• 한국균학회소식:학술대회논문집
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• 2015.05a
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• pp.45-46
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• 2015

Clubroot disease of crucifers has occurred since 1957. It has spread to the whole China, especially in the southwest and nourtheast where it causes 30-80% loss in some fields. The disease has being expanded in the recent years as seeds are imported and the floating seedling system practices. For its effective control, the Ministry of Agriculture of China set up a program in 2010 and a research team led by Dr. Yueqiu HE, Yunnan Agricultural University. The team includes 20 main reseachers of 11 universities and 5 institutions. After 5 years, the team has made a lot of progresses in disease occurrence regulation, resources collection, resistance identification and breeding, biological agent exploration, formulation, chemicals evaluation, and control strategy. About 1200 collections of local and commercial crucifers were identified in the field and by artificiall inoculation in the laboratories, 10 resistant cultivars were breeded including 7 Chinese cabbages and 3 cabbages. More than 800 antagostic strains were isolated including bacteria, stretomyces and fungi. Around 100 chemicals were evaluated in the field and greenhouse based on its control effect, among them, 6 showed high control effect, especially fluazinam and cyazofamid could control about 80% the disease. However, fluzinam has negative effect on soil microbes. Clubroot disease could not be controlled by bioagents and chemicals once when the pathogen Plasmodiophora brassicae infected its hosts and set up the parasitic relationship. We found the earlier the pathogent infected its host, the severer the disease was. Therefore, early control was the most effective. For Chinese cabbage, all controlling measures should be taken in the early 30 days because the new infection could not cause severe symptom after 30 days of seeding. For example, a biocontrol agent, Bacillus subtilis Strain XF-1 could control the disease 70%-85% averagely when it mixed with seedling substrate and was drenching 3 times after transplanting, i.e. immediately, 7 days, 14 days. XF-1 has been deeply researched in control mechanisms, its genome, and development and application of biocontrol formulate. It could produce antagonistic protein, enzyme, antibiotics and IAA, which promoted rhizogenesis and growth. Its The genome was sequenced by Illumina/Solexa Genome Analyzer to assembled into 20 scaffolds then the gaps between scaffolds were filled by long fragment PCR amplification to obtain complet genmone with 4,061,186 bp in size. The whole genome was found to have 43.8% GC, 108 tandem repeats with an average of 2.65 copies and 84 transposons. The CDSs were predicted as 3,853 in which 112 CDSs were predicted to secondary metabolite biosynthesis, transport and catabolism. Among those, five NRPS/PKS giant gene clusters being responsible for the biosynthesis of polyketide (pksABCDEFHJLMNRS in size 72.9 kb), surfactin(srfABCD, 26.148 kb, bacilysin(bacABCDE 5.903 kb), bacillibactin(dhbABCEF, 11.774 kb) and fengycin(ppsABCDE, 37.799 kb) have high homolgous to fuction confirmed biosynthesis gene in other strain. Moreover, there are many of key regulatory genes for secondary metabolites from XF-1, such as comABPQKX Z, degQ, sfp, yczE, degU, ycxABCD and ywfG. were also predicted. Therefore, XF-1 has potential of biosynthesis for secondary metabolites surfactin, fengycin, bacillibactin, bacilysin and Bacillaene. Thirty two compounds were detected from cell extracts of XF-1 by MALDI-TOF-MS, including one Macrolactin (m/z 441.06), two fusaricidin (m/z 850.493 and 968.515), one circulocin (m/z 852.509), nine surfactin (m/z 1044.656~1102.652), five iturin (m/z 1096.631~1150.57) and forty fengycin (m/z 1449.79~1543.805). The top three compositions types (contening 56.67% of total extract) are surfactin, iturin and fengycin, in which the most abundant is the surfactin type composition 30.37% of total extract and in second place is the fengycin with 23.28% content with rich diversity of chemical structure, and the smallest one is the iturin with 3.02% content. Moreover, the same main compositions were detected in Bacillus sp.355 which is also a good effects biocontol bacterial for controlling the clubroot of crucifer. Wherefore those compounds surfactin, iturin and fengycin maybe the main active compositions of XF-1 against P. brassicae. Twenty one fengycin type compounds were evaluate by LC-ESI-MS/MS with antifungal activities, including fengycin A $C_{16{\sim}C19}$, fengycin B $C_{14{\sim}C17}$, fengycin C $C_{15{\sim}C18}$, fengycin D $C_{15{\sim}C18}$ and fengycin S $C_{15{\sim}C18}$. Furthermore, one novel compound was identified as Dehydroxyfengycin $C_{17}$ according its MS, 1D and 2D NMR spectral data, which molecular weight is 1488.8480 Da and formula $C_{75}H_{116}N_{12}O_{19}$. The fengycin type compounds (FTCPs $250{\mu}g/mL$) were used to treat the resting spores of P. brassicae ($10^7/mL$) by detecting leakage of the cytoplasm components and cell destruction. After 12 h treatment, the absorbencies at 260 nm (A260) and at 280 nm (A280) increased gradually to approaching the maximum of absorbance, accompanying the collapse of P. brassicae resting spores, and nearly no complete cells were observed at 24 h treatment. The results suggested that the cells could be lyzed by the FTCPs of XF-1, and the diversity of FTCPs was mainly attributed to a mechanism of clubroot disease biocontrol. In the five selected medium MOLP, PSA, LB, Landy and LD, the most suitable for growth of strain medium is MOLP, and the least for strains longevity is the Landy sucrose medium. However, the lipopeptide highest yield is in Landy sucrose medium. The lipopeptides in five medium were analyzed with HPLC, and the results showed that lipopeptides component were same, while their contents from B. subtilis XF-1 fermented in five medium were different. We found that it is the lipopeptides content but ingredients of XF-1 could be impacted by medium and lacking of nutrition seems promoting lipopeptides secretion from XF-1. The volatile components with inhibition fungal Cylindrocarpon spp. activity which were collect in sealed vesel were detected with metheds of HS-SPME-GC-MS in eight biocontrol Bacillus species and four positive mutant strains of XF-1 mutagenized with chemical mutagens, respectively. They have same main volatile components including pyrazine, aldehydes, oxazolidinone and sulfide which are composed of 91.62% in XF-1, in which, the most abundant is the pyrazine type composition with 47.03%, and in second place is the aldehydes with 23.84%, and the third place is oxazolidinone with 15.68%, and the smallest ones is the sulfide with 5.07%.

• Radiation Oncology Journal
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• v.19 no.2
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• pp.171-180
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• 2001

Purpose : Expression of TIMP, intrinsic inhibitor of MMP, is regulated by signal transduction in response to genotoxins and is likely to be an important step in metastasis, angiogenesis and wound healing after ionizing radiation. Therefore, we studied radiation mediated TIMP expression and its mechanism in head and neck cancer cell lines. Materials and Methods : Human head and neck cancer cell lines established at Asan Medical Center were used and radiosensitivity $(D_0)$, radiation cytotoxicity and metastatic potential were measured by clonogenic assay, n assay and invasion assay, respectively. The conditioned medium was prepared at 24 hours and 48 hours after 2 Gy and 10 Gy irradiation and expression of TIMP protein was measured by Elisa assay with specific antibodies against human TIMP. hTIMP1 promoter region was cloned and TIMP1 luciferase reporter vector was constructed. The reporter vector was transfected to AMC-HN-1 and -HN-9 cells with or without expression vector Ras, then the cells were exposed to radiation or PMA, PKC activator. EMSA was peformed with oligonucleotide (-59/-53 element and SP1) of TIMP1 promoter. Results : $D_0$ of HN-1, -2, -3, -5 and -9 cell lines were 1.55 Gy, 1.8 Gy, 1.5 Gt, 1.55 Gy and 2.45 Gy respectively. n assay confirmed cell viability, over $94\%$ at 24hrs, 48hrs after 2 Gy irradiation and over 73% after 10 Gy irradiation. Elisa assay confirmed that cells secreted TIMP1, 2 proteins continuously. After 2 Gy irradiation, TIMP2 secretion was decreased at 24hrs in HN-1 and HN-9 cell lines but after 10 Gy irradiation, it was increased in all cell lines. At 48hrs after irradiation, it was increased in HN-1 but decreased in HN-9 cells. But the change in TIMP secretion by RT was mild. The transcription of TIMP1 gene in HN-1 was induced by PMA but in HN-9 cell lines, it was suppressed. Wild type Ras induced the TIMP-1 transcription by 20 fold and 4 fold in HN-1 and HN-9 respectively. The binding activity to -59/-53, AP1 motif was increased by RT, but not to SP1 motif in both cell lines. Conclusions : We observed the difference of expression and activity of TIMPs between radiosensitive and radioresistant cell line and the different signal transduction pathway between in these cell lines may contribute the different radiosensitivity. Further research to investigate the radiation response and its signal pathway of TIMPs is needed.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.65 no.3
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• pp.199-213
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• 2020

Grain size and shape are the two important components contributing to rice yield and quality. To analyze traits related to grain-shape, a total of 272 varieties derived from japonica, japonica black and Tongil-type rice accession in Korea were evaluated in this study. The traits, grain length (GL), grain width (GW), grain thickness (GT), length to width ratio (RLW), and 1000-grain weight (TGW) were measured and replicated 10 times. Genes (GW2, GS3, qGL3, qSW5, GS5, TGW6, GW7, and GW8) related to grain-shape were validated in the accessions using specific DNA marker sets. K-mean clustering of the accession based on phenotypic data revealed three groups: group 1 was classified by GW and GT and included most of japonica type, group 2 was classified by RLW and GL reached a medium size and possessed a half spindle-shaped type, and group 3 was classified by TGW, reached a long size and possessed a semi-round shape. In validation tests using the marker sets, both gw2 and tgw6 were validated in less than 1% of the tested accessions and two allelic types, qgl3 and gw8, were only verified in Tongil-type accessions. For GW8 and GW2, any different amplicons were not amplified in any japonica or Tongil-type accessions, respectively. In order to suggest the representative grain-shape gene combinations for each ecotype, the allelic combinations were evaluated by PCR analysis. Cj1 and 2 in japonica (Cj1-7), Cj_b1 and 2 in japonica-black (Cj_b1-3), and CT3 in Tongil-type (CT1-13) turned out to be the dominant combination in each ecotype, respectively. In addition, the results revealed that introgression of four genes (gw2, gs3, qSW5, and GS5) would expand the diversity of grain shape in Korean japonica varieties. The gene combinations information could be utilized practically to understand or enhance grain shape in japonica rice breeding program.

• Childhood Kidney Diseases
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• v.9 no.2
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• pp.175-182
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• 2005

Purpose : Interleukin 1 receptor antagonist(IL-1ra) is an endogenous antiinflammatory agent that binds to IL-1 receptor and thus competitively inhibits the binding of IL-1$\alpha$ and IL-1$\beta$. Allele 2 in association with various autoimmune diseases has been reported. In order to evaluate the influence of IL-1ra gene VNTR polymorphism on the susceptibility to HSP and its possible association with disease severity, manifested by severe renal involvement and renal sequelae, we studied the incidence of carriage rate and allele frequency of the 2 repeats of IL-1ra allele 2($IL1RN^{*}2$) of the IL-1ra gene in children with HSP with and without renal involvement. Methods : The IL-1ra gene polymorphisms were determined in children with HSP with(n=40) or without nephritis(n=34) who had been diagnosed at Busan Paik Hospital and the control groups(n=163). Gene polymorphism was identified by PCR amplification of the genomic DNA. Results : The allelic frequency and carriage rate of $IL1RN^{*}1$ were found most frequently in patients with HSP and in controls. The allelic frequency of $IL1RN^{*}2$ was higher in patients with HSP compared to that of controls($4.7\%\;vs.\;2.5\%$, P=0.794). The carriage rate of $IL1RN^{*}2$ was higher In patients with HSP compared to that of controls($8.1\%\;vs.\;6.8\%$, P=0.916). The allelic frequency of $IL1RN^{*}2$ was higher in patients with HSP nephritis compared to that of HSP($5.3\%\;vs.\;2.9\%$, P=0.356). The carriage rate of $IL1RN^{*}2$ was higher in Patients with HSP nephritis compared to that of HSP($10.0\%\;vs.\;5.9\%$, P=0.523). Among 13 patients with heavy proteinuria(>1.0 g), 11 had $IL1RN^{*}1$, 1 had $IL1RN^{*}2$ and the others had $IL1RN^{*}4$. At the time of last follow up 4 patients had sustained proteinuria and their genotype was $IL1RN^{*}1$. Conclusion : The allelic frequency and carriage rate of $IL1RN^{*}1$ were found most frequently in patients with HSP and in controls. Our study suggests that the carriage rate and allele frequency of the 2-repeats of IL-1lra allele 2($IL1RN^{*}2$) of the IL-1ra gene may not be associated with susceptibility and severity of renal involvement in children with HSP (J Korean Soc Pediatr Nephrol 2005;9:175-182)

• Tuberculosis and Respiratory Diseases
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• v.54 no.2
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• pp.178-190
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• 2003

Background : The committee of tuberculosis(TB) survey planning for the year 2000 decided to construct the Korean Tuberculosis Surveillance System (KTBS), based on a doctor's routine reporting method. The successful keys of the KTBS rely on the precision of the recorded TB notification forms. The purpose of this study was to determine that the accuracy of the TB notification form written at a private general hospital given to the corresponding health center and to improve the comprehensiveness of these reporting systems. Materials and Methods : 291 adult TB patients who had been diagnosed from August 2000 to January 2001, were enrolled in this study. The lists of TB notification forms were compared with the medical records and the various laboratory results; case characteristics, history of previous treatment, examinations for diagnosis, site of the TB by the international classification of the disease, and treatment. Results : In the list of examinations for a diagnosis in 222 pulmonary TB patients, the concordance rate of the 'sputum smear exam' was 76% but that of the 'sputum culture exam' was only 23%. Among the 198 cases of the sputum culture exam labeled 'not examined', 43(21.7%) cases proved to be true 'not examined', 70 cases(35.4%) were proven to be 'culture positive', and 85(43.0%) cases were proven to be 'culture negative'. In the list of examinations for a diagnosis in 69 extrapulmonary TB patients, the concordance rate of the 'smear exam other than sputum' was 54%. In the list of treatments, the overall concordance rate of the 'type of registration' in the TB notification form was 85%. Among the 246 'new' cases on the TB notification form, 217(88%) cases were true 'new' cases and 13 were proven to be 'relapse', 2 were proven to be 'treatment after failure', one was proven to be 'treatment after default', 12 were proven to be 'transferred-in' and one was proven to be 'chronic'. Among the 204 HREZ prescribed regimen, 172(84.3%) patients were taking the HREZ regimen, and the others were prescribed other drug regimens. Conclusion : Correct recording of the TB notification form at the private sectors is necessary for supporting the effective TB surveillance system in Korea.

• Clinical and Experimental Pediatrics
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• v.49 no.3
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• pp.317-325
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• 2006

Purpose : This study was carried out to elucidate the effects of nitric oxide synthase(NOS) inhibitor, NG-monomethyl-L-arginine(L-NMMA) and nitric oxide precursor, L-arginine(L-Arg) on cerebral hemodynamics and energy metabolism during reoxygenation-reperfusion(RR) after hypoxia-ischemia(HI) in newborn piglets. Methods : Twenty-eight newborn piglets were divided into 4 groups; Sham normal control(NC), experimental control(EC), L-NMMA(HI & RR with L-NMMA), and L-Arg(HI & RR with L-Arg) groups. HI was induced by occlusion of bilateral common carotid arteries and simultaneously breathing with 8 percent oxygen for 30 mins, and followed RR by release of carotid occlusion and normoxic ventilation for one hour. All groups were monitored with cerebral hemodynamics and cytochrome $aa_3$ (Cyt $aa_3$) using near infrared spectroscopy(NIRS). $Na^+$, $K^+$-ATPase activity, lipid peroxidation products, and tissue high energy phosphate levels were determined biochemically in the cerebral cortex. Results : In experimental groups, mean arterial blood pressure, $PaO_2$, and pH decreased, and base excess and blood lactate level increased after HI compared to NC group(P<0.05). These variables subsequently returned to baseline after RR except pH. There were no differences among the experimental groups. In NIRS, oxidized hemoglobin($HbO_2$) decreased and hemoglobin(Hb) increased during HI(P<0.05) but returned to base line immediately after RR; 40 min after RR, the $HbO_2$ had decreased significantly compared to NC group(P<0.05). Changes of Cyt $aa_3$ decreased significantly compared to NC after HI and recovered at the end of the experiment. Significantly reduced cerebral cortical cell membrane $Na^+$, $K^+$-ATPase activity and increased lipid peroxidation products(P<0.05) were not improved with L-NMMA or L-Arg. Conclusion : These findings suggest that NO is not involved in the mechanism of HI and RR brain damage during the early acute phase of RR.

• Development and Reproduction
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• v.13 no.4
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• pp.257-264
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• 2009

A neurotoxin, 6-hydroxydopamine (6-OHDA) has been widely used to create animal model for Parkinson's disease (PD) due to its specific toxicity against dopaminergic (DA) neurons. Since DA signals modulate a broad spectrum of CNS physiology, one can expect profound alterations in neuroendocrine activities of both PD patients and 6-OHDA treated animals. Limited applications of 6-OHDA injection model, however, have been made on the studies of hypothalamuspituitary neuroendocrine circuits. The present study was performed to examine whether blockade of brain catecholamine (CA) biosynthesis with 6-OHDA can make any alteration in the transcriptional activities of hypothalamus-pituitary hormone genes in adult male rats. Three-month-old male rats (SD strain) were received 6-OHDA ($200{\mu}g$ in $10{\mu}\ell$ of saline/animal) by intracerebroventricular (icv) injection, and sacrificed after two weeks. To determine the mRNA levels of hypothalamuspituitary hormone genes, total RNAs were extracted and applied to the semi-quantitative RT-PCRs. The mRNA levels of tyrosine hydroxylase (TH), the rate-limiting enzyme for the catecholamine biosynthesis, were significantly lower than those from the control group (control:6-OHDA=1:0.72${\pm}$0.02AU, p<0.001), confirming the efficacy of 6-OHDA injection. The mRNA levels of gonadotropin-releasing hormone (GnRH) and corticotropin releasing hormone (CRH) in the hypothalami from 6-OHDA group were significantly lower than those from the control group (GnRH, control:6-OHDA=1:0.39${\pm}$0.03AU, p<0.001; CRH, control:6-OHDA=1:0.76${\pm}$0.07AU, p<0.01). There were significant decreases in the mRNA levels of common alpha subunit of glycoprotein homones (Cg$\alpha$), LH beta subunit (LH-$\beta$), and FSH beta subunit (FSH-$\beta$) in pituitaries from 6-OHDA group compared to control values (Cg$\alpha$, control:6-OHDA=1:0.81${\pm}$0.02AU, p<0.001; LH-$\beta$, control:6-OHDA=1:0.68${\pm}$0.04AU, p<0.001; FSH-$\beta$, control:6-OHDA=1:0.84${\pm}$0.05AU, p<0.001). Similarly, the level of adrenocorticotrophic hormone (ACTH) transcripts from 6-OHDA group was significantly lower than that from the control group (control: 6-OHDA=1:0.86${\pm}$0.04AU, p<0.01). The present study demonstrated that centrally injected DA neurotoxin could downregulate the transcriptional activities of the two hypothalamus-pituitary neuroendocrine circuits, i.e., GnRH-gonadotropins and CRH-ACTH systems. These results suggested that hypothalamic CA input might affect on the activities of gonad and adrenal through modulation of hypothalamus-pituitary function, providing plausible explanation for frequent occurrence of sexual dysfunction and poor stress-response in PD patients.

• The Korean Journal of Nuclear Medicine
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• v.38 no.1
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• pp.85-98
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• 2004

Purpose: Cellular uptake of $^{99}mTc$-sestamibi (MIBI) and $^{99}mTc$-tetrofosmin (TF) is low in cancer cells expressing multidrug resistance(MDR) by p-glycoprotein(Pgp) or multidrug related protein(MRP). Verapamil is known to increase cellular uptake of MIBI in MDR cancer cells, but is recently reported to have different effects on tracer uptake in certain cancer cells. This study was prepared to evaluate effects of verapamil on cellular uptake of MIBI and TF in several cancer cells. Materials and Methods: Celluar uptakes of Tc-99m MIBI and TF were measured in erythroleukermia K562 cell, breast cancer MCF7 cell, and human ovarian cancer SK-OV-3 cells, and data were compared with those of doxorubicin-resistant K562(Ad) cells. RT-PCR and Western blot analysis were used for the detection of mdr1 mRNA and Pgp expression, and to observe changes in isotypes of PKC enzyme. Effects of verapamil on MIBI and TF uptake were evaluated at different concentrations upto $200{\mu}M\;at\;1{\times}10^6\;cells/ml\;at\;37^{\circ}C$. Radioactivity in supernatant and pellet was measured with gamma counter to calculate cellular uptake ratio. Toxicity of verapamil was measured with MTT assay. Results: Cellular uptakes of MIBI and TF were increased by time in four cancer cells studied. Co-incubation with verapamil resulted in an increase in uptake of MIBI and TF in K562(Adr) cell at a concentration of $100{\mu}M$ and the maximal increase at $50{\mu}M$ was 10-times to baseline. In contrast, uptakes of MIBI and TF in K562, MCF7, SK-OV3 cells were decreased with verapamil treatment at a concentration over $1{\mu}M$. With a concentration of $200{\mu}M$ verapamil, MIBI and TF uptakes un K562 cells were decreased to 1.5 % and 2.7% of those without verapamil, respectively. Cellular uptakes of MIBI and TF in MCF7 and SK-OV-3 cells were not changed with $10{\mu}M$, but were also decreased with verapamil higher than $10{\mu}M$, resulting 40% and 5% of baseline at $50{\mu}M$. MTT assay of four cells revealed that K562, MCF7, SK-OV3 were not damaged with verapamil at $200{\mu}M$. Conclusion: Although verapamil increases uptake of MIBI and TF in MDR cancer cells, cellular uptakes were further decreased with verapamil in certain cancer cells, which is not related to cytotoxicity of drug. These results suggest that cellular uptakes of both tracers might differ among different cells, and interpretation of changes in tracer uptake with verapamil in vitro should be different when different cell lines are used.

• Nuclear Medicine and Molecular Imaging
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• v.42 no.5
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• pp.383-393
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• 2008

Purpose: Cancer specific killing can be achieved by therapeutic gene activated by cancer specific promotor. Expression of sodium iodide symporter (NIS) gene causes transportation and concentration of iodide into the cell, therefore radioiodine treatment after NIS gene transfer to cancer cell could be a form of radionuclide gene therapy. luciferase (Luc) gene transfected cancer cell can be monitored by in vivo optical imaging after D-luciferin injection. Aims of the study are to make vector with both therapeutic NIS gene driven by AFP promoter and reporter Luc gene driven by CMV promoter, to perform hepatocellular carcinoma specific radiodiodine gene therapy by the vector, and assessment of the therapy effect by optical imaging using luciferase expression. Materials and Methods: A Vector with AFP promoter driven NIS gene and CMV promoter driven Luc gene (AFP-NIS-CMV-Luc) was constructed. Liver cancer cell (HepG2, Huh-7) and non liver cancer cell (HCT-15) were transfected with the vector using liposome. Expression of the NIS gene at mRNA level was elucidated by RT-PCR. Radioiodide uptake, perchlorate blockade, and washout tests were performed and bioluminescence also measured by luminometer in these cells. In vitro clonogenic assay with 1-131 was performed. In vivo nuclear imaging was obtained with gamma camera after 1-131 intraperitoneal injection. Results: A Vector with AFP-NIS-CMV-Luc was constructed and successfully transfected into HepG2, Huh-7 and HCT-15 cells. HepG2 and Huh-7 cells with AFP-NIS-CMV-Luc gene showed higher iodide uptake than non transfected cells and the higher iodide uptake was totally blocked by addition of perchlorate. HCT-15 cell did not showed any change of iodide uptake by the gene transfection. Transfected cells had higher light output than control cells. In vitro clonogenic assay, transfected HepG2 and Huh-7 cells showed lower colony count than non transfected HepG2 and Huh-7 cells, but transfected HCT-15 cell did not showed any difference than non transfected HCT-15 cell. Number of Huh-7 cells with AFP-NIS-CMV-Luc gene transfection was positively correlated with radioidine accumulation and luciferase activity. In vivo nuclear imaging with 1-131 was successful in AFP-NIS-CMV-Luc gene transfected Huh-7 cell xenograft on nude mouse. Conclusion: A Vector with AFP promoter driven NIS and CMV promoter driven Luc gene was constructed. Transfection of the vector showed liver cancer cell specific enhancement of 1-131 cytotoxicity by AFP promoter, and the effect of the radioiodine therapy can be successfully assessed by non-invasive luminescence measurement.

• Journal of Life Science
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• v.19 no.6
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• pp.765-769
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• 2009

Recent studies have reported that the distribution of HPV-16 sequence variation differs geographically, and more specifically that HPV-16 E6/E7 intratypic variants might carry a high risk for development of ICC (invasive cervical cancer) and CIN (cervical intraepithelial neoplasia) in a given population. To investigate the genetic diversities of HPV-16 E6/E7 oncogene by region, we collected nineteen HPV-16 isolates from sexually high-risk women in Busan, and analyzed the HPV-16 E6/E7 coding regions (nt 34 to 880) with HPV-16 E6/E7 specific PCR amplification. At the nucleotide levet eleven variants of the E6 genes and nine variants of the E7 genes were identified as follows: E6 T178G (n=l1), E6 T178A (n=l), E6 T350G (n=3), E6 A442C (n=2), E6 AI04T, E6 All1G, E6 C116T, E6 G145T, E6 T183G, E6 C335T, E6 G522C and E7 A647G (n=12), E7 A645C, E7 A777C, E7 G663A, E7 T732C, E7 T760C, E7 A775T, E7 T789C and E7 T795G, respectively. At the amino acid levet the isolated HPV-16 E6 and E7 genes showed eleven E6 variants: E6 D25E (n=12), E6 L83V (n=4), E6 E113D (n=2), E6 MIL, E6 Q3R, E6 P5S, E6 Q14H, E6 D25N, E6 127R, E6 H78Y, E6 C140S and three E7 variants: N29S (n=12), L28F, T72S. HPV16 E6 L83V, the dominant variant in the Caucasian population, showed relatively low frequencies in our study population. We elucidated that the dominant HPV-16 E6/E7 variants were HPV-16 E6 D25E (63.2%) and HPV-16 E7 N29S (63.2%), which were phylogenetically included in Asian lineage. Further study is needed to evaluate the risk of cervical cancer related HPV-16 E6/E7 intratypic variants in the Korean population.