Serine/threonine phosphatases regulate platelet 伪_IIb尾₃ integrin receptor outside-in signaling mechanisms and clot retraction

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• 作者：Antonio Moscard贸 ; Mar铆a Teresa Santos ; Ana Latorre ; Isabel Madrid ; Juana Vall茅s
• 关键词：PSTPs ; protein serine/threonine phosphatases ; OA ; okadaic acid ; FAK ; Focal adhesion kinase
• 刊名：Life Sciences
• 出版年：6 November, 2013
• 年：2013
• 卷：93
• 期：20
• 页码：707-713
• 全文大小：705 K

文摘

Aims

We studied the role of serine/threonine phosphatases (PSTPs) on 伪_IIb尾₃ signaling and the potential selectivity of the level of PSTP inhibition with okadaic acid (OA) on 伪_IIb尾₃ signaling for regulation of platelet aggregation and clot retraction.

Main methods

We used washed platelets from normal donors and OA as inhibitor of PSTPs. Clot retraction was induced by 1 U/mL of thrombin. Reorganized cytoskeleton was isolated from Triton X-100 lysed platelets. The presence of proteins incorporated to the cytoskeleton was assayed by immunoblotting with specific antibodies.

Key findings

We found that both 100 and 500 nM OA blocked platelet mediated clot retraction. In contrast, only 500 nM OA inhibited thrombin-induced inside-out 伪_IIb尾₃ activation, platelet aggregation, and cytoskeletal reorganization. Among markers of 伪_IIb尾₃ outside-in signaling, 500 nM OA inhibited the incorporation to the cytoskeleton of syk, src, and FAK (Focal Adhesion Kinase) tyrosine kinases and the incorporation and phosphorylation at Tyr⁷⁵⁹ of the 尾₃ chain of 伪_IIb尾₃, while 100 nM OA only inhibited the FAK translocation and its tyrosine phosphorylation.

Significance

The level of inhibition of PSTPs by low or high OA concentration (33% and 73% inhibition, respectively) in intact whole cells differentially regulates platelet aggregation and integrin signaling, but have a common effect in blocking clot retraction. The latter may be associated with the presence of phosphorylated FAK in the cytoskeleton. This study reveals a novel target for anti-platelet treatment to block clot retraction without affecting the platelet hemostatic function by a partial inhibition of PSTPs.