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Methylphenidate-triggered ROS generation promotes caveolae-mediated transcytosis via Rac1 signaling and c-Src-dependent caveolin-1 phosphorylation in human brain endothelial cells
- Source :
- Cellular and Molecular Life Sciences. 73:4701-4716
- Publication Year :
- 2016
- Publisher :
- Springer Science and Business Media LLC, 2016.
-
Abstract
- Methylphenidate (MPH) is an amphetamine-like stimulant commonly prescribed for attention deficit hyperactivity disorder. Despite its widespread use, the cellular/molecular effects of MPH remain elusive. Here, we report a novel direct role of MPH on the regulation of macromolecular flux through human brain endothelial cells (ECs). MPH significantly increased caveolae-mediated transcytosis of horseradish peroxidase through ECs without affecting paracellular permeability. Using FRET-based live cell imaging, together with pharmacological inhibitors and lentiviral-mediated shRNA knockdown, we demonstrate that MPH promoted ROS generation via activation of Rac1-dependent NADPH oxidase (NOX) and c-Src activation at the plasma membrane. c-Src in turn was shown to mediate the phosphorylation of caveolin-1 (Cav1) on Tyr14 leading to enhanced caveolae formation and transendothelial transport. Accordingly, the inhibition of Cav1 phosphorylation by overexpression of a phosphodefective Cav1Y14F mutant or knocking down Cav1 expression abrogated MPH-induced transcytosis. In addition, both vitamin C and inhibition of NOX blocked MPH-triggered vesicular transport. This study, therefore, identifies Rac1/NOX/c-Src-dependent signaling in MPH-induced increase in transendothelial permeability of brain endothelial cell monolayers via caveolae-mediated transcytosis.
- Subjects :
- rac1 GTP-Binding Protein
0301 basic medicine
Caveolin 1
Biology
Caveolae
Models, Biological
CSK Tyrosine-Protein Kinase
Capillary Permeability
03 medical and health sciences
Cellular and Molecular Neuroscience
0302 clinical medicine
mental disorders
Humans
Phosphorylation
Transport Vesicles
cdc42 GTP-Binding Protein
Molecular Biology
Horseradish Peroxidase
Pharmacology
NADPH oxidase
Brain
Endothelial Cells
NADPH Oxidases
Biological Transport
Cell Biology
Oxidants
Cell biology
Endothelial stem cell
Vesicular transport protein
src-Family Kinases
030104 developmental biology
Biochemistry
Transcytosis
Paracellular transport
Methylphenidate
biology.protein
Molecular Medicine
Reactive Oxygen Species
rhoA GTP-Binding Protein
human activities
030217 neurology & neurosurgery
Signal Transduction
Subjects
Details
- ISSN :
- 14209071 and 1420682X
- Volume :
- 73
- Database :
- OpenAIRE
- Journal :
- Cellular and Molecular Life Sciences
- Accession number :
- edsair.doi.dedup.....ff929c69424ec10b5bc77a098a247285