Calcineurin Activation and Hair Development Figure 3. CnAß-FK mRNA and NFATc2 Localizations in rat skin. (A) Time-scale of the hair cycle in male Wister rats. Each phase was determined using criteria described previously [42]. The intensity of the gray shading indicates the rate of proliferation of follicular keratinocytes: white,proliferation activity,black. (B–F) Distribution of CnAß-FK mRNA. B and C, telogen stage. D, late phase of anagen stage. E and F, transition stage from catagen to telogen. (C and F) Sections were counter-stained using hematoxylin (blue staining). (G–J) Results of immunohistochemical analysis of NFATc2 (green) and NFATc1 (red) in hair follicles. Cell nuclei were visualized by Hoechst staining (blue in G–I and K). (G) NFATc2 expression in hair follicle at postnatal day 28. (H) NFATc2 and NFATc1 expression in bulge and SG cells at postnatal day 28. Arrows, NFATc1 expression in bulge cells. Arrowheads, nuclear localization of NFATc2 in SG cells. (I) NFATc2 expression at postnatal day 49. Arrowheads indicate localization of NFATc2 in nuclei of SG cells. (J and K) Higher magnification image of the boxed region in G. (J) NFATc2 expression. (K) Hoechst staining. (L) Hematoxylin-eosin (H.E.) staining of hair follicles at postnatal day 28. After immunofluorescence staining, H.E. staining was performed in the same section to reveal the histology of hair follicle. DP, dermal papillae; IRS, inner root sheath; ORS, outer root sheath; SG, sebaceous gland. Scale bars in B–F, 100 mm; in B, C, and G–K, 50 mm. doi:10.1371/journal.pone.0017685.g003 protein was not observed in proliferating keratinocytes in the proximal part of the hair shaft (arrows) at the first anagen stage (Fig. 5E). In HF at the telogen stage, cyclin G2 expression was weak in the hair germ region and was undetectable in bulge stem cells and in DP cells. In contrast, cyclin G2 was detected in SG cells where CnAß-FK mRNA was detected at the telogen stage PLoS ONE | www.plosone.org 7 March 2011 | Volume 6 | Issue 3 | e17685