo effect of four compounds in a human fibrosarcoma mouse model (HT1080) was evaluated and compared to that of a reference compound, Prinomastat. Inhibition of tumour growth was observed for all four compounds.     

Hemogenic endothelium: origins, regulation, and implications for vascular biology

The study of endothelial development has been intertwined with hematopoiesis since the early 20th century when a bi-potential cell (hemangioblast) was noted to produce both endothelial and hematopoietic cells. Since then, ideas regarding the nature of connection between the vascular and hematopoietic systems have ranged from a tenuous association to direct lineage origination. In this review, historical data that spans hematopoietic development is examined within the context of hemogenic endothelium. Hemogenic endothelium, a specialized endothelial population capable of hematopoiesis, is an emerging theory that has recently gained momentum. Evidence across species and decades are reviewed, as are the possible modulators of the phenomenon, which include pathways that specify definitive hematopoiesis (Runx1), arterial identity (Notch1), as well as physiological and developmental factors.     

The bimodal regulation of vascular function by superoxide anion: role of endothelium

Reactive oxygen species (ROS) are implicated in vascular homeostasis. This study investigated whether O(2) (*-), the foundationmolecule of all ROS, regulates vasomotor function. Hence, vascular reactivity was measured using rat thoracic aortas exposed to an O(2) (*-) generator (pyrogallol) which dose-dependently regulated both alpha-adrenergic agonist-mediated contractility to phenylephrine and endothelium-dependent relaxations to acetylcholine. Pyrogallol improved and attenuated responses to acetylcholine at its lower (10 nM - 1 microM) and higher (10 - 100 microM) concentrations, respectively while producing the inverse effects with phenylephrine. The endothelial inactivation by L-NAME abolished acetylcholine-induced vasodilatations but increased phenylephrine and KCl-induced vasoconstrictions regardless of the pyrogallol dose used. Relaxant responses to sodium nitroprusside, a nitric oxide donor, were not affected by pyrogallol. Other ROS i.e. peroxynitrite and H(2)O(2) that may be produced during experiments did not alter vascular functions. These findings suggest that the nature of O(2) (*-)-evoked vascular function is determined by its local concentration and the presence of a functional endothelium.     

The adherence of Candida yeasts to human and bovine vascular endothelium and subendothelial extracellular matrix

Abstract The adherence of Candida albicans yeasts and other Candida species to human umbilical vein endothelial cells (HUVEC), bovine aortic endothelial cells (BAEC) and their respective subendothelial extracellular matrices (ECM) was studied. Yeast adherence to confluent HUVEC and BAEC appeared to occur at intercellular junctions and edges of endothelial cells in preference to the contoured surface of the endothelial cell. Both endothelial cell lines characteristically resisted yeast adherence. The resistance of endothelium to yeast adherence was especially pronounced in the presence of serum. On the other hand, there was avid yeast binding to the subendothelial ECMs, on the order of 200–500% greater than to monolayers of syngeneic endothelial cells.     

Biopsy coupled to quantitative immunofluorescence: a new method to study the human vascular endothelium.

Limited availability of endothelial tissue is a major constraint when investigating the cellular mechanisms of endothelial dysfunction in patients with metabolic and cardiovascular diseases. We propose a novel approach that combines collection of 200-1,000 endothelial cells from a superficial forearm vein or the radial artery, with reliable measurements of protein expression by quantitative immunofluorescence analysis. This method was validated against immunoblot analysis in cultured endothelial cells. Levels of vascular endothelial cell activation, oxidative stress, and nitric oxide synthase expression were measured and compared in five patients with severe chronic heart failure and in four healthy age-matched subjects. In summary, vascular endothelial biopsy coupled with measurement of protein expression by quantitative immunofluorescence analysis provides a novel approach to the study of the vascular endothelium in humans.     

Sequencing and characterization of the soybean leaf metalloproteinase : structural and functional similarity to the matrix metalloproteinase family

A novel zinc endoproteinase has been sequenced and characterized from soybean leaves (Glycine max var Williams 82) and has been designated as Protein Identification Resource accession No. A41820 SMEP1 (soybean metalloendoproteinase 1). Comparison of the primary amino acid sequence with other zinc proteinases revealed the enzyme to be a new member of the matrix metalloproteinase (MMP) family of enzymes. SMEP was found to have MMP cleavage specificity toward peptide substrates and the enzyme is specifically inhibited by naturally occurring tissue inhibitors of MMPs through a high-affinity interaction (inhibitor concentration resulting in an approximate 50% decrease in enzyme activity = 23 x 10(-9) molar). Together, these results suggest that the origin of the MMP family of enzymes and their cognate inhibitors predates the divergence of plants and animals.     

Epithelial organization: may the force be with you

Multicellular organization is determined by the balance of forces between cells as much as by the expression of genes. A recent study in Drosophila combines physical modeling with experimental measurements and mechanical perturbations to shed light on the processes that influence cell patterns in vivo.     

X-ray structure of a novel matrix metalloproteinase inhibitor complexed to stromelysin

A new class of matrix metalloproteinase (MMP) inhibitors has been identified by screening a collection of compounds against stromelysin. The inhibitors, 2,4,6-pyrimidine triones, have proven to be potent inhibitors of gelatinases A and B. An X-ray crystal structure of one representative compound bound to the catalytic domain of stromelysin shows that the compounds bind at the active site and ligand the active-site zinc. The pyrimidine triones mimic substrates in forming hydrogen bonds to key residues in the active site, and provide opportunities for placing appropriately chosen groups into the S1' specificity pocket of MMPS: A number of compounds have been synthesized and assayed against stromelysin, and the variations in potency are explained in terms of the binding mode revealed in the X-ray crystal structure.