2.10 Animal model studies
All animal care and experimental techniques for anesthetics, hair
regrowth experiments and assays were accomplished with the permission of
Animal Research Ethics Committee of Shenzhen University (permit number
AEWC-201412003). Animal studies are stated in amenability with the
ARRIVE guidelines and with the commendations made by the British Journal
of Pharmacology (Lilley et al., 2020) Specific pathogen-free C57BL/6
mice (6 weeks old) were purchased from Nanjing Biomedical Research
Institute of Nanjing University, (Nanjing, China) and acclimatized for 1
week. After anesthesia via an intraperitoneal injection of
2,2,2-tribromoethanol (Sigma-Aldrich, St. Louis, USA), the skin hairs on
the dorsal side of 7-week-old mice (whose hair follicles were in the
telogen phase) were clean-shaven by a hair clipper. The mice were
randomly separated into three groups(Lee et al., 2012).. For every
mouse, 200 μL of each drug was topically applied at an appropriate
concentration everyday (as described in figure legends) for 25 days. The
control group received vehicle alone, the ISX9 group received vehicle
containing 1% of ISX9, and the minoxidil group received 2% of
minoxidil. All reagents used for the hair growth test were dissolved in
a vehicle composed of 50% (v/v) ethanol, 30% water, and 20% propylene
glycol. The back skin of the mice was photographed at 0, 7, 14, 21, 25
days, and analyses was done regarding the weight of regrown hair in
different groups treated for 25 days. After killing mice, skin tissues
were gathered and saved at -80 °C.