Fig. 4: BMP2 expression and activation of TGFβ-BMP signaling on TO-derived epithelia.

a IF microscopy of BMP2 staining on bronchoscopic biopsies collected from non-TO control and TO cases. Epithelial compartments were visualized by E-Cadherin co-staining. Scale bar, 100 μm. b Representative IF imaging of BMP2 on TBBC-derived ALIs, with E-Cadherin outlining the structures. BMP2 expressing cells were highlighted by white arrows. Scale bar, 20 μm. c Detection of phosphorylated SMAD 1/5 via IF staining on bronchoscopic biopsies collected from control and TO cases. Epithelial compartments were visualized by E-Cadherin co-staining. Scale bar, 50 μm. a–c n = 3 biologically independent experiments. d TO-TBBCs were treated with LDN-193189, a selective BMP signaling inhibitor, at a range of concentrations as indicated during ALI differentiation, and structures of cross-sections were visualized by HE staining (n = 3 independent experiments). Scale bar, 20 μm. e Ace-Tub and p-smad1/5 were assessed on ALIs with or without LDN-193189 treatments via IF staining (n = 1 dataset to verify LDN inhibition efficacy). Scale bar, 20 μm. f TO-derived pedigrees were treated with Noggin (200 ng/ml) constantly for 13 days throughout the course of ALI (n = 2 pedigrees). Mucociliary differentiation efficiency was assessed via IF staining and representative top-view images were shown. Scale bar, 50 μm. g Gene expression profiles of ALIs with or without Noggin treatment were examined by RNA-Seq, results of gene set enrichment analysis were shown as indicated.