MicroRNA-146a is a therapeutic target and biomarker for peripartum cardiomyopathy
Julie Halkein, Sebastien P. Tabruyn, Melanie Ricke-Hoch, Arash Haghikia, Ngoc-Quynh-Nhu Nguyen, Michaela Scherr, Karolien Castermans, Ludovic Malvaux, Vincent Lambert, Marc Thiry, Karen Sliwa, Agnes Noel, Joseph A. Martial, Denise Hilfiker-Kleiner, Ingrid Struman
Julie Halkein, Sebastien P. Tabruyn, Melanie Ricke-Hoch, Arash Haghikia, Ngoc-Quynh-Nhu Nguyen, Michaela Scherr, Karolien Castermans, Ludovic Malvaux, Vincent Lambert, Marc Thiry, Karen Sliwa, Agnes Noel, Joseph A. Martial, Denise Hilfiker-Kleiner, Ingrid Struman
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Research Article

MicroRNA-146a is a therapeutic target and biomarker for peripartum cardiomyopathy

Abstract

Peripartum cardiomyopathy (PPCM) is a life-threatening pregnancy-associated cardiomyopathy in previously healthy women. Although PPCM is driven in part by the 16-kDa N-terminal prolactin fragment (16K PRL), the underlying molecular mechanisms are poorly understood. We found that 16K PRL induced microRNA-146a (miR-146a) expression in ECs, which attenuated angiogenesis through downregulation of NRAS. 16K PRL stimulated the release of miR-146a–loaded exosomes from ECs. The exosomes were absorbed by cardiomyocytes, increasing miR-146a levels, which resulted in a subsequent decrease in metabolic activity and decreased expression of Erbb4, Notch1, and Irak1. Mice with cardiomyocyte-restricted Stat3 knockout (CKO mice) exhibited a PPCM-like phenotype and displayed increased cardiac miR-146a expression with coincident downregulation of Erbb4, Nras, Notch1, and Irak1. Blocking miR-146a with locked nucleic acids or antago-miRs attenuated PPCM in CKO mice without interrupting full-length prolactin signaling, as indicated by normal nursing activities. Finally, miR-146a was elevated in the plasma and hearts of PPCM patients, but not in patients with dilated cardiomyopathy. These results demonstrate that miR-146a is a downstream-mediator of 16K PRL that could potentially serve as a biomarker and therapeutic target for PPCM.

Authors

Julie Halkein, Sebastien P. Tabruyn, Melanie Ricke-Hoch, Arash Haghikia, Ngoc-Quynh-Nhu Nguyen, Michaela Scherr, Karolien Castermans, Ludovic Malvaux, Vincent Lambert, Marc Thiry, Karen Sliwa, Agnes Noel, Joseph A. Martial, Denise Hilfiker-Kleiner, Ingrid Struman

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Figure 3

miR-146a can be exported from ECs in exosomes that can be transferred to cardiomyocytes and impair their metabolism.

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miR-146a can be exported from ECs in exosomes that can be transferred to...
(A) Dynamic light scattering analysis of conditioned medium of 16K PRL–treated HUVECs (50 nM, 24 hours). (B) Flow cytometry analysis of exosomes purified from HUVEC medium and labeled with CD63. (C) miR-146a level in exosomes from HUVECs treated with 16K PRL or not treated (NT). (D) Exosome production by HUVECs treated or not with 16K PRL (50 nM, 48 hours). (E) miR-146a level in NRCMs treated or not with 16K PRL. (F) Fluorescence microscopy detecting fusion of miR-146a–loaded endothelial exosomes labeled with the green fluorescent PHK67 membrane linker with NRCMs (α-actinin, red; DAPI, blue). Scale bars: 50 μm. (G) Higher-magnification views of boxed regions in F. Scale bars: 15 μm. (H) Electron micrographs of NRCM sections showing vesicles (arrows); after a 16-hour incubation with HUVEC exosomes, NRCMs showed larger multivesicular vesicles containing the exosomes (inset; enlarged ×2-fold). Scale bars: 500 nm. (I) miR-146a level in NRCMs exposed to miR-146a-exosomes or control-exosomes or transfected with pre-miR-146a or pre-miR-control. (J) Expression level of pri-miR-146a in NRCMs exposed to miR-146a- or control-exosomes. (K) miR-146a expression level, (L) metabolic activity, assessed by MTS assay, and (M) Erbb4 level in NRCMs exposed to control exosomes, miR-146a exosomes, and miR-146a exosomes cotransfected with anti-miR-control or anti-miR-146a. All data are mean ± SD (n ≥ 3). *P < 0.05 vs. respective control; #P < 0.05 vs. miR-146a-exosomes with anti-miR-control. See also Supplemental Figure 3.