1. Slug/Snai2 Is a Downstream Mediator of Epidermal Growth Factor Receptor-Stimulated Reepithelialization 
Donna F. Kusewitt, Changsun Choi, Kimberly M. Newkirk, Pascale Leroy, Yafan Li, Miquella G. Chavez, Laurie G. Hudson 
Journal of Investigative Dermatology 
Volume 129, Issue 2, Pages (February 2009)
DOI: /jid
Copyright © 2009 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
EGFR activation promotes ex vivo reepithelialization. Explants from wild-type mice were cultured for 5 or 7 days as described in “Materials and Methods”. Explants were maintained in basal medium without EGF or in basal medium supplemented with EGF (1nM) with or without 5μM AG1478. Epithelial outgrowth was measured from digital images. Values represent the mean of 2–4 explants per mouse per treatment group from a total of four individual mice ± standard deviation. *P<0.05 EGF treatment compared to untreated control cultures.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2009 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Induction of Slug and Snail by growth factors. (a) SCC 12F cells were grown to confluence, rinsed with phosphate-buffered saline, and placed in serum-free medium containing 0.1% (w/v) bovine serum albumin for 48 hours before addition of the indicated concentrations of growth factors. RNA was collected after 2 hours and mRNA levels were measured as described in “Materials and Methods”. Values were normalized to GAPDH with the baseline level of Slug (filled bars) or Snail (open bars) in untreated cells defined as 1.0 (dotted line) and other values expressed as fold increase. Data shown represent the mean of three independent samples, each analyzed in three separate PCR reactions, ± standard deviation. *P<0.05 compared to Slug level in control group; #P<0.05 compared to Snail level in control group. (b) SCC12F cells were grown as in (a), treated with 1nM EGF or 1nM TGF-α for the indicated times (upper panels) or with the indicated concentrations of EGF or TGF-α for 24 hours (lower panels). Slug protein was detected by immunoblot analysis. Data shown are representative of three independent experiments. (c) Explants from wild-type mice were treated as indicated for 5 days (AG=5μM AG1478). Activity of the β-galactosidase–Slug fusion protein (blue stain) was detected by histochemistry.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2009 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
EGF-dependent epithelial outgrowth is dependent on Slug expression. Explants from sex- and age-matched wild-type (+/+) or homozygous Slug null (-/-) littermates were cultured in complete medium containing 5nM EGF (+) or complete medium containing 5μM AG1478 to inhibit EGF receptor activity (-). Epithelial outgrowth was measured as described in “Materials and Methods” and the legend to Figure 1. *P<0.05 comparing activated EGFR versus inactivated EGFR at the corresponding time point.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2009 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Inhibition of EGFR does not disrupt keratinocyte outgrowth in cells expressing exogenous Slug. SCC 12F cells were infected with either AdSlug or AdGFP as described in “Materials and Methods”. An in vitro wound was introduced into cultures treated with 5μM AG1478 or untreated, and outgrowth into the wounded area was monitored by phase contrast microscopy. As previously reported, ectopic Slug expression enhanced in vitro reepithelialization and cell spreading (Savagner et al., 2005), but note that the Slug-associated keratinocyte outgrowth (arrows) was evident even when EGFR activity was inhibited by AG1478.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2009 The Society for Investigative Dermatology, Inc Terms and Conditions