1. Global Gene Expression Analysis in PKCα−/− Mouse Skin Reveals Structural Changes in the Dermis and Defective Wound Granulation Tissue 
Nichola H. Cooper, Jeya P. Balachandra, Matthew J. Hardman 
Journal of Investigative Dermatology 
Volume 135, Issue 12, Pages (December 2015)
DOI: /jid
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Experimental microarray design. Tissue isolated from WT and KO skin (a, b) and wounds (24hours post wounding, d–g). Microarray analysis was carried out on and two data sets generated from the expression analysis; direct comparison of genes differentially regulated between WT and KO normal skin (c) and ANOVA/unsupervised cluster analysis across all experimental groups (h). ANOVA, analysis of variance; KO, Knock-out; NS, normal skin; W, wound; WT, wild type.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Dermal extracellular matrix (ECM) changes. Immunoblotting reveals reduced SLRP protein expression in the KO skin (a, b). Transmission electron micrographs depicting dermal collagen fibrils with an increased variance, diameter, and interfibrillar spacing (c, arrows) in the KO skin (c, d). Kolmogorov–Smirnov and F-tests performed on the data distribution suggest that the data are significantly different (P≤0.0001; d). Load force at the point of failure was lower in the KO skin compared with WT (e). Representative stress–strain curves for the WT and KO skin show reduced tensile strength in the KO skin loaded to failure at a constant rate of 20mmmin−1. Curves taken from samples with median failure load (f). Scale bar=100nm (c); n=2,300–2,600 fibrils taken from three mice (c); n=4–5 6-week-old female mice (e, f). *P≤0.05 (e), P≤ (d). KO, Knock-out; SLRP, small leucine-rich proteoglycan; WT, wild type.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Altered dermal architechture in Knock-out (KO) skin. Masson’s Trichrome staining and collagen 1 immunofluorescence reveal a comparable dermal architecture and ECM composition in young WT and KO skin (a–d) but highlight differences in the mature skin (h–k). In the KO skin (g, h), despite no differences in skin thickness (j), age results in a reduced dermal collagen content (blue staining, Masson’s Trichrome) and reduced collagen I expression compared with age-matched WT (h, i). In addition, the KO skin is more prone to the development of spontaneous wounds (e, f). Masson’s Trichome and collagen I immunofluorescence (red, Collagen I; blue, 4',6-diamidino-2-phenylindole) on the lesional KO skin (j, k) reveal a clear wound area and an absence of Collagen I expression (k, l). N=6 WT and KO; Scale bars=1cm (a, b), 500μm (c, d). WT, wild type.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Gene profiling reveals complex expression changes in Knock-out (KO) mice. K-means unsupervised clustering analysis reveals four key patterns of gene expression with associated eisen plots (a–d). Analysis revealed genes regulated by wounding alone (a, b), genotype alone (c), and by interaction between wound and genotype (d). Analysis using DAVID identified highly changed gene ontology groups and associated key genes (boxed text) for each cluster. Eisen plots, red, gene induction; green, gene repression. Fold-change>1.5, q≤0.05. Multiple testing correction Benjamini P-value≤0.05 (Gene ontology groups). KOS, knock-out skin; KOW, knock-out wound; WTS, wild-type skin; WTW, wild-type wound. Scale bar=1cm.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Decreased matrix production in Knock-out (KO) wounds. Masson’s Trichrome staining (a–d) of WT (a, c) and KO (b, d) wounds reveals a delay in the replenishment of collagen (blue staining) at both 3 and 7days post wounding in KO mice. Picrosirius Red staining of the extracellular matrix reveals reduced overall collagen (red/yellow) and reduced newly synthesized collagen (green) at 3 and 7days post wounding (e–h) in KO wounds compared with WT wounds. Quantification of collagen content (i–j). Scale bars=1μm (e, g); *P<0.05, Student's t-test based on total collagen content. WT, wild type.
Journal of Investigative Dermatology  , DOI: ( /jid )
Copyright © 2015 The Society for Investigative Dermatology, Inc Terms and Conditions