Fig. 6.

Under the overexpression of KGF, stimulation of progenitor cell proliferation was inhibited by SU5402, an inhibitor for tyrosine kinase of KGFR. a Schematic description of the method for administering an inhibitor for tyrosine kinase of KGFR (SU5402) in vivo and the injection of two thymidine analogues in KGF-transfected mouse ear skin. BrdU was injected intraperitoneally at 24 h prior to vector transfection and EdU was injected intraperitoneally at 2 h prior to each euthanization. Two millimolars SU5402 was administered via eardrops each day every 24 h from the day of vector transfection to the day before euthanization. b Immunohistochemical detection of p-ERK and double immunofluorescence detection of BrdU (red)/EdU (green) in sections of ear skin tissue treated with SU5402 or 2 % DMSO in PBS (vehicle) after KGF gene transfection (KGF, day 4) or vector-alone transfection (Empty, day 4). The ear tissue treated with SU5402 shows no staining with the anti-p-ERK antibody. In the section of KGF gene-transfected ear tissue treated with a vehicle, many p-ERK-positive cells were stained brown (black arrows). Large numbers of BrdU(+)EdU(+) cells (yellow arrows) were detected in the thickened epithelium of KGF gene-transfected ears treated with a vehicle but not detected in the SU5402-treated specimen. Red arrows: BrdU(+)EdU(−) cells. Scale bars: 20 μm. c Dots plot showing the LI of BrdU(−) EdU(+) positive cells or BrdU(+)EdU(+) positive cells of the ear epithelium treated with SU5402 or 2 % DMSO in PBS (vehicle) after vector-alone transfection (n = 3 for each) or KGF gene transfection (n = 3 for each). BrdU(−) EdU(+) LI: *p < 0.0001 as determined by a two-way ANOVA F(11, 24) = 13.23 with a Tukey’s post hoc test. BrdU(+)EdU(+) LI: *p < 0.0001 as determined by a two-way ANOVA F(11, 24) = 68.55 with a Tukey’s post hoc test