Effectiveness of human amnion preserved long-term in glycerol as a temporary biological dressing
Introduction
Temporary biological closure of large burns or open wounds has generally proven to be a life-saving measure [1], [2]. Various materials, such as human skin allografts, pig skin xenografts and human amniotic membranes, have proved effective as temporary wound dressings. These materials have the beneficial effects of reducing loss of water, protein and evaporative heat at the wound surface, and also prevent further bacterial contamination. Fresh human cadaver skin allograft is presumed to be most effective at achieving these effects. However, legal situations concerning organ donation in some countries and the cost of procuring and preparing allografts and xenografts may preclude their extensive use in burn treatment. In contrast, the use of human amnion still remains a good, cost-effective means of treating burns in developing countries, nations whose high incidence of burn injuries is further complicated by financial constraints [3], [4].
Davis introduced the use of human fetal membranes in skin transplantation in 1910, and 3 years later Sabella described it for burned and ulcerated skin surfaces [5]. The object of these early attempts was to achieve durable wound coverage and, although the body ultimately rejected the membranes, lack of infections and alleviation of pain was noted. In 1952, Douglas reported the use of amniotic membranes to temporarily cover burn wounds [5]. Since then, this material has yielded good results as a temporary biological wound dressing and is mostly used to treat burns [3], [4], [5], [6], [7], [8]. Fresh amnion has generally been used, but frozen [6], dried [9], irradiated [9] and lyophilized [10] preparations have also been described.
In order to be practical for clinical use, amnion must be easily obtainable and be able to be conveniently stored long-term, free of contamination and without biodegrading. Studies have shown that amnion can be maintained in viable condition for up to 6 weeks if stored aseptically at 4°C in 0.5% silver nitrate solution [3], in 20% glycerin solution [6], or in sterile saline after passage through one rinse of 0.025% sodium hypochlorite solution [5]. Frozen preservation of amnion allows it to be stored at −70 to −90°C for over 6 months [4], but in this case the material must be thawed before use. Amnion retains its structural properties and, when these conservation processes are used, is comparable to fresh amnion in its effectiveness as a biological dressing.
Long-term preservation and storage of biological materials not only offers the advantage of good availability but also reduces the risk of disease transmission since there is time to check donors material for the presence of pathogens. Glycerol serves the basic mainstay for tissue preservation, glycerolization of skin tissue was first implemented by Basile in 1982 to preserve porcine skin for long periods at low cost [11]. Glycerol dehydrates tissue by physically replacing most of the intracellular water but does not change the cells' ionic concentration, thus, it is an efficient agent that preserves tissue by protecting cell integrity [11], [12].
This experiment was designed to determine whether human amnion that has undergone long-term storage in glycerol is an effective biological dressing compared to fresh amnion and glycerol-preserved human skin. We compared these biological dressings in terms of their performance on rat skin wounds and their ability to reduce bacterial counts in infected rat burn wounds. We also ran a small clinical trial to observe the action of glycerol-preserved amnion on split-thickness graft donor sites.
Section snippets
Materials and methods
Human amniotic membranes were procured at the time of delivery from mothers who had no history of premature membrane rupture, endometritis, or meconium ileus. The amnion was separated from the chorion and was cleansed of blood by flushing with copious amounts of tap water. The human skin samples were obtained from residual tissues from aesthetic surgical operations. Amnion and skin pieces were placed in individual sterile bottles containing 85% glycerol solution. They were kept at 37°C
Results
The qualitative bacterial culture studies of the preserved tissues demonstrated that there was no bacterial growth in any specimen after a storage period of more than a year. Gross examination of the preserved skin and amnion tissue showed no signs of lysis or dissolution. Following rehydration, comparison of glycerol-preserved and fresh amnion showed only minor gross qualitative differences (Fig. 1). The glycerol-preserved membrane was soft and pliable and differed from fresh amnion only in
Discussion
The advantages of amnion use as a biological dressing for burns have been well-documented in the literature [3], [4], [5], [6], [7], [8], [9]. Amnion eliminates pain, allows the wound to dry faster and promotes early epithelialization. The tissue is readily available, inexpensive, easily procured and stored, of low antigenicity and has antimicrobial potential, all of which make it an ideal dressing, especially in countries where economic factors preclude the use of other dressing options [3],
Acknowledgements
We wish to thank to Isil Maral, MD, from Gazi University, Department of Public Health, who performed the statistical tests in this study.
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