2.10. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR)

To assess the modulation of fibrotic genes on HSFs undergoing shikonin HA MN treatment, cells underwent the treatments mentioned above for 24 h and suspended and lysed with TRIzol. Following RNA isolation through ethanol precipitation, cDNA conversion was performed using the iScript Adv cDNA kit according to the manufacturer's protocol. Primer sequences for TGFβ1, FAP-α, and COL1A1 mRNA were obtained and verified from previous studies16^,17 (Supporting Information Table S1). CT values were obtained through qPCR with CFX Connect PCR System (Biorad) and 2^−ΔΔCT expression was utilized to calculate the change in fold expression.