• Journal of Korean Neurosurgical Society
• /
• v.48 no.1
• /
• pp.20-30
• /
• 2010

Objective : We determined whether the expression of GRIM-19 is correlated with pathologic types and malignant grades in gliomas, and determined the function of GRIM-19 in human gliomas. Methods : Tumor tissues were isolated and frozen at $-80^{\circ}C$ just after surgery. The tissues consisted of normal brain tissue (4), astrocytomas (2), anaplastic astrocytomas (2), oligodendrogliomas (13), anaplastic oligodendrogliomas (11), and glioblastomas (16). To profile tumor-related genes, we applied RNA differential display using a $Genefishing^{TM}$ DEG kit, and validated the tumor-related genes by reverse transcription polymerase chain reaction (RT-PCR). A human glioblastoma cell line (U343MG-A) was used for the GRIM-19 functional studies. The morphologic and cytoskeletal changes were examined via light and confocal microscopy. The migratory and invasive abilities were investigated by the simple scratch technique and Matrigel assay. The antiproliferative activity was determined by thiazolyl blue Tetrazolium bromide (MTT) assay and FACS analysis. Results : Based on RT-PCR analysis, the expression of GRIM-19 was higher in astrocytic tumors than oligodendroglial tumors. The expression of GRIM-19 was higher in high-grade tumors than low-grade tumors or normal brain tissue; glioblastomas showed the highest expression. After transfection of GRIM-19 into U343MG-A, the morphology of the sense-transfection cells became larger and more spindly. The antisensetransfection cells became smaller and rounder compared with wild type U343MG-A. The MTT assay showed that the sense-transfection cells were more sensitive to the combination of interferon-$\beta$ and retinoic acid than U343MG-A cells or antisense-transfection cells; the antiproliferative activity was related to apoptosis. Conclusion : GRIM-19 may be one of the gene profiles which regulate cell death via apoptosis in human gliomas.

• Korean Journal of Veterinary Service
• /
• v.43 no.1
• /
• pp.23-29
• /
• 2020

Hepatitis E Virus (HEV) infection is a worldwide disease and the primary cause of acute viral hepatitis in the world. It can be isolated from many different species including pigs. HEV is a zoonotic pathogen and foodborne disease. The main animal reservoir is domestic pigs. It is usually asymptomatic in pig but it is a public health concern, causing acute hepatitis in humans of varying severity. This study focused on the presence of HEV in pig and pork product. One hundred feces and one hundred fifty serum samples were randomly collected from pigs in slaughterhouses in Gwangju from November in 2018 to February in 2020. In addtion, seventy-five pork products were collected from markets in Gwangju. Feces and pork product samples were examined for the presence of HEV RNA using an reverse-transcription realtime PCR (RT-qPCR) assay. Serum samples were tested for the presence of HEV-specific IgG antibodies using Enzyme-linked immunosorbent assay (ELISA). HEV antigen and antibody positive rates were 3.0% (3/100) and 19.3% (29/150), respectively, in Gwangju and nearby areas such as Jeonnam and Jeonbuk. However, HEV antigen was not detected from any of pork product in this study. In conclusion, the prevalence of HEV should be continuously monitored because HEV was sporadically detected in Gwangju and nearby areas.

• Journal of Aquaculture
• /
• v.21 no.2
• /
• pp.63-69
• /
• 2008

We isolated complementary DNA(cDNA) encoding prolactin receptor(PRLR) from gill of black porgy Acanthopagrus schlegeli. Its PRLR cDNA consists of 1,611 base pairs and encodes the protein of 536 amino acids. To investigate the osmoregulatory abilities of black porgy in different salinities(35, 10 and 0 psu), we examined the expression of PRLR mRNA in osmoregulatory organs(gill, kidney and intestine) using reverse transcription(RT)-PCR. In gill and intestine, PRLR mRNA levels were high in 10 psu, and then decreased in 0 psu, but there is no changes in kidney. Also, plasma osmolality, $Na^+\;and\;Cl^-$ levels decreased during the experimental period. These results suggest that PRLR plays an important role in hormonal regulation in osmoregulatory organs during freshwater acclimation, thereby improving the hyper-osmoregulatory ability of black porgy in hypoosmotic environments.

• Journal of Periodontal and Implant Science
• /
• v.24 no.3
• /
• pp.472-482
• /
• 1994

Cytokines expressed specifically in leukocytes subsets and in activated cells, which are involved in chemotaxis and activation of leukocytes, are recently defined as chemokines. Macrophage inflammatory $protein-1{\alpha}(MIP-1{\alpha})$ and $MIP-1{\beta}$ are members of C-C chemokine subfamily which produces wide immunomodulatory, proinflammatory, and hematopoietic modulatory actions. We have studied their gene expression by using Northern blot analysis in various blood cells such as cytolytic T lymphocyte(CTL), helper T lymphocyte(HTL), macrophage, and B lymphocyte. Resting CTL line CTLL-R8 expressed $MIP-1{\alpha}$ mRNA which was downregulated by ConA stimulation. Both of resting and ConA stimulated HTL line Hut78 and Jurkat did not express $MIP-1{\alpha}$ mRNA. There was detectable $MIP-1{\alpha}$ transcript in HTL hybridoma 2B4.11 which was a little upstimulated by ConA stimulation. B cell line 230, and macrophage cell line RAW264.7 and WR19M.1 showed distinct $MIP-1{\alpha}$ message which were induced after LPS stimulation. Expression pattern of $MIP-1{\beta}$ in all cell lines or cell were almost identical to that of $MIP-1{\alpha}$. Also strategies employed to identify and characterize the biological functions was preceded by receptor cloning to trace the shorcut to the final goal of cytokine research. For the cloning of $MIP-1{\alpha}$ receptor(R), we used synthetic oligonucleotides of transmembrane(T) conserved sequences of already cloned human(h) IL-8-R, and performed reverse transcription-polymerase chain reaction(RT-PCR) amplification using murine(m) macrophage cell line mRNA. Among 5RT-PCR products, we isolated a homologous cDNA with hIL-8-R which were shown to be putative mIL-8-R cDNA.

• Proceedings of the Korean Society of Applied Pharmacology
• /
• 2001.11a
• /
• pp.102-102
• /
• 2001

Taurine (2-ethaneaminosulfonic acid) is one of the major intracellular ${\beta}$ -amino acids in mammals and is required for a number of biological processes including membrane stabilization, osmoregulation, antioxidation, detoxification, modulation of calcium flux and neurornodulation. The taurine transporter (TAUT) which contains 12 hydrophobic membrane-spanning domains has been cloned from dog kidney, rat brain, mouse brain, human thyroid, placenta and retina. In this study, The TAUT cDNA from the human intestinal epithelial cell, HT-29 was cloned and sequenced. Reverse-transcription polymerase chain reaction (RT-PCR) was performed to amplify partial cDNA encoding human intestinal TAUT. The coding region of the PCR product was 732 bp long. The primers were designed to encode highly conserved amino acid sequences near the transmembrane domains III (IPYFIFLF) and Ⅵ (KYKYNSYR) both in human and mouse. The TAUT cDNA amplified was ligated into the pGEX 4T-1 expression vector. The resulting sequence of human intestinal TAUT cDNA (Accession number of NCBI Genebank is AF346763) was identical to the sequences of the TAUTs previously determined in the human placenta and retina except 3 base pairs from that of the reported human thyroid. TAUT specific antibodies were generated to use them as biological tools in the studies of the biological role of TAUT. Peptides of 149-162 amino acid residue (14 amino acids) of the TAUT were synthesized. The synthetic peptide used in this study was LFQSFQKELPWAHC. This region was chosen not only to avoid putative glycosylation sites but also to exclude regions of known homology with GABA transporters in the extracellular hydrophilic domains. The synthetic peptide, TAUT-1 was conjugated with carrier protein, kehole lympet hemocyanin (KLH) to use as an antigen. When used for immunization on a rabbit to produce polyclonal antiserum, the conjugates elicited high -titered specific anti-TAUT-1 antibodies, which reacted well with the ovalbumin (OVA) conjugated peptides in ELISA. The KLH-conjugated peptide was also used as immunizing antigen in BALB/c mice to produce TAUT specific monoclonal antibodies. From the culture supernatant of the hybridoma, the specificity of anti-TAUT-1 monoclonal antibodies was confirmed by ELISA. Further applications of more tools in TAUT expression analysis will be performed such as western blotting and flow cytometry.

• Asian-Australasian Journal of Animal Sciences
• /
• v.20 no.9
• /
• pp.1334-1341
• /
• 2007

This study identified hepatic differentially expressed genes (DEGs) affecting the marbling of muscle. Most dietary nutrients bypass the liver and produce plasma lipoproteins. These plasma lipoproteins transport free fatty acids to the target tissue, adipose tissue and muscle. We examined hepatic genes differentially expressed in a differential-display reverse transcription-polymerase chain reaction (ddRT-PCR) analysis comparing high- and low-marbled Hanwoo steers. Using 60 arbitrary primers, we found 13 candidate genes that were upregulated and five candidate genes that were downregulated in the livers of high-marbled Hanwoo steers compared to low-marbled individuals. A BLAST search for the 18 DEGs revealed that 14 were well characterized, while four were not annotated. We examined four DEGs: ATP synthase F0, complement component CD, insulin-like growth factor binding protein-3 (IGFBP3) and phosphatidylethanolamine binding protein (PEBP). Of these, only two genes (complement component CD and IGFBP3) were differentially expressed at p<0.05 between the livers of high- and low-marbled individuals. The mean mRNA levels of the PEBP and ATP synthase F0 genes did not differ significantly between the livers of high- and low-marbled individuals. Moreover, these DEGs showed very high inter-individual variation in expression. These informative DEGs were assigned to the bovine chromosome in a BLAST search of MS marker subsets and the bovine genome sequence. Genes related to energy metabolism (ATP synthase F0, ketohexokinase, electron-transfer flavoprotein-ubiquinone oxidoreductase and NADH hydrogenase) were assigned to BTA 1, 11, 17, and 22, respectively. Syntaxin, IGFBP3, decorin, the bax inhibitor gene and the PEBP gene were assigned to BTA 3, 4, 5, 5, and 17, respectively. In this study, the in silico physical maps provided information on the specific location of candidate genes associated with economic traits in cattle.

• Asian Pacific Journal of Cancer Prevention
• /
• v.15 no.8
• /
• pp.3581-3586
• /
• 2014

Aim: To investigate the effects of tetramethypyrazine (TMP) on proliferation and apoptosis of the human gastric carcinoma cell line 7901 and its possible mechanism of action. Methods: The viability of TMP-treated 7901 cells was measured with a 3-(4, 5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT) and cell apoptosis was analyzed by flow cytometry. The distribution of cells in different phases of cell cycle after exposure of TMPs was analyzed with flow cytometry. To investigate the molecular mechanisms of TMP-mediated apoptosis, the expression of NF-${\kappa}Bp65$, cyclinD1 and p16 in SGC-7901 cells was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. Results: TMP inhibited the proliferation of human gastric carcinoma cell line 7901 in dose and time dependent manners. Cell growth was suppressed by TMP at different concentrations (0.25, 0.5, 1.0, 2.0 mg/ml), the inhibition rate is 0.46%, 4.36%, 14.8%, 76.1% (48h) and 15.5%, 18.5%, 41.2%, 89.8% (72h) respectively. When the concentration of TMPs was 2.0mg/ml, G1-phase arrest in the SGC-7901 cells was significant based on the data for cell cycle distribution. RT-PCR demonstrated that NF-${\kappa}Bp65$ and cyclin D1 mRNA expression was significantly down-regulated in 7901 cells treated with 2.0 mg/ml TMP for 72h (p<0.05), while the p16 mRNA level was up-regulated (p<0.05). The protein expression of NF-${\kappa}Bp65$ and cyclin D1 decreased gradually with the increase in TMP concentration, compared with control cells (p<0.05), while expression of protein p16 was up-regulated (p<0.01). Conclusion: TMP exhibits significant anti-proliferative and pro-apoptotic effects on the human gastric carcinoma cell line SGC-7901. NF-${\kappa}Bp65$, cyclinD1 and p16 may also play important roles in the regulation mechanisms.

• Asian-Australasian Journal of Animal Sciences
• /
• v.21 no.9
• /
• pp.1244-1251
• /
• 2008

The aim of the present study was to delineate the expression and secretion of insulin-like growth factor (IGF) system components by pig liver cells. Hepatocytes were prepared from 3-wk-old weanling piglets following a two-step collagenase perfusion procedure, after which the cells were incubated for 24 or 48 h at a density of $2{\pm}10^5$ cells per 35-mm dish in 2-ml Williams' medium E. The cells were found to express the genes encoding IGF-I, IGF-binding proteins (IGFBPs)-2 and -3 and acid-labile subunit (ALS) by reverse transcription-polymerase chain reaction (RT-PCR) following the culture. However, IGF-I was localized to hepatocytes by immunohistochemical analysis, whereas IGFBP-3 was localized to endothelial cells, but not to hepatocytes. This indicated that the IGFBP-3 gene expression detected by RT-PCR was likely to have been contributed by unidentified non-parenchymal cells that had not been removed during the hepatocyte preparation. The conditioned culture medium (CCM) of the cells contained immunoreactive IGF-I and IGF-II, with the latter being seven-fold more abundant than the former. The CCM also contained 43-, 40-, 34-, 31-kDa doublet and 26-kDa IGFBPs as examined by Western ligand blotting. The 40-, 34- and 31-kDa doublet IGFBPs were approximately three-fold as abundant as the 43- and 26-kDa IGFBPs. Moreover, the 43- and 40-kDa doublet and the 34-kDa IGFBPs were immunoprecipitable with IGFBP-3 and IGFBP-2 antibodies, respectively. Overall, these results are similar to those known in the rat, which suggests that the IGF system components are likely to be expressed and secreted in pig liver in a manner similar to that in rat liver.

• Korean Journal of Microbiology
• /
• v.45 no.2
• /
• pp.105-111
• /
• 2009

Norovirus (NV) with a variety of genotypes, a member of the family Caliciviridae, causes acute nonbacterial gastroenteritis in humans. We determined the nucleotide sequence of three open reading frames (ORFs) of a NV Korean strain and characterized the genetic relationship with others. The Korean strain designated Hu/NLV/Gunpo/2006/KO was isolated from the stool specimen of a 2-year-old female suffering from gastroenteritis. By performing reverse transcription and PCR amplification, three overlapping cDNAs were synthesized and used for direct sequencing. We found that like other NVs, this strain contains three ORFs: ORF1, 5,100 bp; ORF2, 1,647 bp; ORF3, 765 bp. Of 35 NVs, ORF1 had a level of genetic diversity lower than ORF2 and ORF3, of which the C-termini of the ORF2 and ORF3 showed a relatively high degree of genetic diversity. Phylogenetic analyses indicated that the Korean strain belonged to genogroup II, with Saitama U1, Gifu'96, Mc37, and Vietnam 026 being formed a single genetic cluster. The nucleotide sequence information of three ORFs of a NV Korean isolate will be useful not only for the development of a diagnostic tool and understanding of genetic relationship, but also provide important basic information for the functional analysis of their gene products.

• The Plant Pathology Journal
• /
• v.18 no.5
• /
• pp.259-265
• /
• 2002

Apple mosaic virus (ApMV), a member of the genus Ilarvirus, was detected and isolated from diseased 'Fuji' apple (Malus domestica) in Korea. The coat protein (CP) genes of two ApMV strains, denoted as ApMV-Kl and ApMV-K2, were amplified by using the reverse transcription and polymerase chain reaction (RT-PCR) and were analyzed thereafter. The objectives were to define the molecular variability of genomic information of ApMV found in Korea and to develop virus-derived resistant gene source for making virus-resistant trans-genic apple. RT-PCR amplicons for the APMVS were cloned and their nucleotide sequences were determined. The CPs of ApMV-Kl and ApMV-K2 consisted of 222 and 232 amino acid residues, respectively. The identities of the CPs of the two Korean APMVS were 93.1% and 85.6% at the nucleotide and amino acid sequences, respectively. The CP of ApMV-Kl showed 46.1-100% and 43.2-100% identities to eight different ApMV strains at the nucleotide and amino acid levels, respectively. When ApMV-PV32 strain was not included in the analysis, ApMV strains shared over 83.0% and 78.6% homologies at the nucleotide and amino acid levels, respectively. ApMV strains showed heterogeneity in CP size and sequence variability. Most of the amino acid residue differences were located at the N-termini of the strains of ApMV, whereas, the middle regions and C-termini were remarkably conserved. The APMVS were 17.(1-54.5% identical with three other species of the genus Ilarviyus. ApMV strains can be classified into three subgroups (subgroups I, II, and III) based on the phylogenetic analysis of CP gene in both nucleotide and amino acid levels. Interestingly, all the strains of subgroup I were isolated from apple plants, while the strains of subgroups II and III were originated from peach, hop, or pear, The results suggest that ApMV strains co-evolved with their host plants, which may have resulted in the CP heterogeneity.