Researchers reveal new molecular mechanism for stimulating hairgrowth nature
mice were plated onto 3T3 fibroblast feeder layer cells, pre-treated with mitomycin C to induce cell cycle arrest. Cells were co-cultured at 37 °C in William’s E medium supplemented with calcium and antibiotic–antimycotic. Medium was replaced after 48 h, and the attachment rate was evaluated following an additional 12 h of culture. Attached cells were passaged upon confluence, which was achieved every 4–6 days. Calcium-supplemented culture medium was changed every 2–3 days.
For single-cell RNA-seq, cells were captured using the Fluidigm C1 chips as per the manufacturer’s protocol. A concentration of 200,000–350,000 cells per millilitre was used for chip loading. After cell capture, chips were examined visually under the microscope to determine the capture rate, and empty chambers or chambers with multiple cells were excluded from the analysis.
For both bulk and single-cell RNA-seq, reads were first aligned using STAR v.2.4.2a with parameters ‘--outFilterMismatchNmax 10 --outFilterMismatchNoverReadLmax 0.07 --outFilterMultimapNmax 10’ to the reference mouse genome . Gene expression levels were quantified using RSEM v.1.2.25 with expression values normalized into fragments per kilobase of transcript per million mapped reads .
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