Fig 2: IL-17A Trans-Signaling through Cardiac Fibroblasts Downregulates MerTK expression on Monocytes and MDMs(A and B) Representative EM images showing (A) apoptotic cells or apoptotic cellular debris internalized by Ly6Chi MDMs (arrowheads) and (B) engulfed cellular debris were largely absent in Ly6Clo MDMs. Scale bars: 2 µm.(C) Macrophage phagocytic index was calculated using the following formula: (number of engulfed apoptotic cells/total number of macrophages) × (number of macrophages with engulfed apoptotic cells/total number of macrophages) × 100.(D–G) Hearts from day 21 EAM mice.(D) Frequencies of F4/80hiCD64+ macrophages out of viable CD45+Ly6G–CD11b+ cells were assessed by flow cytometry.(E and F) MerTK MFI of F4/80hiCD64+ macrophages (E) and F4/80–CD64+ monocytes (F) in the hearts.(G) ELISA showing soluble Mer (sMer) in WT and IL-17Ra-/- EAM mice sera.(H and I) Cardiac fibroblasts were harvested from WT naive mice, whereas monocytes were sorted from EAM IL-17Ra-/- mice.(H) MerTK MFI of Ly6Chi or Ly6Clo monocytes and MDMs in vitro after 160 h post-co-culture with cardiac fibroblasts stimulated with or without IL-17A.(I) sMer detected in supernatants of the monocyte-fibroblast co-culture by ELISA.(J) Flow cytometric analysis of the frequencies of FITC+F4/80hiCD64+ macrophages in the myocardium of WT, IL-17Ra-/-, and non-treated controls.(K) Percentages of FITC+F4/80hiCD64+ macrophages out of viable CD45+Ly6G–CD11b+ cells.(L) MerTK MFI in patients with either myocarditis or ischemic cardiomyopathy.(D–G) Data are representative of five independent experiments. n = 8 – 9. (H and I) Data are representative of three independent experiments with technical triplicates. n = 3. (J and K) Data are representative of two independent experiments. n = 3. (C–G, I, K, and L) Groups were compared using Student’s t test. *p < 0.05; **p < 0.01. (H) Groups were compared using one-way ANOVA followed by Dunnett test. **p < 0.01; ****p < 0.0001. All data were presented as mean ± SD.See also Figure S7 and Tables S2 and S3.

Fig 3: Neutrophils in Atg7 cKO lungs contribute to impaired antimicrobial responses, exaggerated pathological inflammation, and increased cell death during NTM infection.a Atg7 cWT and Atg7 cKO mice were infected with Mabc-R (2 × 106 CFU) for 10 days. Representative flow cytometry plots (left) show live CD45+MerTK-CD64- cells gated for Ly6G+ neutrophils in lung tissues. Quantification of Ly6G+ neutrophils is shown on the right (n = 4). b, c Atg7 cWT and Atg7 cKO mice were infected with Mabc-R (2 × 106 CFU) for 14 days. Representative 3D images of CitH3 (green), MPO (red), and DAPI (blue) in lung tissues were generated using Aivia (b). The percentage of Ly6G+ cells that were also CitH3+ (n = 21, independent biological replicates) (c). d Atg7 cWT and Atg7 cKO mice were infected with Mabc-R (2 × 106 CFU) or Mav (1 × 107 CFU) for 10 days. Representative TEM images of decondensed chromatin in lung tissues. e–h Atg7 cWT and Atg7 cKO mice were infected with Mabc-S (1 × 107 CFU) for 7 days. Mice were administered with anti-IgG2a or anti-Ly6G antibodies (Ab) via daily intraperitoneal injection until sacrifice. In vivo bacterial loads in lung tissues (e Atg7 cWT: n = 3; Atg7 cKO: n = 4). IL-6 (f IgG2a: n = 6, Ly6G: n = 8) and TNF (g n = 4) levels in lung tissues were measured by ELISA. Expression of GSDME was assessed by immunoblotting in lung tissue lysates. Dot plots show quantification of the GSDME-NT normalized to GSDME-FL, based on Western blot (h). Statistical significance was determined by a two-sided unpaired t-test with Mann–Whitney U-test (a, c), and two-tailed t-test (e–g). Data shown (means ± SEM) represent the combined results of two or three independent experiments (a, c, e, f, g). Images are representative of three independent experiments (b, d). dpi days post infection, CitH3 citrullinated histone H3, NTN-terminal, FL full length, ONM outer nuclear membrane, INM inner nuclear membrane. Source data are provided as a Source Data file.