• Journal of Life Science
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• v.10 no.4
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• pp.408-421
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• 2000

We tried to establish the culture method of the chondrocyte isolated from the epiphyseal cartilage and to investigate morphological changes of chondrocyte cultured with enzyme-digested costal cartilage, the perichondrium and experimentally damaged meniscus of rabbit. De novo chondrocyte pellets were prepared from epiphyseal plates by culturing isolated epiphyseal chondrocytes from 4 week. old rabbits. We morphologically assessed the cartilage formation of the chondrocyte culture with enzyme-digested costal carilage, the perichondrial culture, the cultured chondrocytes transplants into experimentally damaged meniscus of rabbits, the perichondrial culture, the cultured chondrocytes transplants into experimentally damaged meniscus of rabbit. In the 24 days, the epiphyseal chondrocytes maintained the typical phenotypes of the partial nodular cell formation. The 30 days cryopreserved chondrocytes showed abnormal and irregular shape. In the type II collagen added culture, the chondrocytes showed expanded rough endoplasmic reticulum and small and large round-like vesicles of processes. In the type IV collagen added culture, the chondrocytes showed large perinuclear vaculoes and abundant well-developed rough endoplasmic reticulum of processes. In the culture with enzyme- digested costal cartilage and the perichondrial culture, the chondrocytes showed a few swelling rough endoplasmic reticulum and vacuoles. The cultured epiphyseal chondrocytes maintained typical phenotype and the chondrocytes were grown faster and maintained more typical phenotype in the type II and IV collagen added culture. The transformed chondrocytes secreted abundant extracellular matrix in the type II collagen added culture, and showed processes in the type IV collagen added culture. The perichondrial chondrocytes were grown faster and their implants were able to transplant. The cultured chondrocytes transplanted into experimentally damaged meniscus were adapted between the meniscus tissues. And the immunocyto-chemical reaction of the type II collagen of the chondrocytes were found to be maintained. The chondrocytes cultured cartilage. The chondrocytes secreted abundantly. The cultured chondrocytes transplanted into experimentally damaged meniscus changed immature cells into enlarged mature cells with extracellular secretion.

• Journal of Microbiology
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• v.38 no.4
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• pp.218-223
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• 2000

Using two drugs, tunicamycin and brefeldin A, which affect protein processing, we investigated the intracellular processing mechanism of secreted aspartyl proteinase 1 (SAPl) of Candide albicans. Three intracellular forms of SAPI were detected by immunoblotting using menoclonal antibody (MAb) CAPl. Their molecular weights were approximately 40, 41 and 45 kDa, respectively. The 41 kDa protein is a glycoprotein and may be the same as the extracellular form judging by its molecular mass. The 40 kDa protein was the unglycosylated form and its molecular mass coincided with deglycosylated SAPl and the 45 kDa protein was also the unglycosylated form. Neither the 40 and 45 kDa proteins were detected in the culture supernatant of C. albicans. These suggested that the 40 and 45 kDa proteins might be intracellular precursor forms of SAPI. These results show that SAPI is translated as a 45 kDa precusor form in the endoplasmic reticulum and the 45 kDa precursor farm undergoes proteolytic cleavage after translocation into the Golgi apparatus, generating the 40 kDa precursor form. This 40 kDa precursor is converted into a 41 kDa mature form through glycosylation in the Golgi apparatus. The mature form of the 41 kDa protein is sorted into secretary vesicles and finally released into the extracellular space through membrane fusion. When the glycan region of SAPl was digested with N-glycosidase F, both stability and activity of the enzyme decreased. These results indicate that the glycan attached to the enzyme may, at least in parti be related to enzyme stability and activity.

• IMMUNE NETWORK
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• v.23 no.1
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• pp.3.1-3.14
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• 2023

Microvilli are outer membrane organelles that contain cross-linked filamentous actin. Unlike well-characterized epithelial microvilli, T-cell microvilli are dynamic similar to those of filopodia, which grow and shrink intermittently via the alternate actin-assembly and -disassembly. T-cell microvilli are specialized for sensing Ags on the surface of Ag-presenting cells (APCs). Thus, these finger-shaped microprotrusions contain many signaling-related proteins and can serve as a signaling platforms that induce intracellular signals. However, they are not limited to sensing external information but can provide sites for parts of the cell-body to tear away from the cell. Cells are known to produce many types of extracellular vesicles (EVs), such as exosomes, microvesicles, and membrane particles. T cells also produce EVs, but little is known about under what conditions T cells generate EVs and which types of EVs are released. We discovered that T cells produce few exosomes but release large amounsts of microvilli-derived particles during physical interaction with APCs. Although much is unanswered as to why T cells use the same organelles to sense Ags or to produce EVs, these events can significantly affect T cell fate, including clonal expansion and death. Since TCRs are localized at microvilli tips, this membrane event also raises a new question regarding long-standing paradigm in T cell biology; i.e., surface TCR downmodulation following T cell activation. Since T-cell microvilli particles carry T-cell message to their cognate partner, these particles are termed T-cell immunological synaptosomes (TISs). We discuss the potential physiological role of TISs and their application to immunotherapies.

• Journal of Sensor Science and Technology
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• v.28 no.6
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• pp.355-359
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• 2019

Exosomes, known as nanoscale extracellular vesicles in the range of 30-150 nm, are known to contain clinically significant information. However, there is still insufficient information on exosomal membrane proteins for cancer diagnosis. In this work, we investigated the characteristics of the membrane proteins of exosomes shed by cultured breast cancer cell lines using a surface plasmon resonance (SPR) spectroscopy and pre-activated alkanethiols modified sensor chips. The antibodies of breast cancer biomarkers such as MCU-16, EpCAM, CD24, ErbB2, and CA19-9 were immobilized on the pre-activated alkanethiols surfaces without any activation steps. The purified exosomes were loaded onto each antibody surface. The affinity rank of the antibody surfaces was decided by the relative capture efficiency factors for the exosomes. In addition, an antibody with a relative capture efficiency close to 100% was tested with exosome concentration levels of 10⁴/µl, 10⁵/µl, and 10⁶/µl for quantitative analysis.

• Journal of Microbiology and Biotechnology
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• v.25 no.2
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• pp.302-306
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• 2015

VvpM, one of the extracellular metalloproteases produced by Vibrio vulnificus, induces apoptotic cell death via a pathway consisting of ERK activation, cytochrome c release, and activation of caspases-9 and -3. VvpM-treated cells also showed necrotic cell death as stained by propidium iodide (PI). The percentage of PI-stained cells was decreased by pretreatment with Necrostatin-1, indicating that VvpM-mediated cell death occurs through necroptosis. The appearance of autophagic vesicles and lipidated form of light-chain-3B in rVvpM-treated cells suggests an involvement of autophagy in this process. Therefore, the multifarious action of VvpM might be one of the factors responsible for V. vulnificus pathogenesis.

• Korean Journal of Veterinary Research
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• v.35 no.2
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• pp.263-270
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• 1995

Creatine(Cr) and phosphocreatine(PCr), the important mediators of intracellular high-energy phosphate buffer system, were found in the tissues of mouse seminal vesicle and also in the extracellular fluids of seminal vesicle secretion. This study was performed m confirm that the secretion and accumulation of Cr and PCr is regulated by testosterone and its $5{\alpha}$-reduced metabolite, $5{\alpha}$-dihydrotestosterone(DHT). In addition, the effect of nandrolone decanoate(ND), a synthetic anabolic steroid, on the levels of Cr and PCr in the seminal vesicle was compared with those of testosterone propionate(TP) and DHT. Male Swiss-Webster mice were castrated and three groups of the castrates were treated with daily injection(sc) of same molar dose($1.45{\times}10^{-8}mol/g\;BW$) of TP, DHT, or ND. All three androgens rapidly increased weights of seminal vesicle tissue and fluid, and also increased concentrations of Cr and PCr in the tissue and fluid. However, ND was least effective in increasing seminal vesicle weights, whereas ND was as effective as, or in some cases, more effective than, TP or DHT in increasing Cr and PCr levels in the tissue and fluid. The results confirm that the accumulation of Cr and PCr in the seminal vesicles is regulated by testosterone and DHT, and also suggest that the effects of androgens on seminal vesicle growth and secretory activity may be differentiated.

• Journal of Sensor Science and Technology
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• v.30 no.2
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• pp.119-124
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• 2021

Isolation and separation of biological nanoparticles, such as cells and extracellular vesicles, are important techniques for their characterization. Dielectrophoresis (DEP) based on microfluidic chips is an effective method to isolate and separate the nanoparticles. However, the electrodes of the DEP chips are electrolyzed by the electrical signals applied to the nanoparticles. Thus, the isolation/separation efficiency of the nanoparticles is reduced considerably. Through this study, we developed a microfluidic DEP chip for reliable isolation/ separation of nanoparticles and developed a passivation technique for the protection of the DEP chip electrodes. The electrode passivation process was designed using a hydrogel and the stability of the hydrogel passivation layer was verified. The fabricated DEP chip and the proposed passivation technique were used for the collection and dispersion of the fluorescent polystyrene nanoparticles. The proposed chip and the technique for isolation and separation of nanoparticles can be leveraged in various bioelectronic applications.

• BMB Reports
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• v.55 no.1
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• pp.30-38
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• 2022

Cardiovascular disease, especially ischemic heart disease, is a major cause of mortality worldwide. Cardiac repair is one of the most promising strategies to address advanced cardiovascular diseases. Despite moderate improvement in heart function via stem cell therapy, there is no evidence of significant improvement in mortality and morbidity beyond standard therapy. The most salutary effect of stem cell therapy are attributed to the paracrine effects and the stem cell-derived exosomes are known as a major contributor. Hence, exosomes are emerging as a promising therapeutic agent and potent biomarkers of cardiovascular disease. Furthermore, they play a role as cellular cargo and facilitate intercellular communication. However, the clinical use of exosomes is hindered by the absence of a standard operating procedures for exosome isolation and characterization, problems related to yield, and heterogeneity. In addition, the successful clinical application of exosomes requires strategies to optimize cargo, improve targeted delivery, and reduce the elimination of exosomes. In this review, we discuss the basic concept of exosomes and stem cell-derived exosomes in cardiovascular disease, and introduce current efforts to overcome the limitations and maximize the benefit of exosomes including engineered biomimetic exosomes.

• Journal of Microbiology and Biotechnology
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• v.32 no.12
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• pp.1506-1514
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• 2022

Allergic contact dermatitis (ACD) is an allergen-specific T-cell-mediated inflammatory response, albeit with unclear pathogenesis. Exosomes are nanoscale extracellular vesicles secreted by several cell types and widely distributed in various biological fluids. Exosomes affect the occurrence and development of ACD through immunoregulation among other ways. Nevertheless, the role of exosomes in ACD warrants further clarification. This review examines the progress of research into exosomes and their involvement in the pathogenesis, diagnosis, and treatment of ACD and provides ideas for exploring new diagnostic and treatment methods for this disease.

• IMMUNE NETWORK
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• v.22 no.5
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• pp.38.1-38.24
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• 2022

Exosomes, which are well-known nanoscale extracellular vesicles, are multifunctional biomaterials derived from endosomes and perform various functions. The exosome is a critical material in cell-cell communication. In addition, it regulates the pathophysiological conditions of the tumor microenvironment in particular. In the tumor microenvironment, exosomes play a controversial role in supporting or killing cancer by conveying biomaterials derived from parent cells. Innate immunity is a crucial component of the host defense mechanism, as it prevents foreign substances, such as viruses and other microbes and tumorigenesis from invading the body. Early in the tumorigenesis process, the innate immunity explicitly recognizes the tumor via Ags and educates the adaptive immunity to eliminate it. Recent studies have revealed that exosomes regulate immunity in the tumor microenvironment. Tumor-derived exosomes regulate immunity against tumor progression and metastasis. Furthermore, tumor-derived exosomes regulate polarization, differentiation, proliferation, and activation of innate immune cells. Exosomes produced from innate immune cells can inhibit or support tumor progression and metastasis via immune cell activation and direct cancer inhibition. In this study, we investigated current knowledge regarding the communication between tumor-derived exosomes and innate immune cell-derived exosomes (from macrophages, dendritic cells, NK cells, and neutrophils) in the tumor microenvironment. In addition, we discussed the potential development of exosomal immunotherapy using native or engineered exosomes against cancer.