Table 1.
Gene names and primer sequence.
Table 2.
Reaction systems of quantitative real-time PCR.
Table 3.
Reaction conditions of quantitative real-time PCR.
Fig 1.
Expression of keratin 26 in skin was detected by in situ hybridization in experimental group (100×).
Fig 1A. Longisection of primary follicle in experimental group. Fig 1B. Longisection of secondary follicle in experimental group. Fig 1C. Transection of primary follicle in experimental group. Fig 1D. Transection of secondary follicle in experimental group.
Fig 2.
Expression of keratin 26 in skin was detected by in situ hybridization in control group (100×).
Fig 2A. Longisection of primary follicle in control group. Fig 2B. Longisection of secondary follicle in control group. Fig 2C. Transection of primary follicle in control group. Fig 2D. Transection of secondary follicle in control group.
Fig 3.
Expression of keratin 26 in skin was detected by immunohistochemistry in experimental group (100×).
Fig 3A. Longisection of primary follicle in experimental group. Fig 3B. Longisection of secondary follicle in experimental group. Fig 3C. Transection of primary follicle in experimental group. Fig 3D. Transection of secondary follicle in experimental group.
Fig 4.
Expression of keratin 26 in skin was detected by immunohistochemistry in control group (100×).
Fig 4A. Longisection of primary follicle in control group. Fig 4B. Longisection of secondary follicle in control group. Fig 4C. Transection of primary follicle in control group. Fig 4D. Transection of secondary follicle in control group.
Fig 5.
Expression quantity of keratin 26 was detected during anagen, catagen and telogen by quantitative real-time PCR.
Fig 5A. Relative keratin 26 expression quantity was 1.08 × greater in secondary follicles than in primary follicles in anagen (p > 0.05). Fig 5B. Relative keratin 26 expression quantity was 3.3 × greater in secondary follicles than in primary follicles in catagen (p < 0.01.). Note: * p < 0.05, ** p < 0.01. Fig 5C. Relative keratin 26 expression quantity was 0.017884 ×, 0.019129 ×, 0.19 × and 0. 64 × less in primary and secondary follicles in anagen and catagen than in skin during telogen (p < 0.01).
Fig 6.
Skin cells which were infected with lentivirus for 72 h were observed under bright field of the fluorescence microscope in the preliminary experiment.
Fig 6A. Skin cells which were added polybrene when MOI was 0. Fig 6B. Skin cells which were not added polybrene when MOI was 0. Fig 6C. Skin cells which were added polybrene when MOI was 1. Fig 6D. Skin cells which were not added polybrene when MOI was 1. Fig 6E. Skin cells which were added polybrene when MOI was 10. Fig 6F. Skin cells which were not added polybrene when MOI was 10. Fig 6G. Skin cells which were added polybrene when MOI was 30. Fig 6H. Skin cells which were not added polybrene when MOI was 30. Fig 6I. Skin cells which were added polybrene when MOI was 50. Fig 6J Skin cells which were not added polybrene when MOI was 50. Fig 6K. Skin cells which were added polybrene when MOI was 100. Fig 6L. Skin cells which were not added polybrene when MOI was 100.
Fig 7.
Expression quantity change of keratin 26 after Noggin expression interference and Noggin after keratin 26 overexpression was detected by quantitative real-time PCR.
Fig 7A. Expression quantity of keratin 26 was 0.17 × (p < 0.01), 0.33 × (p < 0.01) less in blank control group and experimental group than in negative control group. Fig 7B. Expression quantity of Noggin was 0.81 × (p > 0.05) less, 3.41 × (p < 0.01) greater in blank control group and experimental group than in negative control group.
Fig 8.
Expression quantity change of keratin 26 after MT treatment respectively was detected by quantitative real-time PCR.
Fig 8A. Expression quantity of keratin 26 was 2.31 × (p < 0.05), 13.21 × (p < 0.01) greater, 0.41 × (p < 0.05) less in the groups which cells were treated with 0.02 g/L, 0.2 g/L, 1 g/L MT than the group which cells were treated with 0 g/L MT for 24 h. Fig 8B. Expression quantity of keratin 26 was 1.67 × (p < 0.05), 1.52 × (p > 0.05) and 1.07 × (p> 0.05) greater in the groups which cells were treated with 0.02 g/L, 0.2 g/L, 1 g/L MT than the group which cells were treated with 0 g/L MT for 48 h. Fig 8C. Expression quantity of keratin 26 was 15.99 × (p < 0.01), 35.15 × (p < 0.01), 513.36 × (p < 0.01) greater in the groups which cells were treated with 0.02 g/L, 0.2 g/L, 1 g/L MT than the group which cells were treated with 0 g/L MT for 72 h.
Fig 9.
Expression quantity change of keratin 26 after FGF5 treatment respectively was detected by quantitative real-time PCR.
Fig 9A. Expression quantity of keratin 26 was 1.88 × (p < 0.05), 129.91 × (p < 0.01), 11.95 × (p < 0.01) greater in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L FGF5 than the group which cells were treated for 0 g/L FGF5 for 24 h. Fig 9B. Expression quantity of keratin 26 was 10.93 × (p < 0.01), 7.42 × (p < 0.01), 12.11 × (p < 0.01) greater in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L FGF5 than the group which cells were treated for 0 g/L FGF5 for 48 h. Fig 9C. Expression quantity of keratin 26 was 3.21 × (p < 0.01), 22.3 × (p < 0.01), 17.31 × (p < 0.01) greater in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L FGF5 than the group which cells were treated for 0 g/L FGF5 for 72 h.
Fig 10.
Expression quantity change of keratin 26 after IGF-I treatment was detected by quantitative real-time PCR.
Fig 10A. Expression quantity of keratin 26 was 0.00011595 × (p < 0.01), 0.0000083 × (p < 0.01), 0.00004491 × (p < 0.01) less in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L IGF-I than the group which cells were treated for 0 g/L IGF-I for 24 h. Fig 10B. Expression quantity of keratin 26 was 0.00005124 × (p < 0.01), 0.00009017 × (p < 0.01), 0.0001944 × (p < 0.01) less in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L IGF-I than the group which cells were treated for 0 g/L IGF-I for 48 h. Fig 10C. Expression quantity of keratin 26 was 0.00009369 × (p < 0.01), 0.00012564 × (p < 0.01), 0.00015526 × (p < 0.01) less in the groups which cells were treated with 10−6 g/L, 10−5 g/L, 10−4 g/L IGF-I than the group which cells were treated for 0 g/L IGF-I for 72 h.
Fig 11.
Expression quantity change of keratin 26 after 0.2 g/L MT and 10−5 g/L FGF5 treatment together for 24 h or 1 g/L MT and 10−5 g/L FGF5 treatment together for 72 h was detected by quantitative real-time PCR.
Fig 11A. Expression quantity of keratin 26 was 13.21 × (p < 0.01) greater in the groups which cells were treated with 0.2 g/L MT than the group which cells were treated with 0 g/L MT for 24 h, 129.91 × (p < 0.01) greater in the groups which cells were treated with 10−5 g/L FGF5 than the group which cells were treated for 0 g/L FGF5 for 24 h, 9.09 × (p < 0.01) more in the group which cells were treated with 0.2 g/L MT and 10−5 g/L FGF5 together than the group which cells were treated with 0 g/L FGF5 and 0 g/L MT for 24 h. Fig 11B. Expression quantity of keratin 26 was 513.36 × (p < 0.01) greater in the groups which cells were treated with 1 g/L MT than the group which cells were treated with 0 g/L MT for 72 h, 22.3 × (p < 0.01) greater in the groups which cells were treated with 10−5 g/L FGF5 than the group which cells were treated for 0 g/L FGF5 for 72 h, 9.74 × (p < 0.01) more in the group which cells were treated with 1 g/L MT and 10−5 g/L FGF5 together than the group which cells were treated with 0 g/L FGF5 and 0 g/L MT for 72 h.
Fig 12.
Expression quantity change of keratin 26 after 0.2 g/L MT and 10−5 g/L IGF-I treatment together for 24 h or 1 g/L MT and 10−6 g/L IGF-I treatment together for 72h was detected by quantitative real-time PCR.
Fig 12A. Expression quantity of keratin 26 was 13.21 × (p < 0.01) greater in the groups which cells were treated with 0.2 g/L MT than the group which cells were treated with 0 g/L MT for 24 h, 0.0000083 × (p < 0.01) less in the groups which cells were treated with 10−5 g/L IGF-I than the group which cells were treated for 0 g/L IGF-I for 24 h, 2.41 × (p < 0.01) greater in the group which cells were treated with 0.2 g/L MT and 10−5 g/L IGF-I together than the group which cells were treated with 0 g/L IGF-I and 0 g/L MT for 24 h. Fig 12B. Expression quantity of keratin 26 was 513.36 × (p < 0.01) greater in the groups which cells were treated with 1 g/L MT than the group which cells were treated with 0 g/L MT for 72 h, 0.00009369 × (p < 0.01) less in the groups which cells were treated with 10−6 g/L IGF-I than the group which cells were treated for 0 g/L IGF-I for 72 h, 3.92 × (p < 0.01) greater in the group which cells were treated with 1 g/L MT and 10−6 g/L IGF-I together than the group which cells were treated with 0 g/L IGF-I and 0 g/L MT for 72 h.
Fig 13.
Skin cells which were treated with 0.2 g/L MT, 10−5 g/L FGF5, or 10−6 g/L IGF-I respectively for 72 h were observed under bright field of the laser scanning confocal microscope.
Fig 13A. Skin cells which were treated without MT, FGF5 or IGF-I (25×). Fig 13B. Skin cells which were treated without MT, FGF5 or IGF-I (100×). Fig 13C. Skin cells which were treated with 0.2 g/L MT (25×). Fig 13D. Skin cells which were treated with 0.2 g/L MT (100×). Fig 13E. Skin cells which were treated with 10−5 g/L FGF5 (25×). Fig 13F. Skin cells which were treated with 10−5 g/L FGF5 (100×). Fig 13G. Skin cells which were treated with 10−6 g/L IGF-I (25×). Fig 13H. Skin cells which were treated with 10−6 g/L IGF-I (100×).
Table 4.
The results of gray analysis after treatment with MT, FGF5, IGF-I in immunofluorescence detection preliminary experiment.