Preparation Of Bionic Scaffold Based Bacterial Cellulose And Its Application In Urethral Repair

Urethral stenosis caused by various reasons such as trauma,infection,and iatrogenic procedures is a common disease in urology.With the development and application of tissue engineering,it is expected to achieve the repair and reconstruction of complex urethral strictures through this technology.As an important part of tissue engineering,tissue engineering scaffolds are designed to simulate the microenvironment of cells.Natural extracellular matrix is a micro/nano structure composed of interwoven nanofibers,which combines micron scale pores and nanofibers.Micron scale pores can enhance cell proliferation,adhesion,and migration,and provide spatial structure and mechanical support for nutrient exchange and metabolic excretion.Its nanofibers help regulate cell behavior and signal expression.In terms of macroscopic morphology,the spatial characteristics of the scaffold enable the geometric assembly of cells to not only conform to natural tissue structure,but also allow cell/scaffold composites to be transplanted in the optimal tissue form.Therefore,the preparation of tissue engineering scaffolds with both macroscopic morphology and micro/nano structure is of practical significance.This article is based on the study of the structure of human urethral acellular matrix,and designs and prepares a urethral biomimetic tissue engineering scaffold that combines the macroscopic morphology and micro/nano structure of urethral acellular matrix,with the aim of applying it to the repair and reconstruction of urethral stricture.The main research content is as follows:The two-dimensional imaging information of the human urethra was obtained using ultrasound scanning,and a digital model of urethral tissue was constructed using3D reconstruction software.A silicone mold with macroscopic morphology of the urethra was obtained through indirect 3D printing technology.Gelatin（Gel）was mixed with Silk Fibroin（SF）aqueous solution to prepare Gel/SF tubular porous scaffolds with macroscopic morphology of urethral tissue using freeze-drying method.Based on the successful simulation of macroscopic morphology,the relationship between preparation conditions and structural properties was explored,and porous scaffolds close to the urethral acellular matrix were selected,The three-dimensional nano network structure of bacterial cellulose（BC）was introduced using in-situ fermentation method,and the Gel/SF/BC urethral biomimetic scaffold was finally obtained.The microstructure,chemical structure,mechanical properties and biocompatibility were further tested.The above multi-scale construction method was used to prepare the canine urethral bionic stent,and the biocompatibility of the urethral bionic stent in vivo and the zoology evaluation of urethral defect were carried out.The main conclusions are as follows:（1）The effects of different pre cooling temperature and different proportion of gelatin silk protein on the pore structure,mechanical properties and porosity of porous scaffolds were studied by freeze drying method.Under a constant temperature field,the pore size of the support decreases with the decrease of pre cooling temperature,but exhibits an oriented structure generated along the temperature field at-196℃.Pore size（100～300μm）of porous scaffolds with pre freezing temperature of-20℃.The porosity（73%～83%）is close to that of the urethral acellular matrix.The porous scaffold with a pre freezing temperature of-20℃exhibits a honeycomb like morphology with high tensile and compressive strength and modulus,but low fracture growth rate as the proportion of gelatin increases;Silk accounts for a large proportion,with a layered porous structure and low tensile and compressive strength and modulus,but high fracture growth rate;There may be some intermolecular hydrogen bonds in the volume mixing of Gel/SF,thus enhancing its water absorption performance.When the proportion of Gel is high,there may be some intramolecular hydrogen bonds,thereby reducing its water absorption performance.The porous scaffold of G₅S₅ has good comprehensive morphology,structure,and mechanical properties.（2）The Gel/SF/BC urethral biomimetic scaffold has a macroscopic morphology of urethral tissue,forming a dense inner layer and a porous outer layer.The inner layer is composed of a simple BC membrane,and the outer layer is made of Gel/SF porous scaffold as the skeleton.BC nanofibers are evenly distributed on the surface of the porous pore wall;The Gel/SF/BC urethral biomimetic stent has good mechanical and water absorption properties,and has demonstrated good biocompatibility through cytotoxicity,skin sensitization,and skin irritation experiments.（3）The canine urethral bionic stent was prepared by the multi-scale construction method of human urethral bionic stent.The biocompatibility of the urethral bionic stent in vivo and the zoology evaluation of urethral defect were carried out.The results showed that the Gel/SF/BC urethral biomimetic scaffold combines the macroscopic morphology and micro/nano structure of canine urethral tissue,has good in vivo biocompatibility,and successfully repaired a 7 mm complete urethral defect in Beagle dogs without seed cells.