Total RNA was extracted from cells or tissues by TRIzol reagent according to the manufacturer’s instructions. According to the manufacturer’s protocol, mRNA was reversely transcribed to cDNA using PrimeScript RT Master Mix (TAKARA, Japan). Then the expression levels of mRNA in each sample was measured by TB Green Premix Ex Taq according to the supplier’s protocol in the StepOnePlus system (Applied Biosystems, United States). All tests were run for 40 cycles consisting of denaturation at 95°C for 30°s, annealing at 60°C for 30°s, and an extension at 73°C for 30°s. The relative expression of each gene was evaluated using the 2−ΔΔCt method, and Gapdh was used as the internal control. The primer pairs were used previously (Xiao et al., 2019 (link); Zhang J. et al., 2020 (link)) and are listed below: Tnf-α F: 5-CCT​GTA​GCC​CAC​GTC​GTA​G-3, R: 5-GGG​AGT​AGA​CAA​GGT​ACA​ACC​C-3; Il-6 F: 5-AGT​TGC​CTT​CTT​GGG​ACT​GA-3, R: 5-TCC​ACG​ATT​TCC​CAG​AGA​AC-3; Il-1β F: 5-GGG​CCT​CAA​AGG​AAA​GAA​TC-3; R: 5-TAC​CAG​TTG​GGG​AAC​TCT​GC-3; Gapdh F: 5-ATT​CAA​CGG​CAC​AGT​CAA​G-3, R: 5-CTT​CTG​GGT​GGC​AGT​GAT-3; Mcp-1 F: 5-ACT​GAA​GCC​AGC​TCT​CTC​TTC​CTC-3, R: 5-TTC​CTT​CTT​GGG​GTC​AGC​ACA​GAC-3.
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