scholarly journals Endothelial barrier protection by FTY720 under hyperglycemic condition: involvement of focal adhesion kinase, small GTPases, and adherens junction proteins

2009 ◽  
Vol 297 (4) ◽  
pp. C945-C954 ◽  
Author(s):  
Kei Sarai ◽  
Kenichi Shikata ◽  
Yasushi Shikata ◽  
Kazuyoshi Omori ◽  
Naomi Watanabe ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Biological Effects ◽  
Adherens Junction ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Endothelial Barrier ◽  
Cortical Actin ◽  
Sphingosine 1 Phosphate ◽  
Interfering Rna

Recently, sphingosine 1-phosphate (S1P) has been highlighted as an endothelial barrier-stabilizing mediator. FTY720 is a S1P analog originally developed as a novel immunosuppressant. The phosphorylated form of FTY720 binds to S1P receptors to exert S1P-like biological effects, suggesting endothelial barrier promotion by FTY720. To elucidate whether FTY720 induces signaling events related to endothelial barrier enhancement under hyperglycemic conditions, human microvascular endothelial cells (HMVECs) preincubated with hyperglycemic (30 mM) medium were treated with 100 nM FTY720 for 3 h. Immunofluorescent microscopy and coprecipitation study revealed FTY720-induced focal adhesion kinase (FAK)-associated adherens junction (AJ) assembly at cell-cell contacts coincident with formation of a prominent cortical actin ring. FTY720 also induced transmonolayer electrical resistance (TER) augmentation in HMVEC monolayers in both normoglycemic and hyperglycemic conditions, implying endothelial barrier enhancement. Similar to S1P, site-specific FAK tyrosine phosphorylation analysis revealed FTY720-induced FAK [Y576] phosphorylation without phosphorylation of FAK [Y397/Y925]. Furthermore, FTY720 conditioned the phosphorylation profile of FAK [Y397/Y576/Y925] in hyperglycemic medium to the same pattern observed in normoglycemic medium. FTY720 challenge resulted in small GTPase Rac activation under hyperglycemic conditions, whereas increased Rho activity in hyperglycemic medium was restored to the basal level. Rac protein depletion by small interfering RNA (siRNA) technique completely abolished FTY720-induced FAK [Y576] phosphorylation. These findings strongly suggest the barrier protective effect of FTY720 on HMVEC monolayers in hyperglycemic medium via S1P signaling, further implying the possibility of FTY720 as a therapeutic agent of diabetic vascular disorder.

Edaravone mimics sphingosine-1-phosphate-induced endothelial barrier enhancement in human microvascular endothelial cells

2007 ◽  
Vol 293 (5) ◽  
pp. C1523-C1531 ◽  
Author(s):  
Kazuyoshi Omori ◽  
Yasushi Shikata ◽  
Kei Sarai ◽  
Naomi Watanabe ◽  
Jun Wada ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Protective Effect ◽  
Focal Adhesion ◽  
Small Gtpase ◽  
Endothelial Barrier ◽  
Microvascular Endothelial Cells ◽  
Cell Periphery ◽  
Acute Cerebral Ischemia ◽  
Sphingosine 1 Phosphate ◽  
Microvascular Endothelial

Edaravone is a potent scavenger of hydroxyl radicals and is quite successful in patients with acute cerebral ischemia, and several organ-protective effects have been reported. Treatment of human microvascular endothelial cells with edaravone (1.5 μM) resulted in the enhancement of transmonolayer electrical resistance coincident with cortical actin enhancement and redistribution of focal adhesion proteins and adherens junction proteins to the cell periphery. Edaravone also induced small GTPase Rac activation and focal adhesion kinase (FAK; Tyr576) phosphorylation associated with sphingosine-1-phosphate receptor type 1 (S1P1) transactivation. S1P1 protein depletion by the short interfering RNA technique completely abolished edaravone-induced FAK (Tyr576) phosphorylation and Rac activation. This is the first report of edaravone-induced endothelial barrier enhancement coincident with focal adhesion remodeling and cytoskeletal rearrangement associated with Rac activation via S1P1 transactivation. Considering the well-established endothelial barrier-protective effect of S1P, endothelial barrier enhancement as a consequence of S1P1 transactivation may at least partly be the potent mechanisms for the organ-protective effect of edaravone and is suggestive of edaravone as a therapeutic agent against systemic vascular barrier disorder.

scholarly journals Bionic Silk Fibroin Film Promotes Tenogenic Differentiation of Tendon Stem/Progenitor Cells by Activating Focal Adhesion Kinase

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Kang Lu ◽  
Xiaodie Chen ◽  
Hong Tang ◽  
Mei Zhou ◽  
Gang He ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Progenitor Cells ◽  
Silk Fibroin ◽  
Focal Adhesion ◽  
Cell Morphology ◽  
Biological Activities ◽  
Biological Effects ◽  
Tendon Injuries ◽  
Regeneration Ability ◽  
Gene And Protein Expression

Background. Tendon injuries are common musculoskeletal disorders in clinic. Due to the limited regeneration ability of tendons, tissue engineering technology is often used as an effective approach to treat tendon injuries. Silk fibroin (SF) films have excellent biological activities and physical properties, which is suitable for tendon regeneration. The present study is aimed at preparing a SF film with a bionic microstructure and investigating its biological effects. Methods. A SF film with a smooth surface or bionic microstructure was prepared. After seeding tendon stem/progenitor cells (TSPCs) on the surface, the cell morphology, the expression level of tenogenic genes and proteins, and the focal adhesion kinase (FAK) activation were measured to evaluate the biological effect of SF films. Results. The TSPCs on SF films with a bionic microstructure exhibited a slender cell morphology, promoted the expression of tenogenic genes and proteins, such as SCX, TNC, TNMD, and COLIA1, and activated FAK. FAK inhibitors blocked the enhanced expression of tenogenic genes and proteins. Conclusion. SF films with a bionic microstructure may serve as a scaffold, provide biophysical cues to alter the cellular adherence arrangement and cell morphology, and enhance the tenogenic gene and protein expression in TSPCs. FAK activation plays a key role during this biological response process.

Src and focal adhesion kinase mediate mechanical strain-induced proliferation and ERK1/2 phosphorylation in human H441 pulmonary epithelial cells

2007 ◽  
Vol 292 (5) ◽  
pp. C1701-C1713 ◽  
Author(s):  
Lakshmi S. Chaturvedi ◽  
Harold M. Marsh ◽  
Marc D. Basson
Keyword(s):  
Epithelial Cells ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Mechanical Strain ◽  
A549 Cells ◽  
Cyclic Strain ◽  
Short Interfering Rna ◽  
Erk Activation ◽  
Pulmonary Epithelial Cells ◽  
Interfering Rna

Pulmonary epithelial cells are exposed to repetitive deformation during physiological breathing and mechanical ventilation. Such deformation may influence pulmonary growth, development, and barotrauma. Although deformation stimulates proliferation and activates extracellular signal-regulated kinases (ERK1/2) in human pulmonary epithelial H441 cells, the upstream mechanosensors that induce ERK activation are poorly understood. We investigated whether c-Src or focal adhesion kinase (FAK) mediates cyclic mechanical strain-induced ERK1/2 activation and proliferation in human pulmonary epithelial (NCI-H441) cells. The H441 and A549 cells were grown on collagen I-precoated membranes and were subjected to an average 10% cyclic mechanical strain at 20 cycles/min. Cyclic strain activated Src within 2 min by increasing phosphorylation at Tyr418, followed by rapid phosphorylation of FAK at Tyr397 and Tyr576 and ERK1/2 at Thr202/Tyr204 ( n = 5, P < 0.05). Twenty-four (A549 cells) and 24–72 h (H441 cells) of cyclic mechanical strain increased cell numbers compared with static culture. Twenty-four hours of cyclic strain also increased H441 FAK, Src, and ERK phosphorylation without affecting total FAK, Src, or ERK protein. The mitogenic effect was blocked by Src (10 μmol/l PP2 or short interfering RNA targeted to Src) or MEK (50 μmol/l PD-98059) inhibition. PP2 also blocked strain-induced phosphorylation of FAK-Tyr576 and ERK-Thr202/Tyr204 but not FAK-Tyr397. Reducing FAK by FAK-targeted short interfering RNA blocked mechanical strain-induced mitogenicity and significantly attenuated strain-induced ERK activation but not strain-induced Src phosphorylation. Together, these results suggest that repetitive mechanical deformation induced by ventilation supports pulmonary epithelial proliferation by a pathway involving Src, FAK, and then ERK signaling.

scholarly journals Human ether-a-go-go-related Gene 1 Channels Are Physically Linked to β1 Integrins and Modulate Adhesion-dependent Signaling

2005 ◽  
Vol 16 (6) ◽  
pp. 2972-2983 ◽  
Author(s):  
Alessia Cherubini ◽  
Giovanna Hofmann ◽  
Serena Pillozzi ◽  
Leonardo Guasti ◽  
Olivia Crociani ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Surface Membrane ◽  
Neuroblastoma Cells ◽  
Small Gtpase ◽  
Related Gene ◽  
Signaling Complex ◽  
Current Activation ◽  
Membrane Compartments ◽  
Herg Channels

Adhesive receptors of the integrin family are primarily involved in cell–extracellular matrix adhesion. Additionally, integrins trigger multiple signaling pathways that are involved in cell migration, proliferation, survival, and differentiation. We previously demonstrated that the activation of integrins containing the β1 subunit leads to a selective increase in potassium currents carried by the human ether-a-go-go-related gene (hERG) channels in neuroblastoma and leukemia cells; this current activation modulates adhesion-dependent differentiation in these cells. We hypothesized that the cross-talk between integrins and hERG channels could be traced back to the assembly of a macromolecular signaling complex comprising the two proteins. We tested this hypothesis in both SH-SY5Y neuroblastoma cells and in human embryonic kidney 293 cells stably transfected with hERG1 and, therefore, expressing only the full-length hERG1 protein on the plasma membrane. The β1 integrin and hERG1 coprecipitate in these cells and colocalize in both intracellular and surface membrane compartments. The two proteins also coprecipitate with caveolin-1, suggesting the localization of the complex in lipid rafts/caveolae. hERG1-transfected cells undergo an activation of hERG currents after β1 integrin-mediated adhesion to fibronectin; concomitant with this activation, the focal adhesion kinase associates with the hERG1 protein and becomes tyrosine phosphorylated. Using hERG1-specific inhibitors, we show that the tyrosine phosphorylation of focal adhesion kinase is strictly dependent on hERG channel activity. Similarly, the activity of the small GTPase Rac1 turned out to be dependent on hERG currents. On the whole, these data indicate that the hERG1 protein associates with β1 integrins and modulates adhesion receptor signaling.

scholarly journals Focal Adhesion Kinase Targeting Using In vivo Short Interfering RNA Delivery in Neutral Liposomes for Ovarian Carcinoma Therapy

2006 ◽  
Vol 12 (16) ◽  
pp. 4916-4924 ◽  
Author(s):  
Jyotsnabaran Halder ◽  
Aparna A. Kamat ◽  
Charles N. Landen ◽  
Liz Y. Han ◽  
Susan K. Lutgendorf ◽  
...  
Keyword(s):  
Ovarian Carcinoma ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Short Interfering Rna ◽  
Interfering Rna

scholarly journals Sphingosine 1-phosphate stimulates Rho-mediated tyrosine phosphorylation of focal adhesion kinase and paxillin in Swiss 3T3 fibroblasts

1997 ◽  
Vol 324 (2) ◽  
pp. 481-488 ◽  
Author(s):  
Fang WANG ◽  
Catherine D. NOBES ◽  
Alan HALL ◽  
Sarah SPIEGEL
Keyword(s):  
Focal Adhesion Kinase ◽  
Tyrosine Phosphorylation ◽  
Focal Adhesion ◽  
Stress Fibre ◽  
3T3 Fibroblasts ◽  
Fibre Formation ◽  
Sphingosine 1 Phosphate ◽  
Swiss 3T3 Fibroblasts ◽  
Stress Fibre Formation ◽  
Stimulation Of

Sphingosine 1-phosphate (SPP), a sphingolipid second messenger implicated in the mitogenic action of platelet-derived growth factor [Olivera, A. and Spiegel, S. (1993) Nature (London) 365, 557–560], induced rapid reorganization of the actin cytoskeleton resulting in stress-fibre formation. SPP also induced transient tyrosine phosphorylation of focal adhesion kinase (p125FAK), a cytosolic tyrosine kinase that localizes in focal adhesions, and of the cytoskeleton-associated protein paxillin. Exoenzyme C3 transferase, which ADP-ribosylates Rho (a Ras-related small GTP binding protein) on asparagine-41 and renders it biologically inactive, inhibited both stress-fibre formation and protein tyrosine phosphorylation induced by SPP. Thus Rho may be an upstream regulator of both stress-fibre formation and tyrosine phosphorylation of p125FAK and paxillin. Pretreatment with PMA, an activator of protein kinase C (PKC), inhibited the stimulation of stress-fibre formation induced by 1-oleoyl-lysophosphatidic acid (LPA) but not that by SPP. Similarly, PMA also decreased LPA-induced tyrosine phosphorylation of p125FAK and paxillin without abrogating the response to SPP. Thus PKC is involved in LPA- but not SPP-dependent signalling. The polyanionic drug suramin, a broad-specificity inhibitor of ligand–receptor interactions, did not inhibit either the mitogenic effect of SPP or its stimulation of tyrosine phosphorylation of p125FAK. However, suramin markedly inhibited these responses induced by LPA. These results suggest that in contrast with LPA, SPP may be acting intracellularly in Swiss 3T3 fibroblasts to stimulate tyrosine phosphorylation of p125FAK and paxillin and cell growth.

scholarly journals Endothelial Barrier Strengthening by Activation of Focal Adhesion Kinase

2003 ◽  
Vol 278 (15) ◽  
pp. 13342-13349 ◽  
Author(s):  
Sadiqa K. Quadri ◽  
Mrinal Bhattacharjee ◽  
Kaushik Parthasarathi ◽  
Tatsuo Tanita ◽  
Jahar Bhattacharya
Keyword(s):  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Endothelial Barrier
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