Endothelialization of acellular scaffold from sterilized bovine pericardium / 中国组织工程研究

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.