Quantitative real-time PCR
Total RNA was prepared from the specimens with a Tri Zol reagent
(Invitrogen). Complementary DNA was synthesized using Superscript
reverse transcriptase (Invitrogen) and oligo(dT) primers. For the
analysis of α-SMA, FAP and GAPDH, the reaction was performed using a
Step One Plus™ real-time PCR System (Applied Biosystems). Primer
sequences are presented as follows:
α-SMA (Forward primers 5’-ACT GCC TTG GTG TGT GAC AA-3’; Reverse primers
5’-TCC CAG TTG GTG ATG ATG CC-3’)
FAP (Forward primers 5’-AGA ACC ATG CTT TGG AGA TAC T-3’; Reverse
primers 5’-GGT GGA TCT CCT GGT CTT TGT-3’)
GAPDH (Forward primers 5’-AAT CCC ATC ACC ATC TTC-3’; Reverse primers
5’-AGG CTG TTG TCA TAC TTC-3’)