Poli(3-hidroksi bütirat)/Poli(bütilen adipat-ko-tereftalat)/İpek- Bazlı Tendon Doku İskeleleri: Karakterizasyon ve Tenojenik Aktivite

Abstract

Tendon ruptures are frequently seen in tendons due to inflammation and lesions caused by factors such as mechanical stress accumulation, aging, genetic predisposition, and trauma. Tendon tissue engineering approaches are required for healthy regeneration in the damaged area due to the low number of cells in the tendon tissue, poor vascularization, and low regeneration potential. In the presented thesis, the development of silk fibroin/poly(3-hydroxybutyrate) (SF/P3HB) and silk fibroin/poly(butylene adipate-co terephthalate) (SF/PBAT) aligned nanofibrous matrices that can be used as a patch for tendon regeneration were aimed. The investigation of the in vitro tenogenic activity of rat adipose-derived mesenchymal stem cells (rAdMSC) in these matrices in the presence of growth factor and dynamic culture conditions were also performed.

In the first part of the thesis study, SF/P3HB and SF/PBAT nanofibers were produced using the conventional electrospinning method. Optimum electrospinning conditions were determined by the "response surface methodology" (RSM) approach for both materials. Aligned SF/P3HB and SF/PBAT nanofibrous matrices were obtained with an average diameter of 700 nm.

In the second part, it was found that the transition of the α-helix structure of silk fibroin to the β-sheet structure was induced after the stabilization process applied to the aligned nanofibrous matrices, and both nanofibrous matrices were suitable for use in tendon tissue in terms of mechanical strength, but SF/PBAT nanofibrous matrix was superior in terms of similarity to tendon tissue as mechanically. It was obtained that stabilization and sterilization processes did not affect the surface wettability and crystallinity of the matrices, but cell viability was higher in autoclave sterilized SF/P3HB nanofibrous matrices compared to the ethylene oxide method. It has been determined that SF/PBAT nanofibrous matrices are not suitable for heat treatment. In addition, SF/P3HB nanofibrous matrices were found to have a bacteriostatic effect.

In the third part of the thesis study, a 21-day static culture study was performed with rAdMSCs in aligned SF/P3HB and SF/PBAT nanofibrous matrices in the presence/absence of growth/differentiation factor-5 (GDF-5), and tenogenic differentiation was comparatively examined. Cells were found to be highly viable throughout the culture and organized on the matrices because of alignment. A decrease in mechanical properties was detected in both scaffolds due to the matrix degradation at the end of the culture. It was determined that rAdMSCs cultured on SF/PBAT scaffolds were more suitable for tenogenic activity compared to SF/P3HB due to higher COL1A1, COL3A1, SCX, TNC, and low PPAR gene expressions. In addition, the GDF-5, growth factor that is induced cytoskeletal regulation and differentiation, leading to increased gene expressions. Although the aligned topography is highly effective in the tenogenic differentiation of rAdMSC cells, it was not found to be sufficient in terms of gene expression. In the studies performed with mechanical stimulation bioreactor, it was determined that the SF/P3HB scaffolds did not have sufficient mechanical properties for dynamic culture studies.

It was concluded that aligned SF/PBAT nanofibrous matrices showed superior properties in terms of tenogenic activity compared to SF/P3HB nanofibrous matrices and could be used as patch material in tendon regeneration.