**1. Introduction**

In idiopathic pulmonary fibrosis (IPF) the biomechanics and composition of the extracellular matrix (ECM) are altered causing a pathological phenotype associated with increased tissue stiffness and disorganized structures of the lung [1]. Today there is no real effective treatment for IPF with poor long-term survival, although there are treatments that may slow progression of IPF somewhat. Lung transplantation remains the most valid option for some patients, however, not all patients can be offered this treatment due to recipient co-morbidities or donor shortage. Although largely unknown, a combination of factors is believed to play a role in IPF, including ageing, epigenetic modifications, dysfunctional alveolar epithelium, along with persistent activation of lung fibroblasts that contributes to an increased accumulation of ECM with subsequent detrimental remodeling resulting in the loss of lung function and eventually end-stage lung disease [2–4]. Local signals from the ECM, e.g., sti ffness and bound growth factors and cytokines, have been shown to influence cellular behavior such as migration, di fferentiation and proliferation, activities that are altered due to changes in the local microenvironment [5–7]. In a fibrotic lung, there is an imbalance in the turnover of ECM proteins causing excessive production and deposition of ECM proteins, forming a disease-specific organization and composition of the matrix [8–10]. The pathological mechanism underlying the initiation and progression of IPF is not fully understood, and there are no e ffective treatment options, highlighting the need to identify e ffective molecular targets for therapeutic interventions [11]. Lung tissue slices, decellularized for cellular removal, can serve as human structural matrices to study the important and complex interaction between cells and matrix [12,13]. In comparison to other cell culture systems, decellularized tissue (sca ffolds) comprise a unique ex vivo system that more closely mimics the original intricate 3D milieu of the lung. Through this ex vivo model a better understanding of unknown key cellular mechanisms can be obtained in order to understand which ECM properties drive the formation of fibrotic tissue and which role the ECM of IPF sca ffolds has in disease progression. The matrisome protein classification system defined by Naba et al., clearly describes the ECM components, subgrouping ECM matrisome core proteins (collagens, glycoproteins and proteoglycans) and ECM associated proteins such as ECM-a ffiliated proteins, ECM regulators, and secreted factors [14]. Distal lung tissue is mainly composed of fibrillar collagens I, III, V, and VI and the basement membrane (BM) collagen type IV [7]. Intertwined with collagen type IV are nidogens, perlecan, and laminins, which comprise the BM network, a protein complex facilitating epithelial and endothelial cell attachment and regulating cellular behavior [15]. Alterations of the BM structure and other ECM components affect both morphology and biomechanical properties of the tissues, identifying matrix sti ffness as an important biomechanical signal for cell responses [16]. Tissue sti ffening of the lung, caused by increased ECM deposition in the alveoli that leads to a loss in tissue elasticity, induces di fferentiation of fibroblasts into myofibroblasts, a response that in part could be de-activated when changing culture conditions in vitro from sti ff to softer substrates [17]. In IPF, fibroblasts demonstrate an increased cellular sti ffness, perhaps functioning as a positive feedback loop contributing to the formation of a non-compliant sti ff lung tissue [18].

In this study we focused on the distal parenchymal matrisome properties of lung sca ffolds derived from healthy donors and IPF patients in a unique 3D-ex vivo setting, mimicking pulmonary physiological conditions. Our hypothesis was that the matrisome properties, i.e., biomechanical properties and matrisome composition, of the ECM have a fundamental impact on cellular responses and may act as a mechanism in disease progression.
