The Study of Expression of E-selectin of Fetal Skin Vessel and Cultured Fetal Dermal Microvascular Endothelial Cell

Rah, Dong Kyun; Yoo, Won Min; Park, Beyoung Yun; Lee, Kwang Hoon

Abstract

Inflammation is the characteristics of scar formation which is abscent in fetal wound healing. The adhesion molecules such as selectin groups are believed to have key roles for migration of inflammatory cells through the microvascular endothelial cells to the wound 77re purpose of this study was to evaluate the expression of E-selectin on the cultured human fetal and neonate dermal microvascular endothelial cells. The back skin of spontaneously delivered dead fetus (IUP 18-22 wks) and circumcised prepuce skin of neonate were used Human fetal dermal microvascular endothelial cells (HFDMEC) were isolated by extracting microvascular segment pom trypsin treated fetal and neonate skin tissue and isolated by sieving with nylon mesh and then by 35% Perocoll gradient centrifugation. Further purification was done with the Ulex europaeus I coated magnetic dynabead To confirm the fetal and neonatal endothelial cells, expression of factor VIII antigen on cell surface and uptake of acetylated low-density lipoprotein were checked Expression of E-selectin on cultured fetal and neonatal endothelial cells in response to IL-1alpha TNF-alpha INF-gamma was examined by ELISA. And the expression of E-selectin on fetal and neonatal dermal microvascular endothelial cells was examined by immunohistochemical study using monoclonal 3B7 anti E-selectin antibody in cultured fetal and neonatal skin. The expression of E-selectin on endothelial cells was not significantly digerent between fetal and neonatal endothelial cells. This expression was augmented 10 times more by IL-1alpha, TNF-alpha, INF-gamma. Augmented endothelial E-selectin expression by IL-1alpha, TNF-alpha, INF-gamma showed peak level 4 hours after stimulation and return to baseline level after 48 hrs. This time course was similar in both fetal and neonatal endothelial cells. Immunohistochemically, the expression of E-selectin molecule of unstimulated fetal and neonatal tissue was not observed However, on both fetal and neonatal tissue cultured for 4 hours after stimulation by 100 u/ml of IL-1 and 100 u/ml of TNF, expression of E-selectin molecule in microvasculature of upper dermis was observed and this expression persisted for up to 16 hours of culture. Also after culturing for 48hrs with 500 qlml of IFN, expression of E-selectin was observed in the microvessels of upper dermis. In conclusion, we could not find any digerences between the fetal and neonate skin in the expression of E-selectin on the endothelial cells spontaneously or stimulated by IL-1alpha, TNF-alpha or INF-gamma in vivo and vitro which means the expression of E-selectin vny not be an important mechanism of scarless wound healing in fetus.