P2Y2 receptor modulates shear stress-induced cell alignment and actin stress fibers in human umbilical vein endothelial cells
(2017) In Cellular and Molecular Life Sciences 74(4). p.731-746- Abstract
Endothelial cells release ATP in response to fluid shear stress, which activates purinergic (P2) receptor-mediated signaling molecules including endothelial nitric oxide (eNOS), a regulator of vascular tone. While P2 receptor-mediated signaling in the vasculature is well studied, the role of P2Y2 receptors in shear stress-associated endothelial cell alignment, cytoskeletal alterations, and wound repair remains ill defined. To address these aspects, human umbilical vein endothelial cell (HUVEC) monolayers were cultured on gelatin-coated dishes and subjected to a shear stress of 1 Pa. HUVECs exposed to either P2Y2 receptor antagonists or siRNA showed impaired fluid shear stress-induced cell alignment, and actin... (More)
Endothelial cells release ATP in response to fluid shear stress, which activates purinergic (P2) receptor-mediated signaling molecules including endothelial nitric oxide (eNOS), a regulator of vascular tone. While P2 receptor-mediated signaling in the vasculature is well studied, the role of P2Y2 receptors in shear stress-associated endothelial cell alignment, cytoskeletal alterations, and wound repair remains ill defined. To address these aspects, human umbilical vein endothelial cell (HUVEC) monolayers were cultured on gelatin-coated dishes and subjected to a shear stress of 1 Pa. HUVECs exposed to either P2Y2 receptor antagonists or siRNA showed impaired fluid shear stress-induced cell alignment, and actin stress fiber formation as early as 6 h. Similarly, when compared to cells expressing the P2Y2 Arg-Gly-Asp (RGD) wild-type receptors, HUVECs transiently expressing the P2Y2 Arg-Gly-Glu (RGE) mutant receptors showed reduced cell alignment and actin stress fiber formation in response to shear stress as well as to P2Y2 receptor agonists in static cultures. Additionally, we observed reduced shear stress-induced phosphorylation of focal adhesion kinase (Y397), and cofilin-1 (S3) with receptor knockdown as well as in cells expressing the P2Y2 RGE mutant receptors. Consistent with the role of P2Y2 receptors in vasodilation, receptor knockdown and overexpression of P2Y2 RGE mutant receptors reduced shear stress-induced phosphorylation of AKT (S473), and eNOS (S1177). Furthermore, in a scratched wound assay, shear stress-induced cell migration was reduced by both pharmacological inhibition and receptor knockdown. Together, our results suggest a novel role for P2Y2 receptor in shear stress-induced cytoskeletal alterations in HUVECs.
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- author
- Sathanoori, Ramasri LU ; Bryl-Gorecka, Paulina LU ; Müller, Christa E. ; Erb, Laurie ; Weisman, Gary A. ; Olde, Björn LU and Erlinge, David LU
- organization
- publishing date
- 2017
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- FAK, Integrins, P2Y receptors, Shear stress, Wound repair
- in
- Cellular and Molecular Life Sciences
- volume
- 74
- issue
- 4
- pages
- 731 - 746
- publisher
- Birkhäuser
- external identifiers
-
- pmid:27652381
- wos:000393694900011
- scopus:84988644296
- ISSN
- 1420-682X
- DOI
- 10.1007/s00018-016-2365-0
- language
- English
- LU publication?
- yes
- id
- feebd9c2-419e-45bc-bc33-32e551288cac
- date added to LUP
- 2016-11-02 14:46:55
- date last changed
- 2024-07-26 21:10:21
@article{feebd9c2-419e-45bc-bc33-32e551288cac, abstract = {{<p>Endothelial cells release ATP in response to fluid shear stress, which activates purinergic (P2) receptor-mediated signaling molecules including endothelial nitric oxide (eNOS), a regulator of vascular tone. While P2 receptor-mediated signaling in the vasculature is well studied, the role of P2Y<sub>2</sub> receptors in shear stress-associated endothelial cell alignment, cytoskeletal alterations, and wound repair remains ill defined. To address these aspects, human umbilical vein endothelial cell (HUVEC) monolayers were cultured on gelatin-coated dishes and subjected to a shear stress of 1 Pa. HUVECs exposed to either P2Y<sub>2</sub> receptor antagonists or siRNA showed impaired fluid shear stress-induced cell alignment, and actin stress fiber formation as early as 6 h. Similarly, when compared to cells expressing the P2Y<sub>2</sub> Arg-Gly-Asp (RGD) wild-type receptors, HUVECs transiently expressing the P2Y<sub>2</sub> Arg-Gly-Glu (RGE) mutant receptors showed reduced cell alignment and actin stress fiber formation in response to shear stress as well as to P2Y<sub>2</sub> receptor agonists in static cultures. Additionally, we observed reduced shear stress-induced phosphorylation of focal adhesion kinase (Y397), and cofilin-1 (S3) with receptor knockdown as well as in cells expressing the P2Y<sub>2</sub> RGE mutant receptors. Consistent with the role of P2Y<sub>2</sub> receptors in vasodilation, receptor knockdown and overexpression of P2Y<sub>2</sub> RGE mutant receptors reduced shear stress-induced phosphorylation of AKT (S473), and eNOS (S1177). Furthermore, in a scratched wound assay, shear stress-induced cell migration was reduced by both pharmacological inhibition and receptor knockdown. Together, our results suggest a novel role for P2Y<sub>2</sub> receptor in shear stress-induced cytoskeletal alterations in HUVECs.</p>}}, author = {{Sathanoori, Ramasri and Bryl-Gorecka, Paulina and Müller, Christa E. and Erb, Laurie and Weisman, Gary A. and Olde, Björn and Erlinge, David}}, issn = {{1420-682X}}, keywords = {{FAK; Integrins; P2Y receptors; Shear stress; Wound repair}}, language = {{eng}}, number = {{4}}, pages = {{731--746}}, publisher = {{Birkhäuser}}, series = {{Cellular and Molecular Life Sciences}}, title = {{P2Y<sub>2</sub> receptor modulates shear stress-induced cell alignment and actin stress fibers in human umbilical vein endothelial cells}}, url = {{http://dx.doi.org/10.1007/s00018-016-2365-0}}, doi = {{10.1007/s00018-016-2365-0}}, volume = {{74}}, year = {{2017}}, }