Platelet-derived miR-223 promotes a phenotypic switch in arterial injury repair
Zhi Zeng, … , John Hwa, Wai Ho Tang
Zhi Zeng, … , John Hwa, Wai Ho Tang
Published March 1, 2019; First published January 15, 2019
Citation Information: J Clin Invest. 2019;129(3):1372-1386. https://doi.org/10.1172/JCI124508.
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Research Article Vascular biology

Platelet-derived miR-223 promotes a phenotypic switch in arterial injury repair

Abstract

Upon arterial injury, endothelial denudation leads to platelet activation and delivery of multiple agents (e.g., TXA2, PDGF), promoting VSMC dedifferentiation and proliferation (intimal hyperplasia) during injury repair. The process of resolution of vessel injury repair, and prevention of excessive repair (switching VSMCs back to a differentiated quiescent state), is poorly understood. We now report that internalization of APs by VSMCs promotes resolution of arterial injury by switching on VSMC quiescence. Ex vivo and in vivo studies using lineage tracing reporter mice (PF4-cre × mT/mG) demonstrated uptake of GFP-labeled platelets (mG) by mTomato red–labeled VSMCs (mT) upon arterial wire injury. Genome-wide miRNA sequencing of VSMCs cocultured with APs identified significant increases in platelet-derived miR-223. miR-223 appears to directly target PDGFRβ (in VSMCs), reversing the injury-induced dedifferentiation. Upon arterial injury, platelet miR-223–KO mice exhibited increased intimal hyperplasia, whereas miR-223 mimics reduced intimal hyperplasia. Diabetic mice with reduced expression of miR-223 exhibited enhanced VSMC dedifferentiation and proliferation and increased intimal hyperplasia. Our results suggest that horizontal transfer of platelet-derived miRNAs into VSMCs provides a novel mechanism for regulating VSMC phenotypic switching. Platelets thus play a dual role in vascular injury repair, initiating an immediate repair process and, concurrently, a delayed process to prevent excessive repair.

Authors

Zhi Zeng, Luoxing Xia, Xuejiao Fan, Allison C. Ostriker, Timur Yarovinsky, Meiling Su, Yuan Zhang, Xiangwen Peng, Yi Xie, Lei Pi, Xiaoqiong Gu, Sookja Kim Chung, Kathleen A. Martin, Renjing Liu, John Hwa, Wai Ho Tang

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Figure 1

Platelets were internalized by VSMCs and induced VSMC differentiation.

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Platelets were internalized by VSMCs and induced VSMC differentiation. (...
(A) Expression of markers for differentiation (ACTA2, CNN1, TAGLN) and dedifferentiation (KLF4, KLF5, OPN) in control VSMCs (Ctrl) and in VSMCs after coculture with RPs or APs was determined by Western blot analysis (n = 4). Cell proliferation was assessed by CCK8 assays (n = 7) (B) and BrdU incorporation assays (n = 4) (C). Data are presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001 vs. Ctrl; #P < 0.05, ##P < 0.01, ###P < 0.001 vs. RPs. (D) Representative images of VSMCs cocultured with CMFDA-labeled platelets (green) for 1, 2, 4, or 24 hours in the presence of thrombin. VSMCs were stained with ACTA2 (red) and nuclei visualized with DAPI (n = 6). Scale bars: 2.5 μm. (E) 3D reconstruction of confocal Z-stack images of the whole VSMCs cocultured with CMFDA-labeled platelets (green). (F–I) Transmission electron microscopy imaging of VSMCs cocultured with APs. Panels provide sequence of platelet internalization and incorporation into VSMCs. The red arrow indicates the entering and internalized platelet. Lower-magnification source electron micrographs are shown in Supplemental Figure 3. Scale bars: 1 μm. (J) Representative images of sections from uninjured and injured femoral arteries of PF4-mT/mG and WT mice (n = 7). Arrows indicated multiple VSMCs with incorporated green platelets. The injured femoral arteries were harvested on the seventh day after injury. Scale bars: 20 μm. Statistical significance was determined using 1-way ANOVA followed by Tukey-Kramer multiple-comparisons test (A–C).