Thanh Huyen Phan

Introduction: Chronic obstructive pulmonary disease (COPD), the third leading global cause of death, lacks innovative therapeutics due to the absence of efficient lung models. This study presents an in vitro lung mimic for healthy and COPD conditions. Methods: (1) Employing CNBio's PhysioMimix, we developed a microfluidic 3D system to emulate dynamic human lung flow conditions, ensuring unidirectional cell flow throughout the culture. (2) Recognizing extracellular matrix (ECM)'s influence on cellular activity, we employed collagen type I and fibronectin to functionalize substrates. (3) Comparing co-cultures of normal human lung fibroblasts (NHLFs) with either normal or diseased human bronchial epithelial cells (NHBE, DHBE) created healthy and COPD lung mimics. To assess model reproducibility, we implemented a comprehensive QC strategy involving real-time and end-point measurements, including light microscopy, scanning electron microscopy (SEM), impedance spectroscopy with transepithelial electrical resistance (TEER), immunofluorescence (IF) staining, and cytokine profiling. Results: (1) Dynamic conditions showcased increased phenotype heterogeneity (light microscopy) and hairy-like ciliated cell structures (SEM), indicating enhanced barrier protection and mucociliary clearance. (2) Impedance spectroscopy revealed collagen-I's superior impact on cell adhesion, growth, and differentiation compared to fibronectin. (3) DHBEs exhibited compromised barrier integrity (TEER), goblet cell hypertrophy (IF), reduced cilia (SEM), and an inflammatory phenotype (cytokine). Conclusion: Our study emphasizes technological considerations in constructing lung models. We developed a representative COPD model with comprehensive QC, facilitating COPD therapeutic discovery.

Nanoparticles hold a great potential for therapeutic targeting due to their ability to improve the stability of encapsulated cargo and promote the transport of cargo across membranes to reach to the target site. Most commercially available nanomedicines are simple synthetic liposomes, however, there are numerous side effects due to their off-target delivery and rapid clearance from the bloodstream. Recently, attention has moved toward extracellular vesicles (EVs)–lipid bilayer enclosed particles released by cells (size ranging from 30 to 10,000 nm in diameter). EVs carry and transport lipids, proteins, and nucleic acids from their parental cells to recipient cells, hence they play a key role in intercellular communication. The ability of EVs to cross biological barriers including the blood brain barrier has generated significant attention to explore them as potential biomarkers and natural drug delivery vehicles for various therapeutics and small molecules. EVs have also been implicated in disease pathogenesis by transmitting pathogenic proteins between cells, making them promising biomarkers for disease diagnosis and monitoring. In this review, we will focus on the potential and challenges of EVs as biomarkers, drug delivery vehicles and next-generation therapeutics. Finally, we will explore misfolded protein disorders, amyloidosis, as a case study for how EVs may contribute to disease pathology and how EVs could be applied in the clinic as diagnostic and prognostic biomarkers of amyloid diseases.

Current medicine has only taken us so far in reducing disease and tissue damage. Extracellular vesicles (EVs), which are membranous nanostructures produced naturally by cells, have been hailed as a next-generation medicine. EVs deliver various biomolecules, including proteins, lipids and nucleic acids, which can influence the behaviour of specific target cells. Since EVs not only mirror composition of their parent cells but also modify the recipient cells, they can be used in three key areas of medicine: regenerative medicine, disease detection and drug delivery. In this Review, we discuss the transformational and translational progress witnessed in EV-based medicine to date, focusing on two key elements: the mechanisms by which EVs aid tissue repair (for example, skin and bone tissue regeneration) and the potential of EVs to detect diseases at an early stage with high sensitivity and specificity (for example, detection of glioblastoma). Furthermore, we describe the progress and results of clinical trials of EVs and demonstrate the benefits of EVs when compared with traditional medicine, including cell therapy in regenerative medicine and solid biopsy in disease detection. Finally, we present the challenges, opportunities and regulatory framework confronting the clinical application of EV-based products.

Extracellular vesicles (EVs) are nanoparticles released by cells that contain a multitude of biomolecules, which act synergistically to signal multiple cell types. EVs are ideal candidates for promoting tissue growth and regeneration. The tissue regenerative potential of EVs raises the tantalizing possibility that immobilizing EVs on implant surfaces could potentially generate highly bioactive and cell-instructive surfaces that would enhance implant integration into the body. Such surfaces could address a critical limitation of current implants, which do not promote bone tissue formation or bond bone. Here, we developed bioactive titanium surface coatings (SurfEV) using two types of EVs: secreted by decidual mesenchymal stem cells (DEVs) and isolated from fermented papaya fluid (PEVs). For each EV type, we determined the size, morphology, and molecular composition. High concentrations of DEVs enhanced cell proliferation, wound closure, and migration distance of osteoblasts. In contrast, the cell proliferation and wound closure decreased with increasing concentration of PEVs. DEVs enhanced Ca/P deposition on the titanium surface, which suggests improvement in bone bonding ability of the implant (i.e., osteointegration). EVs also increased production of Ca and P by osteoblasts and promoted the deposition of mineral phase, which suggests EVs play key roles in cell mineralization. We also found that DEVs stimulated the secretion of secondary EVs observed by the presence of protruding structures on the cell membrane. We concluded that, by functionalizing implant surfaces with specialized EVs, we will be able to enhance implant osteointegration by improving hydroxyapatite formation directly at the surface and potentially circumvent aseptic loosening of implants.

Extracellular vesicles (EVs) have been lauded as next generation medicines, but very few EV-based therapeutics have progressed to clinical use. Limited clinical translation is largely due to technical barriers that hamper our ability to mass-produce EVs, i.e. to isolate, purify and characterise them effectively. Technical limitations in comprehensive characterisation of EVs leads to unpredicted biological effects of EVs. Here, using a range of optical and non-optical techniques, we showed that the differences in molecular composition of EVs isolated using two isolation methods correlated with the differences in their biological function. Our results demonstrated that the isolation method determines the composition of isolated EVs at single and sub-population levels. Besides the composition, we measured for the first time the dry mass and predicted sedimentation of EVs. These parameters were shown to correlate well with the biological and functional effects of EVs on single cell and cell cultures. We anticipate that our new multiscale characterisation approach, which goes beyond traditional experimental methodology, will support fundamental understanding of EVs as well as elucidate the functional effects of EVs in in vitro and in vivo studies. Our findings and methodology will be pivotal for developing optimal isolation methods and establishing EVs as mainstream therapeutics and diagnostics. This innovative approach is applicable to a wide range of sectors including biopharma and biotechnology as well as to regulatory agencies.