ABSTRACT
BACKGROUND: In the research of tissue engineering, there are inevitably various kinds of microorganisms attached to animal tissues, and asepsis is a basic requirement of clinical application of tissue engineering materials. OBJECTIVE: To investigate the effect of 75% ethanol sterilization on the properties and biocompatibility of bovine pericardium. METHODS: Bovine pericardial tissue was sterilized with sterile PBS (control group), PBS containing 1% antibiotic (penicillin/streptomycin/ amphotericin B solution), chlorhexidine and 75% ethanol. LB solid medium was used to evaluate the bactericidal effect of four methods. VB staining was used to evaluate the effect of four sterilization treatments on the tissue structure of bovine pericardium. The cytotoxicity of four sterilized extracts was determined by the CCK-8 assay. The bovine pericardium was sterilized with 75% ethanol and then used to make acellular scaffold, which was co-cultured with human umbilical vein endothelial cells to observe the effect of cell adhesion and endothelialization. RESULTS AND CONCLUSION: (1) The bovine pericardium treated with 75% ethanol and chlorhexidine for 24 hours met the requirements of complete sterilization, and significant colony formation was observed in 1% antibiotic-treated and control groups. (2) VB staining revealed that the collagen fibers of bovine pericardium treated with 75% ethanol, chlorhexidine and 1% antibiotic were arranged in wavy pattern, with compact structure, less elastic fiber content but clear structure.(3) Bovine pericardium treated with 75% ethanol did not affect the proliferation activity of L929 cells, and the cell survival rate within 1-3 days was more than 100%. Chlorhexidine-sterilized bovine pericardium had strong cytotoxicity, leading to cell death. (4) Human umbilical vein endothelial cells grew and adhered normally on the surface of acellular scaffold. During the 20-day implantation period, the highest number of cells adhering to acellular scaffold appeared on days 8-12. These results suggest that 75% ethanol could effectively eliminate all microorganisms attached to the bovine pericardium without affecting the histological integrity and biocompatibility.
ABSTRACT
The chemical extraction method was used to prepare the rat uterine decellularized scaffolds, and to investigate the feasibility of preparing the extracellular matrix (ECM) hydrogel. The rat uterus were collected and extracted by 1%sodium dodecyl sulfate (SDS), 3% TritonX-100 and 4% sodium deoxycholate (SDC) in sequence. Scanning electron microscopy, histochemical staining and immunohistochemistry was used to assess the degree of decellularization of rat uterine scaffold. The prepared decellularized scaffold was digested with pepsin to obtain a uterine ECM hydrogel, and the protein content of ECM was determined by specific ELISA kit. Meanwhile, the mechanical characteristic of ECM hydrogel was measured. The results showed that the chemical extraction method can effectively remove the cells effectively in the rat uterine decellularized scaffold, with the ECM composition preserved completely. ECM hydrogel contains a large amount of ECM protein and shows a good stability, which provides a suitable supporting material for the reconstruction of endometrium .
ABSTRACT
The purpose of this study was to fabricate decelluarized valve scaffold modified with polyethylene glycol nanoparticles loaded with transforming growth factor-β1 (TGF-β1),by which to improve the extracellular matrix microenvironment for heart valve tissue engineering in vitro.Polyethylene glycol nanoparticles were obtained by an emulsion-crosslinking method,and their morphology was observed under a scanning electron microscope.Decelluarized valve scaffolds,prepared by using trypsinase and TritonX-100,were modified with nanoparticles by carbodiimide,and then TGF-β1 was loaded into them by adsorption.The TGF-β1 delivery of the fabricated scaffold was measured by asing enzyme-linked immunosorbent assay.Whether unseeded or reseeded with myofibroblast from rats,the morphologic,biochemical and biomechanical characteristics of hybrid scaffolds were tested and compared with decelluarized scaffolds under the same conditions.The enzyme-linked immunosorbent assay revealed a typical delivery of nanoparticles.The morphologic observations and biological data analysis indicated that fabricated scaffolds possessed advantageous biocompatibility and biomechanical property beyond decelluarized scaffolds.Altogether this study proved that it was feasible to fabricate the hybrid scaffold and effective to improve extracellular matrix microenvironment,which is beneficial for an application in heart valve tissue engineering.