scholarly journals LPHN2 inhibits vascular permeability by differential control of endothelial cell adhesion

2021 ◽  
Vol 220 (11) ◽  
Author(s):  
Chiara Camillo ◽  
Nicola Facchinello ◽  
Giulia Villari ◽  
Giulia Mana ◽  
Noemi Gioelli ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Focal Adhesion ◽  
Nuclear Localization ◽  
Molecular Mechanisms ◽  
Transcriptional Regulators ◽  
Dependent Manner ◽  
Knock Out ◽  
Dynamic Modulation ◽  
Differential Control ◽  
G Protein Coupled

Dynamic modulation of endothelial cell-to-cell and cell–to–extracellular matrix (ECM) adhesion is essential for blood vessel patterning and functioning. Yet the molecular mechanisms involved in this process have not been completely deciphered. We identify the adhesion G protein–coupled receptor (ADGR) Latrophilin 2 (LPHN2) as a novel determinant of endothelial cell (EC) adhesion and barrier function. In cultured ECs, endogenous LPHN2 localizes at ECM contacts, signals through cAMP/Rap1, and inhibits focal adhesion (FA) formation and nuclear localization of YAP/TAZ transcriptional regulators, while promoting tight junction (TJ) assembly. ECs also express an endogenous LPHN2 ligand, fibronectin leucine-rich transmembrane 2 (FLRT2), that prevents ECM-elicited EC behaviors in an LPHN2-dependent manner. Vascular ECs of lphn2a knock-out zebrafish embryos become abnormally stretched, display a hyperactive YAP/TAZ pathway, and lack proper intercellular TJs. Consistently, blood vessels are hyperpermeable, and intravascularly injected cancer cells extravasate more easily in lphn2a null animals. Thus, LPHN2 ligands, such as FLRT2, may be therapeutically exploited to interfere with cancer metastatic dissemination.

scholarly journals Transcellular migration of leukocytes is mediated by the endothelial lateral border recycling compartment

2009 ◽  
Vol 206 (12) ◽  
pp. 2795-2808 ◽  
Author(s):  
Zahra Mamdouh ◽  
Alexei Mikhailov ◽  
William A. Muller
Keyword(s):  
Endothelial Cell ◽  
Adhesion Molecule ◽  
Molecular Mechanisms ◽  
Vascular Endothelial Cell ◽  
Dependent Manner ◽  
Vascular Endothelial ◽  
Lateral Border ◽  
Endothelial Surface ◽  
Platelet Endothelial Cell Adhesion ◽  
Intracellular Adhesion Molecule

Leukocyte migration across endothelial cell borders (paracellular) and through endothelial cells (transcellular) appear to be distinct processes. During paracellular migration, membrane from a parajunctional reticulum of interconnected vesicles, the endothelial lateral border recycling compartment (LBRC), moves to surround the leukocyte in a kinesin-mediated, microtubule-dependent manner. We show that transcellular migration likewise requires targeted trafficking of LBRC membrane. We show that in addition to platelet/endothelial cell adhesion molecule (PECAM; CD31), CD99 and junctional adhesion molecule A (JAM-A), but apparently not vascular endothelial cell–specific cadherin (cadherin 5, CD144), are components of the LBRC. During transcellular migration, LBRC membrane invests the transmigrating leukocyte. Intracellular adhesion molecule 1 (ICAM-1) on the apical endothelial surface is enriched around adherent leukocytes. Depolymerization of microtubules has no effect on ICAM-1 enrichment but blocks targeted trafficking of LBRC membrane and transcellular migration by >90%. Similar to their effects on paracellular transmigration, antibodies against PECAM or CD99, but not JAM-A, block transcellular migration. We conclude that similar molecular mechanisms promote both para- and transcellular migration.

scholarly journals Impact of Epithelial–Stromal Interactions on Peritumoral Fibroblasts in Ductal Carcinoma in Situ

2019 ◽  
Vol 111 (9) ◽  
pp. 983-995 ◽  
Author(s):  
Carina Strell ◽  
Janna Paulsson ◽  
Shao-Bo Jin ◽  
Nicholas P Tobin ◽  
Artur Mezheyeuski ◽  
...  
Keyword(s):  
Carcinoma In Situ ◽  
Poor Prognosis ◽  
Molecular Mechanisms ◽  
Statistical Tests ◽  
Hazard Ratio ◽  
Dependent Manner ◽  
Knock Out ◽  
General Terms ◽  
Increased Risk

AbstractBackgroundA better definition of biomarkers and biological processes related to local recurrence and disease progression is highly warranted for ductal breast carcinoma in situ (DCIS). Stromal–epithelial interactions are likely of major importance for the biological, clinical, and pathological distinctions between high- and low-risk DCIS cases.MethodsStromal platelet derived growth factor receptor (PDGFR) was immunohistochemically assessed in two DCIS patient cohorts (n = 458 and n = 80). Cox proportional hazards models were used to calculate the hazard ratios of recurrence. The molecular mechanisms regulating stromal PDGFR expression were investigated in experimental in vitro co-culture systems of DCIS cells and fibroblasts and analyzed using immunoblot and quantitative real-time PCR. Knock-out of JAG1 in DCIS cells and NOTCH2 in fibroblasts was obtained through CRISPR/Cas9. Experimental data were validated by mammary fat pad injection of DCIS and DCIS-JAG1 knock-out cells (10 mice per group). All statistical tests were two-sided.ResultsPDGFRα(low)/PDGFRβ(high) fibroblasts were associated with increased risk for recurrence in DCIS (univariate hazard ratio = 1.59, 95% confidence interval [CI] = 1.02 to 2.46; P = .04 Wald test; multivariable hazard ratio = 1.78, 95% CI = 1.07 to 2.97; P = .03). Tissue culture and mouse model studies indicated that this fibroblast phenotype is induced by DCIS cells in a cell contact-dependent manner. Epithelial Jagged1 and fibroblast Notch2 were identified through loss-of-function studies as key juxtacrine signaling components driving the formation of the poor prognosis-associated fibroblast phenotype.ConclusionsA PDGFRα(low)/PDGFRβ(high) fibroblast subset was identified as a marker for high-risk DCIS. The Jagged-1/Notch2/PDGFR stroma–epithelial pathway was described as a novel signaling mechanism regulating this poor prognosis-associated fibroblast subset. In general terms, the study highlights epithelial–stromal crosstalk in DCIS and contributes to ongoing efforts to define clinically relevant fibroblast subsets and their etiology.

Regulation of endothelial cell morphogenesis by the protein kinase D (PKD)/glycogen synthase kinase 3 (GSK3)β pathway

2012 ◽  
Vol 303 (7) ◽  
pp. C743-C756 ◽  
Author(s):  
Sejeong Shin ◽  
Laura Wolgamott ◽  
Sang-Oh Yoon
Keyword(s):  
Endothelial Cell ◽  
Protein Kinase ◽  
Molecular Mechanisms ◽  
Glycogen Synthase ◽  
Collagen I ◽  
Glycogen Synthase Kinase ◽  
Protein Kinase D ◽  
Dependent Manner ◽  
Vascular Morphogenesis ◽  
Tube Morphogenesis

Vascular morphogenesis is a key process for development, reproduction, and pathogenesis. Thus understanding the mechanisms of this process is of pathophysiological importance. Despite the fact that collagen I is the most abundant and potent promorphogenic molecule known, the molecular mechanisms by which this protein regulates endothelial cell tube morphogenesis are still unclear. Here we provide strong evidence that collagen I induces tube morphogenesis by inhibiting glycogen synthase kinase 3β (GSK3β). Further mechanistic studies revealed that GSK3β activity is regulated by protein kinase D (PKD). PKD inhibited GSK3β activity, which was required for collagen I-induced endothelial tube morphogenesis. We also found that GSK3β regulated trafficking of integrin α2β1 in a Rab11-dependent manner. Taken together, our studies highlight the important role of PKD in the regulation of collagen I-induced vascular morphogenesis and show that it is mediated by the modulation of GSK3β activity and integrin α2β1 trafficking.

scholarly journals Identification of a putative nuclear localization signal in maspin protein shed light into its nuclear import regulation

2018 ◽  
Author(s):  
Jeffrey Reina ◽  
Lixin Zhou ◽  
Marcos R.M. Fontes ◽  
Nelly Panté ◽  
Nathalie Cella
Keyword(s):  
Subcellular Localization ◽  
Nuclear Localization ◽  
Hela Cells ◽  
Nuclear Import ◽  
Nuclear Localization Signal ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Biological Activities ◽  
Dependent Manner ◽  
Localization Signal

AbstractMaspin (SERPINB5) is a potential tumor suppressor gene with pleiotropic biological activities, including regulation of cell proliferation, death, adhesion, migration and gene expression. Several studies suggest that subcellular localization plays an essential role on maspin tumor suppression activity. In this study we investigated the molecular mechanisms underlying maspin nucleocytoplasmic shuttling. Anin vitronuclear-import assay using digitonin-permeabilized HeLa cells demonstrated that maspin enters the nucleus by an energy-and carrier-independent mechanism. However, previous studies indicated that maspin subcellular localization is regulated in the cell. Using a nuclear localization signal (NLS) prediction software, we identified a putative NLS in the maspin amino acid sequence. To distinguish between passive and regulated nuclear translocation, maspinNLS or the full-length protein (MaspinFL) were fused to 5GFP, rendering the construct too large to enter the nucleus passively. Unexpectedly, 5GFP-maspinNLS, but not maspinFL-5GFP, entered the nucleus of HeLa cells. Dominant-negative Ran-GTPase mutants RanQ69L or RanT24N, suppressed 5GFP-maspinNLS nuclear localization. In summary, we provide evidence that maspin translocates to the nucleus passively. In addition, we identified a peptide in the maspin protein sequence, which is able to drive a 5GFP construct to the nucleus in an energy-dependent manner.

N-3-oxododecanoyl homoserine lactone exacerbates endothelial cell death by inducing receptor-interacting protein kinase 1-dependent apoptosis

2021 ◽  
Author(s):  
Jungho Shin ◽  
Sun Hee Ahn ◽  
Su Hyun Kim ◽  
Dong-Jin Oh
Keyword(s):  
Endothelial Cells ◽  
Cell Death ◽  
Endothelial Cell ◽  
Protein Kinase ◽  
Quorum Sensing ◽  
Molecular Mechanisms ◽  
Homoserine Lactone ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Dependent Manner ◽  
Interacting Protein

Endothelial dysfunction is associated with the initiation of sepsis-associated organ failure. Bacterial quorum-sensing molecules act as pathogen-associated molecular patterns; however, the effects of quorum-sensing molecules on endothelial cells remain less understood. This study investigated the molecular mechanisms of quorum sensing molecule-induced cell death and their interaction with lipopolysaccharide (LPS) in human umbilical vein endothelial cells. Endothelial cells were treated with N-3-oxododecanoyl homoserine lactone (3OC12-HSL) and LPS derived from Pseudomonas aeruginosa. Treatment with 3OC12-HSL reduced cell viability in a dose-dependent manner, and co-treatment with 3OC12-HSL and LPS enhanced cell death. Terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling assay revealed an increase in apoptotic cell death following 3OC12-HSL treatment; furthermore, co-treatment with 3OC12-HSL and LPS enhanced apoptosis. Western blotting revealed that treatment with 3OC12-HSL activated the receptor-interacting protein kinase 1 (RIPK1) pathway, leading to an increase in the levels of cleaved caspase 8 and 3. In addition, we found that treatment with necrostatin-1, an RIPK1 inhibitor, reduced cell death and ameliorated the activation of the RIPK1-dependent apoptotic pathway in 3OC12-HSL-treated cells. In conclusion, 3OC12-HSL induced endothelial cell apoptosis via the activation of the RIPK1 pathway, independent of LPS toxicity. Inhibition of RIPK1 may act as a therapeutic option for preserving endothelial cell integrity in patients with sepsis by disrupting the mechanism by which quorum-sensing molecules mediate their toxicity.

scholarly journals Immune Cell Induced Migration of Osteoprogenitor Cells Is Mediated by TGF-β Dependent Upregulation of NOX4 and Activation of Focal Adhesion Kinase

2018 ◽  
Vol 19 (8) ◽  
pp. 2239 ◽  
Author(s):  
Sabrina Ehnert ◽  
Caren Linnemann ◽  
Romina Aspera-Werz ◽  
Daria Bykova ◽  
Sara Biermann ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Immune Cell ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Conditioned Media ◽  
Dependent Manner ◽  
Osteoprogenitor Cells ◽  
Nadph Oxidase 4

The cytokines secreted by immune cells have a large impact on the tissue, surrounding a fracture, e.g., by attraction of osteoprogenitor cells. However, the underlying mechanisms are not yet fully understood. Thus, this study aims at investigating molecular mechanisms of the immune cell-mediated migration of immature primary human osteoblasts (phOBs), with transforming growth factor beta (TGF-β), nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX4) and focal adhesion kinase (FAK) as possible regulators. Monocyte- and macrophage (THP-1 cells ± phorbol 12-myristate 13-acetate (PMA) treatment)-conditioned media, other than the granulocyte-conditioned medium (HL-60 cells + dimethyl sulfoxide (DMSO) treatment), induce migration of phOBs. Monocyte- and macrophage (THP-1 cells)-conditioned media activate Smad3-dependent TGF-β signaling in the phOBs. Stimulation with TGF-β promotes migration of phOBs. Furthermore, TGF-β treatment strongly induces NOX4 expression on both mRNA and protein levels. The associated reactive oxygen species (ROS) accumulation results in phosphorylation (Y397) of FAK. Blocking TGF-β signaling, NOX4 activity and FAK signaling effectively inhibits the migration of phOBs towards TGF-β. In summary, our data suggest that monocytic- and macrophage-like cells induce migration of phOBs in a TGF-β-dependent manner, with TGF-β-dependent induction of NOX4, associated production of ROS and resulting activation of FAK as key mediators.

scholarly journals Murine glomerular transcriptome links endothelial cell-specific molecule-1 deficiency with susceptibility to diabetic nephropathy

2017 ◽  
Author(s):  
Xiaoyi Zheng ◽  
Fariborz Soroush ◽  
Jin Long ◽  
Evan T. Hall ◽  
Puneeth K. Adishesha ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
Endothelial Cell ◽  
Molecular Mechanisms ◽  
Disease Susceptibility ◽  
Unmet Need ◽  
Critical Factor ◽  
Dependent Manner ◽  
Non Invasive ◽  
First Time ◽  
Dose Dependent

AbstractDiabetic nephropathy (DN) is the leading cause of kidney disease; however, there are no early biomarkers and no cure. Thus, there is a large unmet need to predict which individuals will develop nephropathy and to understand the molecular mechanisms which govern this susceptibility. We compared the glomerular transcriptome from mice with distinct susceptibilities to DN, and identified differential regulation of genes that modulate inflammation. From these genes, we identified endothelial cell specific molecule-1 (Esm-1), as a glomerular-enriched determinant of resistance to DN. Glomerular Esm-1 mRNA and protein were lower in DN-susceptible, DBA/2, compared to DN-resistant, C57BL/6, mice. We demonstrated higher Esm-1 secretion from primary glomerular cultures of diabetic mice, and high glucose was sufficient to increase Esm-1 mRNA and protein secretion in both strains of mice. However, induction was significantly attenuated in DN-susceptible mice. Urine Esm-1 was also significantly higher only in DN-resistant mice. Moreover, using intravital microscopy and a biomimetic microfluidic assay, we showed that Esm-1 inhibited rolling and transmigration in a dose-dependent manner. For the first time we have uncovered glomerular-derived Esm-1 as a potential non-invasive biomarker of DN. Esm-1 inversely correlates with disease susceptibility and inhibits leukocyte infiltration, a critical factor in protecting the kidney from DN.

Interaction of Shear Stress, Myosin II, and Actin in Dynamic Modulation of Endothelial Cell Microrheology

2008 ◽  
Author(s):  
Jhanvi H. Dangaria ◽  
Peter J. Butler
Keyword(s):  
Endothelial Cell ◽  
Focal Adhesion ◽  
Molecular Motors ◽  
Myosin Ii ◽  
Polymer Network ◽  
Biological Functions ◽  
Dynamic Modulation ◽  
Actomyosin Interaction ◽  
Dissipation Mechanism ◽  
Intracellular Organelles

The endothelial cell (EC) cytoskeleton mediates several biological functions such as adhesion, migration, phagocytosis, cell division, and mechanosensitivity. These functions are carried out in part through dynamic cytoskeletal polymerization, modulation of crosslinking, and development of tension between intracellular organelles and the extracellular matrix via focal adhesion plaques. One important component of the cytoskeleton is actin which polymerizes into filaments and is thought to be prestressed by virtue of crosslinking proteins such as α-actinin, filamin and myosin II molecular motors. Additionally, actomyosin interaction has been hypothesized to act as a stress dissipation mechanism by virtue of dynamic crossbridging which facilitates actin diffusion through the polymer network of the cytoplasm (Humphrey et al., 2002).

Mechanisms of ionomycin-induced endothelial cell barrier dysfunction

1997 ◽  
Vol 273 (1) ◽  
pp. L172-L184 ◽  
Author(s):  
J. G. Garcia ◽  
K. L. Schaphorst ◽  
S. Shi ◽  
A. D. Verin ◽  
C. M. Hart ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Protein Kinase ◽  
Tyrosine Kinase ◽  
Protein Kinase A ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Cell Permeability ◽  
Dependent Manner ◽  
Barrier Dysfunction ◽  
Kinase A

Myosin light chain (MLC) phosphorylation catalyzed by the Ca(2+)- calmodulin-dependent MLC kinase (MLCK) is critical to thrombin-mediated endothelial cell gap formation and barrier dysfunction. We have tested the hypothesis that the Ca2+ ionophore ionomycin stimulates MLCK-dependent endothelial cell contraction and permeability. Ionomycin significantly increased albumin clearance and decreased electrical resistance across confluent bovine pulmonary microvascular and macrovascular endothelial cell monolayers in a concentration-dependent manner that was temporally similar to that produced by thrombin. In contrast, however, ionomycin produced a significant Ca(2+)-dependent reduction in the levels of phosphorylated MLC with evidence of serine/threonine phosphatase activation. Potential MLCK-independent mechanisms of endothelial cell permeability were examined with little evidence to support a role for stimulated nitric oxide synthase or phospholipase A2 activities. Importantly, ionomycin produced 1) reductions in the activities of the barrier protective adenylate cyclase and the adenosine 3',5'-cyclic monophosphate-dependent protein kinase A, 2) dramatic dose- and time-dependent inhibition of endothelial cell tyrosine kinase activities, and 3) marked decreases in the phosphotyrosine content of the p125 focal adhesion kinase. These data indicate that ionomycin produces endothelial cell barrier dysfunction by mechanisms that are independent of MLCK activation and may involve reductions in endothelial cell tethering forces via inhibition of protein kinase A and tyrosine kinase activities, especially the p125 focal adhesion kinase.

scholarly journals MAML1/2 promote YAP/TAZ nuclear localization and tumorigenesis

2020 ◽  
Vol 117 (24) ◽  
pp. 13529-13540 ◽  
Author(s):  
Jiyoung Kim ◽  
Hyeryun Kwon ◽  
You Keun Shin ◽  
Gahyeon Song ◽  
Taebok Lee ◽  
...  
Keyword(s):  
Nuclear Localization ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Human Cancer ◽  
Hippo Pathway ◽  
Ectopic Expression ◽  
Dependent Manner ◽  
The Hippo Pathway

The Hippo pathway plays a pivotal role in tissue homeostasis and tumor suppression. YAP and TAZ are downstream effectors of the Hippo pathway, and their activities are tightly suppressed by phosphorylation-dependent cytoplasmic retention. However, the molecular mechanisms governing YAP/TAZ nuclear localization have not been fully elucidated. Here, we report that Mastermind-like 1 and 2 (MAML1/2) are indispensable for YAP/TAZ nuclear localization and transcriptional activities. Ectopic expression or depletion of MAML1/2 induces nuclear translocation or cytoplasmic retention of YAP/TAZ, respectively. Additionally, mutation of the MAML nuclear localization signal, as well as its YAP/TAZ interacting region, both abolish nuclear localization and transcriptional activity of YAP/TAZ. Importantly, we demonstrate that the level ofMAML1messenger RNA (mRNA) is regulated by microRNA-30c (miR-30c) in a cell-density-dependent manner. In vivo and clinical results suggest that MAML potentiates YAP/TAZ oncogenic function and positively correlates with YAP/TAZ activation in human cancer patients, suggesting pathological relevance in the context of cancer development. Overall, our study not only provides mechanistic insight into the regulation of YAP/TAZ subcellular localization, but it also strongly suggests that the miR30c–MAML–YAP/TAZ axis is a potential therapeutic target for developing novel cancer treatments.

Sign in / Sign up

Export Citation Format

Share Document