Fig 1: TL1A deficiency (KO and CKO) decreased the levels of SDC-1 and Tjp-3 and increased the expression and activity of Ctse. A Volcano plots for screening DEPs with thresholds of p < 0.05 and fold change > 1.5. B Gene heatmaps show upregulated and downregulated genes. C The top 20 significantly enriched function after TL1A KO. D The top 10 disease associations enriched after TL1A KO. E Changes in Ctse, SDC-1, and Tjp-3 were verified by Western blot analysis. F Ctse activity in lungs was observed by ELISA. G Protein intensity (Ctse, SDC-1, and Tjp-3) analysis of LPS-stimulated mice. Immunofluorescence staining of H Ctse, I SDC-1, and J Tjp-3 in primary alveolar epithelial cells from mice. Ctse (green), SDC-1 (red), and Tjp-3 (red). Scale bar, 50 µm. K Fluorescence intensity analysis of the primary alveolar epithelial cells from mice. L Changes in TL1A and Ctse in hPAEPICs as determined by Western blot analysis. M hPAEPIC TL1A/GAPDH ratio. N hPAEPIC Ctse/GAPDH ratio. O Ctse activity in hPAEPICs was observed by ELISA. Effect of TL1A OE on LPS-induced P SDC-1 and Q Tjp-3 as demonstrated by immunofluorescence. SDC-1 (red) and Tjp-3 (green). Scale bars, 50 µm. R SDC-1 fluorescence intensity analysis of hPAEPICs. S Tjp-3 fluorescence intensity analysis of hPAEPICs. T FITC-dextran in the Transwell assay was used to assess to cell permeability. Data are expressed as the means ± SD of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. NS not significant
Fig 2: DR3 deficiency decreased the levels of SDC-1 and Tjp-3 by increasing the expression of Ctse. A Changes in Ctse, SDC-1, and Tjp-3 were detected by Western blot analysis. B Mouse Ctse /GAPDH ratio. C Mouse SDC-1/GAPDH ratio. D Mouse Tjp-3/GAPDH ratio. E Immunofluorescence staining for the detection of Ctse in DR3 CKO lung sections. Ctse (red), SP-C (green), and DAPI (blue). Scale bar, 50 µm. F Immunofluorescence staining for the detection of SDC-1 in DR3 CKO lung sections. SDC-1 (red), SP-C (green), and DAPI (blue). Scale bar, 50 µm. G Immunofluorescence staining for the detection of Tjp-3 in DR3 CKO lung sections. Tjp-3 (green), SP-C (red), and DAPI (blue). Scale bar, 50 µm. H Ctse, I SDC-1, and J Tjp-3 fluorescence intensity analysis of lungs. K Effects of DR3 OE in hPAEPICs on LPS-induced Ctse were detected by Western blot analysis. L DR3/GAPDH ratio in hPAEPICs. M Ctse/GAPDH ratio in hPAEPICs. Effects of DR3 OE in hPAEPICs on LPS-induced N SDC-1 and O Tjp-3 observed by immunofluorescence. SDC-1 (red), Tjp-3 (green). Scale bars, 50 µm. P SDC-1 and Q Tjp-3 fluorescence intensity analysis of hPAEPICs. R The effects of DR3 OE in hPAEPICs on LPS-induced SDC-1 and Tjp-3 were detected by Western blot analysis. S SDC-1/GAPDH ratio in hPAEPICs. T Tjp-3/GAPDH ratio in hPAEPICs. U FITC-dextran in the Transwell assay was used to assess the cell permeability. Data are expressed as the means ± SD of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. NS not significant
Fig 3: Ctse OE in the mouse alveolar epithelium decreased the levels of SDC-1 and Tjp-3 in LPS-induced ARDS and active human recombinant Ctse protein decreased SDC-1 and Tjp-3 in hPAEPICs. A AT-II-overexpressing cells in mouse lungs were constructed. B Changes of Ctse, SDC-1, and Tjp-3 were detected by Western blot analysis. C Pathological damage was observed by HE. Scale bar, 50 µm. D Glycocalyx microstructures observed by electron microscopy. Scale bars, 1 µm. E Tight junction microstructures observed by electron microscopy. Scale bars, 500 nm. F, G, and H Protein intensity analysis of Ctse, SDC-1, and Tjp-3. I Intensity of FITC-dextran in BALF. Effects of active human recombinant proteins Ctse, MMP-9, and HPA on hPAEPICs. J SDC-1 and K Tjp-3 were detected by immunofluorescence. SDC-1 (red) and Tjp-3 (green). Scale bars, 50 µm. L, M SDC-1 and Tjp-3 fluorescence intensity analysis. N FITC-dextran in the Transwell assay was used to assess the cell permeability. Data are expressed as the means ± SD of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. NS not significant
Fig 4: Scheme showing that a deficiency in the TL1A/DR3 axis exerts negative effects on SDC-1 and Tjp-3 by increasing Ctse signaling in the alveolar epithelium
Supplier Page from Abcam for Cathepsin E Activity Assay Kit (Fluorometric)