• Asian Pacific Journal of Cancer Prevention
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• v.17 no.1
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• pp.249-254
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• 2016

Molecular detection methods such as RT-PCR for detecting breast cancer-associated gene expression in the peripheral blood have the potential to modify breast cancer (BC) staging and therapy. In this regard, we evaluated the potential of erb-B2 molecular marker in BC detection and analyzed the expression of erb-B2 mRNA in the peripheral blood and fresh tissue samples of 50 pretreated female BC patients and 50 healthy females by reverse transcription-PCR (RT-PCR) method. We also assessed the correlation of erb-B2 mRNA marker positivity in peripheral blood and tumor tissue samples with clinical and pathological factors in BC patients in order to evaluate its prognostic value. It was shown that there is a significant difference between healthy females and BC patients with expression of the erb-B2 molecular marker (p<0.01). A significant difference between the expression of erb-B2 in the peripheral blood and tissue samples of BC patients (p<0.01) and the frequency of circulating erb-B2 mRNA expression in peripheral blood and in tissue was detected by RT-PCR. No correlation was found between erb-B2 mRNA expression in blood or tumor tissue samples and lymph node, tumor grade, tumor stage, tumor size, patient's age, ki67, estrogen receptor (ER), progesterone receptor (PGR), P53, and HER-2 status. However, in a small subset of 31 BC patients we found that expression of erb-B2 in peripheral blood or in both peripheral blood and tumor tissue was directly correlated with lympho-vascular invasion and perineural invasion as poor prognostic features. The highest rates of erb-B2 expression in peripheral blood or tumor tissue were in the ER and PR negative and HER-2 positive group. This study suggests that the application of the RT-PCR and immunohistochemical methods for erb-B2 molecular marker detection would provide a higher detection rate, especially in early stage BC.

• Biomedical Science Letters
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• v.6 no.2
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• pp.141-149
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• 2000

The etiologic agents of haemorrhagic fever with renal syndrome (HFRS) in Korea are Hantaan and Seoul virus in the genus Hantavirus, family Bunyaviridae. Antibody titers of sera from HFRS patients against Hantaan virus were measured by immunofluorescent antibody technique (IFAT), enzyme-linked immunosorbent assay (ELISA), high density composite particle agglutination (HDPA) and plaque reduction neutralization test (PRNI). PRNT and nested reverse transcriptase polymerase chain reaction (nested RT-PCR) was used for serotypic differentiation of Hantaviruses against Hantaan and Seoul virus. Eight doubtful HFRS patients showed higher fluorescent, IgG ELISA, agglutination and neutralizing antibody titer by IFAT, ELISA IgG, HDPA and PRNT, respectively Five out of them showed high IgM antibody titer by IgM capture ELISA against Hantaan virus, remarkably. Fifteen HFRS patients showed higher fluorescent antibody titer by IFAT. In PRNT, 12 out of them showed high neutralizing antibody titer against HTNV, 2 against SEOV and 1 against both viruses. In nested RT-PCR using serotype specific-primer, 3 out of them showed positive against HTNV and 1 against SEOV.

• Journal of Life Science
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• v.17 no.7 s.87
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• pp.996-1001
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• 2007

To investigate whether sulindac, sulindac sulfone, and sulindac sulfide could affect cancer cell viabilities, human colorectal HCTl16 cells were treated with 10 ${\mu}M$ of each NSAID. Among treated NSAms, sulindac sulfide dramatically decreased the cell viabilities detected by MTS and the cytotoxic effect showed dose-dependent manner. To understand the molecular mechanism of cell death in response to sulindac sulfide treatment, we performed oligo DNA microarray analysis. We found that 23 genes were up-regulated more than 2 folds, whereas 33 genes were down-regulated more than 2 folds by treatment of 10 ${\mu}M$ sulindac sulfide. Among the up-regulated genes, we selected 3 genes (NAG-1, DDIT3, PCK2) and performed RT-PCR and quantitative real-time PCR to cofirm microarray data. The results of RT-PCR and real-time PCR were highly accorded with those of microarray experiment. As NAG-1 is well-known gene as tumor suppressor, we detected changes of NAG-1 expression by 10 ${\mu}M$ of sulindac, sulindac sulfone, and sulindac sulfide. The results of RT-PCR and quantitacve real-time PCR indicated that sulindac sulfide was the strongest inducer of NAG-1 among treated NSAIDS. This result implies that induction of NAG-1 by sulindac sulfide plays important role in cell death of colorectal cancer. Overall, we speculate that these results may be helpful in understanding the molecular mechanism of the cancer chemoprevention by sulindac sulfide in human colorectal cancer.

• The Journal of Korean Society of Virology
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• v.28 no.2
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• pp.147-155
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• 1998

Hantavirus is a genus of the Bunyaviridae family consisting following serotype groups: Hantaan, Seoul, Puumala, Prospect Hill, Thailand, Belgrade, Thotta palayam, Sin Nombre. Most of Hantavirus group have been associated with many clinically similar disease known collectively as hemorrhagic fever with renal syndrome (HFRS). Hantaan virus is the prototype of the genus hantavirus, originally isolated from Apodemus agrarius. Bat was found as a natural host for Hantaan virus in Lee's lab for the first time. Then, Hantaan-like virus was isolated Hantaan-like virus from bat. To identify hantaviruses that are present in Korea among bats, bats were collected from Jeong-Sun, Won-Joo, Chung-Ju and Hwa-Cheon area, RNA was isolated from lung and serum. RT-PCR was performed with a universal primer from M segment. Nested RT-PCR was carried out to differentiate Hantaan, Seoul and Puumala virus using serotype specific primers. As we expected, Hantaan viruses were detected in bats and Seoul virus was not detected. Interestingly, Puumala viruses were also detected in bats from Won-Ju, but not in other areas. Puumala virus is originally isolated from Clethrinomys glareolus, and cause light HFRS. Recently, Paradoxomis webbiana, a wild bird turn out to be a reservoir for Puumala virus in Korea. These data indicate that bat is a new natural reservoir of Puumala virus.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.6
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• pp.455-460
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• 2009

Glutamate-induced cobalt uptake reveals that non-NMDA glutamate receptors (GluRs) are present in rat taste bud cells. Previous studies involving glutamate induced cobalt staining suggest this uptake mainly occurs via kainate type GluRs. It is not known which of the 4 types of taste bud cells express subunits of kainate GluR. Circumvallate and foliate papillae of Sprague-Dawley rats (45~60 days old) were used to search for the mRNAs of subunits of non-NMDA GluRs using RT-PCR with specific primers for GluR1-7, KA1 and KA2. We also performed RT-PCR for GluR5, KA1, $PLC\beta2$, and NCAM/SNAP 25 in isolated single cells from taste buds. Taste epithelium, including circumvallate or foliate papilla, express mRNAs of GluR5 and KA1. However, non-taste tongue epithelium expresses no subunits of non-NMDA GluRs. Isolated single cell RT-PCR reveals that the mRNAs of GluR5 and KA1 are preferentially expressed in Type II and Type III cells over Type I cells.

• Korean Journal of Veterinary Service
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• v.40 no.1
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• pp.15-20
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• 2017

A two-tube reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was designed for the rapid visual detection of the M gene of all subtypes of avian influenza virus (AIV) and the H5 gene of the H5 subtype of highly pathogenic AIV (HPAIV). The reaction carried out in two tubes in a single step at $58^{\circ}C$ for 40 min, and the assay results could be visually detected by using hydroxynaphthol blue dye. Using M or H5 gene-specific primers, the assay successfully detected all subtypes or H5 subtypes of AIVs, including the Korean representative H5N1 and H5N8 HPAIVs. The detection limit of the assay was approximately $10^{2.0}$ $EID_{50}/reaction$ for the M and H5 genes of H5N1 HPAIV, respectively, and was more sensitive than that of previously reported RT-LAMP and comparable to that of real-time RT-PCR. These results suggest that the present RT-LAMP assay, with its high specificity, sensitivity, and simplicity, will be a useful diagnostic tool for surveillance of currently circulating H5 HPAIVs and other subtypes of AIV in bird population, even in under-equipped laboratories.

• Journal of Pharmacopuncture
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• v.3 no.1
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• pp.35-51
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• 2000

The purpose of this study is to prove the analgesic effects of apitoxin and its mechanism via jaw-opening reflex(JOR) and measuring expression of mRNA in Phospholipase and Tryptophan hydroxylase(TPH) using RT-PCR. The experiments were carried out on Sprague-Dawley rats(300-400g) and mastocytoma(P-185 HTR) for JOR and RT-PCR, respectively. Rats anesthetized with thiopental sodium (80mg/kg) were used in the Tooth Pulp stimulation induced JOR. The amplitude of a digastric electromyogram (dEMG) was recorded during the stimulation at an intensity of 1.5 times the threshold for JOR. Apitoxin used in this experiment was diluted with normal saline by 1:1000. Apitoxin was injected intravenously into the test group while normal saline to the control group. However, it was injected directly into the cell of mastocytoma. We referred to base sequence registered in Genbank in designing primers for RT-PCR. The results were as follows; (1)Compared with control group, analgesic effect started to show right after Sprague-Dawely rats were treated with apitoxin($71.50{\pm}8.08$) and lasted for 50 minutes. (2)As a result of the experiment of RT-PCR, we witnessed significant changes in the degree of expression of phospholipase or rate-limiting enzyme of biosynthesis of prostaglandins with $10{\mu}g/ml$ apitoxin.($31.74{\pm}18.98%$, P<0.05) (3)As a result of the experiment of RT-PCR, we witnessed significant changes in the degree of expression of TPH or rate-limiting enzyme in biosynthesis of serotonin with $10{\mu}g/ml$ apitoxin.($131.37{\pm}16.87%$, P<0.05). These results suggest that $10{\mu}g/ml$ apitoxin have the most analgesic effects. This study showed that apitoxin has analgesic effects and held good for 50 minutes. The injection of apitoxin has brought out changes in the degree of expression of phospholipase and TPH. These results strongly suggest that analgesic mechanism by apitoxin is closely related to prostaglandins and serotonin.

• KSBB Journal
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• v.23 no.4
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• pp.303-310
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• 2008

Bovine blood, cell, tissue, and organ are used as raw materials for manufacturing biologics such as biopharmaceuticals, tissue-engineered products, and cell therapy. Manufacturing processes for the biologics have the risk of viral contamination. Therefore viral validation is essential in ensuring the safety of the products. Bovine parainfluenza virus type 3 (BPIV3) is one of the common bovine pathogens and has widely been known as a contaminant of biologics. In order to establish the validation system for the BPIV3 safety of biologics, a real-time RT-PCR method was developed for quantitative detection of BPIV3 contamination in raw materials, manufacturing processes, and final products. Specific primers for amplification of BPIV3 RNA was selected, and BPIV3 RNA was quantified by use of SYBR Green I. The sensitivity of the assay was calculated to be 2.8 $TCID_{50}/mL$. The real-time RT-PCR method was validated to be reproducible and very specific to BPIV3. The established real-time RT-PCR assay was successfully applied to the validation of Chinese hamster ovary (CHO) cell artificially infected with BPIV3. BPIV3 RNA could be quantified in CHO cell as well as culture supernatant. Also the real-time RT-PCR assay could detect 7.8 $TCID_{50}/mL$ of BPIV3 artificially contaminated in bovine collagen. The overall results indicated that this rapid, specific, sensitive, and robust assay can be reliably used for quantitative detection of BPIV3 contamination during the manufacture of biologics.

• Journal of Pharmacopuncture
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• v.7 no.2
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• pp.43-56
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• 2004

Objectives : In order to investigate effects and immune improvement of distilled red-ginseng herbal Acupuncture, expression of Cox-1, Cox-2, and mRNA of Bcl-2 and Bax were analyzed in A549 cell in vivo. Survival time and expression of cytokine mRNA were measured for the mice with Sarcoma-180 induced abdominal cancer. Methods : Balb/c mouse was treated with distilled red-ginseng Herbal Acupuncture at Wisu($BL_{21}$) and Chung- wan($CV_{12}$) to investigate anti-cancer effects and immune response. Results : 1. For expression of mRNA of Cox-1 using RT-PCR, the control group and the experiment groups didn't show significant differences. For Cox-2, both experiment groups and the normal group showed significant differences. 2. For expression of mRNA of Bcl-2 using RT-PCR, experiment groups showed slight decrease compared to the control group. For Bax, no significant changes were shown between the control group and experiment groups. 3. For survival time, all of experiment groups showed 11.1% increase compared to the control group. 4. For IL-2 and IL-4 productivity using Flow cytometry, all of experiment groups didn't show any significance. 5. For IL-2 productivity using ELISA, all of experiment groups didn't show any significance. 6. For expression of cytokine mRNA using RT-PCR, significant increase of IL.-2 and IL-4 were witnessed in the experiment group II compared to the control group. Significant increase of IL-10 was shown in all off experiment groups compared to the control group. Conclusion : According to the results, we can expect that distilled red-ginseng Herbal Acupuncture may be further effccts in anti-cancer and immune improvement if increasing concentration.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1997.04a
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• pp.94-94
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• 1997

The objective of this study was to generate and characterize monoclonal antibodies against rat airway mucins, and therefore, should serve as a useful tool in studying the regulation of airway mucins using various in vivo rat models that are currently available. As an antigen, we used a high molecular mass mucin preparation purified from the spent media of rat tracheal surface epithelial cells in primary culture. Seven hybridomas were obtained which secrete monoclonal antibodies against the rat mucin among which mAbRT03 showed the highest immunoreactivity against the mucin based on ELISA. All of the antibodies secreted by these hybridomas recognized carbohydrate epitopes but not sialic acid residues since their immunoreactivity was completely abolished by treatment of the mucin with 20 mM periodate but not with neuraminidase. Further characterization of mAbRT03 showed that: (1) it belongs to the IgM type, (2) it binds to high molecular mass mucins based on both Western blot analysis and indirect immunoprecipitation, (3) it binds to the luminal side of tracheal epithelium as well as some goblet cells based on immunohistochemistry, and (4) it also recognizes in vive airway mucins from rats but not from human nor hamsters which have been purified from the airway lavage fluids. This is the first anti-rat airway mucin monoclonal antibody which has been developed against purified rat airway mucins. Therefore, mAbRT03 should be able to serve as an invaluable tool in studying the regulation of airway mucins using various intact rat models that are currently available.