• Toxicological Research
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• v.33 no.2
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• pp.107-118
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• 2017

Although alternative test methods based on the 3Rs (Replacement, Reduction, Refinement) are being developed to replace animal testing in reproductive and developmental toxicology, they are still in an early stage. Consequently, we aimed to develop alternative test methods in male animals using mouse spermatogonial stem cells (mSSCs). Here, we modified the OECD TG 489 and optimized the in vitro comet assay in our previous study. This study aimed to verify the validity of in vitro tests involving mSSCs by comparing their results with those of in vivo tests using C57BL/6 mice by gavage. We selected hydroxyurea (HU), which is known to chemically induce male reproductive toxicity. The 50% inhibitory concentration ($IC_{50}$) value of HU was 0.9 mM, as determined by the MTT assay. In the in vitro comet assay, % tail DNA and Olive tail moment (OTM) after HU administration increased significantly, compared to the control. Annexin V, PI staining and TUNEL assays showed that HU caused apoptosis in mSSCs. In order to compare in vitro tests with in vivo tests, the same substances were administered to male C57BL/6 mice. Reproductive toxicity was observed at 25, 50, 100, and 200 mg/kg/day as measured by clinical measures of reduction in sperm motility and testicular weight. The comet assay, DCFH-DA assay, H&E staining, and TUNEL assay were also performed. The results of the test with C57BL/6 mice were similar to those with mSSCs for HU treatment. Finally, linear regression analysis showed a strong positive correlation between results of in vitro tests and those of in vivo. In conclusion, the present study is the first to demonstrate the effect of HU-induced DNA damage, ROS formation, and apoptosis in mSSCs. Further, the results of the current study suggest that mSSCs could be a useful model to predict male reproductive toxicity.

• Restorative Dentistry and Endodontics
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• v.32 no.2
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• pp.151-161
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• 2007

Dental caries is the most common disease in the oral cavity However, the mechanism and treatment of dental caries is not completely understood since many complex factors are involved. Especially the effect of pH on remineralization of early stage of dental caries is still controversial In this study, dental caries in dentin was induced by using lactic acidulated buffering solutions and the loss or inorganic substance was measured. Also decalcified specimens were remineralized by three groups of solution with different pH (group of pH 4.3, 5.0, and 5.5). Then, the amount and the area of inorganic substance precipitation was quantitatively analyzed with microradiograph. Also a qualitative comparison of the normal phase the demineralized phase, and the remineralized phase of hydroxyapatite crystal was made under SEM. The results were as follows, 1. In microradiograghic analysis, as the pH increased, the amount of remineralization in decalcified dentin tended to increase significantly As the pH decreaced, deeper decalcification, however, occurred along with remineralization. The group of pH 5.5 had a tendency to be remineralized without demineralization (p<0.05). 2. In SEM view, the remineralization in dentine caries occurred from the hydroxyapatite crystal surface surrounding the mesh of organic matrix, and eventually filled up the demineralized area. 3. 5 days after remineralization, hydroxyapatite crystal grew bigger with deposition of inorganic substance in pH 4.3 and 5.0 group, and the crystal in the remineralized area appeared to return to normal. After 10 days, the crystals in group of pH 4.3 and 5.0, which grew bigger after 5 days of remineralization, turned back to their normal size, but in group of pH 5.5, some crystals were found to double their size. In according to the results of this experiment, the decalcifying and remineralizing process of dentine is neither simple nor independent, but a dynamic process in which decalcification and remineralization occur simultaneously. The remineralization process occurred from the hydroxyapatite crystal surface.

• Korean Journal of Food Science and Technology
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• v.31 no.2
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• pp.465-474
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• 1999

RNase activity of Saccharomyces cerevisiae ATCC 7754 was investigated to obtain strains with high ribonucleic acid (RNA) content. The yeast strain contained two RNase activities; an acidic RNase with a optima of pH $3{\sim}4$ and an alkaline RNase with a optima pH 9. The acidic RNase activity was inhibited by $0.08\;M\;HgCl_{2}$ most drastically. The alkaline RNase activity was inhibited by 2.0 M NaCl or KCl, while enhanced by addition of $0.05\;M\;CaCl_{2},\;0.02\;M\;ZnSO_{4},\;or\;0.008\;M\;HgCl_{2}$. Various mutants of Saccharomyces cerevisiae ATCC 7754 were isolated by ethylmethane sulfonate (EMS) treatment or $\gamma$-ray/ultra violet irradiation. Among the mutants that were sensitive to high concentration of KCl which inhibits alkaline RNase, B24 was selected for high RNA content per culture volume. Growth characteristics of the mutant were comparable to those of the mother strain with optimum growth at pH $4.5{\sim}5.5$. The mutant accumulated higher content of RNA than the mother strain when glucose was used as the carbon source. However, both growth rate and total RNA content of the mutant were higher in molasses medium than in glucose medium. RNA content of the mutant increased rapidly during the early stage of growth, and then decreased gradually until the culture reached stationary phase by a fed-batch culture in a 5 L jar fermenter. Maximal cell harvest and the final RNA content using the mutant B24 were 69.6 g/L culture broth and 19.8 g/100 g of the dry cell while those using the mother strain were 68 g/L culture broth and 16.1 g/100 g of dry cell, respectively.

• Tuberculosis and Respiratory Diseases
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• v.46 no.4
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• pp.542-547
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• 1999

Background: Decision in mediastinal lymph node involvement of lung cancer by CT scan is very important and valuable for the treatment planning and prognosis prediction. In general, long diameter of mediastinal lymph node more than 15mm is used as criterion of lung cancer involvement. Adenocarci-noma has a tendency of early distant metastasis and micrometastasis, so adenocarcinoma may involve lymph node earlier and cannot be detected before lymph nodes are enlarged enough. The authors tried to determine the difference between two size criteria(15mm, 10mm) in adenocarcinoma for the detection of cancer involvement. Methods: Numbers of sample are 60 cases(male 46, female 14, median age: 61.5 years). According to pathology, squamous cancer 41, large cell cancer 2, adenocarcinoma 17. According to TNM stage, I 23, III 24, IIIA 13. Results : Mean long diameter of lymph node involvement is 16.0($\pm8.0$) mm in non-adenocarcinoma group, and that of adenocarcinoma group is 12.0($\pm3.2$) mm(p<0.05). If long diameter of lymph node larger than 15mm as involvement criterion is applied, sensitivity, specificity, positive predictive index, negative predictive index, accuracy of nonadenocarcinoma group are 54%, 100%, 100%, 83%, 86%, and those of adenocarcinoma group are 43%, 90%, 75%, 69%, 71%. If long diameter of lymph node larger than 10mm as involvement criterion is applied, sensitivity, specificity, positive predictive index. negative predictive index. accuracy of nonadenocarcinoma group are 65%, 77%, 61%, 92%, 79%, and those of adenocarcinoma group are 100%, 80%, 78%, 100%, 88%. Conclusion: Long diameter of lymph node larger than 10mm is more valuable criterion as lymph node involvement in adenocarcinoma of lungs.

• The Journal of Dong Guk Oriental Medicine
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• v.4
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• pp.1-23
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• 1995

The aim of this study is to compare the diseases which are in the Sun Woo-Yees charts with the diseases which are in the medical books excavated in MA WANG TEI(馬王堆), "YOUNG COO(靈樞)" and "SO MUN(素問)", and to investigate their interactions. The results obtained are as follows : 1. JOE(疽) which is made in stomach and bowels is not found at the medical books excavated in MA WANG TUI(馬王堆), "YOUNG COO(靈樞)" and "SO MUN(素問)". JOE(疽) which is made on the breast is similar to JOE(疽) which is made on the chest of "YOUNG COO(靈樞)". 2. It is said in Sun Woo-Yee(淳于意)'s charts that mental depression blocks up the circulation of Kl(氣), and it is the same viewpoint of "YOUNG COO(靈樞)"and "SO MUN(素問)". 3. The POONG GYUL(風厥) and the YOUL GYUL(熱厥) that are found in the Sun Woo-Yee(淳于意)'s charts is similar to those of "SO MUN(素問)", but different from those of "YOUNG COO(靈樞)". 4. It is regarded that YONG SAN(涌疝) in the Sun Woo-Yee(淳于意)'s charts is similar to the COONG SAN(퇴산) in "SO MUN(素問)", KI SAN(氣疝) in the Sun Woo-Yee(淳于意)'s charts is related to the TUI SAN(癡疝) of the medical books excavated in MA WANG TUI(馬王堆), "YOUNG COO(靈樞)" and "SO MUN(素問)", MO SAN(牡疝) in the Sun Woo-Yee(淳于意)'s charts is considered PE SAN(肺疝) and PE POONG SAN(肺風疝) in "SO MUN(素問)", but more and deeper study is required. In the change of the terms ; from TUI SAN(癡疝) to TUI SAN(퇴산), from PYUN SAN(偏疝) to HO SAN(狐疝), it can be guessed that the terms changed, as new doctrines were introduced and reorganiged with the development of the medical theory. 5. POONG BI in the Sun Woo-Yee(淳于意)'s charts is not found in "YOUNG COO(靈樞)", but it is similar to DAN FEVER(단열) in "SO MUN(素問)". It is regareded that PE SO DAN(폐소단) in the Sun Woo-Yee(淳于意)'s charts is the same as that of "YOUNG COO(靈樞)". 6. DONG PUNG in Sun Woo-Yee(淳于意)'s charts is not found in "YOUNG COO(靈樞)"and "SO MUN(素問)", but it seems like DONG SOEL(洞泄) in "YOUNG COO(靈樞)". It is regareded that the DONC PUNG(동풍) in the Sun Woo-Yee(淳于意)'s charts was at first changed into DONG SOEL(洞泄), and later differentiated into DOHG SOEL(洞泄) and SON SOEL. 7. In the Sun Woo-Yee(淳于意)'s charts, the treatment of the decayed tooth had the classification of the right and left, seeing cauterization with moxa on SU YANG MYOUNG MAEK(手陽明脈), it is considered that the tooth was decayed in the lower right. region. A tooth was related to QI MAEK(齒脈) and BI YANG MYUNG MAEK(臂陽明脈) in the early stage, but gradually was related to not only SU YANG MYUNG MAEK(手陽明脈) but also JOK YANG MYUNG MAEK(足陽明脈), JOK SO YIN MAEK(足少陰脈), JOK TAE YIN MAEK(足太陰脈), and in regards to the tooth and KYUNG MAEK(經脈), "YOUNG COO(靈樞)" emphasised the course of the KYUNG MAEK(經脈), "SO MUN(素問)" emphasised the attachment of the five elements. 8. In regards to BI(痺), the importance of the five elements theory given in the Sun Woo-Yee(淳于意)'s charts is similar to that of "SO MUN(素問)", and "YOUNG COO(靈樞)" and the medical books excavated in MA WANG TUI(馬王堆) give the same importance to the Kyung MAEK(經脈) theory. 9. The explanation of the irregular menstruation in the Sun Woo-Yee(淳于意)'s charts was fundamentally similar to that of "SO MUN(素問)", but it shows the another theory that can not found in "SO MUN(素問)".

• Journal of the Korean Society of Food Science and Nutrition
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• v.30 no.4
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• pp.657-661
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• 2001

Osteoporosis that is associated with ovarian hormone deficiency following menopause (postmenopausal osteoporosis) is by far the most common cause of age-related bone loss. Isoflavone has been reported as a natural substance that possibly minimizes bone loss in postmenopausal women. This study was conducted to investigate the preventing, treating effects of isoflavone on bone loss in ovariectomized rats. 120 Sprague Dawley rats of 13 week-old were devided into 2 groups, a treatment group and prevention group. Each group was consisted of six subgroups; control (CON), sham operated (SH) or ovariectomized (OVX) and isoflavone supplemented goups: OVX+0.25mg isoflavone/kg diet (OL), OVX+0.8mg isoflavone/kg diet(OM) and OVX+2.5mg isoflavone/kg diet(OH). to study the preventing effects of isoflavone on bone loss, OL, OM and OH groups were fed with isoflavone from 4 days after ovariectomization. Treating effects of isoflavone on bone metabolism were investigated with OL, OM, OH groups supplemented with isoflavone from 8 weeks after ovariectomization. Isoflavone supplementation continued for 8 weeks. At 8 weeks after ovariectomization significant increase in alkaline phosphatase occurred comparing with CON and SH group. By isoflavone supplementation from 4 days after ovariectomy alkaline phosphatase and urinary hydroxyproline were lowered and bone mineral density, bone strength of the femur and tibia and bone dry weight were slightly enhanced with no significant difference. Isoflavone supplemented group at the level of 0.8mg/kg diet (OM group) had significantly lower serum alkaline phosphatase, urinary hydroxyproline, and higher strength of femur than OVX group. Groups with isoflavone supplementation fro 8 weeks after ovariectomy had lower level of serum alkaline phosphatase, urinary hydroxyproline than OVX group. Bone mineral density, bone dry weight and bone strength of the femur and tibia were slightly enhanced by isoflavone supplementation. However there was no significanct difference between OVS ad isoflavone supplementation groups. The results suggest that isoflavone might have potential role for preventing postmenopausal bone loss. Isoflavone supplementation at early stage of postemenopause may be beneficial to age-related bone health.

• Journal of Periodontal and Implant Science
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• v.26 no.2
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• pp.469-490
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• 1996

The initial events required for periodontal regeneration is the attachment, spreading, and proliferation of appropriated cells at the healing sites. These have been reported that minocycline stimulates the attachment of periodontal ligament cells, and also $TGF-{\beta}1$ enhances the proliferation of periodontal ligament cells. The purpose of the present study was to evaluate the effects of $TGF-{\beta}1$ on the cellular activity of minocycline treated human periodontal ligament cells. Periodontal ligament cells were obtained from the explants of healthy periodontal ligaments of extracted 3rd molars or premolar teeth extracted from the patients for orthodontic treatment. The cells were cultured in minimal essential medium(${\alpha}-MEM$) supplemented with 10.000units/ml penicillin, $10,000{\mu}g/ml$ streptomycin and 10% FBS(fetal bovine serum) at $37^{\circ}C$ in a humidified atmosphere of 5% carbon dioxide and the 5th to the 8th passages of the cells were used. To evaluate the effect of minocycline on cell attachment, the cells were seeded at a cell density of $1.5{\times}10^4$ cells/well in 24-well culture plates and treated with $20{\mu}g/ml$ and $100{\mu}g/ml$ of minocycline for 1.5 h. After trypsinization, the cells were counted with hemocytometer and were taken photographs for observation of cellular morphology. To evaluate the effect of $TGF-{\beta}1$ on minocycline-pretreated periodontal ligament cells, the cells were seeded at a cell density of $1{\times}10^4$ cells/ well in 24-well culture plates and treated with $20{\mu}g/ml$ and $100{\mu}g/ml$ of minocycline for 1.5 h. After incubation, 1 and 10ng/ml of $rh-TGF-{\beta}1$ were also added to the each well and incubated for 1 and 2 days, respectively. Then, MTT assay, DNA synthesis($^3H-thymidine\;assay$), and protein and collagen assay(3H-proline assay) were carried out. In the MIT assay, after 200ul MTT solutionlconeentration of 5mg/ml) were added to the each well of the 24-well plates and incubated for 3 hours, and 200 ul DMSO were added so as to dissolve insoluble blue formazan crystals which was formed in incubated period. Then it read plates on a ELISA reader. For mitogenic assay, 1 uCi/ml $^3H-thymidine$ was added to each well for the final 2 hours of the incubation periods. After labeling, the wells were washed 3 times with ice cold PBS and 4 times with 5% TCA to remove unincorporated label and precipitate the cellular DNA. DNA, with the incorporated $^3H-thymidine$, was solubilized with 500 ul of 0.1% NaOH/0.1% SDS. A 250 ul aliquot was removed from each well and placed in a scintillation vial with 4ml of scintillation cocktail. Using an liguid scintillation counter, counts per minute(CPM) were determined for each samples. 3 uCi/ml $^3H-proline$ was added to each well for the final 4 hours of the incubation periods and total protein and percent collagen synthesis were carried out. The results indicate that minocycline treated group with $100{\mu}g/ml$ concentration for 1.5 hours significantly increased than that of control in cell attachment, and cell process is also evident compared with that of control in cell morphology, and the cellular activity and DNA synthesis rate of cells treated minocycline and $TGF-{\beta}1$ significantly increased than that of control values, but were below to values of the $TGF-{\beta}1$ only treated group in MIT assay and $^3H-thymidine\;assay$, and the total protein synthesis of minocycline and $TGF-{\beta}1$ treated group also significantly increased than that of control values, but the percent collagen synthesis of tested group significantly decreased to compared with control. On the above the findings, the tested group of minocycline and $TGF-{\beta}1$ did not increase the effect on the cell activity than $TGF-{\beta}1$ only tested group and the tested group of minocycline inhibited cell activity. This results indicate that minocycline was effective on cell attachment in early stage, but it is harmful to cell activity, that inhibitory effect of minocycline was compensated with stimulatory effect of $TGF-{\beta}1$.

• Tuberculosis and Respiratory Diseases
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• v.45 no.4
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• pp.776-784
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• 1998

Background: The cyclin D1 gene is one of the most frequently amplified chromosomal regions(11q13) in human carcinomas. In laryngeal and head and neck carcinomas, its overexpression has been shown to be associated with advanced local invasion and presence of lymph node metastases. Cyclin D1 may therefore playa key role in cell growth regulation and tumorigenesis. Lung cancer is a worldwide problem and in many contries it is the most lethal malignancy. As relapse is frequent after resection of early stage non-small cell lung cancer, there is an urgent need to define prognostic factors. Purpose: This study was undertaken to evaluate the prognostic value of the cyclin D1, that is one the G1 cyclins which control cell cycle progression by allowing G1 to S phase transition, on the patients in radically resected non-small cell lung cancer. Method: Total 81 cases of formalin-fixed paraffin-embedded blocks from resected primary non-small cell lung cancer from January 1, 1983 to July 31, 1995 at Hanyang University Hospital were available for both clinical follow-up and immunohistochemical staining using monoclonal antibodies for cyclin D1. Results : The histologic classification of the tumor was based on WHO criteria, and the specimens included 45 squamous cell carcinomas, 25 adenocarcinomas and 11 large cell carcinomas. Cyclin D1 overexpression was noted in 26 cases of 81 cases tested (30.9%). Cyclin D1 expression was not significantly associated with cell types of the tumor, pathological staging and the size of the tumor. But cyclin D1 overexpression was significantly correlated with positive lymph node metastasis(p=0.035). The mean survival duration was $22.76{\pm}3.50$ months in cyclin D1 positive group and $45.38{\pm}5.64$ months in eyclin D1 negative group. There was a nearly significant difference in overall survival between cyclin D1 positive and negative groups(p=0.0515) in radically resected non-small cell lung cancer. Conclusion: Based on this study, cyelin D1 overexpression appears an important poor prognostic indicator in non-small cell lung cancer and may have diagnostic and prognostic importance in the treatment of resectable non-small cell lung cancer.

• Tuberculosis and Respiratory Diseases
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• v.60 no.6
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• pp.645-652
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• 2006

Objective: Patients with lung cancer have a relative high risk of developing secondary primary lung cancers. This study examined the additional value of autofluorescence bronchoscopy (AFB) for diagnosing synchronous lung cancers and premalignant lesions. Methods: Patients diagnosed with lung cancer from January 2005 to December 2005 were enrolled in this study. The patients underwent a lung cancer evaluation, which included white light bronchoscopy (WLB), followed by AFB. In addition to the primary lesions, any abnormal or suspicious lesions detected during WLB and AFB were biopsied. Results: Seventy-six patients had non-small cell lung cancer (NSCLC) and 23 had small cell lung cancer (SCLC). In addition to the primary lesions, 84 endobronchial biopsies were performed in 46 patients. Five definite synchronous cancerous lesions were detected in three patients with initial unresectable NSCLC and in one with SCLC. The secondary malignant lesions found in two patients were considered metastatic because of the presence of mediastinal nodes or systemic involvement. One patient with an unresectable NSCLC, two with a resectable NSCLC, and one with SCLC had severe dysplasia. The detection rate for cancerous lesions by the clinician was 6.0% (6/99) including AFB compared with 3.0% (3/99) with WLB alone. The prevalence of definite synchronized cancer was 4.0% (4/99) after using AFB compared with 2.0% (2/99) before, and the staging-up effect was 1.0% (1/99) after AFB. Since the majority of patients were diagnosed with advanced disease, the subjects with newly detected cancerous lesions did not have their treatment plans altered, except for one patient with a stage-up IV NSCLC who did not undergo radiotherapy. Conclusions: Additional AFB is effective in detecting early secondary cancerous lesions and is a more precise tool in the staging workup of patients with primary lung cancer than with WLB alone.

• Journal of Genetic Medicine
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• v.7 no.2
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• pp.160-164
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• 2010

Structural abnormalities of the Y chromosome affect normal testicular differentiation and spermatogenesis. The present case showed a rare monocentric derivative Y chromosome with partial duplication of Yp including the SRY gene and deletion of Yq12 heterochromatin. The karyotype was 46,X,der(Y)(pter${\rightarrow}$q11.23::p11.2${\rightarrow}$pter).ish der(Y)(DYZ3+,DYZ1-,SRY++), confirmed through a FISH study. Even though the patient possessed an abnormal Y chromosome, testicular biopsy showed normal testicular volumes in the proband, with gonadal hormonal levels in the normal range but bilateral varicocele and hypospermatogenesis. We speculate that the abnormal Y chromosome arose from sister chromatids during Y chromosome recombination or intra chromosomal NAHR (non-allelic homologous recombination) during meiosis in the patient's father or in the very early stages of embryogenesis. The derivative Y chromosome might interfere in the meiotic stage of spermatogenesis, leading to the developmental arrest of germ cells. The present case illustrates that the infertility phenotype can have various causes. Also, it emphasizes the importance of accurate and various genetic analyses and could aid in male infertility treatment.