• Molecules and Cells
• /
• v.45 no.5
• /
• pp.284-290
• /
• 2022

Exosome, a type of nanoparticles also known as small extracellular vesicles are gaining attention as novel therapeutics for various diseases because of their ability to deliver genetic or bioactive molecules to recipient cells. Although many pharmaceutical companies are gradually developing exosome therapeutics, numerous hurdles remain regarding manufacture of clinical-grade exosomes for therapeutic use. In this mini-review, we will discuss the manufacturing challenges of therapeutic exosomes, including cell line development, upstream cell culture, and downstream purification process. In addition, developing proper formulations for exosome storage and, establishing good manufacturing practice facility for producing therapeutic exosomes remains as challenges for developing clinical-grade exosomes. However, owing to the lack of consensus regarding the guidelines for manufacturing therapeutic exosomes, close communication between regulators and companies is required for the successful development of exosome therapeutics. This review shares the challenges and perspectives regarding the manufacture and quality control of clinical grade exosomes.

• International Journal of Industrial Entomology and Biomaterials
• /
• v.33 no.2
• /
• pp.85-91
• /
• 2016

Exosomes are homogenous vesicles of 40-100 nm diameter produced endogenously. Exosomes are generated by inward budding into multi-vesicular bodies (MVB) and then released to extracellular space. Exosomes contain various nucleic acid and protein cargoes from their cells of origin and this endosomal cellular molecules are used for intracellular communication and for both promotion and suppression of immune responses. Recently, they are also considered as delivery vehicle for therapeutic proteins due to their characteristics of stability in body fluids and ability for target uptake. Also, they show less immune reactivity because the isolated exosome harboring therapeutic proteins can be from the same host. White-spotted flower chafer, Protaetia brevitarsis is one of the major insect commercially reared in Korea. There are bacterial and fungal pathogens causing diseases in the beetle, and these diseases incur economic loss to the larva-rearing farms. Due to their endosomal cargoes, exosomes are good candidates in use of disease diagnosis. In this study, we isolated insect exosome from the hemolymph of P. brevitarsis, and verified it by analysis of the exosome-specific surface proteins and RNA.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.47 no.2
• /
• pp.139-145
• /
• 2021

Plant-derived exosome-like nanovesicles (PELNs) are known to include various biological activities and possess high biocompatibility. Because PELNs can influence immune responses, cell differentiation, and proliferation regulation, they can be applied in multiple industries. However, the studies on the skin physiological of exosome-like nanovesicles derived from plant callus are insignificant compared to nanovesicles derived from mammalian cells. In this study, callus was induced from apple fruit (Malus domestica), and exosome-like nanovesicles (ACELNs) were isolated for improving skin barrier and anti-aging. The yield of ACELNs was 6.42 × 10⁹ particles/mL, and the particle size was ranged from 100 to 200 nm. HDF cells and HaCaT cells were concentration-dependent, increased in cell, and decreased in cytotoxicity. The cornified envelope formation was significantly increased compared to the control group. The COL1A1 expression and the FBN1 expression in HDF cells were increased. In addition, the ACELNs promoted collagen biosynthesis in UVA-irradiated HDF cells. These results might be considered as potential materials that could improve skin barrier and prevent skin aging.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.42 no.4
• /
• pp.359-366
• /
• 2016

Exosome, a small vesicle secreted from cells, has diverse functions depending on cell origins and tissue types and plays a important role in cell viability and intercellular communication. Recently, many researchers have demonstrated the use of exosomes for the treatment of cancers and immune diseases, and the development of diagnostic biomarker. However, the secretion mechanism of exosome from skin cell and its physiological functions in skin remain unclear. Thus, this study aimed to explore whether keratinocyte-derived exosome affects proliferation and migration in HaCaTs. Exosomes were isolated from HaCaTs by ExoQuick-TC and then boiled or unbolied. Boiled and unboiled exosome induced proliferation in HaCaTs in a dose-dependant manner ($0.1{\sim}20{\mu}g/mL$), respectively. Boiled and unboiled exosome at concentration of $20{\mu}g/mL$ increased proliferation level in HaCaTs by $186.96{\pm}3.87%$ and $193.48{\pm}10.48%$ compared with control group. Unboiled exosome stimulated migration in HaCaTs in a dose-dependent manner ($0.1{\sim}20{\mu}g/mL$), which reached a maxium at concentration of $20{\mu}g/mL$ ($179.39{\pm}4.89%$ of control), but boiled exosome did not affect HaCaT migration. In addition, unboiled exosome ($0.1{\sim}20{\mu}g/mL$) dose-dependently stimulated sprout outgrowth in HaCats. These results demonstrate that in exosome from HaCaTs, heat-stable components such as lipid may induce HaCaT proliferation and heat-unstable components such as protein may stimulate migration and sprout outgrowth in HaCaTs, thereby leading to reepithelialization and skin-wound healing activities. It is concluded that exosomes from HaCaTs may be used as cosmetic materials.

• Biomolecules & Therapeutics
• /
• v.31 no.3
• /
• pp.253-263
• /
• 2023

The biogenesis and biological roles of extracellular vesicles (EVs) in the progression of liver diseases have attracted considerable attention in recent years. EVs are membrane-bound nanosized vesicles found in different types of body fluids and contain various bioactive materials, including proteins, lipids, nucleic acids, and mitochondrial DNA. Based on their origin and biogenesis, EVs can be classified as apoptotic bodies, microvesicles, and exosomes. Among these, exosomes are the smallest EVs (30-150 nm in diameter), which play a significant role in cell-to-cell communication and epigenetic regulation. Moreover, exosomal content analysis can reveal the functional state of the parental cell. Therefore, exosomes can be applied to various purposes, including disease diagnosis and treatment, drug delivery, cell-free vaccines, and regenerative medicine. However, exosome-related research faces two major limitations: isolation of exosomes with high yield and purity and distinction of exosomes from other EVs (especially microvesicles). No standardized exosome isolation method has been established to date; however, various exosome isolation strategies have been proposed to investigate their biological roles. Exosome-mediated intercellular communications are known to be involved in alcoholic liver disease and nonalcoholic fatty liver disease development. Damaged hepatocytes or nonparenchymal cells release large numbers of exosomes that promote the progression of inflammation and fibrogenesis through interactions with neighboring cells. Exosomes are expected to provide insight on the progression of liver disease. Here, we review the biogenesis of exosomes, exosome isolation techniques, and biological roles of exosomes in alcoholic liver disease and nonalcoholic fatty liver disease.

• Mass Spectrometry Letters
• /
• v.12 no.3
• /
• pp.93-105
• /
• 2021

Exosomes have gained the attention of the scientific community because of their role in facilitating intercellular communication, which is critical in disease monitoring and drug delivery research. Exosome research has grown significantly in recent decades, with a focus on the development of various technologies for isolating and characterizing exosomes. Among these efforts is the use of matrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS), which offers high-throughput direct analysis while also being cost and time effective. MALDI is used less frequently in exosome research than electrospray ionization due to the diverse population of extracellular vesicles and the impurity of isolated products, both of which necessitate chromatographic separation prior to MS analysis. However, MALDI-MS is a more appropriate instrument for the analytical approach to patient therapy, given it allows for fast and label-free analysis. There is a huge drive to explore MALDI-MS in exosome research because the technology holds great potential, most notably in biomarker discovery. With methods such as fingerprint analysis, OMICs profiling, and statistical analysis, the search for biomarkers could be much more efficient. In this review, we highlight the potential of MALDI-MS as a tool for investigating exosomes and some of the possible strategies that can be implemented based on prior research.

• Journal of Ginseng Research
• /
• v.48 no.2
• /
• pp.211-219
• /
• 2024

Background: Recently, plant-derived exosome-like nanoparticles (PDENs) have been isolated, and active research was focusing on understanding their properties and functions. In this study, the characteristics and molecular properties of ginseng root-derived exosome-like nanoparticles (GrDENs) were examined in terms of skin protection. Methods: HPLC-MS protocols were used to analyze the ginsenoside contents in GrDENs. To investigate the beneficial effect of GrDENs on skin, HaCaT cells were pre-treated with GrDENs (0-2 × 10⁹ particles/mL), and followed by UVB irradiation or H₂O₂ exposure. In addition, the antioxidant activity of GrDENs was measured using a fluorescence microscope or flow cytometry. Finally, molecular mechanisms were examined with immunoblotting analysis. Results: GrDENs contained detectable levels of ginsenosides (Re, Rg1, Rb1, Rf, Rg2 (S), Gyp17, Rd, C-Mc1, C-O, and F2). In UVB-irradiated HaCaT cells, GrDENs protected cells from death and reduced ROS production. GrDENs downregulated the mRNA expression of proapoptotic genes, including BAX, caspase-1, -3, -6, -7, and -8 and the ratio of cleaved caspase-8, -9, and -3 in a dose-dependent manner. In addition, GrDENs reduced the mRNA levels of aging-related genes (MMP2 and 3), proinflammatory genes (COX-2 and IL-6), and cellular senescence biomarker p21, possibly by suppressing activator protein-1 signaling. Conclusions: This study demonstrates the protective effects of GrDENs against skin damage caused by UV and oxidative stress, providing new insights into beneficial uses of ginseng. In particular, our results suggest GrDENs as a potential active ingredient in cosmeceuticals to promote skin health.

• Journal of Animal Science and Technology
• /
• v.63 no.3
• /
• pp.593-602
• /
• 2021

Mammalian milk including microRNAs (miRNAs) as a novel class of noncoding RNAs, that can be transferred to infants and it plays on a critical role in biological functions such as immune regulation and development. However, the origin and functional importance of milk-derived miRNAs are still undetermined. This study applied RNA sequencing to explore the featured profiles of miRNA expression in colostrum and mature milk-originated exosomes from human, bovine, and caprine milk. These dietary exosome-derived miRNAs are highly conserved in human, bovine and caprine milk. Interestingly, abundant miRNAs expressed in human milk are similarly conserved across species. In addition, we confirmed that immune-related miRNAs (miR-30a-5p, miR-22-3p, and miR-26a) are commonly observed in the colostrum and mature milk of cows and caprines as well as humans. Our results provide new insights and resources for investigating the functionality of immune-associated miRNAs and evaluating physiological and biological condition in human, bovine and caprine milk as biomarkers.

• Asian-Australasian Journal of Animal Sciences
• /
• v.33 no.11
• /
• pp.1824-1836
• /
• 2020

Objective: On the hypothesis that grazing of cattle prompts organs to secrete or internalize circulating microRNAs (c-miRNAs) in parallel with changes in energy metabolism, we aimed to clarify biological events in adipose, skeletal muscle, and liver tissues in grazing Japanese Shorthorn (JSH) steers by a transcriptomic approach. Methods: The subcutaneous fat (SCF), biceps femoris muscle (BFM), and liver in JSH steers after three months of grazing or housing were analyzed using microarray and quantitative polymerase chain reaction (qPCR), followed by gene ontology (GO) and functional annotation analyses. Results: The results of transcriptomics indicated that SCF was highly responsive to grazing compared to BFM and liver tissues. The 'Exosome', 'Carbohydrate metabolism' and 'Lipid metabolism' were extracted as the relevant GO terms in SCF and BFM, and/or liver from the >1.5-fold-altered mRNAs in grazing steers. The qPCR analyses showed a trend of upregulated gene expression related to exosome secretion and internalization (charged multivesicular body protein 4A, vacuolar protein sorting-associated protein 4B, vesicle associated membrane protein 7, caveolin 1) in the BFM and SCF, as well as upregulation of lipolysis-associated mRNAs (carnitine palmitoyltransferase 1A, hormone-sensitive lipase, perilipin 1, adipose triglyceride lipase, fatty acid binding protein 4) and most of the microRNAs (miRNAs) in SCF. Moreover, gene expression related to fatty acid uptake and inter-organ signaling (solute carrier family 27 member 4 and angiopoietin-like 4) was upregulated in BFM, suggesting activation of SCF-BFM organ crosstalk for energy metabolism. Meanwhile, expression of plasma exosomal miR-16a, miR-19b, miR-21-5p, and miR-142-5p was reduced. According to bioinformatic analyses, the c-miRNA target genes are associated with the terms 'Endosome', 'Caveola', 'Endocytosis', 'Carbohydrate metabolism', and with pathways related to environmental information processing and the endocrine system. Conclusion: Exosome and fatty acid metabolism-related gene expression was altered in SCF of grazing cattle, which could be regulated by miRNA such as miR-142-5p. These changes occurred coordinately in both the SCF and BFM, suggesting involvement of exosome in the SCF-BFM organ crosstalk to modulate energy metabolism.

• Applied Microscopy
• /
• v.47 no.3
• /
• pp.171-175
• /
• 2017

Negative staining has been traditionally used for exosome imaging; however, the technique is limited to surface topology only and can cause staining artifacts. Therefore, to analyze the internal structure of exosomes, we employed a method of block preparation, thin sectioning, and electron tomography. In addition, an automatic serial sectioning technique with 15-nm thickness through focused ion beam was employed to observe the three-dimensional structure of exosomes of various sizes. Cryo-transmission electron microscopy revealed the near-to-native structure of exosomes.