RNA extraction and real-time PCR
The spleen was taken from the sacrificed mice for RNA extraction using TRIzol™ Reagent (ThermoFisher, 15596026). RNA was reverse transcribed into cDNA using the RevertAid First Strand cDNA Synthesis Kit (Thermo Fisher Scientific, K1622). The concentration and integrity of RNA and cDNA were measured using a NanoDrop (Thermo Fisher, 2000). The mRNA expression values of the genes were normalized to the mRNA expression ofactin . The results were calculated using the comparative cycle threshold (ΔΔCt) method. The primers used were as follows: IL-4(F): GGTCTCAACCCCCAGCTAGT; IL-4(R): GCCGATGATCTCTCTCAAGTGAT; IL-5(F): CTCTGTTGACAAGCAATGAGACG; IL-5(R): TCTTCAGTATGTCTAGCCCCTG; IL-6(F): TAGTCCTTCCTACCCCAATTTCC; IL-6(R): TTGGTCCTTAGCCACTCCTTC; IL-10(F): GCTCTTACTGACTGGCATGAG; IL-10(R): CGCAGCTCTAGGAGCATGTG; IL-13(F): CCTGGCTCTTGCTTGCCTT; IL-13(R): GGTCTTGTGTGATGTTGCTCA; IL-17(F): TTTAACTCCCTTGGCGCAAAA; IL-17(R): CTTTCCCTCCGCATTGACAC; ICAM-1(F): GTGATGCTCAGGTATCCATCCA; ICAM-1(R): CACAGTTCTCAAAGCACAGCG; IFN-γ(F): ATGAACGCTACACACTGCATC; IFN-γ(R): CCATCCTTTTGCCAGTTCCTC; CCL-11(F): GAATCACCAACAACAGATGCAC; CCL-11(R): ATCCTGGACCCACTTCTTCTT; TGF-β1(F): TCTGCATTGCACTTATGCTGA; TGF-β1(R): AAAGGGCGATCTAGTGATGGA; TGF-β2(F): CCGCTGCATATCGTCCTGTG; TGF-β1(R): AGTGGATGGATGGTCCTATTACA; IKK(F): ACAGCCAGGAGATGGTACG; IKK(R): CAGGGTGACTGAGTCGAGAC; IKB(F): GCGGATGCCGATGAATGGT; IKB(R): TGACGTAGCCAAAGACTAAGGG; JUN(F): CCTTCTACGACGATGCCCTC; JUN(R): GGTTCAAGGTCATGCTCTGTTT; JNK(F): CAGACTGTACCCTCAAGATCCT; JNK(R): ATGCACCCGACAGACCAGA; ERK(F): GGTTGTTCCCAAATGCTGACT; ERK(R): CAACTTCAATCCTCTTGTGAGGG; FOS(F): CGGGTTTCAACGCCGACTA; FOS(R): TTGGCACTAGAGACGGACAGA.