Abstract Rationale Pleuroparenchymal fibroelastosis (PPFE) is a progressive interstitial pneumonia whose molecular pathology remains poorly defined because surgical biopsy is often infeasible. Cancer-associated fibroblast (CAF) programs—especially myofibroblastic CAFs (myCAFs)—are well described in tumors, but their presence in non-malignant lung fibrosis is unclear. We hypothesized that PPFE harbors a CAF-like fibroblast state and that cartilage intermediate layer protein (CILP) marks—and functionally aligns with—this program. Methods We profiled explanted PPFE lungs from three recipients by single-cell RNA sequencing, sampling per patient both severely fibrotic and relatively mild regions across upper and lower lobes (12 samples). Cell lineages and states were annotated by canonical markers. Spatial validation used RNAscope for CILP with histochemistry on adjacent sections and image analysis. Primary fibroblasts from PPFE and normal lungs were assayed for CILP expression. Functional testing used lentiviral CILP overexpression in normal human lung fibroblasts (± TGF-β), followed by collagen-gel contraction; bulk RNA-seq and gene-set enrichment assessed pathways and CAF-related signatures. Mixed-effects models accounted for donor effects. Results In fibrotic regions, proportions of vascular endothelium, smooth muscle, and pericytes increased, whereas macrophages, monocytes, and type I alveolar epithelial cells decreased. Aberrant basaloid cells were also detected. After stromal-enrichment sorting, six stromal clusters were resolved. A peribronchial-lineage fibroblast state co-expressed LRRC15 and MMP11 with broader myCAF markers (e.g., COL10A1, COL11A1, ITGA11, CTHRC1), which we term myCAF-like fibroblasts (myCAF-LFs). myCAF-LFs were detectable across patients and relatively enriched in severe versus mild regions. CILP emerged among genes linked to myCAF-LF enrichment and severity. Spatially, CILP RNAscope localized to proteoglycan-rich, fibroblast-dense stroma with high concordance (r = 0.93, p = 8.1 × 10−4). Primary PPFE fibroblasts expressed higher CILP than normal (log2FC=2.60; p = 0.032). In functional assays, CILP overexpression increased gel contraction versus controls (p = 8.8 × 10−³); TGF-β had an independent effect (p = 4.66 × 10−4) without interaction. Hallmark E2F targets (NES=1.66) and G2M checkpoint (NES=1.71) were enriched (FDR0.05). A dividing-CAF signature was enriched (NES=1.18; FDR=0.04), while a myCAF-LF marker set trended positive (NES=1.16; FDR=0.16). Conclusions Explanted PPFE lungs show region-dependent shifts in cellular composition and harbor a reproducible myCAF-like fibroblast state enriched in severely fibrotic regions. CILP marks these loci and functionally augments fibroblast contractility, linking a CAF-like remodeling program to non-malignant fibrosis. These findings suggest that CILP-associated signatures and myCAF-LF biology could serve as mechanistic leads and potential clinical readouts in PPFE. This abstract is funded by: JSPS KAKENHI
Takumida et al. (Fri,) studied this question.