Sealing and healing of fetal membranes

Der vorzeitige Blasensprung vor der 37 Schwangerschaftswoche, tritt in 1 % aller Schwangerschaften auf und ist die Ursache von 30-40 % aller Frühgeburten. Sowohl der vorzeitige Blasensprung als auch die Frühgeburt erhöhen das Risiko der mütterlichen Morbidität sowie der neugeborenen Morbidität und Mortalität. Das Ziel dieser Arbeit war es, eine Therapieoption für den vorzeitigen Blasensprung mittels einer durch Tissue-Engineering hergestellten Matrix zu entwickeln. Zu Beginn der Arbeit wurde eine reproduzierbare Methodik etabliert, welche die Extraktion und Expansion von Amnionzellen von Früh- und Termingeburten zulässt. In einem in vitro Wundheilungstest, wurde gezeigt, dass das Reparaturpotential von Amnionzellen sowie die Reaktion auf verschiedene, Zellteilung stimulierende Faktoren, vom Gestationsalter abhängig ist. Besonders Amnionmesenchymzellen von Frühgeburten zeigen im Vergleich zu Amnionmesenchymalzellen von Termingeburten ein zweifach gesteigertes Reparaturpotential. Die Proliferation von Amnionmesenchymzellen von Frühgeburten konnte mittels Platelet Derived Growth Factor und Tumor Necrosis Factor α verdoppelt werden. Zum Abschluss der Arbeit wurden 3D Fibrinmatrices als Gerüste für Amnionzellen von Frühgeburten verwendet. Es wurde gezeigt, dass Amnionzellen in 3D Fibrinmatrices ihre natürliche Morphologie und Lebensfähigkeit behalten. Ausserdem wurde Kollagen I als Grundgerüst für eine 3D Matrix untersucht. Ein wesentliches Problem in diesem Modell war die Schrumpfung und folglich die Grössenreduktion der Kollagen I Matrix. Im Gegensatz zur Kollagen I Zellmatrix ergab eine Fibrin basierte Zellmatrix während einer Woche keine Schrumpfung. Die Daten dieser Arbeit, lassen vermuten, dass fibrinbasierte Zellmatrix-Systeme bei der Transplantation von Amnionzellen nützlich sein könnten. Jedoch müssen diese Daten als vorläufig betrachtet werden. Die nächsten Untersuchungen zielen nun daraufhin, dies mittels Tissue-Engeneering hergestellten Amnionzelltransplatate unter komplizierteren, multifaktoriellen Bedingungen zu testen. Premature rupture of the membranes occurring before 37 weeks' gestation is referred to as preterm premature rupture of the membranes (PPROM). PPROM occurs in 1 % of all pregnancies and is associated with 30-40 % of preterm deliveries. Both PPROM and preterm birth carry a high risk of maternal morbidity and neonatal morbidity and mortality. This thesis focuses on a strategy to develop a potential option for the treatment of PPROM patients based on a tissue engineering approach. First, a reproducible methodology that permits to collect and expand human amnion epithelial and mesenchymal cells from preterm and term placenta was established. Furthermore, using an in vitro lesion repair assay, it was demonstrated that the repair potential of amnion cells as well as their reaction towards different proliferation stimulants depends on the gestational age. Especially, preterm amnion mesenchymal cells showed approximately two fold increased repair potential compared to their term counterparts. Moreover, an increase in the proliferation of preterm amnion mesenchymal cells was found after treatment with platelet derived growth factor and tumor necrosis factor α. Furthermore, 3D fibrin matrices were used as scaffold for preterm amnion cells. It was demonstrated that amnion cells acquired their natural morphologies and displayed viability in 3D fibrin based cell-matrix. The use of collagen I as a scaffold was also investigated. Limitations of this approach include contraction and consequent reduction in size of collagen I matrices. In contrast to collagen I cell- matrix, no decrease in size of fibrin cell-matrix was observed over the time course of one week.These initial observations demonstrates this amnion cell-matrix systems may be useful in amnion cell transplantation. However, these data are clearly preliminary and have to be confirmed by exposure of amnion cell-based tissue engineered grafts to more complex multifactorial in vivo conditions

The Young's modulus of fetal preterm and term amniotic membranes

OBJECTIVE: To examine the Young's modulus of the human amniotic membranes, as well as its relationship to gestational age. To determine whether cellular and material-related parameters affect this modulus. STUDY DESIGN: In a prospective study at the Obstetric outpatient clinic of the University Hospital Zurich Young's modulus, thickness and mesenchymal:epithelial cell ratio of amniotic membranes of preterm (N=23) and term (N=40) placentae were examined. Significance (P<0.05) was calculated with the Mann-Whitney two-sample rank sum test and Wilcoxon signed rank test, while correlations were made using the Spearman's correlation. RESULTS: The Young's modulus of preterm amniotic membranes was significantly higher than that of term membranes. It varied within the same amniotic membrane. The thickness of the amnion in both preterm and term membranes did not differ significantly. The thinner the preterm and term amniotic membranes, the higher the Young's modulus was. There was no relation to the mesenchymal:epithelial cell ratio in the amnion. CONCLUSIONS: Preterm amniotic membranes are stiffer than term amniotic membranes. Tentatively, we hypothesise that there may be a correlation between the extracellular matrix components and the elastic properties of the membrane

Injectable candidate sealants for fetal membrane repair: bonding and toxicity in vitro

OBJECTIVE: This study was undertaken to test injectable surgical sealants that are biocompatible with fetal membranes and that are to be used eventually for the closure of iatrogenic membrane defects. STUDY DESIGN: Dermabond (Ethicon Inc, Norderstedt, Germany), Histoacryl (B. Braun GmbH, Tuttlingen, Germany), and Tissucol (Baxter AG, Volketwil, Switzerland) fibrin glue, and 3 types of in situ forming poly(ethylene glycol)-based polymer hydrogels were tested for acute toxicity on direct contact with fetal membranes for 24 hours. For the determination of elution toxicity, extracts of sealants were incubated on amnion cell cultures for 72 hours. Bonding and toxicity was assessed through morphologic and/or biochemical analysis. RESULTS: Extracts of all adhesives were nontoxic for cultured cells. However, only Tissucol and 1 type of poly(ethylene glycol)-based hydrogel, which is a mussel-mimetic tissue adhesive, showed efficient, nondisruptive, nontoxic bonding to fetal membranes. Mussel-mimetic tissue adhesive that was applied over membrane defects that were created with a 3.5-mm trocar accomplished leak-proof closure that withstood membrane stretch in an in vitro model. CONCLUSION: A synthetic hydrogel-type tissue adhesive that merits further evaluation in vivo emerged as a potential sealing modality for iatrogenic membrane defects

Concise review: isolation and characterization of cells from human term placenta: outcome of the first international Workshop on Placenta Derived Stem Cells

Placental tissue draws great interest as a source of cells for regenerative medicine because of the phenotypic plasticity of many of the cell types isolated from this tissue. Furthermore, placenta, which is involved in maintaining fetal tolerance, contains cells that display immunomodulatory properties. These two features could prove useful for future cell therapy-based clinical applications. Placental tissue is readily available and easily procured without invasive procedures, and its use does not elicit ethical debate. Numerous reports describing stem cells from different parts of the placenta, using nearly as numerous isolation and characterization procedures, have been published. Considering the complexity of the placenta, an urgent need exists to define, as clearly as possible, the region of origin and methods of isolation of cells derived from this tissue. On March 23-24, 2007, the first international Workshop on Placenta Derived Stem Cells was held in Brescia, Italy. Most of the research published in this area focuses on mesenchymal stromal cells isolated from various parts of the placenta or epithelial cells isolated from amniotic membrane. The aim of this review is to summarize and provide the state of the art of research in this field, addressing aspects such as cell isolation protocols and characteristics of these cells, as well as providing preliminary indications of the possibilities for use of these cells in future clinical application