Idiopathic pulmonary fibrosis (IPF) is a serious and chronic lung disease marked by progressive scarring of lung tissue. Over time, this scarring thickens and stiffens the lungs, making it increasingly difficult for oxygen to pass into the bloodstream. As lung function declines, patients may experience shortness of breath, persistent dry cough, fatigue, and reduced exercise tolerance.
The exact cause of IPF remains unknown. However, researchers understand that the disease is driven by repeated injury to the cells lining the lungs, followed by abnormal wound healing. Instead of resolving normally, the repair process becomes dysregulated. Fibroblasts—cells responsible for producing structural support proteins—become overactive and transform into myofibroblasts. These cells produce excessive extracellular matrix (ECM), including collagen, leading to permanent scarring.
Two antifibrotic drugs were approved by the U.S. Food and Drug Administration in 2014. While these medications can slow disease progression, they do not reverse established fibrosis or cure the disease. As a result, researchers continue to search for more effective and regenerative treatment strategies.
In this study, Lee et al. investigated the therapeutic potential of human adipose-derived mesenchymal stem cell (AD-MSC) EVs in the IPF model.
Mesenchymal Stem Cells and the Shift Toward Cell-Free Therapies
Mesenchymal stem cells (MSCs) have attracted attention in regenerative medicine for their ability to regulate inflammation, reduce fibrosis, and support tissue repair. In lung disease models, MSC therapy has shown promise due to its immunomodulatory and antifibrotic properties.
However, stem cell therapy presents challenges. Injected cells may differentiate unpredictably, survive poorly, or, in rare cases, raise concerns about tumor formation. Additionally, studies examining MSC treatment in pulmonary fibrosis have produced mixed results, with some showing reduced scarring and others suggesting limited or inconsistent benefit. Differences in dose, timing, and delivery methods also complicate interpretation.
To address these limitations, Lee et al.focused on extracellular vesicles (EVs) derived from MSCs. EVs are nanosized membrane-bound particles naturally released by cells. They contain proteins, lipids, and small RNA molecules that reflect the properties of their parent cells. Importantly, EVs can reproduce many of the beneficial effects of MSCs without introducing living cells into the body.
MSC-Derived Extracellular Vesicles as Cell-Free Therapeutics
Extracellular vesicles function as biological messengers. They travel between cells and deliver molecular signals that influence inflammation, fibrosis, and tissue repair. MSC-derived EVs have demonstrated anti-inflammatory, antifibrotic, and regenerative properties in multiple disease models.
Compared with stem cell transplantation, EV therapy offers several potential advantages. EVs carry fewer safety concerns, are easier to store and transport, and can be administered through multiple routes. Their small size allows them to penetrate tissues efficiently, making them particularly attractive for lung delivery.
Several preclinical studies have shown that EVs derived from bone marrow-, umbilical cord-, and adipose tissue-derived MSCs can reduce inflammation and collagen deposition in animal models of pulmonary fibrosis.
Investigating Inhaled AD-MSC EVs for Pulmonary Fibrosis
In the study summarized here, the authors evaluated extracellular vesicles derived from human adipose-derived mesenchymal stem cells (AD-MSCs). These EVs were isolated using a filtration-based purification system and thoroughly characterized to confirm their size, structure, and molecular composition.
The EVs measured approximately 68-140 nanometers in diameter, a size range consistent with functional extracellular vesicles. When aerosolized with a vibrating-mesh nebulizer, the resulting particles had an average aerodynamic diameter of about 3.3 micrometers. This size is ideal for inhalation because it allows particles to reach deep into the small airways and alveoli, where fibrosis develops.
Lee et al. tested inhaled AD-MSC EVs in a well-established mouse model of bleomycin-induced pulmonary fibrosis, a chemical that causes lung injury and scarring similar to that seen in IPF.
Inhalation as a Targeted Delivery Strategy
Inhalation is an important route for treating respiratory diseases because it delivers therapy directly to the lung tissue. Compared with intravenous injection, inhaled delivery can achieve therapeutic effects at lower doses while reducing systemic side effects. It also makes repeated administration more feasible and convenient.
Extracellular vesicles are well-suited for inhalation due to their structural stability and small size. Early-phase clinical trials are already exploring inhaled EVs for respiratory diseases, including viral infections and inflammatory lung conditions.
In this study, inhaled AD-MSC EVs were delivered directly into the lungs of fibrotic mice. The treatment significantly reduced collagen deposition and decreased α-smooth muscle actin (α-SMA) expression, a marker of activated myofibroblasts. These findings suggest that EV treatment reduced both scarring and fibroblast activation.
Blocking Fibrosis at the Molecular Level
Pulmonary fibrosis is driven in part by overactivation of two key signaling pathways: transforming growth factor beta (TGF-β) and WNT signaling. These pathways regulate fibroblast migration, myofibroblast differentiation, and extracellular matrix production.
When TGF-β signaling is activated, it triggers phosphorylation of proteins called Smad2 and Smad3. This promotes fibroblast transformation into myofibroblasts and increases collagen production. TGF-β signaling also interacts with the WNT pathway, amplifying fibrotic responses.
This study demonstrated that AD-MSC EVs inhibited the activation of both TGF-β and WNT pathways. Specifically, EV treatment reduced phosphorylation of Smad3 and GSK-3β, key downstream regulators of these pathways. As a result, fibroblast migration decreased, myofibroblast differentiation was suppressed, and extracellular matrix production was reduced in both cell culture experiments and animal models.
By targeting these central signaling mechanisms, EV therapy appeared to interrupt the biological cascade that drives progressive lung scarring.
The Role of microRNAs in EV Function
One of the most important components of extracellular vesicles is microRNA (miRNA), a small regulatory RNA that influences gene expression. The authors analyzed the miRNA content of AD-MSC EVs and identified several highly enriched miRNAs associated with antifibrotic effects.
Among these, members of the let-7 family—specifically let-7a and let-7b—were of particular interest. These miRNAs are known to regulate TGF-β and WNT signaling pathways. In laboratory experiments, let-7a and let-7b suppressed the expression of TGFBR1 (a receptor that activates TGF-β signaling) and WNT9A (a protein involved in WNT pathway activation).
By reducing TGFBR1 and WNT9A expression, these miRNAs lowered the activity of downstream fibrotic genes, including α-SMA and collagen-related genes such as COL1A1 and COL3A1. This molecular suppression aligns with the observed reduction in myofibroblast differentiation and collagen deposition in treated animals.
These findings suggest that the therapeutic effect of AD-MSC EVs may be driven, at least in part, by their miRNA cargo, which regulates fibrosis-related signaling pathways at the genetic level.
Implications for Future Therapy
The results of this study demonstrate that inhaled AD-MSC EVs significantly reduced lung fibrosis in an established animal model. By suppressing TGF-β and WNT signaling and delivering antifibrotic miRNAs, EV therapy disrupted the core biological drivers of fibrosis.
While the findings are encouraging, further research is required before translation to clinical practice. Questions remain regarding optimal dosing, treatment frequency, long-term safety, and how EVs behave within the human lungs over time. Studies examining pharmacokinetics and biodistribution will be essential for guiding clinical trial design.
Nonetheless, extracellular vesicles represent an innovative and biologically targeted approach to treating pulmonary fibrosis. Unlike traditional antifibrotic drugs that primarily slow disease progression, EV-based therapies aim to modify the underlying cellular environment and potentially support tissue repair.
Conclusion
Idiopathic pulmonary fibrosis remains a progressive and life-threatening disease with limited treatment options. Abnormal wound healing, persistent fibroblast activation, and excessive extracellular matrix deposition drive irreversible lung scarring.
This study provides evidence that inhaled extracellular vesicles derived from adipose-derived mesenchymal stem cells can reduce fibrosis in a preclinical model. By suppressing TGF-β and WNT signaling pathways and delivering antifibrotic microRNAs such as let-7a and let-7b, AD-MSC EVs decreased myofibroblast activation and collagen production.
Although additional research is necessary to confirm safety and efficacy in humans, these findings highlight the potential of inhaled EV therapy as a promising next-generation strategy for pulmonary fibrosis.
Source: Lee KS, Yeom SH, Kim MK, Woo CH, Choi YC, Choi JS, Cho YW. Therapeutic potential of human stem cell-derived extracellular vesicles in idiopathic pulmonary fibrosis. Extracell Vesicles. 2024;4:100045. doi:10.1016/j.vesic.2024.100045.