• Journal of Chest Surgery
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• v.37 no.12
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• pp.967-977
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• 2004

Background: DNA methylation is one of the important gene expression mechanisms of the cell. When cytosine of CpG dinucleotide in promotor is hypomethylated, expression of some genes that is controlled by this promoter is altered. In this study, the author investigated the effect of DNA demethylating agent, 5-aza-2'-deoxycytidine (ADC), on the expressions of cancer antigen genes, MHC and B7 in 4 lung cancer cell lines, NCIH1703, NCIH522, MRC-5, and A549. Material and Method: After treatment of cell lines, NCIH1703, NCIH522, MRC-5 and A549 with ADC (1 uM) for 48 hours, RT-PCR was performed by using the primers of MAGE, GAGE, NY-ESO-1, PSMA, CEA, and SCC antigen gene. In order to find the optimal ADC treatment condition for induction of cancer antigen, we studied the effect of ADC treatment time and dose on the cancer antigen gene expression. To know the effect of ADC on the expression of MHC or B7 and cell growth, cells were treated with 1 uM of ADC for 72 hours for FACS analysis or cells were treated with 0.2, 1 or 5 uM of ADC for 96 hours for cell counting. Result: After treatment of ADC (1 uM) for 48 hours, the expressions of MAGE, GAGE, NY-ESO-1, and PSMA genes increased in some cell lines. Among 6 MAGE isotypes tested, and gene expression of MAGE-1, -2, -3, -4 and -6 could be induced by ADC treatment. However, CEA gene expression did not change and SCC gene expression was decreased by ADC treatment. Gene expression was generally induced 24 - 28 hours after ADC treatment and expression of MAGE, GAGE, and NY-ESO-1 was maintained at least 14 days after ADC ADC teatment, and expression of MAGE, GAGE, and NY-ESO-1 was maintained at least 14 days after ADC teatment in ADC-Free medium. Most gene expression could be induced at 0.2 uM of ADC, but gene expression increased dependently on ADC treatment dose. The expression of MHC and B7 was not increased by ADC treatment in all four cell lines, and the growth rate of 4 cell lines decreased significantly with the increase of ADC concentrations. Conclusion: Treatment of lung cancer cell lines with ADC increases the gene expression MAGE, GAGE and NY-ESO-1 that are capable of induction of cytotoxic T lymphocyte response. We suggest that treatment with 1 uM of ADC for 48 hours and then culturing in ADC-free medium is optimal condition for induction of cancer antigen. However, ADC has no effect on MHC and B7 induction, additional modification for increase of expression of MHC, B7 and cytokine will be needed for production of efficient cancer cell vaccine.

• The Sea:JOURNAL OF THE KOREAN SOCIETY OF OCEANOGRAPHY
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• v.17 no.4
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• pp.292-302
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• 2012

This paper reviews comparison analysis of current and latest application for stock identification methods of Todarodes pacificus, and the pros and cons of each method and consideration of how to compensate for each other. Todarodes pacificus which migrates wide areas in western North Pacific is important fishery resource ecologically and commercially. Todarodes pacificus is also considered as 'biological indicator' of ocean environmental changes. And changes in its short and long term catch and distribution area occur along with environmental changes. For example, while the catch of pollack, a cold water fish, has dramatically decreased until today after the climate regime shift in 1987/1988, the catch of Todarodes pacificus has been dramatically increased. Regarding the decrease in pollack catch, overfishing and climate changes were considered as the main causes, but there has been no definite reason until today. One of the reasons why there is no definite answer is related with no proper analysis about ecological and environmental aspects based on stock identification. Subpopulation is a group sharing the same gene pool through sexual reproduction process within limited boundaries having similar ecological characteristics. Each individual with same stock might be affected by different environment in temporal and spatial during the process of spawning, recruitment and then reproduction. Thereby, accurate stock analysis about the species can play an efficient alternative to comply with effective resource management and rapid changes. Four main stock analysis were applied to Todarodes pacificus: Morphologic Method, Ecological Method, Tagging Method, Genetic Method. Ecological method is studies for analysis of differences in spawning grounds by analysing the individual ecological change, distribution, migration status, parasitic state of parasite, kinds of parasite and parasite infection rate etc. Currently the method has been studying lively can identify the group in the similar environment. However It is difficult to know to identify the same genetic group in each other. Tagging Method is direct method. It can analyse cohort's migration, distribution and location of spawning, but it is very difficult to recapture tagged squids and hard to tag juveniles. Genetic method, which is for useful fishery resource stock analysis has provided the basic information regarding resource management study. Genetic method for stock analysis is determined according to markers' sensitivity and need to select high multiform of genetic markers. For stock identification, isozyme multiform has been used for genetic markers. Recently there is increase in use of makers with high range variability among DNA sequencing like mitochondria, microsatellite. Even the current morphologic method, tagging method and ecological method played important rolls through finding Todarodes pacificus' life cycle, migration route and changes in spawning grounds, it is still difficult to analyze the stock of Todarodes pacificus as those are distributed in difference seas. Lately, by taking advantages of each stock analysis method, more complicated method is being applied. If based on such analysis and genetic method for improvement are played, there will be much advance in management system for the resource fluctuation of Todarodes pacificus.

• Tuberculosis and Respiratory Diseases
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• v.48 no.2
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• pp.180-190
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• 2000

Background: Genomic instability, which is manifested by the replication error(RER) phenotype, has been proposed for the promotion of genetic alterations necessary for carcinogenesis. Merlo et al. reported frequent microsatellite instability in primary small cell lung cancers. However, Kim et al. found that instability occurred in only 1% of the loci tested and did not resemble the replication error-positive phenotype. The significance of microsatellite instability in the tumorigenesis of small cell lung cancer as well as the relationship between microsatellite instability and its clinical prognosis was investigated in our study. Methods: Fifteen primary small cell lung cancers were chosen for this study. The DNAs extracted from paraffin-embedded tissue blocks with primary tumor and corresponding control tissue were investigated. Forty microsatellite markers on chromosome 1p, 2p, 3p, 5q, 6p, 6q, 9p, 9q, 13q, and 17p were used in the microsatellite analysis. Results: Thirteen(86.7%) of 15 tumors exhibited LOH in at least one of the tested microsatellite markers. Three of 13 tumors exhibiting LOH lost a larger area in chromosome 9p. LOH was shown in 72.7% on chromosome 2p, 40% on 3p, 50% on 5q, 46.7% on 9p, 69.2% on 13q, and 66.7% on 17p(Table 1). Nine(60%) of 15 tumors exhibited shifted bands in at least one of the tested microsatellite markers. Nine cases exhibiting shifted bands showed altered loci ranging 2.5~52.5%(mean $9.4%\pm16.19$)(Table 2). Shifted bands occurred in 5.7% (34 of 600) of the loci tested(Table 2). Nine cases with shifted bands exhibited LOH ranging between 0~83.3%, and the median survival duration of those cases was 35 weeks. Six cases without shifted bands exhibited LOH ranging between 0~83.3%, and the median survival duration of those cases was 73 weeks. There was no significant difference between median survival durations of the two groups(p=0.4712). Conclusion: Microsatellite instability as well as the inactivation of several tumor suppressor genes may play important roles in the development and progression process of tumors. However, the relationship between microsatellite instability and its clinical prognosis in primary small cell lung cancer could not be established.

• Tuberculosis and Respiratory Diseases
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• v.56 no.5
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• pp.465-484
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• 2004

Background : Insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) inhibits the proliferation of non-small cell lung cancer (NSCLC) cells by inducing apoptosis. Methods : In this study, we investigated whether hypermethylation of IGFBP-3 promoter play an important role in the loss of IGFBP-3 expression in NSCLC. We also studied the mechanisms that mediate the silencing of IGFBP-3 expression in the cell lines which have hypermethylated IGFBP-3 promoter. Results : The IGFBP-3 promoter has hypermethylation in 7 of 15 (46.7%) NSCLC cell lines and 16 (69.7%) of 23, 7 (77.8%) of 9, 4 (80%) of 5, 4 (66.7 %) of 6, and 6 (100%) of 6 tumor specimens from patients with stage I, II, IIIA, IIIB, and IV NSCLC, respectively. The methylation status correlated with the level of protein and mRNA in NSCLC cell lines. Expression of IGFBP-3 was restored by the demethylating agent 5'-aza-2'-deoxycytidine (5'-aza-dC) in a subset of NSCLC cell lines. The Sp-1/ Sp-3 binding element in the IGFBP-3 promoter, important for promoter activity, was methylated in the NSCLC cell lines which have reduced IGFBP-3 expression and the methylation of this element suppressed the binding of the Sp-1 transcription factor. A ChIP assay showed that the methylation status of the IGFBP-3 promoter influenced the binding of Sp-1, methyl-CpG binding protein-2 (MeCP2), and histone deacetylase (HDAC) to Sp-1/Sp-3 binding element, which were reversed by by 5'-aza-dC. In vitro methylation of the IGFBP-3 promoter containing the Sp-1/Sp-3 binding element significantly reduced promoter activity, which was further suppressed by the overexpression of MeCP2. This reduction in activity was rescued by 5'-aza-dC. Conclusion : These findings indicate that hypermethylation of the IGFBP-3 promoter is one mechanism by which IGFBP-3 expression is silenced and MeCP2, with recruitment of HDAC, may play a role in silencing of IGFBP-3 expression. The frequency of this abnormality is also associated with advanced stages among the patients with NSCLC, suggesting that IGFBP-3 plays an important role in lung carcinogenesis/progression and that the promoter methylation status of IGFBP-3 may be a marker for early molecular detection and/or for monitoring chemoprevention efforts.

• Journal of Embryo Transfer
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• v.22 no.2
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• pp.121-126
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• 2007

The objective of this study was carried out to examine the selection effects of in vitro matured porcine follicular oocytes with polar body extrusion and early cleavage as non-invasive marker to know the developmental competence in advance. The porcine oocytes matured for 48 h were examined the polar body extrusion. The examined oocytes were matured for additional $16{\sim}18h$ and activated with 7% ethanol and cultured in $5{\mu}g/ml$ cytochalasin B for 5 h for diploid formation. The treated oocytes were cultured and examined the cleavage after 48 h and continued culturing for 5 days. The oocytes of 21.9% were discarded in morphological selection and 32.1% oocytes were discarded by failure of first polar body extrusion. The selected oocytes were matured and activated and then after 48 h the cleavage rates were examined. In morphologically selected oocytes, 15.8% oocytes were not cleaved and 52.6% oocytes were normally cleaved and 31.6% oocytes were hyper-cleaved over 8-cell stage. However in the first polar body extruded oocytes, 7.1% oocytes were not cleaved and 73.1% oocytes were normally cleaved and 19.8% oocytes were hyper-cleaved. The morphologically selected embryos that not cleavage-selected were developed in 16.7% up to blastocyst and the morphologically selected and cleavage-selected embryos were developed in 31.7%. The polar body extruded oocytes that were not carried out cleavage selection were developed in 39.0% and the polar body extruded and cleavage-selected embryos were developed 49.0%. The first cleavage timing was examined with 12 h interval after activation. In $0{\sim}12,\;12{\sim}24,\;24{\sim}36,\;and\;36{\sim}48h$ intervals, 4.1%, 68.6%, 19.1%, and 2.3% oocytes were cleaved and 5.9% oocytes were not cleaved until 48 after activation. The cleaved oocytes in each interval were cultured and developed upto blastocyst with 0, 39.1, 9.5, and 0%, respectively. This results suggests that polar body extruded and cleaved at $12{\sim}36h$ embryo has higher developmental potential than the others.

• The Korean Journal of Nuclear Medicine
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• v.39 no.4
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• pp.252-256
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• 2005

Purpose: Thyroglobulin (Tg) is a valuable and sensitive tool as a marker for diagnosis and follow-up for several thyroid disorders, especially, in the follow-up of patients with differentiated thyroid cancer (DTC). Often, clinical decisions rely entirely on the serum Tg concentration. But the Tg assay is one of the most challenging laboratory measurements to perform accurately owing to antithyroglobulin antibody (Anti-Tg). In this study, we have compared the degree of Anti-Tg effects on the measurement of Tg between availale Tg measuring kits. Materials and Methods: Measurement of Tg levels for standard Tg solution was performed with two different kits commercially available (A/B kits) using immunoradiometric assay technique either with absence or presence of three different concentrations of Anti-Tg. Measurement of Tg for patient's serum was also performed with the same kits. Patient's serum samples were prepared with mixtures of a serum containing high Tg levels and a serum containg high Anti-Tg concentrations. Results: In the measurements of standard Tg solution, presence of Anti-Tg resulted in falsely lower Tg level by both A and B kits. Degree of Tg underestimation by h kit was more prominent than B kit. The degree of underestimation by B kit was trivial therefore clinically insignificant, but statistically significant. Addition of Anti-Tg to patient serum resulted in falsely lower Tg levels with only A kit. Conclusion: Tg level could be underestimated in the presence of anti-Tg. Anti-Tg effect on Tg measurement was variable according to assay kit used. Therefore, accuracy test must be performed for individual Tg-assay kit.

• Tuberculosis and Respiratory Diseases
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• v.53 no.5
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• pp.497-509
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• 2002

Background : The mechanisms through which cellular activation results in intracellular mycobacterial killing is only partially understood. However, in vitro studies of human immunity to Mycobacterium tuberculosis have been largely modeled on the work reported by Crowle, which is complicated by several factors. The whole blood culture is simple and allows the simultaneous analysis of the relationship between bacterial killing and the effect of effector cells and humoral factors. In this study, we attempted to determine the extent to which M. tuberculosis is killed in a human whole blood culture and to explore the role of the host and microbial factor in this process. Methods : The PPD positive subject were compared to the umbilical cord blood and patients with tuberculosis, diabetes and lung cancer. The culture is performed using heparinized whole blood diluted with a culture medium and infected with a low number of M. avium or M. tuberculosis $H_{37}Ra$ for 4 days by rotating the culture in a $37^{\circ}C$, 5% $CO_2$ incubator. In some experiments, methlprednisolone- or pentoxifyline were used to inhibit the immune response. To assess the role of the T-cell subsets, CD4+, CD8+ T-cells or both were removed from the blood using magnetic beads. The ${\Delta}$ log killing ratio was defined using a CFU assay as the difference in the log number of viable organisms in the completed culture compared to the inoculum. Results : 1. A trend was noted toward the improved killing of mycobacteria in PPD+ subjects comparing to the umbilical cord blood but there was no specific difference in the patients with tuberculosis, diabetes and lung cancer. 2. Methylprednisolone and pentoxifyline adversely affected the killing in the PPD+ subjects umbilical cord blood and patients with tuberculosis. 3. The deletion of CD4+ or CD8+ T-lymphocytes adversely affected the killing of M. avium and M. tuberculosis $H_{37}Ra$ by PPD+ subjects. Deletion of both cell types had an additive effect, particularly in M. tuberculosis $H_{37}Ra$. 4. A significantly improved mycobacterial killing was noted after chemotherapy in patients with tuberculosis and the ${\Delta}$ logKR continuously decreased in a 3 and 4 days of whole blood culture. Conclusion : The in vitro bactericidal assay by human whole blood culture model was settled using a CFU assay. However, the host immunity to M. tuberculosis was not apparent in the human whole blood culture bactericidal assay, and patients with tuberculosis showed markedly improved bacterial killing after anti-tuberculous chemotherapy compared to before. The simplicity of a whole blood culture facilitates its inclusion in a clinical trial and it may have a potential role as a surrogate marker in a TB vaccine trial.

• Clinical and Experimental Reproductive Medicine
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• v.35 no.4
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• pp.309-317
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• 2008

Objectives: The aim of this study was to evaluate the usefulness of Anti-mullerian hormone (AMH) as a predictive marker for ovarian response and cycle outcome in IVF cycles. Methods: From Jan., to Aug., 2007, 111 patients undergoing IVF/ICSI stimulated by short or antagonist protocol were selected. On cycle day 3, basal serum AMH level and FSH level were measured. The correlation between basal serum AMH or FSH, and COH outcome was analyzed and IVF outcome was compared according to the AMH levels. To determine the threshold value of AMH for poor- and hyper-response, ROC curve was analyzed. Results: Serum AMH showed higher correlation coefficient (r=0.792, p<0.001) with the number of retrieved mature oocyte than serum FSH (r=-0.477, p<0.001). According to ovarian response, FSH and AMH leves showed significant differences among poor, normal, and hyperresponder. For predicting poor (${\leq}2$ oocytes) and hyperresponse (${\geq}17$ oocyets), AMH cut-off values were 0.5 ng/ml (the sensitivity 88.9% and the specificity 89.5%) and 2.5 ng/ml (sensitivity 85.7%, specificity 87.0%), respectively. According to the AMH level, patients were divided into 3 groups: low (${\leq}0.60\;ng/ml$), normal ($0.60{\sim}2.60\;ng/ml$), and high AMH (${\geq}2.60\;ng/ml$). The number of retrieved mature oocytes was significantly higher ($2.7{\pm}2.2$, $8.1{\pm}4.8$, $16.5{\pm}5.7$) and total gonadotropin dose was lower ($3530.5{\pm}1251.0$, $2957.1{\pm}1057.6$, and $2219.2{\pm}751.9\;IU$) in high AMH group (p<0.001). There was no significant difference in fertilization rates and pregnancy rates (23.8%, 34.0%, 37.5%) among the groups. Conclusions: Basal serum AMH level correlated better with the number of retrieved mature oocytes than FSH level, suggesting its usefulness for predicting ovarian response. However, IVF outcome was not significantly different according to the AMH levels. Serum AMH level presented good cut-off value for poor- or hyper-responders, therefore it could be useful in prediction of cycle cancellation, gonadotropin dose, and OHSS risk in IVF cycles.

• The Journal of Korean Society for Radiation Therapy
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• v.19 no.2
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• pp.99-106
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• 2007

Purpose: The patient's position and anatomy during the treatment course little bit varies to some extend due to setup uncertainties and organ motions. These factors could affected to not only the dose coverage of the gross tumor but over dosage of normal tissue. Setup uncertainties and organ motions can be minimized by precise patient positioning and rigid immobilization device but some anatomical site such as prostate, the internal organ motion due to physiological processes are challenge. In planning procedure, the clinical target volume is a little bit enlarged to create a planning target volume that accounts for setup uncertainties and organ motion as well. These uncertainties lead to differences between the calculated dose by treatment planning system and the actually delivered dose. The purpose of this study was to evaluate the differences of interfractional displacement of organ and GTV based on the tomoimages. Materials and Methods: Over the course of 3 months, 3 patients, those who has applied rectal balloon, treated for prostatic cancer patient's tomoimage were studied. During the treatment sessions 26 tomoimages per patient, Total 76 tomoimages were collected. Tomoimage had been taken everyday after initial setup with lead marker attached on the patient's skin center to comparing with C-T simulation images. Tomoimage was taken after rectal balloon inflated with 60 cc of air for prostate gland immobilization for daily treatment just before treatment and it was used routinely in each case. The intrarectal balloon was inserted to a depth of 6 cm from the anal verge. MVCT image was taken with 5 mm slice thickness after the intrarectal balloon in place and inflated. For this study, lead balls are used to guide the registration between the MVCT and CT simulation images. There are three image fusion methods in the tomotherapy, bone technique, bone/tissue technique, and full image technique. We used all this 3 methods to analysis the setup errors. Initially, image fusions were based on the visual alignment of lead ball, CT anatomy and CT simulation contours and then the radiation therapist registered the MVCT images with the CT simulation images based on the bone based, rectal balloon based and GTV based respectively and registered image was compared with each others. The average and standard deviation of each X, Y, Z and rotation from the initial planning center was calculated for each patient. The image fusions were based on the visual alignment of lead ball, CT anatomy and CT simulation contours. Results: There was a significant difference in the mean variations of the rectal balloon among the methods. Statistical results based on the bone fusion shows that maximum x-direction shift was 8 mm and 4.2 mm to the y-direction. It was statistically significant (P=<0.0001) in balloon based fusion, maximum X and Y shift was 6 mm, 16mm respectively. One patient's result was more than 16 mm shift and that was derived from the rectal expansions due to the bowl gas and stool. GTV based fusion results ranging from 2.7 to 6.6 mm to the x-direction and 4.3$\sim$7.8 mm to the y-direction respectively. We have checked rotational error in this study but there are no significant differences among fusion methods and the result was 0.37$\pm$0.36 in bone based fusion and 0.34$\pm$0.38 in GTV based fusion.

• Tuberculosis and Respiratory Diseases
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• v.59 no.3
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• pp.286-297
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• 2005

Background : Non-small cell lung cancer (NSCLC) is a common cause of cancer-related death in North America and Korea, with an overall 5-year survival rate of between 4 and 14%. The TNM staging system is the best prognostic index for operable NSCLC . However, epidermal growth factor receptor (EGFR), matrix metalloproteinase-9(MMP-9), and C-erbB-2 have all been implicated in the pathogenesis of NSCLC and might provide prognostic information. Methods : Immunohistochemical staining of 81 specimens from a resected primary non-small cell lung cancer was evaluated in order to determine the role of the biological markers on NSCLC . Immunohistochemical staining for EGFR, MMP-9, and C-erbB-2 was performed on paraffin-embedded tissue sections to observe the expression pattern according to the pathologic type and surgical staging. The correlations between the expression of each biological marker and the survival time was determined. Results : When positive immunohistochemical staining was defined as the extent area>20%(more than Grade 2), the positive rates for EGFR, MMP-9, and C-erbB-2 staining were 71.6%, 44.3%, and 24.1% of the 81 patients, respectively. The positive rates of EGFR and MMP-9 stain for NSCLC according to the surgical stages I, II, and IIIa were 75.0% and 41.7%, 66.7% and 47.6%, and 76.9% and 46.2%, respectively. The median survival time of the EGFR(-) group, 71.8 months, was significantly longer than that of the EGFR(+) group, 33.5 months.(p=0.018, Kaplan-Meier Method, log-rank test).. The MMP-9(+) group had a shorter median survival time than the MMP-9(-) group, 35.0 and 65.3 months, respectively (p=0.2). The co-expression of EGFR and MMP-9 was associated with a worse prognosis with a median survival time of 26.9 months, when compared with the 77 months for both negative-expression groups (p=0.0023). There were no significant differences between the C-erbB-2(+) and C-erbB-2 (-) groups. Conclusion : In NSCLC, the expression of EGFR might be a prognostic factor, and the co-expression of EGFR and MMP-9 was found to be associated with a poor prognosis. However, C-erbB-2 expression had no prognostic significance.