Methylphenidate-triggered ROS generation promotes caveolae-mediated transcytosis via Rac1 signaling and c-Src-dependent caveolin-1 phosphorylation in human brain endothelial cells

被引：30

作者：

Coelho-Santos, Vanessa ^{[1
,2
]}

Socodato, Renato ^{[3
,4
]}

Portugal, Camila ^{[3
,4
]}

Leitao, Ricardo A. ^{[1
,2
]}

Rito, Manuel ^{[5
]}

Barbosa, Marcos ^{[5
,6
]}

Couraud, Pierre-Olivier ^{[7
]}

Romero, Ignacio A. ^{[8
]}

Weksler, Babette ^{[9
]}

Minshall, Richard D. ^{[10
,11
]}

Fontes-Ribeiro, Carlos ^{[1
,2
]}

Summavielle, Teresa ^{[3
,4
]}

Relvas, Joao B. ^{[3
,4
]}

Silva, Ana P. ^{[1
,2
]}

机构：

[1] Univ Coimbra, Inst Pharmacol & Expt Therapeut, Fac Med, Coimbra, Portugal

[2] Univ Coimbra, CNC IBILI, Coimbra, Portugal

[3] Univ Porto, Inst Invest & Inovacao Saude I3S, Oporto, Portugal

[4] Univ Porto, IBMC, Oporto, Portugal

[5] CHUC, Neurosurg Serv, Coimbra, Portugal

[6] Univ Coimbra, Fac Med, Coimbra, Portugal

[7] Univ Paris 05, Sorbonne Paris Cite, Inst Cochin, INSERM U1016,CNRS UMR8104, Paris, France

[8] Open Univ, Dept Life Sci, Fac Sci, Milton Keynes, Bucks, England

[9] Cornell Univ, Dept Med, Weill Med Coll, New York, NY 10021 USA

[10] Univ Illinois, Dept Anesthesiol, Chicago, IL USA

[11] Univ Illinois, Dept Pharmacol, Chicago, IL USA

来源：

CELLULAR AND MOLECULAR LIFE SCIENCES | 2016年 / 73卷 / 24期

关键词：

Psychostimulants; Attention deficit hyperactivity disorder; Blood-brain barrier; NADPH oxidase; Vesicular transport; Vitamin C; TRANSCELLULAR TRANSPORT; BARRIER FUNCTION; TYROSINE KINASE; TIGHT JUNCTIONS; IN-VITRO; BLOOD; RHO; ACTIVATION; EXPRESSION; PATHOPHYSIOLOGY;

D O I：

10.1007/s00018-016-2301-3

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

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 Tyr(14) leading to enhanced caveolae formation and transendothelial transport. Accordingly, the inhibition of Cav1 phosphorylation by overexpression of a phosphodefective Cav1(Y14F) 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.

引用

页码：4701 / 4716

页数：16

共 56 条

[1] Dose-related immunohistochemical and ultrastructural changes after oral methylphenidate administration in cerebrum and cerebellum of the rat [J].