Endothelium forms a semi-permeable barrier that separates blood from the underlying tissue. Barrier function is largely determined by cell-cell and cell-matrix adhesions that define the limits of ...cell borders. Yet, such cell-cell and cell-matrix tethering is critically reliant upon the nature of adherence within the cell itself. Indeed, the actin cytoskeleton fulfills this essential function, to provide a strong, dynamic intracellular scaffold that organizes integral membrane proteins with the cell's interior, and responds to environmental cues to orchestrate appropriate cell shape. The actin cytoskeleton is comprised of three distinct, but inter-related structures, including actin cross-linking of spectrin within the membrane skeleton, the cortical actin rim, and actomyosin-based stress fibers. This review addresses each of these actin-based structures, and discusses cellular signals that control the disposition of actin in different endothelial cell phenotypes.
Development and Pathology of Pulmonary Hypertension Tuder, Rubin M., MD; Abman, Steven H., MD; Braun, Thomas, MD, PhD ...
Journal of the American College of Cardiology,
06/2009, Letnik:
54, Številka:
1
Journal Article
Recenzirano
Odprti dostop
The Development and Pathology working group was charged with reviewing the present knowledge, gaps in understanding, and areas for further studies in a broad range of themes. These themes in ...pulmonary vascular biology and pathobiology involved: 1) pulmonary vascular development; 2) pulmonary vascular disease accompanying fetal development and perinatal life; 3) properties of pulmonary vascular endothelial cells; 4) role of bone marrow cells in pulmonary vascular disease; 5) insights into pulmonary thromboembolic disease; 6) role of pathology in the assessment of pulmonary vascular disease; and 7) considerations of tissue banking for research in pulmonary hypertension. These important goals provide a blueprint for future research that may significantly impact our present and future understanding of pulmonary hypertension.
Pain, redness, heat, and swelling are hallmarks of inflammation that were recognized as early as the first century AD. Despite these early observations, the mechanisms responsible for swelling, in ...particular, remained an enigma for nearly two millennia. Only in the past century have scientists and physicians gained an appreciation for the role that vascular endothelium plays in controlling the exudation that is responsible for swelling. One of these mechanisms is the formation of transient gaps between adjacent endothelial cell borders. Inflammatory mediators act on endothelium to reorganize the cytoskeleton, decrease the strength of proteins that connect cells together, and induce transient gaps between endothelial cells. These gaps form a paracellular route responsible for exudation. The discovery that interendothelial cell gaps are causally linked to exudation began in the 1960s and was accompanied by significant controversy. Today, the role of gap formation in tissue edema is accepted by many, and significant scientific effort is dedicated toward developing therapeutic strategies that will prevent or reverse the endothelial cell gaps that are present during the course of inflammatory illness. Given the importance of this field in endothelial cell biology and inflammatory disease, this focused review catalogs key historical advances that contributed to our modern-day understanding of the cell biology of interendothelial gap formation.
Collective migration of endothelial cells is important for wound healing and angiogenesis. During such migration, each constituent endothelial cell coordinates its magnitude and direction of ...migration with its neighbors while retaining intercellular adhesion. Ensuring coordination and cohesion involves a variety of intra- and inter-cellular signaling processes. However, the role of permeation of extracellular Na+ in collective cell migration remains unclear. Here, we examined the effect of Na+ permeation in collective migration of pulmonary artery endothelial cell (PAEC) monolayers triggered by either a scratch injury or a barrier removal over 24 hours. In the scratch assay, PAEC monolayers migrated in two approximately linear phases. In the first phase, wound closure started with fast speed which then rapidly reduced within 5 hours after scratching. In the second phase, wound closure maintained at slow and stable speed from 6 to 24 hours. In the absence of extracellular Na+, the wound closure distance was reduced by >50%. Fewer cells at the leading edge protruded prominent lamellipodia. Beside transient gaps, some sustained interendothelial gaps also formed and progressively increased in size over time, and some fused with adjacent gaps. In the absence of both Na+ and scratch injury, PAEC monolayer migrated even more slowly, and interendothelial gaps obviously increased in size towards the end. Pharmacological inhibition of the epithelial Na+ channel (ENaC) using amiloride reduced wound closure distance by 30%. Inhibition of both the ENaC and the Na+/Ca2+ exchanger (NCX) using benzamil further reduced wound closure distance in the second phase and caused accumulation of floating particles in the media. Surprisingly, pharmacological inhibition of the Ca2+ release-activated Ca2+ (CRAC) channel protein 1 (Orai1) using GSK-7975A, the transient receptor potential channel protein 1 and 4 (TRPC1/4) using Pico145, or both Orai1 and TRPC1/4 using combined GSK-7975A and Pico145 treatment did not affect wound closure distance dramatically. Nevertheless, the combined treatment appeared to cause accumulation of floating particles. Note that GSK-7975A also inhibits small inward Ca2+ currents via Orai2 and Orai3 channels, whereas Pico145 also blocks TRPC4, TRPC5, and TRPC1/5 channels. By contrast, gene silence of Orai1 by shRNAs led to a 25% reduction of wound closure in the first 6 hours but had no effect afterwards. However, in the absence of extracellular Na+ or cellular injury, Orai1 did not affect PAEC collective migration. Overall, the data reveal that Na+ permeation into cells contributes to PAEC monolayer collective migration by increasing lamellipodial formation, reducing accumulation of floating particles, and improving intercellular adhesion.
Exoenzyme Y (ExoY) is a Pseudomonas aeruginosa toxin that is introduced into host cells through the type 3 secretion system (T3SS). Once inside the host cell cytoplasm, ExoY generates cyclic ...nucleotides that cause tau phosphorylation and microtubule breakdown. Microtubule breakdown causes interendothelial cell gap formation and tissue edema. Although ExoY transiently induces interendothelial cell gap formation, it remains unclear whether ExoY prevents repair of the endothelial cell barrier. Here, we test the hypothesis that ExoY intoxication impairs recovery of the endothelial cell barrier following gap formation, decreasing migration, proliferation, and lung repair. Pulmonary microvascular endothelial cells (PMVECs) were infected with P. aeruginosa strains for 6 h, including one possessing an active ExoY (PA103 exoUexoT::Tc pUCPexoY; ExoY(+)), one with an inactive ExoY (PA103ΔexoUexoT::Tc pUCPexoY(K81M); ExoY(K81M)), and one that lacks PcrV required for a functional T3SS (ΔPcrV). ExoY(+) induced interendothelial cell gaps, whereas ExoY(K81M) and ΔPcrV did not promote gap formation. Following gap formation, bacteria were removed and endothelial cell repair was examined. PMVECs were unable to repair gaps even 3-5 days after infection. Serum-stimulated growth was greatly diminished following ExoY intoxication. Intratracheal inoculation of ExoY(+) and ExoY(K81M) caused severe pneumonia and acute lung injury. However, whereas the pulmonary endothelial cell barrier was functionally improved 1 wk following ExoY(K81M) infection, pulmonary endothelium was unable to restrict the hyperpermeability response to elevated hydrostatic pressure following ExoY(+) infection. In conclusion, ExoY is an edema factor that chronically impairs endothelial cell barrier integrity following lung injury.
Transmembrane heparan sulfate proteoglycans regulate multiple aspects of cell behavior, but the molecular basis of their signaling is unresolved. The major family of transmembrane proteoglycans is ...the syndecans, present in virtually all nucleated cells, but with mostly unknown functions. Here, we show that syndecans regulate transient receptor potential canonical (TRPCs) channels to control cytosolic calcium equilibria and consequent cell behavior. In fibroblasts, ligand interactions with heparan sulfate of syndecan-4 recruit cytoplasmic protein kinase C to target serine714 of TRPC7 with subsequent control of the cytoskeleton and the myofibroblast phenotype. In epidermal keratinocytes a syndecan-TRPC4 complex controls adhesion, adherens junction composition, and early differentiation in vivo and in vitro. In Caenorhabditis elegans, the TRPC orthologues TRP-1 and -2 genetically complement the loss of syndecan by suppressing neuronal guidance and locomotory defects related to increases in neuronal calcium levels. The widespread and conserved syndecan-TRPC axis therefore fine tunes cytoskeletal organization and cell behavior.
The plexiform lesions of severe pulmonary arterial hypertension (PAH) are similar in histologic appearance, whether the disease is idiopathic or secondary. Both forms of the disease show actively ...proliferating endothelial cells without evidence of apoptosis. Here, we discuss the pathobiology of the atypical, angioproliferative endothelial cells in severe PAH. The concept of the endothelial cell as a "quasi-malignant" cell provides a new framework for antiproliferative, antiangiogenic therapy in severe PAH.
Idiopathic pulmonary arterial hypertension is a progressive and potentially fatal disease with a limited number of therapeutic options. Two key lesions underlie the pathophysiology of this disease. ...The principal lesion is found in large- and intermediate-sized blood vessels and is characterized by medial and adventitial hypertrophy/hyperplasia, with distal extension of smooth-muscle layers into normally nonmuscularized vessels. The second lesion, found prominently in severe forms of pulmonary hypertension, originates in small precapillary vessel segments, commonly at blood vessel bifurcations. This “plexiform lesion” is a lumen-obliterative lesion comprised, at least in part, of cells that share endothelial cell attributes, but that have lost the “law of the monolayer.” Indeed, the endothelial contribution to the (mal-)adaptive response in pulmonary hypertension is becoming increasingly apparent, with evidence that endothelium plays an important role in promoting the vasoconstriction and hyperproliferation of medial and adventitial cell layers in large- and intermediate-vessel sizes, and lumen obliteration in the plexiform lesion. Recent evidence indicates endothelial cells along the pulmonary artery and precapillary segments are phenotypically distinct and may fulfill different roles in these site-specific lesions. Thus, the present review summarizes our current understanding of pulmonary endothelial cell heterogeneity and discusses the potential role(s) of endothelial cell heterogeneity in the pathogenesis of pulmonary hypertension.
Acidosis is common among critically ill patients, but current approaches to correct pH do not improve disease outcomes. During systemic acidosis, cells are either passively exposed to extracellular ...acidosis that other cells have generated (extrinsic acidosis) or they are exposed to acid that they generate and export into the extracellular space (intrinsic acidosis). Although endothelial repair following intrinsic acidosis has been studied, the impact of extrinsic acidosis on migration and angiogenesis is unclear. We hypothesized that extrinsic acidosis inhibits metabolism and migration but promotes capillary-like network formation in pulmonary microvascular endothelial cells (PMVECs). Extrinsic acidosis was modeled by titrating media pH. Two types of intrinsic acidosis were compared, including increasing cellular metabolism by chemically inhibiting carbonic anhydrases (CAs) IX and XII (SLC-0111) and with hypoxia. PMVECs maintained baseline intracellular pH for 24 h with both extrinsic and intrinsic acidosis. Whole cell CA IX protein expression was decreased by extrinsic acidosis but not affected by hypoxia. When extracellular pH was equally acidic, extrinsic acidosis suppressed glycolysis, whereas intrinsic acidosis did not. Extrinsic acidosis suppressed migration, but increased Matrigel network master junction and total segment length. CRISPR-Cas9 CA IX knockout PMVECs revealed an independent role of CA IX in promoting glycolysis, as loss of CA IX alone was accompanied by decreased hexokinase I and pyruvate dehydrogenase E1α expression and decreasing migration. 2-deoxy-d-glucose had no effect on migration but profoundly inhibited network formation and increased N-cadherin expression. Thus, we report that while extrinsic acidosis suppresses endothelial glycolysis and migration, it promotes network formation.
PDF
