Jing-cong Luo1, Xiu-qun Li, Zhi-ming Yang. 1. Division of Stem Cell and Tissue Engineering, Key Laboratory of Biotherapy of Human Diseases Ministry of Education, West China Hospital, Sichuan University, Chengdu, Sichuan, P. R. China 610041.
Abstract
OBJECTIVE: To prepare human acellular amniotic membrane (HAAM) and to measure its cytocompatibility and biocompatibility. METHODS: HAAM were prepared by chemical detergent-enzymatic extraction. Fresh human amnion was crosslinked with glutaradehyde, shaken in 0.5% SDS for 24 hours, and then treated with 0.25% trypsin for 4 hours. The production were freeze-drying and sterilized using ethylene oxide. Human fibroblasts were isolated from embryo and expanded in vitro. The fibroblasts were seeded in HAAM. HAAM and specimen were stained with HE and Mallory, and observed grossly, under light microscopy and scanning electron microscopy. The HAAM were implanted in the back of SD rats. RESULTS: There were no residues of cells in the HAAM (HE, Mallory staining). One side of HAAM had reticular and porous structure, the other side had compact fibrous structure. Pore size was from 10 to 80 nm. The HAAM could be seeded with expanded fibroblasts in vitro, and fibroblasts had the potential of spread and proliferation. The SD rat in the implant test had no death, convulsions and other abnormal response. CONCLUSION: The detergent-enzymatic extraction process can remove cells and solvable components effectively and preserve the tissue matrix well and keep the reticular structure. The HAAM can be used as an ideal scaffold of biological membrane for tissue engineering.
OBJECTIVE: To prepare human acellular amniotic membrane (HAAM) and to measure its cytocompatibility and biocompatibility. METHODS: HAAM were prepared by chemical detergent-enzymatic extraction. Fresh human amnion was crosslinked with glutaradehyde, shaken in 0.5% SDS for 24 hours, and then treated with 0.25% trypsin for 4 hours. The production were freeze-drying and sterilized using ethylene oxide. Human fibroblasts were isolated from embryo and expanded in vitro. The fibroblasts were seeded in HAAM. HAAM and specimen were stained with HE and Mallory, and observed grossly, under light microscopy and scanning electron microscopy. The HAAM were implanted in the back of SD rats. RESULTS: There were no residues of cells in the HAAM (HE, Mallory staining). One side of HAAM had reticular and porous structure, the other side had compact fibrous structure. Pore size was from 10 to 80 nm. The HAAM could be seeded with expanded fibroblasts in vitro, and fibroblasts had the potential of spread and proliferation. The SD rat in the implant test had no death, convulsions and other abnormal response. CONCLUSION: The detergent-enzymatic extraction process can remove cells and solvable components effectively and preserve the tissue matrix well and keep the reticular structure. The HAAM can be used as an ideal scaffold of biological membrane for tissue engineering.