Fig. 4

Viable ID8 cells enhance IL-1β production by Mettl3-deficient macrophages independent of inflammasome activation. (A) LPS-stimulated BMDMs were treated with viable ID8 cells or their lysate in vitro, and their supernatants were collected to measure IL-1β levels via ELISA. Elevated IL-1β level was measured in Mettl3-deficient BMDMs across all treatment groups compared with WT BMDMs. Moreover, viable ID8 cells treated with Mettl3-deficient BMDMs revealed increased IL-1β levels than those treated with ID8 cell lysate. (B) The transcriptional level of IL-1β mRNA in BMDMs treated with viable ID8 cells or ID8 cell lysate was analyzed by qRT-PCR. Treatment with ID8 cell lysate (4:1 but not 2:1) significantly elevated IL-1β mRNA transcription in Mettl3-deficient BMDMs than in WT BMDMs with or without LPS stimulation. ***p < 0.001. Only LPS stimulation and dead ID8 cells elicited significantly increased IL-1β mRNA transcription in Mettl3-deficient BMDMs than in WT BMDMs. ***p < 0.001. (C) Western blot analysis of inflammasome-related proteins. (D) Cell cytotoxicity was measured by an LDH release assay. No significant difference in cell death or cytotoxicity was observed between WT and Mettl3-deficient BMDMs without LPS stimulation. Data from one of six independent experiments are shown. Bars represent SD; *p < 0.05, **p < 0.01, ***p < 0.005