ABSTRACT
Objective:To investigate the effect of the transcriptional coactivator Mediator 1 (Med1) on mouse hair regeneration, and to explore potential mechanisms.Methods:Med1 flox/flox C57BL/6J mice were mated with K14-Cre mice, and the mice with epidermis-specific knockout of Med1 gene, namely K14-Cre-expressing Med1 flox/flox mice (knockout group) , were obtained by using the Cre-Loxp system, while Med1 flox/flox mice without K14-Cre expression served as control group. Mice in the two groups (3 mice in each group) were raised together for 8 weeks followed by dorsal hair removal. Hair regeneration was observed for 12 consecutive days after hair removal. After 12 days, all mice in the two groups were sacrificed, their depilated and non-depilated dorsal skin tissues were resected, and total RNA was extracted from the tissues. Real-time quantitative PCR was performed to determine the mRNA expression of hair keratin genes, vitamin D receptor/β-catenin pathway-related genes, and genes associated with maintenance of hair follicle stem cell proliferation and quiescence. Paraffin-embedded sections of depilated and non-depilated mouse skin tissues were prepared, and immunofluorescence staining was conducted to determine the number of stem cells in the hair follicle bulge. Two-independent-sample t test was used for comparisons between two groups. Results:From days 0 to 12 after depilation, hair regeneration was delayed in the depilated skin area in the knockout group compared with the control group. Real-time quantitative PCR showed significantly decreased mRNA relative expression levels of hair keratin genes Ha1 and Krt2-16, vitamin D receptor/β-catenin pathway-related genes S100a3, Dlx3 and Tubb3, and genes associated with maintenance of hair follicle stem cell proliferation and quiescence including Lhx2, Sox9 and Nfatc1 in the depilated skin tissues in the knockout group (22.09 ± 12.32, 2.07 ± 0.20, 0.02 ± 0.01, 12.36 ± 2.12, 1.75 ± 0.46, 0.39 ± 0.02, 4.42 ± 0.76, 0.44 ± 0.07, respectively) compared with the control group (70.53 ± 9.46, 7.76 ± 0.49, 0.05 ± 0.01, 26.16 ± 2.96, 2.60 ± 0.14, 0.71 ± 0.09, 11.93 ± 0.42, 0.75 ± 0.04, respectively; t = 5.40, 18.64, 3.89, 6.57, 3.04, 6.10, 15.03, 6.18, respectively, all P < 0.05) . Immunofluorescence staining showed that the number of CD34 +K15 + hair follicle stem cells in the hair follicle bulge in both depilated and non-depilated skin tissues was significantly lower in the knockout group than in the control group. Conclusion:Med1 gene knockout may down-regulate the expression of downstream genes of the vitamin D receptor/β-catenin pathway and genes associated with maintenance of hair follicle stem cell proliferation and quiescence (Sox9, Nfatc1 and Lhx2) , and reduce the number of hair follicle stem cells, leading to hair follicle differentiation disorder and hair regeneration delay.
ABSTRACT
@#To explore the role of autophagy in hair follicle cycle and whether 3-methyladenine(3-MA)could promote hair regeneration in C57BL/6 mice through inhibiting autophagy flux, hair regeneration model of C57BL/6 mice was induced on the dorsal skin by depilation, and 3-MA was intraperitoneally injected to investigate hair regrowth, meanwhile vehicle and rapamycin(RAPA)were used as the controls. Results showed that 3-MA could obviously promote hair regrowth in depilated C57BL/6 mice. Furtherly, haematoxylin-eosin staining, immunohistochemical staining, immunofluorescence and Western blot tests were used to investigate the autophagy signals and the cell proliferation. Results showed that the expression of Beclin1 and LC3B II/I ratio were significantly decreased. Expression of P62 and Ki67 were increased, as well as the CD34 and CD49f double-labeled hair follicle stem cells were obviously increased inside bulge areas in 3-MA group, while contrarily in RAPA group. These results affirmed 3-MA, an autophagy inhibitor, could promote hair regeneration in depilated C57BL/6 mice by facilitating the transformation of hair follicle from telogen to anagen. 3-MA and other analogous autophagy inhibitors probably have a potential usage in future therapy in human telogen effluvium diseases.