Structure And Cell Viability Of Lyophilized Amniotic Membrane Are Equivalent To Those Of Cryopreserved Viable Amnion
Kathryn E. Davis, PhD, Lawrence A. Lavery, DPM, MPH.
UT Southwestern, Dallas, TX, USA.
Background:Human amniotic membrane (AM) has been widely applied in the management of burns, dermatological defects and ocular surface reconstruction [1-5]. Data from use of cryopreserved AM and fluid allograft containing live cells, proteins, cytokines and growth factors healed in patients with foot and ankle wounds suggesting that, in addition to structural matrix and growth factors, retaining AM cell viability is an important indicator of healing efficacy [6-10]. The use of cryopreserved amniotic tissue presents a few challenges, namely the availability of cryo-storage at the site and added process of rinsing the cryopreservation solution from the tissue prior to use. The lyophilized AM appears as a shelf stable paper-like sheet. It can be rehydrated using a saline rinse or the fluid from the patient’s own wound. Methods:We evaluated a lyopreserved viable AM samples for cell viability and compared to the cryopreserved viable AM. For this study, 3 individual samples of cryopreserved and lyopreserved AM derived from 3 different donors were assessed for cell viability. Each section was stained with SYTO 24 green fluorescent dye staining viable cells and Ethidium Homodimer-1 red fluorescent dye staining dead cells. Stained samples were analyzed using fluorescent microscopy. Imaging was performed for 10 microscopic fields for each section at 5x and 10x magnification for viable and non-viable cells. Images of viable and non-viable cells were overlapped and blindly assessed using a semiquantitative scale: non-viable, <50% viable, ~50% viable, or >50% viable. Results:Both preservation techniques resulted in AM tissues that had an average viability of greater than 50% with no statistically significant difference in viability between the cryopreserved and lyopreserved samples. Conclusion:AM tissue allograft that is stable and can be stored in the office without freezing and thawing the product simplifies documentation of storage and makes the product more accessible and easier to apply. These data suggest that a lyopreserved viable AM would produce equivalent clinical outcomes as it was reported for the cryopreserved viable AM product.
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