Introduction: During modern age, transplantation techniques changed reconstructive surgery scenario, making remarkable improvements concerning the functionality of transplanted organs or tissues. Transplantations show also several pitfalls, such as limited tissue availability and multiple surgeries needed. Moreover, one of the main causes of prosthetic transplants failure is the in situ infection of the surgical site, caused by bacterial colonization that can occur during the operating procedures [1]. For these reasons, decellularization of native tissues has gained a significant attention especially for obtaining biological scaffolds that maintain the native tissues’ ECM structure, thus preserving their biomechanical properties. In order to optimize the post-surgical rehabilitation process, the biological substitutes can be enriched with several functional molecules and/or repopulated with different cell types.
Materials and Methods: Our approach is based on antibiotic enrichment of decellularized bovine pericardium scaffolds Antibiotic (4% gentamicin) has been covalently bound on the substitute surface in order to inhibit the colonization of Staphylococcus aureus, a dangerous nosocomial bacterial strain, until complete healing, and thus favour the post-surgical tissue regeneration.
Evaluation of decellularization and enrichment was performed with DAPI, XPS and ToF-SIMS MS analyses.
Sensitivity tests (inhibition halo, XTT assay) towards 4% gentamicin of Staphylococcus aureus were performed. In order to verify cell sensitivity of tissues interacting with the scaffold, toxicity tests on primary porcine vascular endothelial and smooth muscle cells, were performed. The biomechanical and biocompatibility properties of decellularized and enriched matrices have been characterized, with tensile stress and Young modulus analyses, and MTT assay.
Results and Discussion: The DAPI staining of the analyzed scaffold confirmed the absence of nuclei. Analytical chemistry tests confirmed the efficiency of enrichment method. A significant difference, in terms of inhibition halo formation, between gentamicin (4%) coated and grafted decellularized scaffolds, was observed: a large inhibition halo for the 4% gentamicin covalently enriched samples has been shown. XTT assay showed a decrease of bacterial viability for the 4% gentamicin covalently enriched samples towards other samples.
Regarding the comparison between the mechanical properties of native and decellularized matrices, no significant differences were found in Young moduli. MTT assay showed similar cell viability values for gentamicin enriched samples and untreated controls, meaning that treated materials do not inhibit tissue regeneration.
Conclusion: Our tests confirm the efficiency of the “decellularized tissue” method and its ability to maintain adequate mechanical properties. Thus the enriched biological substitutes could represent an innovative engineering approach for surgical tissue regeneration.
We would like to thank the Assut S.p.A for financial support.
References:
[1] Hasse B. et al., Swiss Med Wkly. 143: w13754, 2013