Date Published: February 2, 2017
Publisher: Public Library of Science
Author(s): Jinglei Wu, Priya Ravikumar, Kytai T. Nguyen, Connie C. W. Hsia, Yi Hong, Xiaoming He.
Decellularized extracellular matrix (ECM) contains complex tissue-specific components that work in concert to promote tissue repair and constructive remodeling and has been used experimentally and clinically to accelerate epithelial wound repair, leading us to hypothesize that lung-derived ECM could mitigate acute lung injury. To explore the therapeutic potential of ECM for noninvasive delivery to the lung, we decellularized and solubilized porcine lung ECM, then characterized the composition, concentration, particle size and stability of the preparation. The ECM preparation at 3.2 mg/mL with average particle size <3 μm was tested in vitro on human A549 lung epithelial cells exposed to 95% O2 for 24 hours, and in vivo by tracheal instillation or nebulization into the lungs of rats exposed intermittently or continuously to 90% O2 for a cumulative 72 hours. Our results showed that the preparation was enriched in collagen, reduced in glycosaminoglycans, and contained various bioactive molecules. Particle size was concentration-dependent. Compared to the respective controls treated with cell culture medium in vitro or saline in vivo, ECM inhalation normalized cell survival and alveolar morphology, and reduced hyperoxia-induced apoptosis and oxidative damage. This proof-of-concept study established the methodology, feasibility and therapeutic potential of exogenous solubilized ECM for pulmonary cytoprotection, possibly as an adjunct or potentiator of conventional therapy.
Acute lung injury (ALI) has an incidence from 16 per 100,000 among youths to 306 per 100,000 in the elderly, with 200,000 cases occurring in the United States annually and an in-hospital mortality of 40% [1,2]. Regardless of the specific cause of ALI, the common manifestations include increased permeability of the epithelium and endothelium, as well as recruitment and activation of alveolar macrophages and neutrophils to the lung, which lead to the release of pro-inflammatory and cytotoxic mediators . Furthermore, the increased reactive oxygen species (ROS) damage cell components including proteins, lipids, carbohydrates and DNA, promote apoptosis and reduce endogenous antioxidant capability [4–7].
Most studies of decellularized ECM focused on preserving the whole lung ECM scaffold for re-cellularization and organ replacement. To our knowledge, this is the first study to develop a decellularized lung-derived ECM suspension/solution suitable for inhalational delivery to the lung, and test its effects on injury mitigation. Our major findings are as follows: By enzymatic digestion and filtration, porcine lung ECM was processed into a fine solution/suspension with particle diameters less than 5 μm, making it suitable for nebulization to reach the distal lung. The ECM preparation significantly increased viability of lung epithelial cells exposed to hyperoxia in vitro. A dilute ECM preparation (3.2 mg/mL concentration) was delivered into rat lung by either tracheal instillation or nebulization. Both delivery methods showed similar ameliorative effects against acute lung damage induced by either continuous or intermittent hyperoxia exposure. These data established the feasibility a novel ECM-based formulation for nebulization and non-invasive targeted delivery to the lung, and demonstrated its beneficial effects in alleviating acute lung injury.
We manufactured and characterized a lung-specific decellularized ECM microparticle suspension/solution designed for inhalation delivery, demonstrated its efficacy in ameliorating pulmonary oxidant damage, and identified several bioactive components of this complex mixture that could have mediated the observed effects. These results support the use of decellularized ECM as a novel lung protection strategy, and direct further investigation to clarify its mechanisms of action. Future studies will aim to identify additional ECM components and their interactions, standardize the preparation, optimize dose-response and immune profiles following repeated administration, and examine a possible role for ECM-mediated potentiation of conventional pharmacological agents used in the treatment of acute lung injury.