Abstract
Platelet aggregation at sites of vascular injury is not only essential for hemostasis, but may also cause acute ischemic disease states such as myocardial infarction or stroke. The hemi-immunoreceptor tyrosine-based activation motif-containing C-type lectinlike receptor 2 (CLEC-2) mediates powerful platelet activation through a Src- and spleen tyrosine kinase (Syk)-dependent tyrosine phosphorylation cascade. Thereby, CLEC-2 not only contributes to thrombus formation and stabilization but also plays a central role in blood-lymphatic vessel development, tumor metastasis, and prevention of inflammatory bleeding, making it a potential pharmacologic target to modulate these processes. We have previously shown that injection of the anti-CLEC-2 antibody, INU1, results in virtually complete immunodepletion of platelet CLEC-2 in mice, which is, however, preceded by a severe transient thrombocytopenia thereby limiting its potential therapeutic use. The mechanisms underlying this targeted CLEC-2 downregulation have remained elusive. Here, we show that INU1-induced CLEC-2 immunodepletion occurs through Src-family kinase-dependent receptor internalization in vitro and in vivo, presumably followed by intracellular degradation. In mice with platelet-specific Syk deficiency, INU1-induced CLEC-2 internalization/degradation was fully preserved whereas the associated thrombocytopenia was largely prevented. These results show for the first time that CLEC-2 can be downregulated from the platelet surface through internalization in vitro and in vivo and that this can be mechanistically uncoupled from the associated antibody-induced thrombocytopenia.
Original language | English |
---|---|
Pages (from-to) | 4069-4077 |
Number of pages | 9 |
Journal | Blood |
Volume | 125 |
Issue number | 26 |
Early online date | 20 Mar 2015 |
DOIs | |
Publication status | Published - 25 Jun 2015 |
Keywords
- Animals
- Antibodies, Monoclonal
- Blood Platelets
- Blotting, Western
- Down-Regulation
- Flow Cytometry
- Fluorescent Antibody Technique
- Intracellular Signaling Peptides and Proteins
- Lectins, C-Type
- Male
- Mice
- Mice, Inbred C57BL
- Mice, Knockout
- Platelet Activation
- Protein-Tyrosine Kinases
- Thrombocytopenia