RT Journal Article
SR Electronic
T1 82 Microrna-21 regulates transforming growth factor beta-1 release from platelets
JF Heart
JO Heart
FD BMJ Publishing Group Ltd and British Cardiovascular Society
SP A71
OP A71
DO 10.1136/heartjnl-2018-BCS.82
VO 104
IS Suppl 6
A1 Temo Barwari
A1 Seda Eminaga
A1 Paul Armstrong
A1 Melissa Chan
A1 Ruifang Lu
A1 Javier Barallobre-Barreiro
A1 Xiaoke Yin
A1 Raimund Pechlaner
A1 Sarah Langley
A1 Anna Zampetaki
A1 Peter Santer
A1 Martin Weger
A1 Timothy Warner
A1 Stefan Kiechl
A1 Johann Willeit
A1 Ajay Shah
A1 Manuel Mayr
YR 2018
UL http://heart.bmj.com/content/104/Suppl_6/A71.1.abstract
AB Introduction MicroRNA-21 (miR-21) has been implicated as a regulator of fibrosis. Inhibitors of miR-21 are currently undergoing clinical trials. Our aim was to explore mechanisms of how miR-21 inhibition attenuates fibrosis using a proteomics approach.Methods and results Primary murine cardiac fibroblasts (CF) were transfected with miR-21 mimics and inhibitors. In line with previous reports, miR-21 overexpression increased CF proliferation (n=4 vs. 4; p<0.0001 for 2-way ANOVA). Unexpectedly, proteomic analysis revealed limited effects on extracellular matrix (ECM) secretion. Similarly, the ECM protein composition of hearts from miR-21 null mice was similar to wild-type controls. Thus, we searched for additional explanations as to how miR-21 might regulate fibrosis. In the community-based Bruneck study (n=671), we compared circulating miR-21 with a proteomic panel of 229 proteins associated with cardiovascular disease. Several platelet-derived pro-fibrotic factors significantly correlated with plasma miR-21 levels, including the latency-associated peptide of TGF-β1 (Pearson r 0.68; q<0.0001). Argonaute2 immunoprecipitation demonstrated enrichment of miR-21 in a human megakaryoblastic cell line. Furthermore, TGF-β1 was abundant in megakaryocytes as demonstrated by immunofluorescence staining of murine bone marrow. Platelet immunodepletion in mice (n=5 vs. 7 for control and anti-CD42b antibody injection, respectively) revealed a strong platelet dependency of plasma TGF-β1 levels (fold change: 2.37; p=0.003). Correlation of miR-21 with total TGF-β1 was confirmed in humans by ELISA measurements in platelet-poor plasma collected during the Bruneck 2015 follow-up (n=332; Pearson r: 0.63; q<0.0001). When mice were treated with antagomiR-21, no differences were observed in the platelet count or in their aggregation response to different agonists. However, a proteomics analysis of the platelet releasate revealed TGF-β1 as differentially regulated. An attenuated TGF-β1 release after antagomiR-21 treatment (n=5 vs. 5 for control and antagomiR-21) was validated in platelets activated with collagen and a thrombin receptor-activating peptide (p=0.004 and 0.002, respectively, for 2-way ANOVA). Mechanistically, Wiskott-Aldrich Syndrome protein, a negative regulator of platelet TGF-β1 secretion, was identified as a direct target of miR-21.Conclusions This study reports a previously unrecognised effect of miR-21 inhibition on platelet TGF-β1 release. This novel systemic effect of miR-21 on platelets may contribute to the anti-fibrotic effects of miR-21 inhibitors.