Preterm premature rupture of membranes (PPROM) is the rupture of fetal membranes prior to 37 weeks gestation, and before the onset of labour. PPROM complicates 40% of preterm births, which can result in lifelong disabilities such as respiratory, cardiac and neurological disorders. The causes of PPROM are multifactorial and not well understood. Iatrogenic PPROM is a major complication after invasive fetal interventions and occurs in 6-45% of cases. The high prevalence of iatrogenic PPROM after fetal surgery, due to the absent healing capacity of fetal membranes, reduces the effectiveness of interventions to treat fetal abnormalities demonstrating a need to design therapies with clinical potential. The present study demonstrates that connexin 43 (Cx43) is verexpressed in amniotic membrane (AM) after fetoscopic surgery and artificial in vitro trauma. Cx43 was preferentially distributed in mesenchymal cells compared to epithelial cells, with significant expression in the fibroblast layer compared to the epithelial layer. Polarisation of mesenchymal cell nuclei and collagen fibres at the wound edge is also reported. To investigate mechanotransduction AM weakening mechanisms we used an ex-vivo bioreactor system to study the effect of cyclic tensile strain. Changes in matrix composition (collagen, elastin and GAG), and pro-inflammatory factors (MMPs and PGE2) after 24 hours were studied. Cyclic tensile strain significantly increased GAG synthesis and release of MMPs and PGE2, with an associated reduction of collagen and elastin content, compared to unstrained AM. Furthermore, we demonstrate the reversal of these biochemical changes induced by cyclic tensile strain after AM exposure to pharmacological agents that target the broad group of PI3-kinases and selectively inhibit AKT-1/2 activity. Interestingly, addition of Cx43 and COX-2 inhibiting agents also reversed the biochemical response after cyclic tensile strain. It is suggested that alterations in the ECM composition affects AM integrity and leads to fetal membrane weakening following cyclic tensile strain. Finally, a novel sealing approach based on peptide amphiphile self-assembling gels in the presence of amniotic fluid is developed. By using peptide amphiphiles we were able to seal fetal membrane defects in vitro. This innovative approach provides a new avenue for a tissue engineering approach to prevent PPROM. The results obtained in this study contributes to our understanding on: (1) AM wound healing and repair capacity, the (2) mechanotransduction mechanisms behind AM weakening, and (3) a novel tissue engineering approach to seal fetal membrane defects using self-assembly of peptide amphiphiles for iatrogenic PPROM prevention.

PhD Thesis

https://qmro.qmul.ac.uk/xmlui/handle/123456789/62643