The study was aimed to investigate the optimum conditions of freeze drying preservation of amniotic membrane (AM). The AM from the health puerperal woman was preserved by freeze drying at optimized way. The key factors of freeze drying process, including abstersion aqua, conservation liquor, the curve of freezing temperature, and the ingredient of protective agent, were optimized. All their morphologic structure was observed by light microscope and scanning electron microscope. The degradation rates by collagenase IV and the characterization of biomechanics were analyzed. The radio-immunologic method was used to investigate the cytokines quantity. All properties of freeze dried AM were compared with those of fresh AM. Light micrographs showed that the five structure-layers exist both in the fresh AM and in those preserved by freeze drying, while the fibro-material was tight-structured in the fresh AM, but loose slightly; the thickness of fibro-material was larger slightly in freeze dried AM. Scanning electron micrographs show that the micro-hairs of epithelial cells in fresh AM were decreased slightly in optimized drying AM, the collagen fibre of fresh AM and of optimized drying AM were well in morphological structures and arranged tightly. The degradation rate by collagenase IV was faster in optimized drying AM,compared with that of the fresh AM. There were insignificant diversity in biomechanical characters (tensile strength, elongation at break and elastic modulus) of the optimized drying AM compared with fresh AM. The cytokines quantity in optimized drying AM decreased significantly compared with fresh AM. The improved freeze drying process has better advantage in keeping the morphological structure, preferable biomechanics and biological vitality of AM, compared with the early research.
Amnion ; metabolism ; ultrastructure ; Biomechanical Phenomena ; Collagenases ; metabolism ; Cytokines ; metabolism ; Female ; Freeze Drying ; methods ; Humans