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Spotlight on Lung Fibrosis: Activation of JUN in fibroblasts promotes pro-fibrotic programme and modulates protective immunity.

Pulmonary fibrosis is a serious lung disease, in which scar tissue is built up spontaneously in the absence of an infectious or autoimmune disease. It has a very bad prognosis, with a 3 year survival rate at only 50% and there are no curative treatments other than lung transplantation. Despite the discovery of genetic factors such as telomerase mutations, in most cases, the exact cause remains unknown.

Fibroblasts are known to be at the core of the fibrotic response; however, there is still no comprehensive single-cell data about this heterogeneous population in pulmonary fibrosis with many fibroblast markers only identifying small subsets of fibroblasts. Similarly, monocytes and macrophages are thought to play a critical role in injury and repair due to their role in innate immunity and macrophage heterogeneity has also been implicated in fibrosis progression.

In this study, the authors used single cell mass cytometry and immunostaining to identify the specific immune mechanisms that promote fibrosis, and suggested a therapeutic approach that could be used alongside conventional anti-fibrotic drugs for pulmonary fibrosis.

Fibroblast frequency was 5‑fold higher in fibrotic lungs; 15% in normal lungs compared to 80% in fibrotic lungs. Principal component analysis showed that fibroblasts in fibrotic lungs were not only increased in percentage but also differed phenotypically from control lung fibroblasts.

Consistent with these results, an enrichment of a distinct fibrotic lung-specific fibroblasts was identified. This subpopulation expressed high levels of CD47 and podoplanin.

Immunofluorescent staining detected co-expression of CD47 with fibroblast-specific protein‑1 (FSP1), and PD‑L1 with smooth muscle actin (αSMA) in fibroblasts, suggesting that these cells upregulate either one of the two immune-checkpoint proteins in the fibrotic lung.

High-dimensional mass cytometry also demonstrated co-activation of phospho JUN and AKT in patients with pulmonary fibrosis.

The ratio of interstitial macrophages to alveolar macrophages was significantly different between fibrotic and normal lungs, pointing to a markedly different composition of macrophages during fibrosis. Furthermore, increased regulatory and exhausted T cells in human lungs, suggested an immunosuppressive microenvironment in lung fibrosis.

Assay for Transposase-Accessible Chromatin using sequencing (ATAC‑seq), in addition to doxycycline-inducible overexpression and CRISPR-editing loss-of-function experiments indicated that JUN acts as an enhancer selector that modulates the accessibility of DNA in fibroblasts, potentially involved in profibrotic TGFbeta and Stat3 signalling pathways.

Using the ‘single hit’ bleomycin-induced model of lung fibrosis, authors showed that activation of JUN in lung fibroblasts promotes profibrotic programmes and modulates protective immunity. By contrast, immune checkpoint treatment prevented lung fibrosis.

In the same animal model, authors found two distinct mechanisms by which blocking IL‑6, CD47 and PD‑L1 reversed fibrosis, suggesting that JUN signalling might be a druggable target in pulmonary fibrosis.

Pulmonary fibrosis is a progressive, typically fatal lung disease with few curative treatments and poor understanding of the mechanisms involved in it progression. In this study the authors identify several pathways that may promote progression of the disease. These pathways have been well characterised for other medical indications and therapeutics intervention may be possible. The data presented in this study are highly significant and could form the basis of novel drug development/combination therapies that may be critical in halting the progression of this disease.

Reference

Cui L, Chen SY, Lerbs T, Lee JW, Domizi P, Gordon S, Kim YH, Nolan G, Betancur P, Wernig G. Activation of JUN in fibroblasts promotes pro-fibrotic programme and modulates protective immunity. Nat Commun 11, 2795 (2020)
doi: 10.1038/s41467-020-16466-4

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Epistem is a GCLP-accredited laboratory specialising in providing biomarker, target discovery and personalised medicine information. We offer microarray, qPCR and NGS services (including the 10X genomics platform for single-cell sequencing) for gene expression, whole-genome, and epigenetic analysis in all species.

The Histology and Immunohistochemistry team at Epistem have a wealth of experience in developing IHC protocols for a range of biological targets. Our deep understanding of IHC methodology has allowed us to develop and troubleshoot hundreds of working protocols for both automated (Ventana Discovery Ultra) and manual applications. We offer a bespoke service which can be suited to your project. We can work readily with FFPE, frozen tissues, with chromogenic and fluorescent detection systems as well as RNAScope methodologies.

Dr. Fernando Magdaleno Verduzco has recently joined Epistem to lead our fibrosis development programme, which includes liver, lung, and kidney models. We would be happy to talk to you about any fibrosis-related enquiries you have.

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