Fig. 7: FFAR3 reprograms ILC2s to an anti-inflammatory effector state by increasing EGFR expression. | Nature Communications

Fig. 7: FFAR3 reprograms ILC2s to an anti-inflammatory effector state by increasing EGFR expression.

From: Genetic diversity of Collaborative Cross mice implicates FFAR3 as a target for ILC2 anti-inflammatory reprogramming

Fig. 7

A Schematic for vehicle- vs. AR420626 (10 µM)–treated ILC2s by bulk RNA-sequencing. B Volcano plot of differentially expressed genes from (A). Egfr encircled. Labels = 10 most positively and negatively differentially expressed genes by log2 fold change (p-value threshold = 5 × 10⁻⁴; fold change threshold = 4; n = 5 paired biological replicates, representative of one experiment for (A–C). C Normalized EGFR counts from RNA-seq data in vehicle vs agonist-treated cells. Significance determined for (B, C) by DESeq2 DGE analysis (Wald test (two-tailed) with Benjamini–Hochberg correction); ***p < 2×10−52. (Bar height = mean, error bars = SD). D Representative flow plots of EGFR expression on ILC2s with and without FFAR3 agonist. Secondary-only control: no α-EGFR primary antibody. E EGFR expression by median fluorescence intensity (MFI) (n = 5 paired biological replicates; representative of one experiment) **** p < 0.0001. F Total ILC2s after 6-day culture with DMSO AR420626 (10 µM) and gefitinib (5 µM), groups = A–C left to right. (A-B *** p = 0.0005) (B-C ** p = 0.0018). G IL-10 levels in the supernatant; two vehicle (AR420626-/gefitinib-) samples were below detection. (A-B **** p < 0.0001) (A–C ** p = 0.0070) (B-C ** p = 0.0097). (F, G): n = 5 paired biological replicates; representative of two independent experiments). H Representative intracellular cytokine staining of IL-5 (gefitinib = 10 µM). I Quantification of IL-5 MFI (n = 4 for Vehicle and AR420626 (1 sample each used as FMOs); n = 5 for AR420626 + gefitinib). IL-5 quantification for vehicle and AR420626 is shown in Fig. 5E (A-B *** p = 0.0007) (A–C *** p = 0.0002) (B-C **** p < 0.0001). J Amphiregulin in the supernatant of ILC2s with and without 10 µM AR420626 measured by ELISA (n = 5 paired biological replicates; representative of one experiment) ** p = 0.0012. K IL-10 in supernatant of ILC2s cultured with IL-33, TSLP, IL-2 + AR420626 + α-Amphiregulin antibody or isotype control (n = 5 paired biological replicates; representative of one experiment) * p = 0.033. C, E–G, I, J bar height = mean, error bars = SD). Significance for (E), (J), and (K) was assessed by two-tailed paired Student’s t-test; for (F), (G), and (I) by two-way ANOVA with Tukey’s multiple comparisons test. ns not significant.

Back to article page