태아피부 혈관 및 배양한 태아진피 미세혈관 내피세포에서 E-selectin 발현에 관한 연구

Abstract

Inflammation is the characteristics of scar formation which is abscent in fetal wound healing. The adhesion molecules such as selection groups are believed to have key roles for migration of inflammatory cells through the microvascular endothelial cells to the wound. The purpose of this study was to evaluate the expression of E-selection on the cultured human fetal and neonate dermal microvascular endothelial cells. The back skin of spontaneously delivered dead fetus (IUP 18-22wks) and circumcised prepuce skin of neonate were used. Human fetal dermal microvascular endothelial cells (HFDMEC) were isolated by extracting microvascular segment from 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-selection on cultured fetal and neonatal endothelial cells in response to IL-1αTNF-αINF-ｒwas examined by ELISA. And the expression of E-selection on fetal and neonatal dermal microvascular endothelial cells was examined by immunohistochemical study using monoclonal 3B7 anti E-selection antibody in cultured fetal and neonatal skin. The expression of E-selection on endothelial cells was not significantly different between fetal and neonatal endothelial cells. This expression was augmented 10 times more by IL-1αTNF-αINF-ｒ. Augmented endothelial E-selection expression by IL-1αTNF-αINF-ｒshowed 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-selection molecule of unstimulated fetal and neonatal tissue was not observed. However, on both fetal and neonatal tissue cultured for 4 hours after stimulation by 100u/ml of IL-1 and 100u/ml of TNF, expression of E-selection 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 500u/ml of IFN, expression of E-selection was observed in the microvessels of upper dermis. In conclusion, we could not find any differences between the fetal and neonate skin in the expression of E-selection on the endothelial cell spontaneously or stimulated by IL-1αTNF-αINF-ｒin vivo and vitro which means the expression of E-selection may not be an important mechanism of scarless wound healing in fetus.