Precision-cut lung slices as a model to study Radiation-Induced genome instability and Pulmonary Fibrogenesis - 09/05/26

Idiopathic pulmonary fibrosis (IPF) is a severe and deadly lung disease with limited treatment options. Preliminary data show significantly higher somatic mutation rates in IPF patients compared to controls. We hypothesize that these somatic mutations may actively drive disease progression. We propose to develop a model recapitulating both genome instability and fibrogenesis using human and mice precision-cut lung slices (PCLS).

Fresh human lung tissue ( n = 2) from Marie Lannelongue hospital and lungs from C57Bl/6 mice ( n = 5) were utilized for the generation of PCLS with a thickness of 300 μm. A time-course and radiation dose-response study was conducted following exposure of PCLS to X-ray radiation (0–10 Gy, 0–10 days) four PCLS per condition. At the end of each experiment, PCLS were snap-frozen for RNA or protein extraction, single-nucleus sequencing, and genomic analysis. To validate our model, we assessed radiation-induced DNA damage by staining for γH ₂ AX, along with histological staining to evaluate fibrosis, and RT-qPCR to analyze fibrotic markers and cytokine expression.

In mice PCLS, γ-H ₂ AX staining demonstrated that DNA damage is both dose- and time-dependent, with the proportion of positive cells increasing with radiation dose and culture duration. At 10 Gy, positive cells were sixfold higher than at 0 Gy, and by Day 10 with 5 Gy, the proportion was tenfold higher compared to day 0 without radiation (panel E). Radiation exposure also upregulated DNA damage–associated genes, including Atr, Atm, Brca1, and the telomere maintenance gene Tert, with Atr expression increasing ninefold from day 5–0 Gy to day 5–10 Gy (panel F). Higher radiation doses and longer culture durations further elevated the expression of fibrosis-related genes, including Col1a1 and Tgfb1, with Tgfb1 increasing elevenfold from day 5–0 Gy to day 5–10 Gy (panel F), indicating early tissue remodeling. Preliminary data from human PCLS confirm this finding ( n = 2).

Together, these results demonstrate that radiation triggers dose- and time-dependent transcriptional responses involving both fibrogenic and DNA damage pathways Fig. 1 .