Fig. 3

Blocking ENO1 reduced PBMC migration and immune cells recruitment to the alveolar space and lung interstitium in bleomycin-treated mice. After intratracheal injection of 3 mg/kg bleomycin (BLM) (day 0), mice were treated with ENO1 Ab HL217 (10 mg/kg) intravenously on a 6-day interval from day 1. (A) Pooled PBMCs of each group (n = 5) were collected on day 7 & 14 and subjected to migration assay. BALF (B) was collected from the groups of sham (n = 6), BLM + Vehicle (n = 10), and BLM + HL217 (n = 9) on day 4, which was then assessed by using flow cytometry for the number total cell, neutrophil (CD11c⁻/Ly6G⁺/Ly6B.2⁺ cells), or monocyte (CD11c⁻/Ly6G⁻/Ly6B.2⁺ cells). The perfused lungs (C, D) were collected from the groups of sham (n = 6), BLM + Vehicle (n = 6), and BLM + HL217 (n = 5) on day 7. The collected cell samples were subjected to flow cytometry analysis for tissue-resident alveolar macrophages (TR-AM) (CD45⁺CD11c⁺SigF⁺), neutrophils (CD45⁺CD11b⁺Gr-1⁺), eosinophils (CD45⁺CD11c⁻SigF⁺), constitutive monocytes/macrophages (Gr-1⁻MoMp) (CD45⁺CD11b⁺MHC-II⁻CD64⁺Gr-1⁻), classical MoMp (Gr-1⁺MoMp) (CD45⁺CD11b⁺MHC-II⁻CD64⁺Gr-1⁺), dendritic cells (DC) (CD45⁺CD11b⁺MHC-II⁺CD64⁻CD24⁺), monocyte-derived alveolar macrophage (Mo-AM) (CD45⁺CD11b⁺MHC-II⁺CD64⁺CD11c⁺), and interstitial macrophages (IM) (CD45⁺CD11b⁺MHC-II⁺CD64⁺CD11c⁻). *P < 0.05, **P < 0.01, ***P < 0.001. (A, C, D) Data were representative for two independent experiments. (B) Data were pooled from two independent experiments. Each data point was from one mouse except (A) was shown as fold change and each group contained three technical replicates of pooled blood from five mice