This study aims to observe the effect of hepatocyte growth factor (HGF) on the growth of human hair follicles, and explore its mechanism of promoting hair growth.

Hair follicles were isolated from the normal scalp tissue discarded by 4 rhytidectomy patients in the Plastic Surgery Hospital of the Chinese Academy of Medical Sciences, and the isolated single hair follicles were cultured in vitro. Normal culture condition was used as the control group, and HGF with a concentration of 10 ng/ml was added to the culture medium as the experimental group. The growth length of hair follicles in different culture days was measured under a microscope. The effect of HGF on the growth cycle of hair follicles was evaluated by observing the morphology of hair matrix and dermal papilla of hair follicle. Hair follicle epithelial cells (HFECs) were identified by flow cytometry. The normal cultured HFECs were used as the control groups, while the HFECs treated with 10 ng/ml HGF for 48h as the experimental groups. HFECs were collected to extract RNA and transcriptome sequencing was applied to detect the effects of HGF on gene expression of HFECs. Real-time PCR was used to verify the sequencing results. All quantitative data were displayed as Mean ± SD deviation, the independent sample t test was applied for the comparison between the two groups and the difference was statistically significant when P<0.05.

With the extension of culture time in vitro, hair follicle growth tends to stop. HGF promoted the growth of cultured hair follicles, and the growth length (unit 0.1 mm) of hair follicles were statistically different between the experimental and control group at 7 d (16.700 ± 5.143 vs. 12.210 ± 4.191, t=2.353, P<0.05 ), 10 d (18.800 ± 4.917 vs. 13.710 ± 3.518,t=2.962, P<0.01) and 14 d (23.000 ± 7.196 vs. 14.000 ± 4.057,t=3.910, P<0.01). HFECs we cultured displayed cuboidal morphology and highly expressed epithelial cell surface marker CD49f. RNA-seq showed HFECs highly expressed epithelial cell marker genes including KRT14, KRT5, KRT6A, CDH1, SOX9 and CD49f, the FPKM of which were 6 012 ± 2 141, 4 072 ± 1 369, 3 896 ± 1 991, 95.06 ± 21.48, 101.30± 38.52, 162.00 ± 47.83 respectively. HFECs did not or hardly express mesenchymal markers as THY1, DPP4, CDH2 (N-cadherin), ACTA2, PDGFRA, COL1A1and COL3A1, the FPKM of which were 0.740 ± 0.825, 0.632 ± 0.765, 0.000 ± 0.034, 1.674 ± 1.235, 0.000 ± 0.014, 2.526 ± 3.531, 0.000 ± 0.015, respectively. GO analysis showed the differential expressed genes between HGF treated and normal cultured HFECs were enriched in cell cycle-related biological processes such as nuclear division and chromosome separation. Real-Time PCR further verified that the expression of CENPA、CDC20、UBE2C、CDK1、AURKB、NDC80 in HGF treated HFECs were significantly downregulated to 0.689 ± 0.053(t=10.17, P<0.001)、0.676 ± 0.121 (t=4.652, P<0.01)、0.761 ± 0.148(t=2.785, P<0.05)、0.599 ± 0.153(t=4.530, P<0.05)、0.706 ± 0.113(t=4.507, P<0.05)、0.579 ± 0.092 (t=7.931, P<0.01) of the control group, respectively.

HGF can effectively promote the growth of cultured human hair follicles in vitro, but downregulate the cell cycle-related genes of hair follicle epithelial cells.

Hepatocyte growth factor; Hair follicle; Epithelial cells