• Journal of Marine Life Science
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• v.9 no.1
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• pp.9-21
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• 2024

Paralytic shellfish poisoning (PSP) including Saxitoxin (STX) is caused by harmful algae, and poisoning occurs when the contaminated seafood is consumed. The mouse bioassay (MBA), a standard test method for detecting PSP, is being sanctioned in many countries due to its low detection limit and the animal concerns. An alternative to the MBA is the Neuro-2a cell-based assay. This study aimed to establish various test conditions for Neuro-2a assay, including cell density, culture conditions, and STX treatment conditions, to suit the domestic laboratory environment. As a result, the initial cell density was set to 40,000 cells/well and the incubation time to 24 hours. Additionally, the concentration of Ouabain and Veratridine (O/V) was set to 500/50 μM, at which most cells died. In this study, we identified eight concentrations of STX, ranging from 368 to 47,056 fg/μl, which produced an S-shaped dose-response curve when treated with O/V. Through inter-laboratory variability comparison of the Neuro-2a assay, we established five Quality Control Criteria to verify the appropriateness of the experiments and six Data Criteria (Top and Bottom OD, EC₅₀, EC₂₀, Hill slop, and R² of graph) to determine the reliability of the experimental data. The Neuro-2a assay conducted under the established conditions showed an EC₅₀ value of approximately 1,800~3,500 fg/μl. The intra- & inter-lab variability comparison results showed that the coefficients of variation (CVs) for the Quality Control and Data values ranged from 1.98% to 29.15%, confirming the reproducibility of the experiments. This study presented Quality Control Criteria and Data Criteria to assess the appropriateness of the experiments and confirmed the excellent repeatability and reproducibility of the Neuro-2a assay. To apply the Neuro-2a assay as an alternative method for detecting PSP in domestic seafood, it is essential to establish a toxin extraction method from seafood and toxin quantification methods, and perform correlation analysis with MBA and instrumental analysis methods.

• Journal of Life Science
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• v.26 no.11
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• pp.1313-1319
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• 2016

Cancer is a complex disease heterogeneously composed of various types of cells including cancer stem-like cells responsible for relapse and chemoresistance in the tumor microenvironment. The conventional two-dimensional cell culture-based platform has critical limitations for representing the heterogeneity of cancer cells in the three-dimensional tumor niche in vivo. To overcome this insufficiency, three-dimensional cell culture methods in a scaffold-dependent or -free physical environment have been developed. In this study, we improved and simplified the HCT-8 colon cancer cell-based spheroid culture protocol and evaluated the relationship between cancer stemness and responses of chemosensitivity to 5- Fluorouracil (5-FU), a representative anticancer agent against colon cancer. Supplementation with defined growth factors in the medium and the culture dish of the regular surface with low attachment were required for the formation of constant-sized spheroids containing $CD44^+$ and $CD133^+$ colon cancer stem cells. The chemo-sensitivities of $CD44^+$ cancer stem cells in the spheroids were much lower than those of $CD44^-$ non-stem-like cancer cells, indicating that the chemoresistance to 5-FU is due to the stemness of colon cancer cells. Taken together, the inflammation and oncogenic gut environment-sensitive HCT-8 cell-based colon cancer spheroid culture and comparative evaluation using the simplified model would be an efficient and applicable way to estimate colon cancer stemness and pharmaceutical response to anticancer drugs in the realistic tumor niche.

• Journal of Applied Biological Chemistry
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• v.66
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• pp.416-423
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• 2023

Figs has known to have antioxidant, whitening, anti-inflammatory, and antibacterial effects in their leaves, roots, stems, latex, and fruits. In order to develop cosmetic materials based on natural products, we have studied on the skin activity of the ficin in latex as well as the whitening function of the fruit extract with 70% ethanol, and used it as a raw material for released cosmetic product. However, there is little research on the demand for the development of new eutectic solvent extraction methods and its ability to control skin inflammation and psoriasis regulation. Thus, in this study, we evaluated the effectiveness of fig fruit extracts and fractions using eutectic solvent extraction for skin inflammation control and psoriasis. First, fig fruits were extracted under optimal eutectic solvent conditions and fractionated with n-hexane, dichloromethane, ethyl acetate, and butanol. First, the antioxidant activity and inhibition of nitric oxide (NO) production were confirmed in mouse macrophage RAW264.7 cells. In addition, as a result of observing the mRNA expression through RT-PCR, pro-inflammatory cytokines such as TNF-α, IL1α, and IL-1β were suppressed significantly in the hexane, dichloromethane, and ethyl acetate fractions. In addition, it was confirmed in TNF-α stimulated HaCaT keratinocyte model. Finally, chemokine CC motif ligand 20 (CCL20), marker gene of human psoriasis skin disease, was significantly suppressed in the hexane, dichloromethane, and ethyl acetate fractions. These results suggested its anti-inflammatory and skin soothing effect and the possibility of development as an excellent skin soothing natural cosmetic material in the future through future clinical trials.

• Journal of Life Science
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• v.30 no.12
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• pp.1118-1127
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• 2020

Alzheimer's disease causes progressive neuronal loss that leads to cognitive disturbances. It is not currently curable, and there is no way to stop its progression. However, since medical treatment for Alzheimer's disease is most effective in the early stages, early detection can provide the best chance for symptom management. Biomarkers for the diagnosis of Alzheimer's disease include amyloid β (Aβ) deposition, pathologic tau, and neurodegeneration. Aβ deposition and phosphorylated tau can be detected by cerebrospinal fluid (CSF) analysis or positron emission tomography (PET). However, CSF sampling is quite invasive, and PET analysis needs specialized and expensive equipment. During the last decades, blood-based biomarker analysis has been studied to develop fast and minimally invasive biomarker analysis method. And one of the remarkable findings is the involvement of platelets as a primary source of Aβ in plasma. Aβ can be transported across the blood - brain barrier, creating an equilibrium of Aβ levels between the brain and blood under normal condition. Interestingly, a number of clinical studies have unequivocally demonstrated that plasma Aβ42/Aβ40 ratios are reduced in mild cognitive impairment and Alzheimer's disease. Together, these recent findings may lead to the development of a fast and minimally invasive early diagnostic approach to Alzheimer's disease. In this review, we summarize recent advances in the biomarkers of Alzheimer's disease, especially the involvement of platelets as a source of peripheral Aβ production and its potential as a blood-based biomarker.