Journal: The British journal of dermatology
Article Title: Coexistence of Eph receptor B 1 and ephrin B 2 in port-wine stain endothelial progenitor cells contributes to clinicopathological vasculature dilatation
doi: 10.1111/bjd.15716
Figure Lengend Snippet: Port-wine stain (PWS) endothelial cells (ECs) showed dual arterial and venous identities of co-expression of Eph receptor B1 (EphB1) and ephrin B2 (EfnB2). (a, b) PWS ECs were EphB1+ and EfnB2+. (c, d) A normal dermal venule showed expression of EphB1 but not EfnB2. (e) PWS ECs were EphB4−. (f) Relative mRNA levels of EphB1 and EfnB2 in selected normal human dermal microvascular endothelial cell (hDMVEC) subtypes. (g–i) Forced co-expression of EphB1 and EfnB2 in normal hDMVECs showed a significant increase in (g) branch thickness, (h) branch point area and (i) tube circumference of the capillary tubes formed in vitro compared with controls. (j) PWS blood vessel-like phenotypes were observed in EphB1+/EfnB2+ but not in wild-type and EphB1+/EGFP+ control hDMVECs in an in vitro capillary tube formation assay at 12 h after cell plating. Positive stain is diaminobenzidine (DAB; brown). Scale bar = 100 μm. *P < 0·05, ** P < 0·01 vs. control. The arrows indicate blood vessels. (k) Left panel, detection of expression of EphB1, EphB4, β-actin and EfnB2 by Western blot (W.B.) in various hDMVEC subpopulations. hDMVEC, heterogeneous population prior to EfnB2–Fc selection; EphB1− hDMVEC, the remaining hDMVEC subpopulation after EfnB2–Fc selection; enhanced green fluorescent protein (EGFP) or EfnB2–His, overexpression of EGFP or EfnB2 in the EfnB2–Fc selected hDMVEC subpopulation. Right panel, an anti-EphB1 antibody was used to immunoprecipitate EphB1 from cell lysate and EfnB2 was detected from the immunoprecipitated protein complex using an anti-EfnB2 or anti-His antibody.
Article Snippet: Human dermal microvascular endothelial cells (hDMVECs) were purchased from Cell Applications (San Diego, CA, U.S.A.) and were cultured in EC Basal Medium with growth supplement (Cell Applications).
Techniques: Staining, Expressing, In Vitro, Control, Capillary Tube Formation Assay, Western Blot, Selection, Over Expression, Immunoprecipitation
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