Rmil 10

A total of 48 mice were randomly divided into the control, AR, and IL-10 groups with 8 mice in each group, and 2 independent experiments were performed. OVA (Grade V, Sigma, St. Louis, MO) was used for allergen sensitization and challenge to establish the AR mouse model according to the method previously described [16 (link)]. Briefly, on days 0, 7, and 14, mice were immunized by intraperitoneal injection of 25 μg OVA and 1 mg aluminium hydroxide in 300 μl phosphate-buffered saline (PBS), and followed by daily intranasal challenge (from day 21 to 27) with 2.5% OVA in 40 μl PBS (20 μl for each nose). The control group received PBS sensitization and challenge instead of OVA. The IL-I0 group received 0.1 μg rmIL-10 (PeproTech, USA) administration simultaneously with OVA challenge continuously for 7 days [12 (link)].

To study the responsiveness of NK cells to IL-15, splenocytes were resuspended with RPMI 1640 medium and plated on 24-well plates. Then splenocytes were treated with rmIL-15 (Peprotech, 100 ng/mL) for 1, 2, 3, 5, 7, or 9 days, followed by flow cytometry. To investigate the influence of cytokines on METTL3 expression in NK cells, we treated splenocytes with rmIL-17F (25 ng/mL, Peprotech), rmIL-27 (25 ng/mL, Peprotech), rmIL-10 (25 ng/mL, Peprotech), rmIL-15 (25 ng/mL, Peprotech), rmTGF-β (25 ng/mL, R&D) and rmIL-18 (25 ng/mL, R&D) for 3 days. To study the effect of MC38 cells on METTL3 expression in NK cells, purified NK cells were cocultured with MC38 cells under the stimulation of rmIL-15 (10 ng/mL) with the presence or absence of SIS3 (20 μM, APExBIO, Houston, TX, USA) or GW788388 (20 μM, APExBIO) for 2 days. To observe the effects of SHP-2 on the NK cells, splenocytes were treated with rmIL-15 (50 ng/mL, Peprotech) with or without SHP-2 inhibitor SHP099 (4 μM, Selleck, Houston, TX, USA) for 3 days.