Manufacture of a Cell-free Amnion Matrix Scaffold that Supports Amnion Cell Outgrowth In Vitro
Introduction
The human fetal membranes surrounding the amniotic cavity are composed of the amnion and the chorion [1]. The amnion is the interior part consisting of a single layer of epithelial cells (human amnion epithelial cells, HAECs) on a thicker basement membrane and collagen spongy layer containing mesenchymal cells (human amnion mesenchymal cells, HAMCs). It retains the amniotic fluid, secretes mediators such as prostaglandins, cytokines and protein hormones both into the amniotic fluid and towards the uterus, and guards the fetus against infections ascending the genital tract and from mechanical injury [2].
Spontaneous preterm premature rupture of the amniotic membranes (sPPROM), at less than 37 completed gestational weeks, occurs in 1–2% of all pregnancies and accounts for 30–40% of all preterm deliveries [2], [3].
The etiology of sPPROM is multifactorial [2], [4] and includes, for example, mechanical stress, infection or cervical incompetence [2], [4]. In addition, we are confronted with cases of iatrogenic PPROM (iPPROM), a potential complication and limiting step of invasive prenatal procedures. The incidence is 1.2% after genetic amniocentesis [5], 3–5% after diagnostic fetoscopy [6], >30% after fetoscopic cord ligation [7] and >62% after endoscopic tracheal clipping [8]. Although there is a classification in spontaneous and iatrogenic PPROM, the overall perinatal mortality of previable PPROM managed expectantly is reported to be between 55% and 66% [9], [10]. Actual management differs between immediate delivery or termination of pregnancy and conservative management with or without the use of tocolytics, antibiotics, and steroids in various combinations [3].
Current research is focused on finding methods to anatomically and functionally restore ruptured membranes. Approaches include the amniopatch [11], [12], [13], amniograft [14], intracervical fibrin tissue sealants [15], [16], gelatin sponge plugs [17], [18], collagen plugs [19], [20] and human amnion pieces [20]. Although these attempts possess apparent potential, they are small case-based studies and it is difficult to apply these generally. The search for alternative methods, such as in this study, is mandatory.
The aim of the present study was to explore the utility of both cellular as well as manufactured extracellular components of human amnion for reconstituting defective amnion in PPROM patients, using the methods of tissue engineering (Fig. 1).
Section snippets
Material and methods
Table 1 shows an overview of the different study groups and the used material.
Manufacture cell-free scaffolds from human amnion
Methodology combining enzymatic treatment and mechanical scraping, detailed in Section 2, allowed a reproducible, rapid and cost-efficient way to manufacture cell-free scaffolds from native human amnion collected between 31 and 40 weeks of gestation. The failure rate was very low. On 14 membranes tested, just one single amnion piece could not be worked up into a scaffold because of incomplete removal of the epithelium as detected by light optical microscopy.
Our stratification experiments showed
Discussion
This study of acellular amnion scaffolds for amnion cell outgrowth is part of a broader effort by our laboratory towards development of biomaterial implants for application in PPROM. Attempts to achieve lasting closure of fetal membrane lesions are likely to benefit best from biomaterial plugs that permit not only instant sealing of the membrane leak but also subsequent anatomic repair of the lesion through amnion cells recruited from adjacent tissue. The acellular amnion scaffold presented in
Acknowledgments
We thank Prisca Schär-Zammaretti from the Institute for Biomedical Engineering, ETH & University of Zurich for help and support with CLSM analyses.
This study was supported by the EMDO Stiftung Zurich.
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