• Korean Journal of Pharmacognosy
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• v.46 no.1
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• pp.38-43
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• 2015

Ligularia stenocephala has a wide range of types of constituents with various pharmacological properties. Here in this study, we examined the effect of methanolic extract of L. stenocephala (MLS) on the lifespan and stress tolerance using Caenorhabditis elegans model system. We found that lifespan of wild-type worms was significantly lengthened in the presence of MLS in a dose dependent manner. MLS also elevated the tolerance of worms against osmotic, heat shock, and oxidative stress. We also demonstrated in vivo antioxidant capacity of MLS by checking intracellular reactive oxygen species levels as well as antioxidant enzyme activities such as catalase and superoxide dismutase. We further investigated several aging-related factors, including pharyngeal pumping rate and body length. Here, we showed that MLS exerts longevity effect independent of both factors. In addition, body movement of aged worms was significantly elevated, suggesting MLS could enhance healthspan as well as lifespan.

• Korean Journal of Veterinary Research
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• v.39 no.4
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• pp.797-805
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• 1999

T sergenti cDNA library were constructed to get a more broad information about the structural, functional or antigenic properties of the proteins, and analyzes for their partial cDNA sequences and expression sequences tags(ESTg). The mRNA were purified from T sergenti isolates to identify the information of antigen gene, then first and second strand cDNA was synthesized. EcoR I adaptor ligation and Xho I enzyme restriction were used to the synthesized cDNA, and ligated into a Uni-ZAP XR vector. T sergenti cDNA library was constructed with packaging and amplification in vitro. Antibody screening was performed with constructed T sergenti cDNA library using antisera against T sergenti. Among those clones, eight phagemids were rescued from the recombinant in vivo excision with f1 helper phage. Using the analysis of endonuclease restriction and PCR, the recombinant cDNA were proved having a 0.5-3.0kb of inserts. The eight of partial cDNA clones' sequences were obtained and examined for their homology using BLASTN and BLASTX. The eight of sequenced clones were classified into three groups according to the basis of database searches. A total 3,045bp of partial cDNA sequence were determined from six clones. The putatively identified clones contain a cytochrome c gene, a heat shock protein gene, a cyclophilin gene, and a ribosomal protein gene. The unidentified clones have a homology to ATP-binding protein(mtrA) gene of S argillaceus, DNA-binding protein(DBP) gene of Pseudorabies virus 85kDa merozoite protein gene of B bovis, mRNA spm1 protein of T annulata and glycine-rich RNA-binding protein mRNA of O sativa etc.

• Molecules and Cells
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• v.21 no.3
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• pp.381-388
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• 2006

Heat shock proteins (Hsp) 70 are a ubiquitous family of molecular chaperones involved in many cellular processes. A yeast strain, ssa1/2, with two functionally redundant cytosolic Hsp70s (SSA1 and SSA2) deleted shows thermotolerance comparable to mildly heatshocked wild type yeast, as well as increased protein synthesis and ubiquitin-proteasome protein degradation. Since mRNA abundance does not always correlate well with protein expression levels it is essential to study proteins directly. We used a gel-based approach to identify stress-responsive proteins in the ssa1/2 mutant and identified 43 differentially expressed spots. These were trypsin-digested and analyzed by nano electrospray ionization liquid chromatography tandem mass spectrometry (nESI-LC-MS/MS). A total of 22 non-redundant proteins were identified, 11 of which were confirmed by N-terminal sequencing. Nine proteins, most of which were up-regulated (2-fold or more) in the ssa1/2 mutant, proved to be stress-inducible proteins such as molecular chaperones and anti-oxidant proteins, or proteins related to carbohydrate metabolism. Interestingly, a translational factor Hyp2p up-regulated in the mutant was also found to be highly phosphorylated. These results indicate that the cytosolic Hsp70s, Ssa1p and Ssa2p, regulate an abundance of proteins mainly involved in stress responses and protein synthesis.

• Molecules and Cells
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• v.28 no.4
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• pp.331-339
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• 2009

Capsaicin is a very important secondary metabolite that is unique to Capsicum. Capsaicin biosynthesis is regulated developmentally and environmentally in the placenta of hot pepper. To investigate regulation of capsaicin biosynthesis, the promoter (1,537 bp) of pepper capsaicin synthase (CS) was fused to GUS and introduced into Arabidopsis thaliana (Col-0) via Agrobacterium tumefaciens to produce CSPRO::GUS transgenic plants. The CS was specifically expressed in the placenta tissue of immature green fruit. However, the transgenic Arabidopsis showed ectopic GUS expressions in the leaves, flowers and roots, but not in the stems. The CSPRO activity was relatively high under light conditions and was induced by both heat shock and wounding, as CS transcripts were increased by wounding. Exogenous capsaicin caused strong suppression of the CSPRO activity in transgenic Arabidopsis, as demonstrated by suppression of CS expression in the placenta after capsaicin treatment. Furthermore, the differential expression levels of Kas, Pal and pAmt, which are associated with the capsaicinoid biosynthetic pathway, were also suppressed in the placenta by capsaicin treatment. These results support that capsaicin, a feedback inhibitor, plays a pivotal role in regulating gene expression which is involved in the biosynthesis of capsaicinoids.

• Parasites, Hosts and Diseases
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• v.56 no.3
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• pp.237-245
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• 2018

Toxoplasma gondii can infect all the vertebrates including human, and leads to serious toxoplasmosis and considerable veterinary problems. T. gondii heat shock protein 60 (HSP60) is associated with the activation of antigen presenting cells by inducing initial immune responses and releasing inflammatory cytokines. It might be a potential DNA vaccine candidate for this parasite. A pVAX-HSP60 DNA vaccine was constructed and immune responses was evaluated in Kunming mice in this study. Our data indicated that the innate and adaptive immune responses was elicited by successive immunizations with pVAX-HSP60 DNA, showing apparent increases of CD3e+CD4+ and CD3e+CD8a+ T cells in spleen tissues of the HSP60 DNA-immunized mice ($24.70{\pm}1.23%$ and $10.90{\pm}0.89%$, P<0.05) and higher levels of specific antibodies in sera. Furthermore, the survival period of the immunized mice ($10.53{\pm}4.78day$) were significantly prolonged during the acute T. gondii infection. Decrease of brain cysts was significant in the experimental group during the chronic infection (P<0.01). Taken together, TgHSP60 DNA can be as a vaccine candidate to prevent the acute and chronic T. gondii infections.

• Molecular & Cellular Toxicology
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• v.1 no.4
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• pp.268-274
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• 2005

Bromobenzene (BB) is well known hepatotoxicant. Also, BB is an industrial solvent that arouses toxicity predominantly in the liver where it causes centrilobular necrosis. BB is subjected to Cytochrome P450 mediated epoxidation followed by either conjugation with glutathione, enzymatic hydrolysis or further oxidation. In this study, we focused on BB-induced gene expression at non-hepatotoxic dose. Mice were exposed to two levels of BB, sampled at 24 h, and hepatic gene expression levels were determined to evaluate dose dependent changes. When examining the toxic dose of BB treated group in other previous studies, genes related to heat shock protein, oxidative stress, and drug metabolism are expressed. Compared to these results, our study, in which non-toxic dose of BB was administrated, showed similar patterns as the toxic conditions above. The purpose of the study was to select genes that showed changes in relation to the differing dose through confirmation of the difference within transcriptomic boundaries, but those that are not detected by the existing classic toxicology tools in non-hepatotoxic dose.

• Reproductive and Developmental Biology
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• v.33 no.4
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• pp.237-242
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• 2009

Pregnancy is a unique event in which a fetus develops in the uterus despite being genetically and immunologically different from the mother, and the underlying mechanisms remain poorly understood. To analyze the differential gene expression profiles in nonpregnant and 7 days post coitus (dpc) pregnant uterus of mice, we performed a global proteomic study by 2-D gel electrophoresis (2-DE) and MALDI-TOF-MS. The uterine proteins were separated using 2-DE, Approximately 1,000 spots were detected on staining with Coomassie brilliant blue. An image analysis using Melanie III (Swiss Institute for Bioinformatics) was performed to detect variations in protein spots between pregnant and nonpregnant uterus. Twenty-one spots were identified as differentially expressed proteins, of which 10 were up-regulated proteins such as alpha-fetoprotein, chloride intracellular channel 1, transgelin, heat-shock protein beta-1, and carbonic anhydrase II, while 11 were down-regulated proteins such as X-box binding protein, glutathione S-transferase omega 1, olfactory receptor Olfr204, and metalloproteinase-disintegrin domain containing protein TECADAM. Most of the identified proteins appeared to be related with catabolism, cell growth, metabolism, regulation, cell protection, protein repair, or protection. Our results uncovered key proteins of mouse uterus involved in pregnancy.

• Reproductive and Developmental Biology
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• v.33 no.4
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• pp.243-248
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• 2009

To examine the differential expression of proteins during the cycling (70~80% confluences) and G0/G1 (full confluences) phases in porcine fetal fibroblast cells, we used a global proteomics approach by 2-D gel electrophoresis (2-DE) and MALDI-TOF-MS. Cycling cell were harvested at approximately 70% to 80% confluent state while cells in G0/G1 phase were recovered after maintenance of a confluent state for 48 hr. Cellular proteins with isoelectric points ranging between 3.0~10.0, were analyzed by 2-DE with 2 replicates of each sample. A total of approximately 700 spots were detected by 2.D gels stained with Coomassie brilliant blue. On comparing the cell samples obtained from the cycling and G0/G1 phases, a total of 13 spots were identified as differentially expressed proteins, of which 8 spots were up-regulated in the cycling cell and 5 were up-regulated in the G0/G1 phase. Differentially expressed proteins included K3 keratin, similar to serine protease 23 precursor, protein disulfide-isomerase A3, microsomal protease ER-60, alpha-actinin-2, and heat-shock protein 90 beta. The identified proteins were grouped on the basis of their basic functions such as molecular binding, catabolic, cell growth, and transcription regulatory proteins. Our results show expression profiles of key proteins in porcine fetal fibroblast cells during different cell cycle status.

• The Journal of the Korean Society for Microbiology
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• v.35 no.4
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• pp.299-308
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• 2000

Recently developed cDNA microarray or DNA chip technology allows expression monitoring of expression of hundreds and thousands of genes simultaneously and provides a format for identifying genes as well as changes in their activity. In order to search for changes in gene expression after X irradiation in HL60 cells, cDNA microarray technique was done. In this study, expression of 588 human genes (including oncogenes, tumor suppressor genes, cell cycle regulator genes, intracellular signal transduction modulator genes, apoptosis related genes, transcription factor genes, growth factors and receptor genes, cytokine genes, etc) were analyzed. For cDNA microarray analysis mRNAs were extracted from control and 8 Gy-irradiated HL60 cells. As a result the changes in expression of several genes were observed. This alteration of gene expression was confirmed by reverse transcription-polymerase chain reaction. The expression of heat shock 60 KD protein, c-jun, erythroid differentiation factor, CPP32, myeloid cell nuclear differentiation antigen, MAP kinase-activated protein kinase, interleukin-8, monocyte chemotactic peptide 1 and RANTES genes was increased, but the expression of p55CDC gene was decreased after X irradiation.

• Journal of Veterinary Clinics
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• v.27 no.1
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• pp.23-28
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• 2010

Mycobacterium avium ssp paratuberculosis, intracellular bacteria that can cause chronic granulomatous enteritis in cattle, continues to pose significant economic losses and health problem with high prevalence. The purpose of this study is the polymerase chain reaction (PCR)-base strategy for early detection of M. avium ssp paratuberculosis in whole blood. Blood samples were collected from korean cattles in Jeonbuk, Korea. The 16 out of 88 serum samples were detected M. partuberculosis by ELISA. Then samples of infected 8 Korean cattles were amplified by PCR. The PCR amplified targets are 16s rDNA and heat shock protein 65kDa (hsp 65). The 16s rDNA provided a highly sensitive and specific tool for the direct detection of mycobacteria. In addition M. avium was confirmed characteristically by the hsp65. Finally there were sure to M. avium ssp paratuberculosis by IS900 PCR. The restriction fragment length polymorphism was identified by PCR amplifications and subsequence restriction enzyme digestions with Pst I of a hsp65. These results indicate that confirm M. avium with 16s rDNA, hsp65 and a restriction fragment length polymorphism in the hsp65 gene can be seem the other pattern. Therefore, these results can be used for clinical direct detections of M. avium ssp paratuberculosis in whole blood of Korean cattle and also to be used epidemiological researches.