Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation
Nitric oxide (NO) is a known anti-adhesive molecule that prevents platelet aggregation and leukocyte adhesion to endothelial cells (ECs). The mechanism has been attributed to its role in the regulation of adhesion molecules on leukocytes and the adhesive properties of platelets. Our previous study c...
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Frontiers Media S.A.
2018-01-01
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| Series: | Frontiers in Physiology |
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| Online Access: | http://journal.frontiersin.org/article/10.3389/fphys.2017.01124/full |
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| author | Feng Gao Brandon P. Lucke-Wold Brandon P. Lucke-Wold Xiang Li Aric F. Logsdon Aric F. Logsdon Li-Chong Xu Sulei Xu Kyle B. LaPenna Huaqi Wang Huaqi Wang M. A. Hassan Talukder Christopher A. Siedlecki Jason D. Huber Charles L. Rosen Pingnian He |
| author_facet | Feng Gao Brandon P. Lucke-Wold Brandon P. Lucke-Wold Xiang Li Aric F. Logsdon Aric F. Logsdon Li-Chong Xu Sulei Xu Kyle B. LaPenna Huaqi Wang Huaqi Wang M. A. Hassan Talukder Christopher A. Siedlecki Jason D. Huber Charles L. Rosen Pingnian He |
| author_sort | Feng Gao |
| collection | DOAJ |
| description | Nitric oxide (NO) is a known anti-adhesive molecule that prevents platelet aggregation and leukocyte adhesion to endothelial cells (ECs). The mechanism has been attributed to its role in the regulation of adhesion molecules on leukocytes and the adhesive properties of platelets. Our previous study conducted in rat venules found that reduction of EC basal NO synthesis caused EC ICAM-1-mediated firm adhesion of leukocytes within 10–30 min. This quick response occurred in the absence of alterations of adhesion molecules on leukocytes and also opposes the classical pattern of ICAM-1-mediated leukocyte adhesion that requires protein synthesis and occurs hours after stimulation. The objective of this study is to investigate the underlying mechanisms of reduced basal NO-induced EC-mediated rapid leukocyte adhesion observed in intact microvessels. The relative levels of ICAM-1 at different cell regions and their activation status were determined with cellular fractionation and western blot using cultured human umbilical vein ECs. ICAM-1 adhesiveness was determined by immunoprecipitation in non-denatured proteins to assess the changes in ICAM-1 binding to its inhibitory antibody, mAb1A29, and antibody against total ICAM-1 with and without NO reduction. The adhesion strength of EC ICAM-1 was assessed by atomic force microscopy (AFM) on live cells. Results showed that reduction of EC basal NO caused by the application of caveolin-1 scaffolding domain (AP-CAV) or NOS inhibitor, L-NMMA, for 30 min significantly increased phosphorylated ICAM-1 and its binding to mAb1A29 in the absence of altered ICAM-1 expression and its distribution at subcellular regions. The Src inhibitor, PP1, inhibited NO reduction-induced increases in ICAM-1 phosphorylation and adhesive binding. AFM detected significant increases in the binding force between AP-CAV-treated ECs and mAb1A29-coated probes. These results demonstrated that reduced EC basal NO lead to a rapid increase in ICAM-1 adhesive binding via Src-mediated phosphorylation without de novo protein synthesis and translocation. This study suggests that a NO-dependent conformational change of constitutive EC membrane ICAM-1 might be the mechanism of rapid ICAM-1 dependent leukocyte adhesion observed in vivo. This new mechanistic insight provides a better understanding of EC/leukocyte interaction-mediated vascular inflammation under many disease conditions that encounter reduced basal NO in the circulation system. |
| first_indexed | 2024-12-16T10:08:21Z |
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| last_indexed | 2024-12-16T10:08:21Z |
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| spelling | doaj.art-d35b1ac8ed2c45688e4023d390b951ca2022-12-21T22:35:37ZengFrontiers Media S.A.Frontiers in Physiology1664-042X2018-01-01810.3389/fphys.2017.01124316300Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated PhosphorylationFeng Gao0Brandon P. Lucke-Wold1Brandon P. Lucke-Wold2Xiang Li3Aric F. Logsdon4Aric F. Logsdon5Li-Chong Xu6Sulei Xu7Kyle B. LaPenna8Huaqi Wang9Huaqi Wang10M. A. Hassan Talukder11Christopher A. Siedlecki12Jason D. Huber13Charles L. Rosen14Pingnian He15Department of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Neurosurgery, West Virginia University School of Medicine, Morgantown, WV, United StatesThe Center for Neuroscience, West Virginia University School of Medicine, Morgantown, WV, United StatesDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesThe Center for Neuroscience, West Virginia University School of Medicine, Morgantown, WV, United StatesDepartment of Basic Pharmaceutical Sciences, West Virginia University School of Pharmacy, Morgantown, WV, United StatesDepartment of Surgery, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesRespiratory Department, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, ChinaDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Surgery, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesDepartment of Basic Pharmaceutical Sciences, West Virginia University School of Pharmacy, Morgantown, WV, United StatesDepartment of Neurosurgery, West Virginia University School of Medicine, Morgantown, WV, United StatesDepartment of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey, PA, United StatesNitric oxide (NO) is a known anti-adhesive molecule that prevents platelet aggregation and leukocyte adhesion to endothelial cells (ECs). The mechanism has been attributed to its role in the regulation of adhesion molecules on leukocytes and the adhesive properties of platelets. Our previous study conducted in rat venules found that reduction of EC basal NO synthesis caused EC ICAM-1-mediated firm adhesion of leukocytes within 10–30 min. This quick response occurred in the absence of alterations of adhesion molecules on leukocytes and also opposes the classical pattern of ICAM-1-mediated leukocyte adhesion that requires protein synthesis and occurs hours after stimulation. The objective of this study is to investigate the underlying mechanisms of reduced basal NO-induced EC-mediated rapid leukocyte adhesion observed in intact microvessels. The relative levels of ICAM-1 at different cell regions and their activation status were determined with cellular fractionation and western blot using cultured human umbilical vein ECs. ICAM-1 adhesiveness was determined by immunoprecipitation in non-denatured proteins to assess the changes in ICAM-1 binding to its inhibitory antibody, mAb1A29, and antibody against total ICAM-1 with and without NO reduction. The adhesion strength of EC ICAM-1 was assessed by atomic force microscopy (AFM) on live cells. Results showed that reduction of EC basal NO caused by the application of caveolin-1 scaffolding domain (AP-CAV) or NOS inhibitor, L-NMMA, for 30 min significantly increased phosphorylated ICAM-1 and its binding to mAb1A29 in the absence of altered ICAM-1 expression and its distribution at subcellular regions. The Src inhibitor, PP1, inhibited NO reduction-induced increases in ICAM-1 phosphorylation and adhesive binding. AFM detected significant increases in the binding force between AP-CAV-treated ECs and mAb1A29-coated probes. These results demonstrated that reduced EC basal NO lead to a rapid increase in ICAM-1 adhesive binding via Src-mediated phosphorylation without de novo protein synthesis and translocation. This study suggests that a NO-dependent conformational change of constitutive EC membrane ICAM-1 might be the mechanism of rapid ICAM-1 dependent leukocyte adhesion observed in vivo. This new mechanistic insight provides a better understanding of EC/leukocyte interaction-mediated vascular inflammation under many disease conditions that encounter reduced basal NO in the circulation system.http://journal.frontiersin.org/article/10.3389/fphys.2017.01124/fullconstitutive ICAM-1ICAM-1-mediated leukocyte adhesionbasal nitric oxideconformational change of ICAM-1atomic force microscopy |
| spellingShingle | Feng Gao Brandon P. Lucke-Wold Brandon P. Lucke-Wold Xiang Li Aric F. Logsdon Aric F. Logsdon Li-Chong Xu Sulei Xu Kyle B. LaPenna Huaqi Wang Huaqi Wang M. A. Hassan Talukder Christopher A. Siedlecki Jason D. Huber Charles L. Rosen Pingnian He Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation Frontiers in Physiology constitutive ICAM-1 ICAM-1-mediated leukocyte adhesion basal nitric oxide conformational change of ICAM-1 atomic force microscopy |
| title | Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation |
| title_full | Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation |
| title_fullStr | Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation |
| title_full_unstemmed | Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation |
| title_short | Reduction of Endothelial Nitric Oxide Increases the Adhesiveness of Constitutive Endothelial Membrane ICAM-1 through Src-Mediated Phosphorylation |
| title_sort | reduction of endothelial nitric oxide increases the adhesiveness of constitutive endothelial membrane icam 1 through src mediated phosphorylation |
| topic | constitutive ICAM-1 ICAM-1-mediated leukocyte adhesion basal nitric oxide conformational change of ICAM-1 atomic force microscopy |
| url | http://journal.frontiersin.org/article/10.3389/fphys.2017.01124/full |
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