Figure 3: TNF-α is sufficient to induce HF TAT and is crucial for WIHN.

(a,b) Intracutaneous injection of TNF-α can induce the HF telogen–anagen transition at the injection site in 7-week (W)-old mice (refractory phase) and 9-week-old mice (competent phase) (a), and the number of TNF-α-induced anagen hair follicles in the two different group was quantified (b). Seven-week-old mice, n=6 for each group; 9-week-old mice, n=8 for each group. (c,d) Wounding to the skin induced more anagen HFs in 9-week-old mice than in 7-week-old mice (c), and the number of anagen HFs in the two different groups was quantified (d). n=7 for each age group. (e,f) WIH-A analysis in TNFR1^−/− mice, TNFR2^−/− mice and WT mice (e), and the number of anagen HFs in different groups was quantified (f). Wild-type (C57BL/6) mice, n =7; TNFR1^−/− mice, n=6; TNFR2^−/− mice, n=9. (g,h) Wound-induced anagen HFs in mice constitutively expressing TNF-α (Tg-TNF-α) were significantly increased compared with WT mice, and the number of anagen HFs in the two different groups was quantified (h). Control mice (WT), n=6; Tg-TNF-α mice, n=5. (i) Bioluminescent imaging of Tnf-Luc-eGFP mice at different days (D) after wounding showed changes in the TNF-α level in the wound. Areas with high TNF-α levels (green/red) shifted from the wound (W) periphery to the wound centre with the progression of wound healing. n=15 for Tnf-Luc-eGFP mice, and n=10 for wild-type (WT) mice. (j) IF analysis of sections from the PWD-14 wound showed the presence of F4/80⁺ macrophages in the wound, which largely co-localized with TNF-α. Scale bar, 50 μm. (k,l) WIHN analysis in WT, TNFA^−/− and Tg-TNF-α mice at PWD-30 (k), and the number of neogenic HFs in wounds was quantified (l). n=6 for both wild-type and TNFA^−/− mice; n=12 for Tg-TNF-α mice. Scale bars, 2 mm. Data are expressed as the mean±s.e.m. *P<0.05, **P<0.01, ***P<0.005, unpaired t-test, two-tailed.