• Journal of Life Science
• /
• v.27 no.8
• /
• pp.879-887
• /
• 2017

It is well established that brain-derived neurotrophic factor (BDNF) is expressed not only in the brain but also in skeletal muscle, and is required for normal neuromuscular system function. Histone deacetylases (HDACs) and insulin-like growth factor-I (IGF-I) are potent regulators of skeletal muscle myogenesis and muscle gene expression, but the mechanisms of HDAC and IGF-I in skeletal muscle-derived BDNF expression have not been examined. In this study, we examined the effect of IGF-I and suberoylanilide hydroxamic acid (SAHA), a pan-HDAC inhibitor, on BDNF induction. Proliferating or differentiating C2C12 skeletal muscle cells were treated with increasing concentrations (0-50 ng/ml) of IGF-I in the absence or presence of $5{\mu}M$ SAHA for various time periods (3-24 hr). Treatment of C2C12 cells with IGF-I resulted in a dose- and time-dependent decrease in BDNF mRNA expression. However, inhibition of HDAC led to a significant increase in the expression of BDNF mRNA levels. In addition, immunocytochemistry revealed high BDNF protein levels in undifferentiated C2C12 skeletal muscle cells, whether untreated, IGF-I-treated, or exposed to SAHA. These results represent the first evidence that IGF-I can suppress the mRNA and protein expression of BDNF; conversely, SAHA attenuates the effects of IGF-I. Consequently, SAHA upregulates BDNF expression in C2C12 skeletal muscle cells.

• Korean Journal of Food Science and Technology
• /
• v.46 no.6
• /
• pp.762-768
• /
• 2014

In this study, the ameliorating effects of lactic acid-fermented garlic extract (LAFGE) on non-alcoholic fatty liver were investigated using oleic acid-induced steatotic HepG2 cells. The ameliorating mechanism was analyzed by RT-PCR and Western blot. Treatment with 1 mg/mL LAFGE decreased intracellular lipid accumulation approximately 1.5-fold, compared to that achieved with non-fermented garlic extract. LAFGE reduced fatty acid influx into hepatocytes through down-regulation of FAT/CD36 mRNA expression in the steatotic HepG2 cells. $PPAR{\alpha}$ and CPT-1 mRNA expression was significantly up-regulated by LAFGE treatment of HepG2 cells as a consequence of activation of beta oxidation. Additionally, the treatment with 1 mg/mL LAFGE highly down-regulated mRNA expression of SREBP-1c and FAS to 51% and 35%, respectively. LAFGE showed concentration-dependent down-regulation patterns in protein expression of SREBP-1c and FAS, as determined by Western blot. These results suggest that LAFGE treatment improves hepatic steatosis triggered by the imbalance of hepatic lipid metabolism owing to oleic acid treatment.

• Reproductive and Developmental Biology
• /
• v.32 no.3
• /
• pp.153-158
• /
• 2008

Tissue-specific and temporal regulation of milk protein gene expression is advantageous when creating transgenic animal that produces foreign protein into milk. Gene expression, i.e. protein production, is regulated not only by promoter strength but also mRNA stability. Especially, poly A tail length by polyadenylation affects in vivo and in vitro mRNA stability and translation efficiency of the target gene. In the present study, nucleotide sequence of 3'-UTR was analyzed to evaluate the effects of mRNA stability on the target gene expression. Based on the poly A signal of 3' -untranslated region (UTR), nucleotide sequences of putative cytoplasmic polyadenylation elements (CPEs) and downstream elements (DSEs: U-rich, G-rich, GU-rich) were analyzed and used to construct 15 luciferase reporter vectors. Each vector was transfected to HC11 and porcine mammary gland cell (PMGC) and measured for dual luciferase expression levels after 48 hours of incubation. Luciferase expression was significantly higher in construct #6 (with CPE 2, 3 and DSE 1 of exon 9) and #11 (with CPE 2, 3 and DSE 1, 2 and 3 of exon 9) than construct #1 in the PMGC. These results suggest that expression of target genes in PMGC may be effectively expressed by using the construct #6 and #11 on production of transgenic pig.

• Journal of Life Science
• /
• v.19 no.11
• /
• pp.1637-1643
• /
• 2009

Shikonin, a component of Lithospermum erythrorhizon Sieb. et Zucc, exerts various characteristics such as anti-inflammatory, anti-cancer and anti-obesity functions. To elucidate the molecular mechanism of shikonin-induced inhibition of adipogenesis, we analyzed the mRNA expression level of various adipogenesis-related factors including C/EBPs (CCAAT/enhancerbinding proteins) and $PPAR{\gamma}$ (peroxisome proliferator-activated receptor $\gamma$). The data showed that mRNA expressions of C/$EBP{\beta}$ and C/$EPB{\delta}$ were only slightly changed by shikonin treatment, but mRNA expressions of $PPAR{\gamma}$ and C/$EPB{\alpha}$ were significantly down-regulated. Then, we tested whether upstream regulators of C/$EBP{\beta}$ and $PPAR{\gamma}$ were involved in anti-adipogenesis of shikonin. C/$EBP{\gamma}$ and CHOP (C/EBP homologous protein), which are upstream regulators of C/$EBP{\beta}$, were not affected by shikonin treatment. On the contrary, the mRNA level of KROX20 was markedly down-regulated by shikonin treatment. These results suggest that KROX20 might regulate downstream factors of adipogenesis through C/$EBP{\beta}$-independent pathway. The expression of KLF15 (Kruppel-like factor15), which is a member of KLF family and is a upstream regulator of $PPAR{\gamma}$, was dramatically decreased by shikonin treatment, but KLF2 was not changed. Shikonin had no impact on the expression of KLF5 in the early stage of adipogenesis, but shikonin increased expression of KLF5 in the late stage of adipogenesis. Even though mRNA expression of KLF5 was moderately changed by shikonin treatment, its effect may be small compared with the effect of KLF15, which was markedly inhibited. Taken together, these results suggest that shikonin inhibits adipogenesis through the down-regulation of $PPAR{\gamma}$ and C/$EPB{\alpha}$, which is mediated by the down-regulation of two pro-adipogenic factors, KROX20 and KLF15.

• Journal of Life Science
• /
• v.25 no.10
• /
• pp.1098-1102
• /
• 2015

FABPpm (plasma membrane-bound fatty acid binding protein ) is highly expressed in skeletal muscle. The principal role of this protein is modulating fatty acid uptake and metabolism. The influence of insulin-like growth factor-I (IGF-I), which is a major regulator of skeletal muscle cells, on FABPpm in skeletal muscle cells has not been investigated. To determine the effect of IGF-I on the expression of FABPpm, differentiated C2C12 murine skeletal muscle cells were treated with 20 ng/ml of IGF-I for different times. IGF-I increased the expression of FABPpm in a time-dependent manner. The mRNA level of FABPpm was measured by real-time quantitative PCR to determine whether the IGF-1-induced induction of FABPpm was regulated pretranslationally. The IGF-I treatment resulted in very rapid induction of the FABPpm mRNA transcript in the C2C12 myotubes. After 24 and 48 hr of the IGF-I treatment, FABPpm mRNA increased 130 and 179%, respectively. The increase in the protein expression returned to control levels after 72 hr of the IGF-I treatment, suggesting that IGF-1 regulated the FABPpm gene pretranslationally in skeletal muscle cells. This is the first evidence that IGF-I has a modulatory effect on the expression of FABPpm. In conclusion, IGF-I induced rapid transcriptional modification of the FABPpm gene in C2C12 skeletal muscle cells and exerted modulatory effects on FABPpm.

• Development and Reproduction
• /
• v.4 no.1
• /
• pp.101-108
• /
• 2000

Objectives: The pleiotrophin (PTN) and midkine (MK) are secreted heparin-binding neurokines that share 50％ sequence homology. PTN and MK are expressed in the range of primary human tumors. The association of PTN and MK with carcinogenesis, enhancement of plasminogen activator activity and angiogenic factor are reported. Patients with endometriosis are characterized by the ability of the endometrium to implant; angiogenic and growth factors may play a significant role in the pathogenesis of endometriosis. To test the hypothesis that higher expression of PTN and MK in endometrium from women with endometriosis might be increase angiogenesis and growth ectopic endometriosis implants, we investigated PTN and MK expression by quantitative and competitive polymerase chain reaction (QC-PCR) in endometrium from women with and without endometriosis throughout the menstrual cycle. Design: MK and PTN mRNA expression in endomeoium from women with endometriosis and control patients without endometriosis were determined by QC-PCR throughout the menstrual cycle. Methods: Endometrial tissue was obtained from 25 patients with severe endometriosis and 30 patients without endometriosis undergoing hysterectomy or endometrial biopsy. Stage of endometrial cycle and a diagnosis of endometriosis were confirmed histologically. Total RNA was extracted and reverse transcribed into c-DNA. QC-PCR was performed to evaluate PTN and MK mRNA expression. Results were analysed by Post Hoc test. Results: MK and PTN were expressed throughout the menstrual cycle in both groups. MK expression was higher in follicular phase than luteal phase in endometrium from normal women. endometrium from endometriosis patients showed increased expression of PTN and MK compared to endometrium from normal women in the luteal phase (p<0.05). Conclusion: Our results suggest that uterine endometrium from women with endometriosis expresses higher levels of MK and PTN than endometrium from normal women during luteal phase. Increased MK and PTN expression may be related to the initiation of ectopic endometrial implants and their subsequent peritoneal invasion.

• Journal of Life Science
• /
• v.24 no.11
• /
• pp.1180-1186
• /
• 2014

Autophagy, the lysosomal degradation pathway, is an intracellular recycling system that is necessary for the metabolic benefits of exercise and for producing lasting beneficial effects of exercise in various diseases. However, the most recent studies have only examined the effect of a single bout of exercise or resistance exercise on autophagic responses. To determine the differential effects of acute and chronic exercise on the expression of autophagy-related genes in D. melanogaster, white-eyed mutant D. melanogaster were assigned randomly to four groups: control, acute exercise, 2 hr chronic exercise, and 3 hr chronic exercise. The flies were exercised using a mechanized platform known as the Power Tower. Our results revealed that a single bout of exercise resulted in increased mRNA levels of the Atg8a gene (~20%, p<0.05). However, Atg1 and Atg6 mRNA expression were not induced by acute exercise. Transcript levels of Atg6 (~29%, p<0.05) related to the nucleation of autophagosomes were significantly induced by 2 hr of chronic exercise. However, this chronic exercise was not enough to increase Atg1 and Atg8a mRNA expression. On the other hand, 3 hr of exercise for 7 days significantly increased Atg1, Atg6, and Atg8a gene expression-about 57%, 37%, and 71%, respectively (p<0.05). These results suggest that a single bout of exercise is not enough to induce full activation of selected autophagy-related genes in D. melanogaster. Our results demonstrated that chronic regular exercise induced autophagy-related gene expression, suggesting that chronic regular exercise training might be required to activate autophagic responses important for producing beneficial effects of exercise in various diseases.

• Radiation Oncology Journal
• /
• v.18 no.4
• /
• pp.321-328
• /
• 2000

Purpose :NOS2 induce NO Production and NO activate TGF-${\beta}$. The TGF-${\beta}$ is a inhibitor of NOS2. If this negative feedback mechanism operating in radiation pneumonitis model, NOS2 inhibitor may play a role in TGF-${\beta}$ suppression. We planned this study to evaluate the expression patterns of NO, NOS2 and TGF-${\beta}$ in vivo radiation pneumonitis model. Materials and Methods : Sixty sprague-Dawley rat were irradiated 5 Gy or 20 Gy. They were sacrificed 3, 7, 14, 28 and 56 days after irradiation. During sacrifice, we peformed broncho-alveolar lavage (BAL). The BAL fluids were centrifuged and supernatents were used for measure NO and TGF-${\beta}$, and the cells were used for RT-PCR. Results : After 5 Gy of radiation, NO in BAL fluid increased at 28 days in both lung and TGF-${\beta}$ in left lung at 56 days. NO increased in BAL fluid at 28 days in both lung after irradiation and TGF-${\beta}$ in right lung at 28-56 days after 20 Gy of radiation. After 5 Gy of radiation, NOS2 expression was increased in right lung at 14 days, in both lung at 28 days and in left lung at 56 days. TGF-${\beta}$ expression was reduced in both lung at 28 days and increased in left lung at 56 days. Conclusions :The Proposed feedback mechanism of NO, NOS2 and TGF-${\beta}$ was operated in vivo radiation pneumonitis model. At 56 days, however, NOS2 and TGF-${\beta}$ expressed concurrently in left lung after 5 Gy and in both lung after 20 Gy of radiation.

• Journal of Korean Ophthalmic Optics Society
• /
• v.13 no.3
• /
• pp.95-101
• /
• 2008

Purpose: Hydrogen peroxide which is one of the reactive oxygen species has been seen to cause various diseases, various cellular disinfections, gene transformation and cell death. The goals of this study were to determine the protective effect of EGCG against $H_2O_2$-induced apoptotic death in conjunctival cell lines. Methods: We measured cell viability by MTT assay and analyzed DNA fragmentation to check up a distinctive feature in cell death and measured the removal ability of free radicals by DPPH free radical scavenging assay and evaluated the oxygen free radical's quantity in the cell by DCFH-DA assay. The mRNA expression in the cell were examined by RT-PCR. Results: Cell viability and free radical scavening activites was significantly increased in dose dependently after cell was exposed to EGCG. And DNA fragmentation and intracellular ROS was decreased. It was showed the mRNA expression which increase of bcl-2, bcl-xL expression and decrease of bax expression. Conclusions: From these results, it suggests that EGCG has an antioxidant effect and protects conjunctival cell lines from the $H_2O_2$-mediated apoptosis through the modulation of the mRNA expression.

• Journal of Society of Preventive Korean Medicine
• /
• v.11 no.1
• /
• pp.9-21
• /
• 2007

This study was performed to evaluate the effect of Carthami tinctorius(HH) on osteoblast function and gene expression. The osteoblasts separated from the rat calvariae were cultivated to evaluate the cell function, and MG-63 cell was also cultivated for the test of mRNA synthesis. In this experiments, cell proliferation of rat calvarial cells was increased by HH. PKC activity, intracellular free calcium level and collgen synthesis from calvarial cells were increased by HH, but not PKA activity. And the mRNA of $PLA_2$, COX-2, and $PGE_2$ synthase from MG-63 were decreased by HH, but the mRNA of prostacyclin synthase was increased. It is concluded that HH might increase the proliferation of calvarial cell resulted from augumentation of osteoblast activity and its mRNA synthesis.