Electron microscopy analysis of endothelial-to-mesenchymal transition in native and prosthetic blood vessels and heart valves

Aim. To analyse the electron microscopy signs of endothelial-to-mesenchymal transition (EndoMT) in endothelial cells (ECs) of native blood vessels and medical devices for cardiovascular surgery (including vascular patches, stents, and bioprosthetic heart valves) for assessing the role of EndoMT in the pathogenesis of implantation-associated long-term complications. Materials and Methods. To achieve this task, we interrogated internal thoracic artery used as a coronary bypass conduit, ovine carotid artery with a xenopericardial patch, stented human carotid artery, and bioprosthetic aortic valve. The tissues were stained with heavy metals, embedded into epoxy resin, grinded and polished, sputter coated with carbon, and visualised using backscattered scanning electron microscopy (EM-BSEM technique). Results. All samples contained both physiological ECs and the cells undergoing EndoMT. Physiological ECs retained apicobasal polarity, elongated nucleus, and intact basement membrane. Cells undergoing EndoMT had polymorphic nuclei, loss of apicobasal polarity, degraded and disintegrated basement membrane and internal elastic lamina, and migration into the vascular wall. Numerous immune cells (i.e., neutrophils, eosinophils, and macrophages) in the subendothelial layer indicated distinct stages of inflammation and vascular or valvular remodeling. Cell undergoing EndoMT migrated into the medial layer acquiring a myofibroblastic phenotype. Conclusion. These results confirm the role of EndoMT in the pathogenesis of the complications associated with the implantation of medical devices. Signs of dysfunctional ECs and EndoMT have been accompanied by inflammation, extracellular matrix degradation, and disruption of endothelial barrier altogether promoting intimal hyperplasia and restenosis as well as structural valve deterioration. These data require the detailed investigation of EndoMT molecular mechanisms and therapeutic approaches of its prevention in patients undergoing cardiovascular surgery interventions.

1. Li Y, Lui KO, Zhou B. Reassessing endothelial-to-mesenchymal transition in cardiovascular diseases. Nat Rev Cardiol. 2018;15(8):445–456. https://doi.org/10.1038/s41569-018-0023-y

2. Kovacic JC, Dimmeler S, Harvey RP, Finkel T, Aikawa E, Krenning G et al. Endothelial to Mesenchymal Transition in Cardiovascular Disease: JACC State-of-the-Art Review. J Am Coll Cardiol. 2019;73(2):190–209. https://doi.org/10.1016/j.jacc.2018.09.089

3. Chen PY, Schwartz MA, Simons M. Endothelial-to-Mesenchymal Transition, Vascular Inflammation, and Atherosclerosis. Front Cardiovasc Med. 2020;7:53. https://doi.org/10.3389/fcvm.2020.00053

4. Alvandi Z, Bischoff J. Endothelial-Mesenchymal Transition in Cardiovascular Disease. Arterioscler Thromb Vasc Biol. 2021;41(9):2357–2369. https://doi.org/10.1161/ATVBAHA.121.313788

5. Peng Q, Shan D, Cui K, Li K, Zhu B, Wu H et al. The Role of Endothelial-to-Mesenchymal Transition in Cardiovascular Disease. Cells. 2022;11(11):1834. https://doi.org/10.3390/cells11111834

6. Lu X, Gong J, Dennery PA, Yao H. Endothelial-to-mesenchymal transition: Pathogenesis and therapeutic targets for chronic pulmonary and vascular diseases. Biochem Pharmacol. 2019;168:100–107. https://doi.org/10.1016/j.bcp.2019.06.021

7. Yun E, Kook Y, Yoo KH, Kim KI, Lee MS, Kim J et al. Endothelial to Mesenchymal Transition in Pulmonary Vascular Diseases. Biomedicines. 2020;8(12):639. https://doi.org/10.3390/biomedicines8120639

8. Hashimoto N, Phan SH, Imaizumi K, Matsuo M, Nakashima H, Kawabe T, et al. Endothelial-mesenchymal transition in bleomycin-induced pulmonary fibrosis. Am J Respir Cell Mol Biol. 2010;43(2):161–172. https://doi.org/10.1165/rcmb.2009-0031OC

9. Hu M, Guan XH, Wang LF, Xu HM, Ke SF, Yuan QY et al. Endothelial CD38-induced endothelial-to-mesenchymal transition is a pivotal driver in pulmonary fibrosis. Cell Mol Life Sci. 2024;82(1):30. https://doi.org/10.1007/s00018-024-05548-x

10. Kanno Y, Hirota M, Matsuo O, Ozaki KI. alpha2-antiplasmin positively regulates endothelial-to-mesenchymal transition and fibrosis progression in diabetic nephropathy. Mol Biol Rep. 2022;49(1):205–215. https://doi.org/10.1007/s11033-021-06859-z

11. Hall IF, Kishta F, Xu Y, Baker AH, Kovacic JC. Endothelial to mesenchymal transition: at the axis of cardiovascular health and disease. Cardiovasc Res. 2024;120(3):223–236. https://doi.org/10.1093/cvr/cvae021

12. Di Benedetto P, Ruscitti P, Berardicurti O, Vomero M, Navarini L, Dolo V et al. Endothelial-to-mesenchymal transition in systemic sclerosis. Clin Exp Immunol. 2021;205(1):12–27. https://doi.org/10.1111/cei.13599

13. Gierlinger G, Rech L, Emani SM, Del Nido PJ, Friehs I. A Neonatal Heterotopic Rat Heart Transplantation Model for the Study of Endothelial-to-Mesenchymal Transition. J Vis Exp. 2023;(197). https://doi.org/10.3791/65426

14. Wang Z, Han Z, Tao J, Wang J, Liu X, Zhou W et al. Role of endothelial-to-mesenchymal transition induced by TGF-beta1 in transplant kidney interstitial fibrosis. J Cell Mol Med. 2017;21(10):2359–2369. https://doi. org/10.1111/jcmm.13157

15. Souilhol C, Harmsen MC, Evans PC, Krenning G. Endothelial-mesenchymal transition in atherosclerosis. Cardiovasc Res. 2018;114(4):565–577. https://doi.org/10.1093/cvr/cvx253

16. Good RB, Gilbane AJ, Trinder SL, Denton CP, Coghlan G, Abraham DJ et al. Endothelial to Mesenchymal Transition Contributes to Endothelial Dysfunction in Pulmonary Arterial Hypertension. Am J Pathol. 2015;185(7):1850–1858. https://doi.org/10.1016/j.ajpath.2015.03.019

17. Dahal S, Huang P, Murray BT, Mahler GJ. Endothelial to mesenchymal transformation is induced by altered extracellular matrix in aortic valve endothelial cells. J Biomed Mater Res A. 2017;105(10):2729–2741. https://doi.org/10.1002/jbm.a.36133

18. Immanuel J, Yun S. Vascular Inflammatory Diseases and Endothelial Phenotypes. Cells. 2023;12(12):1640. https://doi.org/10.3390/cells12121640

19. Anaraki KT, Zahed Z, Javid RN, Shafiei S, Beiranvandi F, Kahrizsangi NG et al. Immune response following transcatheter aortic valve procedure. Vascul Pharmacol. 2024;154:107283. https://doi.org/10.1016/j.vph.2024.107283

20. Kostyunin A, Glushkova T, Stasev A, Mukhamadiyarov R, Velikanova E, Bogdanov L et al. Early Postoperative Immunothrombosis of Bioprosthetic Mitral Valve and Left Atrium: A Case Report. Int J Mol Sci. 2022;23(12):6736. https://doi.org/10.3390/ijms23126736

21. Human P, Bezuidenhout D, Aikawa E, Zilla P. Residual Bioprosthetic Valve Immunogenicity: Forgotten, Not Lost. Front Cardiovasc Med. 2022;8:760635. https://doi.org/10.3389/fcvm.2021.760635

22. Veraar C, Koschutnik M, Nitsche C, Laggner M, Polak D, Bohle B et al. Inflammatory immune response in recipients of transcatheter aortic valves. JTCVS Open. 2021;6:85–96. https://doi.org/10.1016/j.xjon.2021.02.012

23. Kostyunin AE, Yuzhalin AE, Rezvova MA, Ovcharenko EA, Glushkova TV, Kutikhin AG. Degeneration of Bioprosthetic Heart Valves: Update 2020. J Am Heart Assoc. 2020;9(19):e018506. https://doi.org/10.1161/JAHA.120.018506

24. Wechsler ME, Munitz A, Ackerman SJ, Drake MG, Jackson DJ, Wardlaw AJ et al. Eosinophils in Health and Disease: A State-of-the-Art Review. Mayo Clin Proc. 2021;96(10):2694–2707. https://doi.org/10.1016/j.mayocp.2021.04.025

25. Ramirez GA, Yacoub MR, Ripa M, Mannina D, Cariddi A, Saporiti N, et al. Eosinophils from Physiology to Disease: A Comprehensive Review. Biomed Res Int. 2018;2018:9095275. https://doi.org/10.1155/2018/9095275

26. Hendrickson MJ, Wallace ZS. Mechanisms and Screening for Atherosclerosis in Adults With Vasculitis. Arterioscler Thromb Vasc Biol. 2025;45(1):3–10. https://doi.org/10.1161/ATVBAHA.124.319982

27. Shao Y, Saredy J, Yang WY, Sun Y, Lu Y, Saaoud F et al. Vascular Endothelial Cells and Innate Immunity. Arterioscler Thromb Vasc Biol. 2020;40(6):e138–e152. https://doi.org/10.1161/ATVBAHA.120.314330

28. Wang L, Luqmani R, Udalova IA. The role of neutrophils in rheumatic disease-associated vascular inflammation. Nat Rev Rheumatol. 2022;18(3):158–170. https://doi.org/10.1038/s41584-021-00738-4

29. Pugh D, Karabayas M, Basu N, Cid MC, Goel R, Goodyear CS et al. Large-vessel vasculitis. Nat Rev Dis Primers. 2022;7(1):93. https://doi.org/10.1038/s41572-021-00327-5

30. Shishkova DK, Sinitskaya AV, Sinitsky MYu, Matveeva VG, Velikanova EA, Markova VE et al. Spontaneous endothelial-to-mesenchymal transition in human primary umbilical vein endothelial cells. Complex Issues of Cardiovascular Diseases = Kompleksnye problemy serdečno-sosudistyh zabolevanij. 2022:11(3):97–114. https://doi.org/10.17802/2306-1278-2022-11-3-97-114.