Figure 1.
Wnt signaling in back skin compared to the ear.
(A) H&E staining of back skin, illustrating hair follicles. (B) Adjacent section from an Axin2LacZ/+ mouse, stained with X-Gal. (C) H&E staining of the epidermis and dermis, and (D) corresponding X-Gal staining. (E) H&E staining of the ear. (F) Adjacent Axin2LacZ/+ section stained with X-Gal. (G) higher magnification of a hair follicle in the ear. (H) X-Gal staining of the epidermis and dermis of the ear. Abbreviations: e, epidermis; df, dermal fibroblasts; HF, hair follicle; dp, dermal papilla; car, cartilage; mc, matrix cells; hs, hair shaft. Scale bars = 50 µm.
Figure 2.
Wnt signaling varies with the hair cycle.
(A) Hemotoxylin staining of 5-week-old ear skin. (B) Gomori trichrome staining of 5-week-old ear skin; anagen-stage hair follicles appear red. (C) X-Gal staining of adjacent tissue from a 5-week-old Axin2LacZ/+ mouse. (D) Hemotoxylin staining, (E) Gomori trichrome staining, and (F) X-Gal staining of ear skin from a 7-week-old Axin2LacZ/+ mouse. (G) Hemotoxylin staining, (H) Gomori trichrome staining, and (I) X-Gal staining of ear skin from a 9-week-old Axin2LacZ/+ mouse. (J) The number of Wnt-responsive dermal cells per ROI was quantified at during anagen, catagen, and telogen (N = 3; 3 sections per mouse, 3 mice per week). All mice were male. Abbreviations as in Fig. 1. Scale bars = 50 µm; asterisk denotes P<0.05.
Figure 3.
Wound healing and Wnt signaling varies with the hair cycle.
On post-wound day 1, H&E staining of injury sites created in (A) anagen or (B) telogen. Post-wound day 2, (C) H&E staining of anagen (D) versus telogen wounds. Dotted yellow line indicates the healing tongue. Post-wound day 3, (E) H&E staining of anagen (F) versus telogen wounds. Dotted yellow line indicates regenerating epithelium. Post-wound day 4, (G) H&E staining of anagen (H) versus telogen wounds. (I) Percentage of healed wounds expressed as a function of the hair cycle (N = 20 per phase). (J) X-Gal staining of a wound bed in anagen versus (K) telogen. Dotted yellow line indicates auricular cartilage. (L) Hair follicles adjacent to the wound edges, in anagen versus (M) telogen. All mice were male. Abbreviations: we, wound edge; re, regenerating epithelium; es, eschar. Scale bar = 50 µm; asterisk denotes P<0.05.
Figure 4.
Elevated Wnt signaling in Axin2LacZ/LacZ mice correlates with improved cutaneous repair.
(A) Wound on post-injury day 1, stained with Gomori trichrome in Axin2LacZ/+ and (B) Axin2LacZ/LacZ mice. (C) Wound edge in Axin2LacZ/+ and (D) Axin2LacZ/LacZ mice. (E) Wound edge stained X-Gal from Axin2LacZ/+ and (F) Axin2LacZ/LacZ mice. Arrows indicate X-Gal+ve dermal fibroblasts. (G) Wound on day 3, stained with Gomori in Axin2LacZ/+ and (H) Axin2LacZ/LacZ mice. (I) Wounds stained with TUNEL in Axin2LacZ/+ and (J) Axin2LacZ/LacZ mice. Dotted line indicates auricular cartilage. (K) X-Gal in Axin2LacZ/+ and (L) Axin2LacZ/LacZ mice. (M) Wounds on day 3, stained with X-Gal in Axin2LacZ/+ and (N) Axin2LacZ/LacZ mice. (O) An eschar covers the wound beds in Axin2LacZ/+ mice. (P) Regenerating epithelium covers the wound beds in Axin2LacZ/LacZ mice. (Q) X-Gal in Axin2LacZ/+ and (R) Axin2LacZ/LacZ mice. (S) BrdU in Axin2LacZ/+ and (T) Axin2LacZ/LacZ mice. (U) Ki67 immunostaining in Axin2LacZ/+ and (V) Axin2LacZ/LacZ mice. (W) Quantification of X-Gal+ve dermal fibroblasts, and proliferating cells in the wound beds of Axin2LacZ/+ and Axin2LacZ/LacZ mice. All mice were male. Scale bar = 50 µm; asterisk denotes P<0.05.
Figure 5.
L-Wnt3a stimulates cutaneous repair.
(A) Fluorescent signal in the wound bed on day 3 treated with DiI-labeled liposomes containing PBS or (B) Wnt3a. Dotted lines indicate boundary between the epithelium and dermis. (C) Fluorescent signal in hair follicles and epithelium adjacent to the wound bed, treated with DiI-labeled liposomes containing PBS or (D) Wnt3a. (E–H) X-Gal staining of adjacent tissue sections shown in A–D. (I) BrdU staining in the wound bed treated with L-PBS or (J) L-Wnt3a. Arrows indicate immunopositive cells. (K) TUNEL staining in the wound bed treated with L-PBS or (L) L-Wnt3a. Arrows indicate immunopositive cells. (M) Telogen wounds treated with L-PBS fail to heal. (N) Telogen wounds treated with L-Wnt3a heal. (O) Quantification of the percent of L-PBS versus L-Wnt3a treated wounds that healed (N = 10 per treatment). All mice were male. Scale bars = 50 µm; asterisk denotes P<0.05.