• The Korean Journal of Food And Nutrition
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• v.26 no.3
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• pp.485-491
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• 2013

Rhodiola spp. and red ginseng have been used for food and medicinal applications in disease chemoprevention in many Asian countries. Increased oxidative stress by reactive oxygen species (ROS) has been proposed to be a major cause of muscle fatigue. The present study was designed to investigate the protective effects of a fermented hot-water extract mixture from Rhodiola sachalinensis and red ginseng (MFR) on cell damage and the antioxidant enzyme system in $H_2O_2$-induced oxidative stress in skeletal muscle cells. C2C12 myoblasts were treated with various concentrations of NFR (non-fermented Rhodiola sachalinensis extract), FR (fermented hot-water extract from Rhodiola sachalinensis) and MFR for up to 5 days after the standard induction of differentiation, followed by semi-quantitative RT-PCR. MFR treatment dose-dependently protected oxidative damage of C2C12 cells. The treatment with MFR also enhanced mRNA expressions of MyoD, Cu/Zn SOD, Mn-SOD and GPX up to 16%. These results indicate that MFR exerts an anti-oxidative effect through a mechanism (s) that may involve the up-regulation of antioxidant enzymes, which may be important for the cellular redox environment in muscle cells.

• Toxicological Research
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• v.6 no.2
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• pp.205-213
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• 1990

The regulation of neurotoxicants has usually been based upon setting reference doses by dividing a no observed adverse effect level (NOAEL) by uncertainty factors that theoretically account for interspecies and intraspecies extraploation of experimental results in animals to humans. Recently, we have proposed a four-step alternative procedure which provides quantitative estimates of risk as a function of dose. The first step is to establish a mathematical relationship between a biological effect or biomarker and the dose of chemical administered. The second step is to determine the distribution (variability) of individual measurements of biological effects or their biomarkers about the dose response curve. The third step is to define an adverse or abnormal level of a biological effect or biomarker in an untreated population. The fourth and final step is to combine the information from the first three steps to estimate the risk (proportion of individuals exceeding on adverse or abnormal level of a biological effect or biomarker) as a function of dose. The primary purpose of this report is to enhance the certainty of the first step of this procedure by improving our understanding of the relationship between a biomarker and dose of administered chemical. Several factors which need to be considered include: 1) the pharmacokinetics of the parent chemical, 2) the target tissue concentrations of the parent chemical or its bioactivated proximate toxicant, 3) the uptake kinetics of the parent chemical or metabolite into the target cell(s) and/or membrane interactions, and 4) the interaction of the chemical or metabolite with presumed receptor site(s). Because these theoretical factors each contain a saturable step due to definitive amounts of required enzyme, reuptake or receptor site(s), a nonlinear, saturable dose-response curve would be predicted. In order to exemplify this process, effects of the neurotoxicant, methlenedioxymethamphetamine (MDMA), were reviewed and analyzed. Our results and those of others indicate that: 1) peak concentrations of MDMA and metabolites are ochieved in rat brain by 30 min and are negligible by 24 hr, 2) a metabolite of MDMA is probably responsible for its neurotoxic effects, and 3) pretreatment with monoamine uptake blockers prevents MDMA neurotoxicity. When data generated from rats administerde MDMA were plotted as bilolgical effect (decreases in hippocampal serotonin concentrations) versus dose, a saturation curve best described the observed relationship. These results support the hypothesis that at least one saturable step is involved in MDMA neurotoxicity. We conclude that the mathematical relationship between biological effect and dose of MDMA, the first step of our quantitative neurotoxicity risk assessment procedure, should reflect this biological model information generated from the whole of the dose-response curve.

• Toxicological Research
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• v.32 no.1
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• pp.73-80
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• 2016

Chronic exposure to cadmium (Cd) is known to adversely affect renal function. Our previous studies indicated that Cd induces p53-dependent apoptosis by inhibiting gene expression of the ubiquitin-conjugating enzyme (Ube) 2d family in both human and rat proximal tubular cells. In this study, the effects of Cd on protein expression of p53 and apoptotic signals in the kidney and liver of mice exposed to Cd for 12 months were examined, as well as the effects of Cd on p53 protein levels and gene expression of the Ube2d family in various cell lines. Results showed that in the kidney of mice exposed to 300 ppm Cd for 12 months, there was overaccumulation of p53 proteins in addition to the induction of apoptosis, which was triggered specifically in the proximal tubules. Interestingly, the site of apoptosis was the same as that of p53 accumulation in the proximal tubules. In the liver of mice chronically exposed to Cd, gene expression of the Ube2d family tended to be slightly decreased, together with slight apoptosis without the accumulation of p53 protein. In rat small intestine epithelial (IEC-6) cells, Cd decreased not only the p53 protein level but also gene expression of Ube2d1, Ube2d2 and Ube2d4. In human brain microvascular endothelial cells (HBMECs), Cd did not suppress gene expression of the Ube2d family, but increased the p53 protein level. In human brain astrocytes (HBASTs), Cd only increased gene expression of UBE2D3. These results suggest that Cd-induced apoptosis through p53 protein is associated with renal toxicity but not hepatic toxicity, and the modification of p53 protein by Cd may vary depending on cell type.

• Journal of Chest Surgery
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• v.33 no.8
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• pp.648-654
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• 2000

Background: Cardiopulmonary bypass during open heart surgery causes systemic inflammatory respose. IL-10 is an anti-inflammatory cytokine that inhibits inflammatory process and protects organ function by down regulation of pro-inflammatory cytokine release and maintenance of blood level balance with pro-inflammatory cytokines. Mateial and Method: Plasma IL-10 levels were measured and analyzed in 22 patients who underwent open heart surgery (11 cases of coronary artery bypass graft, 11 cases of valve replacement) under cardiopulmonary bypass since 1988 January to July at Department of Thoracic and Czardiovascular surgery, Yeungnam University Hospital. 1g of methylprednisolone was administrated to thirteen patients randomly. Blood samp.es were taken and collected at the time of induction of anesthesia, 10 min before cardiopulmonary bypass, 10 min after starting of CPB, 10 min aftr aortic cross clamping, 10 min after ACC release, and 10 min, 2 hours, and `5 hours after CPB respectively. The plasma levels of IL-10 were determined by enzyme-linked immunosorbent assays(ELISA). Wilcoxon-Raule Sum test was used for statistical analysis. Result: In all 22 patients, cardiopulmonary bypass time was used for statistical analysis. Result: In all 22 patients, cardiopulmonary bypass time was 171$\pm$41.4 min and aortic cross clamp time was 118$\pm$36.5 min. Peak IL-10 level was achieved at 10 min after ACC(361.0$\pm$52.81pg/ml) and was decreased sharply at 2 hours after CPB. Peak IL-10 level was correlated positively with aortic cross clamp time(p=0.011); however, it did not correlated with bypass time(p=0.181). In valve replacement group, mean IL-10 level at peak point was 567.89$\pm$107.69 pg/ml and was significantly higher than that of coronary artery bypass group(205.67$\pm$192.70 pg/ml)(p<0.001). ACC time in valve replacement group was significantly longer than that of coronary artery bypass group(p<0.01), however, bypass time was not(p=0.212). Thirteen patients with steroid pretreatment before starting of CPB showed relatively higher plasma IL-10 level than in control group, however, no statistical significance was noted(p=0.19). Conclusion: plasma level of IL-10 was increased in association with cardiopulmonary bypass and revealed peak at 10 min after ACC release. IL-10 level was correlated positively with ACC time. Therefore, systemic inflammatory respeonse in association with cardiopulmonary bypass could be decreased by reducing ACC time during cardiac surgery.

• Development and Reproduction
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• v.11 no.3
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• pp.195-203
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• 2007

Sex determination and sex differentiation are influenced by genotype in many gonochoristic fish. Cytochrome P450 aromatase (CYP19) is the terminal enzyme in steridogenic pathway that converts androgens into estrogens. In this study, partial fragments of aromatase genes (ovarian aromatase, P450aromA and brain aromatase, P450aromB) were cloned and sequenced in Korean rockfish (Sebastes schlegeli), and gene specific primers were designed based on their sequences. Using these primers, aromatase gene expression during sex differentiation was investigated by RT-PCR. Expression of these aromatase genes were detected both in the head and body parts at 35 dab (days after birth). The number of fish that expressed the aromatase genes decreased at 52 dab, implying down-regulation of these genes. However, these genes were expressed at 59 dab in almost all fish studied here. The expression patterns of both genes are similar throughout the investigated period except for 45 dab where the expression of P450aromB was detected in more fish than that of P450aromA both in the head and body parts. Timing of sex differentiation in this species has been shown to be at around $50{\sim}65$ dab by histological analysis. However, the results from this study suggest that sex differentiation of rockfish may take place $1{\sim}2$ weeks earlier than the period proposed previously. The results also suggest that the mechanism of sex differentiation in viviparous fish may be similar to that in oviparous fish in terms of the importance of aromatase action during the critical period.

• Food Science and Biotechnology
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• v.18 no.5
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• pp.1204-1211
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• 2009

Hericium erinaceus is an edible mushroom used as a medicinal food in Asian countries. In this study, the chemopreventive effects of H. erinaceus mycelia hot water extract (HEW) were evaluated. HEW remarkably induced the luciferase activity of the antioxidant response element (ARE), located in the promoter region of phase 2 and antioxidant genes and regulated by nuclear factor E2-related factor 2 (Nrf2). The up-regulation of ARE activity by HEW corresponded with the induction of Nrf2 and the antioxidant enzyme, hemeoxygenase-1. The inhibition of cyclooxygenase-2 (COX-2) activity is a promising effective approach in cancer chemoprevention, and HEW prominently suppressed COX-2 protein expression in HepG2 cells. Furthermore, HEW showed anti-inflammatory activity by modulating inflammatory mediators such as nitric oxide (NO), inducible NO synthase, tumor necrosis factor-${\alpha}$, interleukin-$1{\beta}$, and the transcription factor, nuclear factor-${\kappa}B$, in lipopolysaccharide-stimulated RAW 264.7 cells. These results suggest that H. erinaceus possessed anti-tumor and anti-inflammatory effects via the modulation of Nrf2/ARE and inflammatory signaling pathways, and may therefore have potential use as a natural chemopreventive agent.

• Journal of Life Science
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• v.24 no.1
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• pp.1-7
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• 2014

Eukaryotic gene expression is an important process, which is initiated by several transcription factors and RNA polymerases that occupy the promoter region of genomic DNA. Although there are many experiments to identify the promoter region in a gene, it is time and labor consuming to finalize it. In this study, we utilized bioinformatic programs, including Ensembl, NCBI, and CpG plots, to identify the cloning promoter region in SETDB1 genomic DNA. We performed PCR amplification to obtain the SETDB1 promoter on an approximately 2 kb region upstream from the TSS named SETDB1-P1. The PCR product was ligated with TA cloning vectors, and we confirmed the insert size using restriction enzyme digestion. Sequentially, the insert was subcloned into a pGL3-luc vector to produce pGL3-SETDB1- P1-luc and then confirmed by DNA sequencing. We also obtained a fragmented PCR product called P2 and P3 and performed a luciferase assay using pGL3-SETDB1-P1-luc transfection. We found that several anticancer drugs, including taxol, 4-FU, and doxorubicin, decreased the promoter activity of SETDB1. We obtained consistent data on the regulation of SETDB1 gene expression after anticancer drug treatment using Western blot analysis and RT-PCR. Our results suggest that promoter cloning of the human SETDB1 gene utilizing bioinformatics is a very useful and timesaving approach to study gene expression.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1103-1109
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• 2018

Objective: This study aimed to screen and identify the target genes of miR-375 in pig Sertoli (ST) cells and to elucidate the effect of miR-375 on the proliferation of ST cells. Methods: In this study, bioinformatics software was used to predict and verify miR-375 target genes. Quantitative polymerase chain reaction (PCR) was used to detect the relationship between miR-375 and its target genes in ST cells. Enzyme-linked immunosorbent assay (ELISA) of rearranged L-myc fusion (RLF) and hypoxia-induced gene domain protein 1A (HIGD1A) was performed on porcine ST cells, which were transfected with a miR-375 mimics and inhibitor to verify the results. Dual luciferase reporter gene assays were performed to assess the interactions among miR-375, RLF, and HIGD1A. The effect of miR-375 on the proliferation of ST cells was analyzed by CellTiter 96 AQueous One Solution Cell Proliferation Assay (MTS). Results: Five possible target genes of miR-375, including RLF, HIGD1A, colorectal cancer associated 2, POU class 3 homeobox 1, and WW domain binding protein 1 like, were found. The results of quantitative PCR suggested that mRNA expression of RLF and HIGD1A had a negative correlation with miR-375, indicating that RLF and HIGD1A are likely the target genes of miR-375. The ELISA results revealed that RLF and HIGD1A were negatively correlated with the miR-375 protein level. The luminescence results for the miR-375 group cotransfected with wild-type RLF and HIGD1A vector were significantly lower than those of the miR-375 group co-transfected with the blank vector or mutant RLF and HIGD1A vectors. The present findings suggest that RLF and HIGD1A are target genes of miR-375 and that miR-375 inhibits ST cell proliferation according to MTS analysis. Conclusion: It was speculated that miR-375 affects cell proliferation through its target genes, which play an important role in the development of testicular tissue.

• Clinical and Experimental Reproductive Medicine
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• v.43 no.2
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• pp.82-89
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• 2016

Objective: The long interspersed elements (LINE-1, L1s) are a group of genetic elements found in large numbers in the human genome that can translate into phenotype by controlling genes. Growing evidence supports the role of epigenetic in polycystic ovary syndrome (PCOS). The purpose of this study is to evaluate the DNA methylation levels in LINE-1 in a tissue-specific manner using cumulus cells from patients with PCOS compared with normal controls. Methods: The study included 19 patients with PCOS and 22 control patients who were undergoing controlled ovarian hyperstimulation. After oocyte retrieval, cumulus cells were extracted. LINE-1 DNA methylation levels were analysed by bisulfite treatment, polymerase chain reaction, and restriction enzyme digestion. The Connection Up- and Down-Regulation Expression Analysis of Microarrays software package was used to compare the gene regulatory functions of intragenic LINE-1. Results: The results showed higher LINE-1 DNA methylation levels in the cumulus cells of mature oocytes in PCOS patients, 79.14 (${\pm}2.66$) vs. 75.40 (${\pm}4.92$); p=0.004, but no difference in the methylation of cumulus cells in immature oocytes between PCOS and control patients, 70.33 (${\pm}4.79$) vs. 67.79 (${\pm}5.17$); p=0.155. However, LINE-1 DNA methylation levels were found to be higher in the cumulus cells of mature oocytes than in those of immature oocytes in both PCOS and control patients. Conclusion: These findings suggest that the epigenetic modification of LINE-1 DNA may play a role in regulating multiple gene expression that affects the pathophysiology and development of mature oocytes in PCOS.

• Journal of Ginseng Research
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• v.40 no.4
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• pp.423-430
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• 2016

Background: The induction of cellular defensive genes such as phase II detoxifying and antioxidant enzymes is a highly effective strategy for protection against carcinogenesis as well as slowing cancer development. Transcription factor Nrf2 (nuclear factor E2-related factor 2) is responsible for activation of phase II enzymes induced by natural chemopreventive compounds. Methods: Red ginseng oil (RGO) was extracted using a supercritical $CO_2$ extraction system and chemical profile of RGO was investigated by GC/MS. Effects of RGO on regulation of the Nrf2/antioxidant response element (ARE) pathway were determined by ARE-luciferase assay, western blotting, and confocal microscopy. Results: The predominant components of RGO were 9,12-octadecadienoic acid (31.48%), bicyclo[10.1.0] tridec-1-ene (22.54%), and 22,23-dihydrostigmasterol (16.90%). RGO treatment significantly increased nuclear translocation of Nrf2 as well as ARE reporter gene activity, leading to upregulation of heme oxygenase-1 and NAD(P)H:quinone oxidoreductase 1. Phosphorylation of the upstream kinases such as apoptosis signal-regulating kinase (ASK)1, mitogen-activated protein kinase (MAPK) kinase (MKK)4/7, c-Jun N-terminal kinase (JNK), and p38 MAPK were enhanced by treatment with RGO. In addition, RGO-mediated Nrf2 expression and nuclear translocation was attenuated by JNK inhibitor SP600125 and p38 MAPK inhibitor SB202190. Conclusion: RGO could be used as a potential chemopreventive agent, possibly by induction of Nrf2/ARE-mediated phase II enzymes via ASK1-MKK4/7-JNK and p38 MAPK signaling pathways.