Real-time PCR

Total RNA was extracted from lung tissues using the RNAiso Plus reagent (Takara, Otsu, Japan). First-strand cDNA was synthesized using the reverse transcription kit (Vazyme Biotech, Nanjing, China), according to the manufacturer’s protocols. In the presence of fluorescent dye (SYBR Green I), the real-time PCR reaction was performed using quantitative PCR (Applied Biosystems, Thermo Fisher Scientific, CA, USA). TREM-1 relative mRNA expression was normalized using β-actin and studied using the 2^-ΔΔCt method. The primer sequences (Tsingke Biotechnology, Beijing, Jiangsu, China) are listed below (F – forward primer, R – reverse primer):

Rat TREM-1-F: GAACGACCCTGTTCTGCTCT; R: GTGGAGACACTCGTAGGATCTG

Rat β-actin-F: TGCTGTCCCTGTATGCCTCTG; R: TGATGTCACGCACGATTTCC