Acellular Normal and Fibrotic Human Lung Matrices as a Culture System for <i>In Vitro</i> Investigation

TLDR

Extracellular matrix (ECM) regulates cell phenotype and is essential for organ integrity, yet conventional in vitro cultures fail to replicate the ECM’s composition, architecture, or distensibility, especially in the lung where fibroblasts reside in ECM‑rich interstitial spaces. The study aims to develop an acellular human lung matrix culture system to investigate how disease‑specific ECM influences fibroblast phenotype. Decellularization of normal and fibrotic lungs was performed with detergents, salts, and DNase, producing uniform, dimensionally preserved matrices suitable for reseeding. The resulting matrices retained native stiffness, allowed conventional fibroblast assays, and revealed that fibrotic matrices drive TGF‑β–independent myofibroblast differentiation and exhibit distinct compositional profiles compared with normal matrices.

Abstract

Rationale: Extracellular matrix (ECM) is a dynamic tissue that contributes to organ integrity and function, and its regulation of cell phenotype is a major aspect of cell biology. However, standard in vitro culture approaches are of unclear physiologic relevance because they do not mimic the compositional, architectural, or distensible nature of a living organ. In the lung, fibroblasts exist in ECM-rich interstitial spaces and are key effectors of lung fibrogenesis.Objectives: To better address how ECM influences fibroblast phenotype in a disease-specific manner, we developed a culture system using acellular human normal and fibrotic lungs.Methods: Decellularization was achieved using treatment with detergents, salts, and DNase. The resultant matrices can be sectioned as uniform slices within which cells were cultured.Measurements and Main Results: We report that the decellularization process effectively removes cellular and nuclear material while retaining native dimensionality and stiffness of lung tissue. We demonstrate that lung fibroblasts reseeded into acellular lung matrices can be subsequently assayed using conventional protocols; in this manner we show that fibrotic matrices clearly promote transforming growth factor-β–independent myofibroblast differentiation compared with normal matrices. Furthermore, comprehensive analysis of acellular matrix ECM details significant compositional differences between normal and fibrotic lungs, paving the way for further study of novel hypotheses.Conclusions: This methodology is expected to allow investigation of important ECM-based hypotheses in human tissues and permits future scientific exploration in an organ- and disease-specific manner.