A Combination of Biomesh Processing Techniques Towards Functional Improvement: In Vitro and In Vivo Assessment

Background and Aim: Biomeshes based on decellularized bovinepericardium are the gold standard in reconstructive surgery. They wereintroduced into the area in 1971 by Ionescu et al as bioprostheses ofheart valves. However, their xenogeneic nature shortens their servicelife and reduces implant/host tissue integration efficiency. To the date,significant progress was achieved in biomaterials science concerningthe immunogenicity elimination. Developed approaches involve decellularization, cross-linking, supercritical fluid extraction and geneengineeringtechniques. Each of these techniques possesses their ownadvantages and drawbacks. Aside from immunogenicity, they influenceon functional properties of biomeshes, namely, mechanical and structuralcharacteristics, biodegradability, cytotoxicity, in vivo calcification andvascularization that predetermine the clinical outcome. In our studywe performed a complex characterization of biomeshes prepared usingvarious combinations of decellularization, cross-linking and supercriticalfluid extraction protocols.Methods: Decellularization procedure was based on alkaline treatmentfollowed by cross-linking with a set of chemical agents and supercriticalCO2 treatment as schematically illustrated in Fig. 1. Variouslytreated samples were thoroughly characterized by mechanical trials(nanoindentation, tensile testing), biocompatibility (in vitro extractiontest, contact cytotoxicity study, histological evaluation of explants) andbiodegradability (collagenase A digestion, subcutaneous implantationin mice) evaluation and ultrastructural studies (scanning electronmicroscopy, atomic force microscopy in vitro and optical coherencetomography and multiphoton laser scanning microscopy in vivo).Results: The samples prepared according to the experimental designwere analyzed in a comparative study at each stage of the preparationprocedure. We demonstrated that the cross-linking chemistry stronglyinfluences the biodegradation rate. The values reduced to a differentextent following the cross-linking. Optical properties were examinedboth in vitro and in vivo indicating ultrastructural alterations of thesamples occurring in the series of treatment. Elastic modulus, in general,reduced due to the shrinkage of the material as the result of cross-linking;as well as mechanical anisotropy lowered. In vivo studies indicatedbetter vascularization of cross-linked biomeshes when compared to theintact decellularized pericardium. We attribute this effect to structuralalterations undergoing during cross-linking (increased water swelling,porosity, and proteolytic stability). Additional treatment in supercriticalCO2 reduced acquired cytotoxicity and improved elastic properties.Conclusion: To conclude, our study revealed significant improvementof the functional properties of biomeshes following cross-linking andsupercritical CO2 extraction. These properties are crucial for the successof implant/host tissue integration and enable to tailor the material’sbehavior in vivo that is demanded adequate treatment. This work issupported by the Russian Science Foundation (Grant No. 18-15-00401).

Keywords:

Pericardium , Reconstructive surgery , Xenotransplantation , Tissue-engineering