| Summary: | 碩士 === 國立臺灣科技大學 === 醫學工程研究所 === 107 === Clinically, tissue reconstruction is needed in patients with urethral and ureteral injury with long segment defects. However, there is no customized platform and suitable bio-compatibility materials to produce the tissue regeneration vehicle currently. We hope this customized scaffold could be a useful replacement material in tissue regeneration for urological trauma patients. Electrospinning (ES) system is a new nanofiber process technology in recent years. It can directly and quickly spin the material into nanofibers, improve the specific surface area, porosity and micro/nano-scale pore structure. 3D printing technology can provide uniform and customized brackets in different fields. Our study chooses highly bio-compatibility materials, silk fibroin (SF) and polycaprolactone (PCL) as bio-materials scaffolds respectively. The differences in physicochemical properties and biocompatibility of bio-scaffolds prepared by ES and 3D printing combined with electrospinning (3D-ES) were compared.
SF powder was prepared from dissolved cocoons, then extracted by dialysis and lyophilized before study. Various ratios of PCL and SF nanofiber in formic acid were generated by ES or self-designed 3D-ES platform. The physical and biological characteristics were assayed by SEM, degradation test, tension test and cell attachment assay of fibroblasts and urothelial cells on various nanofibers of PCL-SF. The ex vivo resected ureteral tissue was anastomosed with the PCL-SF scaffolds and cultured in ex vivo bath for two weeks. The cellular growth on scaffold was observed microscopically by HE stain. In the New Zealand white rabbit model, we performed a 1/5 ratio (2 cm) replacement of the unilateral ureter(rabbit ureter is about 10 cm length). After 6 and 7 weeks, the animals were sacrificed and the scaffolds were taken out for tissue sectioning, and the cellular growth was observed by HE and Masson staining.
In physical assay, both the diameter and size of nanofiber holes were increased in the PCL-SF scaffold when the proportion of SF is increased. There is no difference for degradation rate under cell culture medium soaking for 8 weeks which was 10% less in dry weight. The tensile strength of PCL-SF nanofibers increased when the PCL ratio increased. Typical spectrum peaks for PCL and SF were observed in the spectra of PCL/SF blends. MTT result indicates that the incorporation of SF into PCL was beneficial for cell proliferation and parallel to the percentage of SF ratio and 4:6 of PCL-SF scaffold is the best ratio for cellular growth. Higher cellular proliferation was seen in 4:6 of 3D-ES PCL-SF scaffolds than 4:6 of ES PCL-SF scaffolds due to increased diameter of nanofibers and size of holes. The PCL-SF scaffold anastomosis in ex vivo bath showed cellular growth stably along the scaffold for two weeks and most of the cells grow along the outboard. In animal model, after 6 and 7 weeks, different cells can be observed to grow along the outboard of the scaffold, from the lumen outward: Mucosa, Submucosa, muscular layer and the serosa layer, mucosal layer growth in the scaffold inner edge of proximal (kidney) side, Mucosa and muscular layer growth in the scaffold inner edge of distal (bladder) side, However, in the middle of the inner edge of the scaffold, the cells had not grown yet till 7 weeks.
In our study, 3D-ES produced 4:6 PCL-SF nanofiber scaffolds are suitable for cell tissue growth, and achieve the purpose of ureteral reconstruction in animal experiments. Therefore this new form scaffold can be used as clinical urinary tract tissue reconstruction in the future.
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