Institutional review board approval
Clinical samples
In vitro fetal membrane organ culture and stimulation with CSE
Collection of CD-1 amniotic sacs
AEC in vitro culture
AMC in vitro culture
Quantitation of AEC and AMC from amnion membranes
Cell culture treatments
Overexpression of PGRMC2 in AMCs
Scratch assay and cell culture treatments
Transmission electron microscopy
Bright-field microscopy
Confocal microscopy
Immunohistochemistry
Trichrome staining for collagen
Protein extraction and immunoblotting
Quantitative RT-PCR
Immunocytochemical localization of intermediate filaments cytokeratin and vimentin
Crystal violet cell viability assay
TGF-β enzyme-linked immunosorbent assay
Statistics
The cell shape index was determined for AEC and AMC cultures by evaluating one frame from each N (total of three images) per treatment for cell circularity using ImageJ software. The shape index was calculated using the following formula: SI = 4π × area/perimeter2, which is an established method that was originally reported to determine vascular cell shape (64). A circle would have a shape index of 1; a straight line, an index of 0.