Exosomes are extracellular vesicles secreted by most cell types and represent various biological properties depending on their producing cells. They are also known to be important mediators of intercellular communication. Recent data suggest that exosomes can mediate the therapeutic effects of their parental cells; hence, they have been in the spotlight as novel therapeutics. To develop and manufacture effective therapeutic exosomes, customized strategies are needed to use appropriate technologies for exosome engineering and to select suitable production cell lines. In this review, we provide an overview of currently available exosome engineering platform technologies for loading active pharmaceutical ingredient cargo and the types of human cells/cell lines that are being used as exosome-producing cells, particularly focusing on their characteristics, advantages, and disadvantages.

Receptor activator of nuclear factor kappa B-ligand (RANKL), its receptor RANK, and osteoprotegerin which binds RANKL and acts as a soluble decoy receptor, are essential controllers of bone remodeling. They also play important roles in establishing immune tolerance and in the development of the lymphatic system and mammary glands. In bone, RANKL stimulates osteoclast formation by binding RANK on osteoclast precursors and osteoclasts. This is required for bone resorption. Recently, RANKL and RANK have been shown to be functional components of extracellular vesicles (EVs). Data linking RANKL and RANK in EVs to biological regulatory roles are reviewed, and crucial unanswered questions are examined. RANKL and RANK are transmembrane proteins and their presence in EVs allows them to act at a distance from their cell of origin. Because RANKL-bearing osteocytes and osteoblasts are often spatially distant from RANK-containing osteoclasts in vivo, this may be crucial for the stimulation of osteoclast formation and bone resorption. RANK in EVs from osteoclasts has the capacity to stimulate a RANKL reverse signaling pathway in osteoblasts that promotes bone formation. This serves to couple bone resorption with bone formation and has inspired novel bifunctional therapeutic agents. RANKL- and RANK- containing EVs in serum may serve as biomarkers for bone and immune pathologies. In summary, EVs containing RANKL and RANK have been identified as intercellular regulators in bone biology. They add complexity to the central signaling network responsible for maintaining bone. RANKL- and RANK-containing EVs are attractive as drug targets and as biomarkers.

Aim: To confirm the presence of extracellular vesicles (EVs) in cell-free saliva (CFS) of children with asthma and describe the isolated EV population.

Methods: A pooled sample of CFS EVs isolated from 180 participants using ExoQuick-TC was examined in downstream analyses. Transmission electron microscopy (TEM) was used to confirm the presence of EVs. Nanoparticle tracking analysis (NTA) and single particle interferometric reflectance imaging sensing (SP-IRIS) with fluorescence were used for sizing, counting, and phenotyping of EVs. Capillary immunoassays were used for protein quantitation.

Results: TEM confirmed the presence of EVs of diverse sizes, indicating the prep contained a heterogeneous population of EVs. Capillary immunoassays confirmed the presence of EV-associated proteins (CD9, CD63, CD81, ICAM-1, and ANXA5) and indicated limited cellular contamination. As others have also reported, there were discrepancies in the EV sizing and enumeration across platforms. Fluorescent NTA detected particles with a mode diameter of ~90 nm, whereas SP-IRIS reported sizes of ~55-60 nm that more closely approximated the TEM results. Consistent with protein immunoassay results, SP-IRIS with fluorescence showed that the majority of these EVs were CD9- and CD63-positive, with little expression of CD81.

Conclusion: EVs from CFS can be isolated using a high-throughput method that can be scaled to large epidemiological studies. To our knowledge, we are the first to characterize CFS EVs from patients with asthma. The use of CFS EVs as potential novel biomarkers in asthma warrants further investigation and opens a new avenue of research for future studies.

Exosomes are a subtype of extracellular vesicles released from different cell types including those in the nervous system, and are enriched in a variety of bioactive molecules such as RNAs, proteins and lipids. Numerous studies have indicated that exosomes play a critical role in many physiological and pathological activities by facilitating intercellular communication and modulating cells’ responses to external environments. Particularly in the central nervous system, exosomes have been implicated to play a role in many neurological disorders such as abnormal neuronal development, neurodegenerative diseases, epilepsy, mental disorders, stroke, brain injury and brain cancer. Since exosomes recapitulate the characteristics of the parental cells and have the capacity to cross the blood-brain barrier, their cargo can serve as potential biomarkers for early diagnosis and clinical assessment of disease treatment. In this review, we describe the latest findings and current knowledge of the roles exosomes play in various neurological disorders and brain cancer, as well as their application as promising biomarkers. The potential use of exosomes to deliver therapeutic molecules to treat diseases of the central nervous system is also discussed.

Extracellular vesicles (EVs) have been recognized as an evolving biomarker within the liquid biopsy family. While carrying both host cell proteins and different types of RNAs, EVs are also present in sufficient quantities in biological samples to be tested using many molecular analysis platforms to interrogate their content. However, because EVs in biological samples are comprised of both disease and non-disease related EVs, enrichment is often required to remove potential interferences from the downstream molecular assay. Most benchtop isolation/enrichment methods require > milliliter levels of sample and can cause varying degrees of damage to the EVs. In addition, some of the common EV benchtop isolation methods do not sort the diseased from the non-diseased related EVs. Simultaneously, the detection of the overall concentration and size distribution of the EVs is highly dependent on techniques such as electron microscopy and Nanoparticle Tracking Analysis, which can include unexpected variations and biases as well as complexity in the analysis. This review discusses the importance of EVs as a biomarker secured from a liquid biopsy and covers some of the traditional and non-traditional, including microfluidics and resistive pulse sensing, technologies for EV isolation and detection, respectively.