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Mechanical strain triggers endothelial-to-mesenchymal transition of the endocardium in the immature heart

Abstract

Background

Endothelial-to-mesenchymal-transition (EndMT) plays a major role in cardiac fibrosis, including endocardial fibroelastosis but the stimuli are still unknown. We developed an endothelial cell (EC) culture and a whole heart model to test whether mechanical strain triggers TGF-β-mediated EndMT.

Methods

Isolated ECs were exposed to 10% uniaxial static stretch for 8 h (stretch) and TGF-β-mediated EndMT was determined using the TGF-β-inhibitor SB431542 (stretch + TGF-β-inhibitor), BMP-7 (stretch + BMP-7) or losartan (stretch + losartan), and isolated mature and immature rats were exposed to stretch through a weight on the apex of the left ventricle. Immunohistochemical staining for double-staining with endothelial markers (VE-cadherin, PECAM1) and mesenchymal markers (αSMA) or transcription factors (SLUG/SNAIL) positive nuclei was indicative of EndMT.

Results

Stretch-induced EndMT in ECs expressed as double-stained ECs/total ECs (cells: 46 ± 13%; heart: 15.9 ± 2%) compared to controls (cells: 7 ± 2%; heart: 3.1 ± 0.1; p < 0.05), but only immature hearts showed endocardial EndMT. Inhibition of TGF-β decreased the number of double-stained cells significantly, comparable to controls (cells/heart: control: 7 ± 2%/3.1 ± 0.1%, stretch: 46 ± 13%/15 ± 2%, stretch + BMP-7: 7 ± 2%/2.9 ± 0.1%, stretch + TGF-β-inhibitor (heart only): 5.2 ± 1.3%, stretch + losartan (heart only): 0.89 ± 0.1%; p < 0.001 versus stretch).

Conclusions

Endocardial EndMT is an age-dependent consequence of increased strain triggered by TGF- β activation. Local inhibition through either rebalancing TGF-β/BMP or with losartan was effective to block EndMT.

Impact

  • Mechanical strain imposed on the immature LV induces endocardial fibroelastosis (EFE) formation through TGF-β-mediated activation of endothelial-to-mesenchymal transition (EndMT) in endocardial endothelial cells but has no effect in mature hearts.

  • Local inhibition through either rebalancing the TGF-β/BMP pathway or with losartan blocks EndMT.

  • Inhibition of endocardial EndMT with clinically applicable treatments may lead to a better outcome for congenital heart defects associated with EFE.

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Fig. 1: Static stretch triggers EndMT in isolated endothelial cells.
Fig. 2: Stretch-induced EndMT is mediated through activation of the TGF-β pathway.
Fig. 3: Excess strain induces EndMT in immature hearts.
Fig. 4: Excess strain does not induce EndMT in mature hearts.
Fig. 5: EndMT-regulating transcription factors (SLUG/SNAIL) in immature versus mature hearts.

Data availability

The original data set used and analyzed during the current study is readily available from the corresponding author on reasonable request.

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Acknowledgements

This research did not receive any specific grant from funding agencies in the public commercial or not-for-profit sectors but was supported by discretionary departmental funds.

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Contributions

C.V., V.W., and I.F. provided substantial contributions to conception and design, C.V., V.W., M.D., P.H., R.-Z.L., and I.F. contributed to the acquisition and analysis of data. C.V., V.W., M.D., P.H., J.M.M.-M., R.A.-F., P.J.d.N., and I.F. were involved in the interpretation of data. C.V., V.W., and I.F. drafted the article, and R.A.-F., P.H., R.-Z.L., J.M.M.-M., and P.J.d.N. commented for revision of the manuscript. All authors granted final approval of the version to be published.

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Correspondence to Ingeborg Friehs.

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Vorisek, C., Weixler, V., Dominguez, M. et al. Mechanical strain triggers endothelial-to-mesenchymal transition of the endocardium in the immature heart. Pediatr Res 92, 721–728 (2022). https://doi.org/10.1038/s41390-021-01843-6

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