Journal: bioRxiv

Article Title: Local IFNγ signaling contributes to the regenerative decline of aged alveolar progenitor cells

doi: 10.64898/2026.04.07.716929

Figure Lengend Snippet: Immunoproteasome is required for aged AT2 cell regenerative decline and the establishment of IFN γ+ T cells in tertiary lymphoid structures. A) Schematic figure to show the culture of alveolar organoids from young or aged immunoproteasome knockout (IP KO) and control C57BL/6 (WT) mice in feeder-free media at 7 days of growth, including B) organoid forming efficiency (OFE) and (C) diameter. D) RTqPCR of the IFNγ-inducible MHC-I complex component B2m in isolated lung AT2 cells from young or aged immunoproteasome KO and WT mice. E) Representative immunofluorescence staining of Ki-67 to mark proliferating cells with AT2 cell marker SPC and AT1 cell marker Rage/Ager in corresponding feeder-free organoids from aged mice in A-C. F) The ratio of Ki-67+ proliferating cells in aged immunoproteasome knockout (IP KO) versus WT mice. G) Dysregulated pathways in aged immunoproteasome KO epithelium in comparison to WT based on DEG genes (padj < 0.05, LogFC <0), with bar size representing enrichment score. H) Flow cytometry quantifications of CD8+ and CD4+ (I) T cells as % of all CD45+ immune cells in aged immunoproteasome KO and WT control lungs. J) Zoomed-in multiplex fluorescent images of TLS in aged lungs of immunoproteasome KO and WT controls with K) corresponding absolute quantification of CD8+ T cell and IFNγ+ cell (L) densities. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: We used the following primary antibodies: α-SPC (1:500, AB3786, Millipore), α-RAGE/AGER (1:500, MAB1179-100, R&D System), α-Ki-67 (1:200, 550609, BD Bioscience).

Techniques: Knock-Out, Control, Isolation, Immunofluorescence, Staining, Marker, Comparison, Flow Cytometry, Multiplex Assay, Quantitative Proteomics