Poster: 203Generation of KRT7-CreERT2 ferrets for targeting pancreatic duct and acinar cells

X. Sun; Y. Yi; P. Wu; Y. Sun; F. Yuan; Y. Weng; X. Liu; Z. Yan; J. Engelhardt

doi:10.1016/j.jcf.2026.07.234

Life expectancy for people with cystic fibrosis (pwCF) has increased to ∼68 years. As survival improves, cystic fibrosis-related diabetes (CFRD) is emerging as a major complication and may ultimately contribute more to morbidity than CF lung disease. Therefore, a deeper understanding of CFRD pathogenesis is essential to identifying potential therapeutic targets. Using CF ferrets, we have elucidated the central role of CFTR-expressing ductal cells in pancreatic pathologies in CF. However, the in vivo lineage relationships of ductal cells and their responses to injury remain poorly understood in the ferret pancreas. To generate a fate map of pancreatic ductal cells, we created a KRT7-IRES-CreERT2 ferret because Keratin 7 (KRT7) is primarily expressed in centroacinar cells and ductal epithelia of the pancreas. We used the Easi-CRISPR method with a long single-stranded DNA donor fragment to generate knock-in ferrets. Correct targeting was confirmed by PCR and Southern blotting. KRT7-IRES-CreERT2 ferrets were crossed with ROSA26 mTmG reporter ferrets, with or without the CFTR-G551D mutation, to enable lineage tracing under physiological and cystic fibrosis conditions. Tamoxifen was administered by intraperitoneal injection, and tissues were analyzed by immunofluorescence, confocal microscopy, and RNA in situ hybridization to evaluate epithelial lineage labeling across organs, with a focus on the pancreas. We generated two founder lines with precise insertion of the CreERT2 cassette at the endogenous KRT7 locus. After tamoxifen induction, KRT7-derived EGFP⁺ cells were distributed throughout glandular and ductal epithelia across multiple organs. In the pancreas, KRT7 lineage-labeled cells were observed in centroacinar cells and at all levels of the ductal tree under homeostatic conditions. In CF ferrets, KRT7 labeling persisted in ductal cells within dilated terminal ducts and swollen intercalated and intralobar ducts, demonstrating that ductal cell fate can be tracked during early CF disease without loss of KRT7 lineage identity. These KRT7-derived ductal lineages exhibited significant structural remodeling, including dilation and expansion of SOX9⁺ intercalated ducts. Notably, although acinar cells were not lineage-labeled in the healthy pancreas, EGFP⁺/CELA3B⁺ cells emerged in the CF pancreas and in acinar-derived organoid models, indicating activation of KRT7 expression during acinar-to-ductal metaplasia. KRT7 lineage-labeled cells were also frequently associated with pancreatic islets, although direct labeling of β cells was rare in both wild-type and CF conditions. These studies establish the KRT7-IRES-CreERT2 ferret as a versatile model for investigating epithelial lineage dynamics, ductal plasticity, and disease-related remodeling in vivo. This model provides a foundation for studying KRT7⁺epithelial populations in both physiological settings and in ferret disease models that closely mimic human pathology, including cystic fibrosis and related pancreatic disorders.

Poster: 203Generation of KRT7-CreERT2 ferrets for targeting pancreatic duct and acinar cells

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