• Journal of Embryo Transfer
• /
• v.27 no.2
• /
• pp.101-105
• /
• 2012

Linuron is a pesticide with a weak anti-androgenic property, which impacts male reproductive organs. In this study, to clarify whether linuron affects the cellular antioxidant system of ventral prostate, gene expression patterns of the representative antioxidant enzymes such as glutathione peroxidase (GPx), selenoprotein P (SePP), and superoxide dismutase (SOD) were investigated in the rat ventral prostates exposed to linuron using real-time RT-PCR analyses. Sprague-Dawley rats castrated at 6 weeks old were treated with linuron (25, 50, or 100 mg/kg per oral) daily for 10 days after testosterone propionate administration (0.4 mg/kg) subcutaneously. As compared to normal control animals, mRNA levels of phospholipid hydroperoxide GPx (PHGPx), SePP, and Mn SOD significantly increased in the prostates exposed to linuron (25, 50, and 100 mg/kg). However, cytosolic GPx (100 mg/kg) and Cu/Zn SOD (25, 50, and 100 mg/kg) mRNA levels significantly decreased in the ventral prostates. These results indicate that linuron upregulates the expressions of PHGPx, SePP, and Mn SOD mRNAs, but down-regulates the expressions of cytosolic GPx and Cu/Zn SOD in rat prostates, suggesting that linuron may have dual effects in the cellular antioxidant system of prostate.

• Proceedings of the Korean Society of Plant Pathology Conference
• /
• 2003.10a
• /
• pp.69.1-69
• /
• 2003

RSH (relA/spoT homolog) has been known to determine the level of guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp), which are the effector nucleotide of the prokaryotic stringent response and also play a role in antibiotic production and differentiation in Streptomyces species but not a little in eukaryotic organism, especially in plant. Salicylic acid (SA), a critical signal molecule of establishing systemic acquired resistance (SAR), could induce SAR in Pepper (Capcicum annuum) against Phytophthora capsici. And the extent of SAR induction was in proportion to the dosage of SA (or BTH). Suppression subtractive hybridization (SSH), a PCR-based method for cDNA subtraction, was carried out between SA-treated and non-SA-treated pepper leaves to isolate genes which may be responsible for defense signaling against pathogens. Early upregulated gene was selected from reverse northern and kinetics of SSH-genes transcripts in SA-treated pepper leaves upon SA treatment. Full-length cDNA of the gene (PepRSH； Pepper RelA / SpoT homolog) had an open reading frame (ORF) of 2166 bp encoding a protein of 722 amino acids and a significant homology with (p)ppGpp phosphohydrolase or synthetase. Genomic DNA gel blot analysis showed that pepper genome has at least single copy of PepRSH. PepRSH transcripts was very low in untreated pepper leaves but strongly induced by SA and methyljasmonic acid (MeJA), indicating that PepRSH may share common SA and MeJA-mediated signal transduction pathway Functional analysis in E. coli showed PepRSH confers phenotypes associated with (p)ppGpp synthesis through a complementation using active site mutagenesis.

• Biomolecules & Therapeutics
• /
• v.23 no.2
• /
• pp.141-148
• /
• 2015

Epithelial mesenchymal transition (EMT) is the first step in metastasis and implicated in the phenotype of cancer stem cells. Therefore, understanding and controlling EMT, are essential to the prevention and cure of metastasis. In the present study, we examined, by Western blot, reverse transcription polymerase chain reaction (RT-PCR), and confocal microscopy, the effects of cardamonin (CDN) on transforming growth factor-${\beta}1$ (TGF-${\beta}1$)-induced EMT of A549 lung adenocarcinoma cell lines. TGF-${\beta}1$ induced expression of N-cadherin and decreased expression of E-cadherin. CDN suppressed N-cadherin expression and restored E-cadherin expression. Further, TGF-${\beta}1$ induced migration and invasion of A549 cancer cells, which was suppressed by CDN. TGF-${\beta}1$ induced c-Jun N-terminal kinase (JNK) activation during EMT, but CDN blocked it. Protein serine/threonine phosphatase 2A (PP2A) expression in A549 cancer cells was reduced by TGF-${\beta}1$ but CDN restored it. The overall data suggested that CDN suppresses TGF-${\beta}1$-induced EMT via PP2A restoration, making it a potential new drug candidate that controls metastasis.

• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.16
• /
• pp.7277-7284
• /
• 2015

Background: The Wnt/${\beta}$-catenin signaling pathway is an important regulator of cellular functions such as proliferation, survival and cell adhesion. Wnt/${\beta}$-catenin signaling is associated with tumor initiation and progression; ${\beta}$-catenin mutations explain only 30% of aberrant signaling found in breast cancer, indicating that other components and/or regulation of the Wnt/${\beta}$-catenin pathway may be involved. Objective: We evaluated AXIN2 rs2240308 and rs151279728 polymorphisms, and expression profiles of ${\beta}$-catenin destruction complex genes in breast cancer patients. Materials and Methods: We collected peripheral blood samples from 102 breast cancer and 102 healthy subjects. The identification of the genetic variation was performed using PCR-RFLPs and DNA sequencing. RT-qPCR was used to determine expression profiles. Results: We found significant association of AXIN2 rs151279728 and rs2240308 polymorphisms with breast cancer risk. Significant increase was observed in AXIN2 level expression in breast cancer patients. Further analyses showed APC, ${\beta}$-catenin, CK1${\alpha}$, GSK3${\beta}$ and PP2A gene expression to be associated to clinic-pathological characteristics. Conclusions: The present study demonstrated, for the first time, that AXIN2 genetic defects and disturbance of ${\beta}$-catenin destruction complex expression may be found in breast cancer patients, providing additional support for roles of Wnt/${\beta}$-catenin pathway dysfunction in breast cancer tumorigenesis. However, the functional consequences of the genetic alterations remain to be determined.

• Journal of Broadcast Engineering
• /
• v.9 no.1
• /
• pp.83-90
• /
• 2004

DVB-MHP recommends that NPT(normal play time) be used as the times of stream events. NPT is the local time within an event(TV program). But we found that commercial transport stream (TS) generators and middlewares for DVB-MHP settop boxes are not ready to support the use of NPT by applications. In particular, TS generators do not create NPT reference descriptors needed to reconstruct NPT at the TV receiver. This situation is undesirable because program providers cannot experiment with the idea of synchronized applications. So we have implemented a TS generator that inserts NPT reference descriptors to TS and MyGetNPT API to approximately reconstruct NPT. STC (system time clock) is needed to reconstruct NPT, but Xlets are not allowed to read it. So, we approximate STC by using PCR (program clock reference) and the Java system tune. In this method, the stream generator extrats PCRs from an existing TS and inserts them into null TS packets in the form of MPEG sections, which can be read by Xlets. Because PCRs are displaced into new positions in TS, their values should be adjusted based on the time intervals between the original positions and the new positions. We implemented a synchronized application by using our TS generator and MyGetNPT API, where the task of stream events are to display graphic images. We found that graphic images are displayed where 240 ㎳ from their intended time, where 240ms is a human tolerance for the synchronization skew between graphic image and video.

• Proceedings of the PSK Conference
• /
• 2002.10a
• /
• pp.427.2-428
• /
• 2002

Previously we formulated new cationic liposomes, DDC, composed of DOTAP. DOPE, and cholesterol (Chol) in 1 : 0.7 : 0.3 molar ratios, and showed that DDC efficiently deliver the plasmid DNA into ovarian cancer cell lines. Here, wild type p53 DNA was transfected into ovarian cancer cells, using the DOC as a nonviral vector and the expression and activity of p53 gene were evaluated in vitro and in vivo. The complexes of plasmid DNA (pp53-EGFP) and DDC were transfected into OVCAR-3 cells. The gene expression was determined by RT -PCR and western blot analysis. (omitted)

• Journal of Korean Neurosurgical Society
• /
• v.59 no.3
• /
• pp.259-268
• /
• 2016

Objective : Accumulation of advanced glycation end-products (AGE) and mitochondrial glycation is importantly implicated in the pathological changes of the brain associated with diabetic complications, Alzheimer disease, and aging. The present study was undertaken to determine whether sildenafil, a type 5 phosphodiesterase type (PDE-5) inhibitor, has beneficial effect on neuronal cells challenged with AGE-induced oxidative stress to preserve their mitochondrial functional integrity. Methods : HT-22 hippocampal neuronal cells were exposed to AGE and changes in the mitochondrial functional parameters were determined. Pretreatment of cells with sildenafil effectively ameliorated these AGE-induced deterioration of mitochondrial functional integrity. Results : AGE-treated cells lost their mitochondrial functional integrity which was estimated by their MTT reduction ability and intracellular ATP concentration. These cells exhibited stimulated generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential, induction of mitochondrial permeability transition, and release of the cytochrome C, activation of the caspase-3 accompanied by apoptosis. Western blot analyses and qRT-PCR demonstrated that sildenafil increased the expression level of the heme oxygenase-1 (HO-1). CoPP and bilirubin, an inducer of HO-1 and a metabolic product of HO-1, respectively, provided a similar protective effects. On the contrary, the HO-1 inhibitor ZnPP IX blocked the effect of sildenafil. Transfection with HO-1 siRNA significantly reduced the protective effect of sildenafil on the loss of MTT reduction ability and MPT induction in AGE-treated cells. Conclusion : Taken together, our results suggested that sildenafil provides beneficial effect to protect the HT-22 hippocampal neuronal cells against AGE-induced deterioration of mitochondrial integrity, and upregulation of HO-1 is involved in the underlying mechanism.

• Biomolecules & Therapeutics
• /
• v.8 no.2
• /
• pp.153-159
• /
• 2000

A novel serine/threonine protein phosphatase with EF-hand motif, which belongs to PPEF family was partially cloned from rat brain cDNA by employing RT-PCR method. The size of the amplified clone was 1.6kbp. The amplified DNA was subcloned into pGEM-T-Easy vector and the resulting plasmid was maned as pGEM-rPPEF2. The nucleuotide sequence is shared by 88% with that of mouse PPEF-2 cDNA, and the deduced amino acid sequence reveal 92% homology with that of mouse PPEF-2 cDNA. The N-terminal region of the cloned rat brain PPEF contains a putative phosphatase catalytic domain (PP domain) and the C-terminal region contains multiple $Ca^{2+}$ binding sites (EF region). The putative catalytic domin (PP) and the EF-hand motif (EF) regions were subcloned into pGEX4T-1 and were overexpressed in E. coli DH5 as glutathione-S-transferase (GST) fusion proteins. Expression of the desired fusion protein was identified by SDS-PAGE and also by immunoblot analysis using monoclonal antibody against GST. The recombinant proteins were purified by glutathione-agarose chromatography. This report is first to demonstrate the cloning of PPEF family from rat brain tissues. The clone reported here would be invaluable for the investigation of the role of this new type-phosphatase in rat brain.

• Molecules and Cells
• /
• v.39 no.6
• /
• pp.508-513
• /
• 2016

To investigate the potential therapeutic effects of polyphenols in treating Pb induced renal dysfunction and intoxication and to explore the detailed underlying mechanisms. Wistar rats were divided into four groups: control groups (CT), Pb exposure groups (Pb), Pb plus Polyphenols groups (Pb+PP) and Polyphenols groups (PP). Animals were kept for 60 days and sacrificed for tests of urea, serum blood urea nitrogen (BUN) and creatinine. Histological evaluations were then performed. In vitro studies were performed using primary kidney mesangial cells to reveal detailed mechanisms. Cell counting kit-8 (CCK-8) was used to evaluate cell viability. Pb induced cell apoptosis was measured by flow cytometry. Reactive oxygen species (ROS) generation and scavenging were tested by DCFH-DA. Expression level of tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), interleukin-1-${\beta}$ (IL-1-${\beta}$) and IL-6 were assayed by ELISA. Western blot and qPCR were used to measure the expression of ERK1/2, JNK1/2 and p38. Polyphenols have obvious protective effects on Pb induced renal dysfunction and intoxication both in vivo and in vitro. Polyphenols reduced Pb concentration and accumulation in kidney. Polyphenols also protected kidney mesangial cells from Pb induced apoptosis. Polyphenols scavenged Pb induced ROS generation and suppressed ROS-mediated ERK/JNK/p38 pathway. Downstream pro-inflammatory cytokines were inhibited in consistency. Polyphenol is protective in Pb induced renal intoxication and inflammatory responses. The underlying mechanisms lie on the antioxidant activity and ROS scavenging activity of polyphenols.

• Radiation Oncology Journal
• /
• v.16 no.4
• /
• pp.377-388
• /
• 1998

Purpose : To evaluate changes in expression of cell cycle related genes during apoptosis induced in HL60 cells by X-irradiation to understand molecular biologic aspects in mechanism of radiation therapy. Material and Methods : HL-60 cell line (promyelocytic leukemia cell line) was grown in culture media and irradiated with 8 Gr by linear accelerator (6 MV X-ray). At various times after irradiation, ranging from 3 to 48 hours were analyzed apoptotic DNA fragmentation assay for apoptosis and by western blot analysis and semi-quantitative RT-PCR for expression of cell cycle related genes (cyclin A, cyclin B, cyclin C, cyclin Dl, cyclin E, cdc2, CDK2, CDK4, $p16^{INK4a}$, $p21^{WAF1}$, $p27^{KIP1}$, E2F, PCNA and Rb). Results : X-irradiation (8 Gy) induced apoptosis in HL-60 cell line. Cycline A protein increased after reaching its peak 48 h after radiation delivery and cyclin E, E2F, CDK2 and RB protein increased then decreased after radiation. Radiation induced up-regulation of the expression of E2F is due to mostly increase of Phosphorylated retinoblastoma proteins (ppRb). Cyclin Dl, PCNA, CDC2, CDK4 and $p16^{INK4a}$ protein underwent no significant change at any times after irradiation. There was not detected $p21^{WAF1}$ and $p27^{KIP1}$ protein. Cyclin A, B, C mRNA decreased immediately after radiation and then increased at 12 h after radiation. Cyclin Dl mRNA increased immediately and then decreased at 48 h after radiation. After radiation, cyclin E mRNA decreased with the lapse of time. CDK2 mRNA decreased at 3h and increased at eh after radiation. CDK4 mRNA rapidly increased at 6 to 12 h after radiation. There was no change of expression of $p16^{INK4a}$ and not detected in expressin of $p21^{WAF1}$ and $p27^{KIP1}$ mRNA. Conclusion : We suggest that entry into S phase may contribute to apoptosis of HL60 cells induced by irradiation. Increase of ppRb and decrease of pRb protein are related with radiation induced auoptosis of HL60 cells and tosis of HL60 cells induced by irradiation. Increase of ppRb and decrease of PRb protein are related with radiation induced apoptosis of HL60 cells and this may be associated with induction of E2F and cyclinE/CDK2. These results support that $p21^{WAF1}$ and $p27^{KIP1}$ are not related with radiation induced-apoptosis.