# Quantification and characterization of lung fibrosis in ARDS patients using picrosirius red staining

## TL;DR

This study uses picrosirius red staining to quantify and characterize lung fibrosis in ARDS patients, revealing a new collagen fiber subtype linked to poor outcomes.

## Contribution

A novel method to distinguish collagen fiber subtypes in ARDS lung tissue using PSR staining and digital analysis.

## Key findings

- ARDS patients showed significantly increased total collagen deposition compared to controls.
- A new collagen fiber subtype, 'scattered' fibers, was identified and was strongly associated with impaired lung function and adverse outcomes.
- Scattered fibers were not linked to collagen type I or III differences but showed distinct polarized light characteristics.

## Abstract

Evolution toward diffuse lung fibrosis is common in patients with ARDS but remains poorly characterized. In particular, the quantification and characterization of collagen fibers in lung tissue from patients with ARDS has not been performed and remain technically challenging. This study aims to precisely quantify and characterize collagen deposition in lung tissue from patients with ARDS.

Lung samples from ARDS patients and controls were retrospectively analyzed to quantify lung tissue fibrosis using computer-assisted digital processing after picrosirius red (PSR) staining of whole tissue sections. Additional analyses were conducted using immunofluorescence (IF) in conjunction with PSR staining and using examination of PSR stained sections under polarized light (PL).

Compared to controls, patients with ARDS exhibited increased total collagen deposition (p = 0.0006). Morphological segmentation of the collagen fibers and subsequent classification based on their staining intensity identified a subtype of fibers, termed “scattered” fibers, whose deposition was significantly increased in lung tissue of patients with ARDS compared to controls (p < 0.0001). The extent of scattered fibers was independent of ARDS etiology, histology or duration, but was correlated with impaired lung mechanics, impeded gas exchange, and adverse clinical outcomes. The PSR-based distinction between scattered and compact collagen fibers did not seem to reflect differences in the relative amounts of type I and type III collagen. However, the two fiber types were associated with distinct characteristics observable under polarized light microscopy.

Lung fibrosis during ARDS is predominantly driven by the deposition of weakly stained, loose segments of collagen fibers, which is associated with impaired lung mechanics and poorer clinical outcomes. Future studies should define candidates for early anti-fibrotic therapies.