Figure 6.

Scratch wound healing in cell culture of primary keratinocytes: c-Met–positive primary keratinocytes migrate preferentially into scratch wounds. Primary keratinocytes were isolated from newborn skin of control (A) and conditional c-Met mutant mice (B and C). After scratch wounding, cells were further cultured in the presence of HGF/SF or TGFα. Photos were taken 0, 24, 48, and 96 h after scratch wounding. Note that wounds in the cultures derived from conditional mutant mice did only close after 96 h in the presence of HGF/SF. (D) Proliferation of primary keratinocytes from control and conditional c-Met mutant mice 24 h after stimulation with HGF/SF, as assessed by phospho-histone 3 antibody staining (red). A dashed line marks the scratch edge. Counterstaining was performed with phalloidin (green). (E) Quantification of proliferation of primary keratinocytes at wound edges stimulated with HGF/SF in the experiments described in D. Error bars represent the SD. (F and G) Primary keratinocytes isolated from control and conditional c-Met mutant skin were scratch wounded and further cultured with HGF/SF. After 24, 48, and 96 h, cells were stained with anti–phospho-c-Met antibodies (green). Nuclei were visualized by YO-PRO staining (red). Note that many cells in the controls (F) showed a cobblestone pattern of phospho-c-Met staining at the plasma membrane. In populations derived from the skin of conditional mutant mice (G), cells with phospho-c-Met were initially rare (at 24 h), but after 96 h they are abundant in the scratched area (marked by arrows). The original edges of the scratch wounds are marked with a dashed line. Bars: (A–D) 100 μm; (F and G) 50 μm.