Unravelling specific mechanisms of wound healing and pulmonary fibrosis in human ex vivo lung tissue slices

Pulmonary fibrosis comprises a multitude of rapidly progressing interstitial lung diseases assumed to develop from failure in the process of wound healing. Although TGFβ was reported to play a central role in the pathogenesis, it seems evident that also pro-inflammatory mediators contribute to the initiation and progression of lung fibrosis. In this study, the effect of TGFβ and TNFα on the induction of profibrotic mediators was investigated in human ex vivo lung tissue slices (Precision-Cut Lung Slices; PCLS). PCLS were prepared from tumor-free lung tissue sections and were cultured in the presence of TGFβ, TNFα or combination (TGFβ/TNFα) for up to 48h. Stimulation with TGFβ, TNFα or TGFβ/TNFα lead to specific up- or downregulation of distinct mediators in PCLS as compared to control samples, highlighting the necessity of combining key cytokines for the efficient induction of a pro-fibrotic profile. Upon others, TNFα as well as TGFβ/TNFα stimulation of PCLS lead to specific upregulation of IL-1β, while single stimulation with TGFβ showed no effect. IP10 and IFNγ levels remain unchanged in the presence of TNFα, while TGFβ/TNFα, or TGFβ alone lead to down-modulation of the expression. Stimulation of PCLS with either TNFα or TGFβ showed 2- to 3-fold up-regulation of PAI1. The combination of TNFα and TGFβ lead to further amplification of PAI1 mRNA levels to up to 7-fold. Importantly, ex vivo treatment with Pirfenidone efficiently reverts the TGFβ and TNFα-mediated modulation. We describe here the induction of a specific pro-fibrotic biomarker pattern in PCLS by TGFβ and TNFα stimulation, enabling the investigation of fibrotic signaling pathways with high translational relevance.