Background: PSC is a chronic inflammatory condition of the biliary epithelium characterized by concentric periductal fibrosis. As opposed to inflammatory bowel disease (IBD), a frequently overlapping trait, there are currently no drugs that definitively slow disease progression. This study aims to define cellular abundances and pathogenic cross-talk driving periductal fibrosis in PSC using single-cell analysis.

Methods: scRNA-seq was performed on tissue sections from 4 FFPE blocks that included tissue from 2 PSC patients undergoing liver transplant and 2 patients with secondary liver cancer undergoing resection (controls, tumor-free margin). scRNA-seq was performed using the 10X Genomics Next GEM platform. Cells were projected onto reference datasets within CellTypist. Ligand-receptor interactions were assessed with CellChat.

Results: We recovered 34,183 cells from the 4 samples. PSC samples were enriched for T cells, mast cells, and monocyte-derived macrophages and depleted for zone 1 and zone 2 hepatocytes compared to controls as determined by a two-sample proportion test. PSC samples were enriched for an “intestinal macrophage”-like subset defined by expression of CD209, IGF1, and TNFSF13, predicted to interact with memory B cells and plasma cells via TNFSF13B & TNFSF17 (p=2.3x10^-7). 64% of collagen-producing fibroblasts (CPFs) and 30% of ACTA2-expressing myofibroblasts (MFs) in PSC samples expressed the portal fibroblast (PF) marker elastin (ELN). PDGF-PDGFR signaling between portal endothelial cells and CPFs was determined to be active in PSC samples but not in controls (p=5.7x10^-7).

Conclusion: ELN expression in CFPs and MFs in PSC samples implicate PFs as key effectors of periductal fibrosis in PSC. Predicted cell-cell interactions will be validated in intact tissues of selected protein analytes, focusing on periportal regions. The increased abundance of intestinal-like macrophages implicates a shared pathophysiology with IBD, possibly involving aberrant myeloid-stromal cell differentiation and cross-talk.

Figure 1) Study Methodology. FFPE tissue blocks from PSC patients and controls were sectioned, digested, and used to perform single-cell transcriptomics. Notable findings included an increased proportion of “intestinal macrophage” in the PSC patient liver and portal fibroblast cell markers in activated stromal subsets.

Figure 2) Activated Stromal Clusters Express Portal Fibroblast (PF) Markers in the PSC Liver. Col1-hi-producing fibroblasts express the PF markers elastin (ELN) and fibulin-2 (FBLN2). A subset of myofibroblasts express the hepatic stellate cell (HSC) markers desmin (DES) and nestin (NES), suggesting myofibroblasts come from both HSCs and PFs in these patients.

Jake Herb

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