New Tissue-Engineered Vascular Matrix Based on Regenerated Silk Fibroin: in vitro Study

The aim of the study was to make a vascular patch based on regenerated silk fibroin (SF) and study its physical and mechanical characteristics, biocompatibility and matrix properties in comparison with polyhydroxybutyrate/valerate/polycaprolactone with incorporated vascular endothelial growth factor (PHBV/PCL/VEGF) and commercial bovine xenopericardium (XP) flap in experiments in vitro.

Materials and Methods. Tissue-engineered matrices were produced by electrospinning. The surface structure, physical and mechanical characteristics, hemocompatibility (erythrocyte hemolysis, aggregation, adhesion and activation of platelets after contact with the material) and matrix properties of vascular patches (adhesion, viability, metabolic activity of EA.hy926 cells on the material) were studied.

Results. The surface of SF-based matrices and PHBV/PCL/VEGF-based tissue engineered patches had a porous and fibrous structure compared to a denser and more uniform XP flap. The physical and mechanical characteristics of SF matrices were close to those of native vessels. Along with this, tissue-engineered patches demonstrated high hemocompatible properties, which do not differ from those for commercial XP flap. Adhesion, viability, and metabolic activity of EA.hy926 endothelial cells also corresponded to the previously developed PHBV/PCL/VEGF matrix and XP flap, which indicates the nontoxicity and biocompatibility of SF matrices.

Conclusion. Matrices produced from regenerated SF demonstrated satisfactory results, comparable to those for PHBV/PCL/VEGF and commercial XP flap, and in the case of platelet adhesion and activation, they outperformed these patches. In total, SF can be defined as material having sufficient biological compatibility, which makes it possible to consider a tissue-engineered matrix made from it as promising for implantation into the vascular wall.

1. Fukuda S., Shimogonya Y., Yonemoto N., Fukuda M., Watanabe A., Fujiwara K., Enomoto R., Hasegawa K., Yasoda A., Tsukahara T.; NHO Carotid CFD Study Group. Hemodynamic risk factors for the development of carotid stenosis in patients with unilateral carotid stenosis. World Neurosurg 2022; 160: e353–e371, https://doi.org/10.1016/j.wneu.2022.01.019.
2. Messas E., Goudot G., Halliday A., Sitruk J., Mirault T., Khider L., Saldmann F., Mazzolai L., Aboyans V. Management of carotid stenosis for primary and secondary prevention of stroke: state-of-the-art 2020: a critical review. Eur Heart J Suppl 2020; 22(Suppl M): M35–M42, https://doi.org/10.1093/eurheartj/suaa162.
3. AbuRahma A.F., Avgerinos E.D., Chang R.W., Darling R.C. III, Duncan A.A., Forbes T.L., Malas M.B., Murad M.H., Perler B.A., Powell R.J., Rockman C.B., Zhou W. Society for Vascular Surgery clinical practice guidelines for management of extracranial cerebrovascular disease. J Vasc Surg 2022; 75(1S): 4S–22S, https://doi.org/10.1016/j.jvs.2021.04.073.
4. Suroto N.S., Rantam F.A., Al Fauzi A., Widiyanti P., Turchan A., Pangaribuan V. Selection criteria for patch angioplasty material in carotid endarterectomy. Surg Neurol Int 2022; 13: 362, https://doi.org/10.25259/sni_470_2022.
5. Antonova L.V., Krivkina E.O., Sevostianova V.V., Mironov A.V., Rezvova M.A., Shabaev A.R., Tkachenko V.O., Krutitskiy S.S., Khanova M.Y., Sergeeva T.Y., Matveeva V.G., Glushkova T.V., Kutikhin A.G., Mukhamadiyarov R.A., Deeva N.S., Akentieva T.N., Sinitsky M.Y., Velikanova E.A., Barbarash L.S. Tissue-engineered carotid artery interposition grafts demonstrate high primary patency and promote vascular tissue regeneration in the ovine model. Polymers (Basel) 2021; 13(16): 2637, https://doi.org/10.3390/polym13162637.
6. Khanova M.Yu., Velikanova E.A., Glushkova T.V., Matveeva V.G. Development of personalized cell-populated vascular graft in vitro. Kompleksnye problemy serdecno-sosudistyh zabolevanij 2021; 10(2): 89–93, https://doi.org/10.17802/2306-1278-2021-10-2S-89-93.
7. Fang G., Sapru S., Behera S., Yao J., Shao Z., Kundu S.C., Chen X. Exploration of the tight structural-mechanical relationship in mulberry and non-mulberry silkworm silks. J Mater Chem B 2016; 4(24): 4337–4347, https://doi.org/10.1039/c6tb01049k.
8. Agapova O.I., Efimov A.E., Moisenovich M.M., Bogush V.G., Agapov I.I. Comparative analysis of three-dimensional nanostructure of porous biocompatible scaffolds made of recombinant spidroin and silk fibroin for regenerative medicine. Vestnik transplantologii i iskusstvennyh organov 2015; 17(2): 37–44, https://doi.org/10.15825/1995-1191-2015-2-37-44.
9. Cetin Y., Sahin M.G., Kok F.N. Application potential of three-dimensional silk fibroin scaffold using mesenchymal stem cells for cardiac regeneration. J Biomater Appl 2021; 36(4): 740–753, https://doi.org/10.1177/08853282211018529.
10. Gavrilova N.A., Borzenok S.A., Revishchin A.V., Tishchenko O.E., Ostrovkiy D.S., Bobrova M.M., Safonova L.A., Efimov A.E., Agapova O.I., Agammedov M.B., Pavlova G.V., Agapov I.I. The effect of biodegradable silk fibroin-based scaffolds containing glial cell line-derived neurotrophic factor (GDNF) on the corneal regeneration process. Int J Biol Macromol 2021; 185: 264–276, https://doi.org/10.1016/j.ijbiomac.2021.06.040.
11. Partlow B.P., Hanna C.W., Rnjak-Kovacina J., Moreau J.E., Applegate M.B., Burke K.A., Marelli B., Mitropoulos A.N., Omenetto F.G., Kaplan D.L. Highly tunable elastomeric silk biomaterials. Adv Funct Mater 2014; 24(29): 4615–4624, https://doi.org/10.1002/adfm.201400526.
12. Agammedov M.B., Ostrovsky D.S., Sobolev V.P., Ushakova L.I., Agapov I.I., Gavrilova N.A., Borzenok S.A. Pathogenetic and regenerative features of corneal damage repair with biodegradable materials based on silk fibroin. Patogenez 2022; 20(4): 63–68, https://doi.org/10.25557/2310-0435.2022.04.63-68.
13. Meinel L., Hofmann S., Karageorgiou V., Kirker-Head C., McCool J., Gronowicz G., Zichner L., Langer R., Vunjak-Novakovic G., Kaplan D.L. The inflammatory responses to silk films in vitro and in vivo. Biomaterials 2005; 26(2): 147–155, https://doi.org/10.1016/j.biomaterials.2004.02.047.
14. Sun W., Gregory D.A., Tomeh M.A., Zhao X. Silk fibroin as a functional biomaterial for tissue engineering. Int J Mol Sci 2021; 22(3): 1499, https://doi.org/10.3390/ijms22031499.
15. Alessandrino A., Chiarini A., Biagiotti M., Dal Prà I., Bassani G.A., Vincoli V., Settembrini P., Pierimarchi P., Freddi G., Armato U. Three-layered silk fibroin tubular scaffold for the repair and regeneration of small caliber blood vessels: from design to in vivo pilot tests. Front Bioeng Biotechnol 2019; 7: 356, https://doi.org/10.3389/fbioe.2019.00356.
16. Chan A.H.P., Filipe E.C., Tan R.P., Santos M., Yang N., Hung J., Feng J., Nazir S., Benn A.J., Ng M.K.C., Rnjak-Kovacina J., Wise S.G. Altered processing enhances the efficacy of small-diameter silk fibroin vascular grafts. Sci Rep 2019; 9(1): 17461, https://doi.org/10.1038/s41598-019-53972-y.
17. Dingle Y.T.L., Bonzanni M., Liaudanskaya V., Nieland T.J.F., Kaplan D.L. Integrated functional neuronal network analysis of 3D silk-collagen scaffold-based mouse cortical culture. STAR Protoc 2021; 2(1): 100292, https://doi.org/10.1016/j.xpro.2020.100292.
18. Antonova L.V., Mironov A.V., Shabaev A.R., Silnikov V.N., Krivkina E.O., Matveeva V.G., Velikanova E.A., Senokosova E.A., Khanova M.Yu., Sevostyanova V.V., Glushkova T.V., Mukhamadiyarov R.A., Barbarash L.S. Tissue-engineered vascular patches: comparative characteristics and preclinical test results in a sheep model. Vestnik transplantologii i iskusstvennyh organov 2022; 24(4): 94–108, https://doi.org/10.15825/1995-1191-2022-4-94-108.
19. Antonova L.V., Sevost’yanova V.V., Rezvova M.A., Krivkina E.O., Kudryavtseva Yu.A., Barbarash O.L., Barbarash L.S. Technology of producing functionally active biodegradable small-diameter vascular prostheses with drug coating. Patent RU 2702239. 2019.
20. Ye X., Wang Z., Zhang X., Zhou M., Cai L. Hemocompatibility research on the micro-structure surface of a bionic heart valve. Biomed Mater Eng 2014; 24(6): 2361–2369, https://doi.org/10.3233/bme-141049.
21. Wang Z., Lin M., Xie Q., Sun H., Huang Y., Zhang D., Yu Z., Bi X., Chen J., Wang J., Shi W., Gu P., Fan X. Electrospun silk fibroin/poly(lactide-co-ε-caprolactone) nanofibrous scaffolds for bone regeneration. Int J Nanomedicine 2016; 11: 1483–1500, https://doi.org/10.2147/ijn.s97445.
22. Zhang X., Xiao L., Ding Z., Lu Q., Kaplan D.L. Fragile-tough mechanical reversion of silk materials via tuning supramolecular assembly. ACS Biomater Sci Eng 2021; 7(6): 2337–2345, https://doi.org/10.1021/acsbiomaterials.1c00181.
23. Di Nardo A., Louvelle L., Romero D.A., Doyle M., Forbes T.L., Amon C.H. A comparison of vessel patch materials in tetralogy of Fallot patients using virtual surgery techniques. Ann Biomed Eng 2023; 1, https://doi.org/10.1007/s10439-023-03144-x.
24. Jolee Bartrom B.S. ASTM hemolysis. NAMSA; 2008; p. 1–12.
25. Sevostianova V.V., Antonova L.V., Mironov A.V., Yuzhalin A.E., Silnikov V.N., Glushkova T.V., Godovikova T.S., Krivkina E.O., Bolbasov E., Akentyeva T.N., Khanova M.Y., Matveeva V.G., Velikanova E.A., Tarasov R.S., Barbarash L.S. Biodegradable patches for arterial reconstruction modified with RGD peptides: results of an experimental study. ACS Omega 2020; 5(34): 21700–21711, https://doi.org/10.1021/acsomega.0c02593.

Prokudina E.S., Senokosova E.A., Antonova L.V., Krivkina E.O., Velikanova E.A., Akentieva T.N., Glushkova T.V., Matveeva V.G., Kochergin N.A. New Tissue-Engineered Vascular Matrix Based on Regenerated Silk Fibroin: in vitro Study. Sovremennye tehnologii v medicine 2023; 15(4): 41, https://doi.org/10.17691/stm2023.15.4.04