Regulation of Th2/Th17 cell balance in asthma

Magnetic bead separation (130–049-201, Miltenyi Biotec, Germany) isolated mice splenic CD4+ T cells. The asthma model transfected BECs were pretreated with 10 nM of DHT [41 (link)], 1 nM of E2 [42 (link)], and 10 : 1 nM of DHT:E2 (see subsection 4.2.3) in severe asthma + DHT, severe asthma +E2, and severe asthma +DHT/E2 groups, respectively, for 24 h, cocultured with CD4+ T cells at a ratio of 10 : 1 (TCs: BECs) for 24 h in complete RPMI 1640 culture medium (Gibco, Australia) supplemented with 10% FBS, 1% penicillin and streptomycin, soluble anti-CD28 (1.0 μg/ml, eBioscience), soluble anti-cd3e (0.5 μg/ml, eBioscience), and IL-2 (20 ng/ml, eBioscience). To analyze T cell subsets, cells were collected after 24 h, and flow cytometry determined the concentration of IL-4 and IL-17A to obtain the ratio of Th2 to Th17 cells. BECs MBD2 was extracted by western blotting.