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Antrum Formation and Growth of Mouse Pre-antral Follicles Cultured in Two Different Culture Media without Hormones

  • Kim, Ju-Hwan;Kim, Hwan-Tae;Park, Kee-Sang;Song, Hai-Bum;Chun, Sang-Sik
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.8-8
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    • 2001
  • Mouse follicles require the addition of gonadotropins (Gns) to complete maturation and ovulation of oocyte and antrum formation of follicles in vitro. However, we tried examination of in vitro growth of mouse pre-antral follicles in medium without Gns and physiological factors. And also, pre-antral follicles were isolated from ovaries by mechanical method. Our present studies were conducted to evaluate on the growth of follicles and intra-follicular oocytes and antrum formation in vitro of mouse pre-antral follicles in two different media. Pre-antral follicles (91-120${\mu}{\textrm}{m}$) were isolated mechanically by fine 30G needles not using enzymes from ovary of 3-6 weeks old female ICR mice. Isolated pre-antral follicles were cultured in 20 ${mu}ell$ droplets of TCM (n=17; follicles: 107.8 $\pm$ 1.58 ${\mu}{\textrm}{m}$; oocytes: 59.9$\pm$1.2 ${\mu}{\textrm}{m}$) or MEM (n=12; follicles: 109.3$\pm$2.53 ${\mu}{\textrm}{m}$; oocytes: 55.4 $\pm$1.6${\mu}{\textrm}{m}$) under mineral oil on the 60mm culture dish. All experimental media was supplemented with 10% FBS but without Gns and/or physiological factors. Pre-antral follicles were individually cultured in drops for 8 days. Antrum formation and growth of pre-antral follicles and intra-follicular oocytes were evaluated using a precalibrated ocular micrometer at $\times$200 magnifications during in vitro culture. Results between different groups were analyzed using combination of Student's t-test and Chi-square, and considered statistically significant when P<0.05. Antrum formation of pre-antral follicles had started in two culture media on day-2. On day-8, antrum formation had occurred in 58.3%(7/12) of pre-antral follicles cultured in MEM, but only in 23.5% (4/17) of those cultured in TCM (P=0.0364). Growth of pre-antral follicles and intra-follicular oocytes were observed on day-4 and -8. On day-4, follicular diameters was similar (P=0.1338) in TCM (119.4$\pm$2.58 ${\mu}{\textrm}{m}$) and MEM (125.4$\pm$4.52 ${\mu}{\textrm}{m}$). However, on day-8, diameters of pre-antral follicles cultured in MEM (168.9$\pm$17.29 ${\mu}{\textrm}{m}$) was significantly (P=0.0248) bigger than that in TCM (126.7$\pm$4.28 ${\mu}{\textrm}{m}$). On day-4 and -8, diameters of intra-follicular oocytes were similar TCM (67.1$\pm$1.3 and 72.4$\pm$0.9${\mu}{\textrm}{m}$) and MEM (65.2$\pm$1.7 and 73.3$\pm$1.5 ${\mu}{\textrm}{m}$), respectively. We can conform that medium not supplemented with Gns and/or physiological factors can be used for in vitro antrum formation and growth of mouse pre-antral follicles and intra-follicular oocytes. In conclusion, MEM supplemented with FBS can be used for growth in vitro of mouse pre-antral follicles isolated mechanically.

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β-elemene Induces Caspase-dependent Apoptosis in Human Glioma Cells in vitro through the Upregulation of Bax and Fas/FasL and Downregulation of Bcl-2

  • Li, Chen-Long;Chang, Liang;Guo, Lin;Zhao, Dan;Liu, Hui-Bin;Wang, Qiu-Shi;Zhang, Ping;Du, Wen-Zhong;Liu, Xing;Zhang, Hai-Tao;Liu, Yang;Zhang, Yao;Xie, Jing-Hong;Ming, Jian-Guang;Cui, Yu-Qiong;Sun, Ying;Zhang, Zhi-Ren;Jiang, Chuan-Lu
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.23
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    • pp.10407-10412
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    • 2015
  • Background: ${\beta}$-elemene, extracted from herb medicine Curcuma wenyujin has potent anti-tumor effects in various cancer cell lines. However, the activity of ${\beta}$-elemene against glioma cells remains unclear. In the present study, we assessed effects of ${\beta}$-elemene on human glioma cells and explored the underlying mechanism. Materials and Methods: Human glioma U87 cells were used. Cell proliferation was determined with MTT assay and colony formation assay to detect the effect of ${\beta}$-elemene at different doses and times. Fluorescence microscopy was used to observe cell apoptosis with Hoechst 33258 staining and change of glioma apoptosis and cell cycling were analyzed by flow cytometry. Real-time quantitative PCR and Western-blotting assay were performed to investigated the influence of ${\beta}$-elemene on expression levels of Fas/FasL, caspase-3, Bcl-2 and Bax. The experiment was divided into two groups: the blank control group and ${\beta}$-elemne treatment group. Results: With increase in the concentration of ${\beta}$-elemene, cytotoxic effects were enhanced in the glioma cell line and the concentration of inhibited cell viability ($IC_{50}$) was $48.5{\mu}g/mL$ for 24h. ${\beta}$-elemene could induce cell cycle arrest in the G0/G1 phase. With Hoechst 33258 staining, apoptotic nuclear morphological changes were observed. Activation of caspase-3,-8 and -9 was increased and the pro-apoptotic factors Fas/FasL and Bax were upregulated, while the anti-apoptotic Bcl-2 was downregulated after treatment with ${\beta}$-elemene at both mRNA and protein levels. Furthermore, proliferation and colony formation by U87 cells were inhibited by ${\beta}$-elemene in a time and does-dependent manner. Conclusions: Our results indicate that ${\beta}$-elemene inhibits growth and induces apoptosis of human glioma cells in vitro. The induction of apoptosis appears to be related with the upregulation of Fas/FasL and Bax, activation of caspase-3,-8 and -9 and downregulation of Bcl-2, which then trigger major apoptotic cascades.

Heparanase mRNA and Protein Expression Correlates with Clinicopathologic Features of Gastric Cancer Patients: a Meta-analysis

  • Li, Hai-Long;Gu, Jing;Wu, Jian-Jun;Ma, Chun-Lin;Yang, Ya-Li;Wang, Hu-Ping;Wang, Jing;Wang, Yong;Chen, Che;Wu, Hong-Yan
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.18
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    • pp.8653-8658
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    • 2016
  • Background: Heparanase is believed to be involved in gastric carcinogenesis. However, the clinicopathologic features of gastric cancer with high heparanase expression remain unclear. Aim : The purpose of this study was to comprehensively and quantitatively summarize available evidence for the use of heparanase mRNA and protein expression to evaluate the clinicopathological associations in gastric cancer in Asian patients by meta-analysis. Materials and Methods: Relevant articles listed in MEDLINE, CNKI and the Cochrane Library databases up to MARCH 2015 were searched by use of several keywords in electronic databases. A meta-analysis was performed to clarify the impact of heparanase mRNA and protein on clinicopathological parameters in gastric cancer. Combined ORs with 95%CIs were calculated by Revman 5.0, and publication bias testing was performed by stata12.0. Results: A total of 27 studies which included 3,891 gastric cancer patients were combined in the final analysis. When stratifying the studies by the pathological variables of heparanase mRNA expression, the depth of invasion (633 patients) (OR=4.96; 95% CI=2.38-1.37; P<0.0001), lymph node metastasis (639 patients) (OR=6.22; 95%CI=2.70-14.34, P<0.0001), and lymph node metastasis (383 patients) (OR=6.85; 95% CI=2.04-23.04; P=0.002) were all significant. When stratifying the studies by the pathological variables of heparanase protein expression, this was the case for depth of invasion (1250 patients) (OR=2.76; 95% CI=1.52-5.03; P=0.0009), lymph node metastasis (1178 patients) (OR=4.79 ; 95% CI=3.37-6.80, P<0.00001), tumor size (727 patients) (OR=2.06 ; 95% CI=1.31-3.23; P=0.002) (OR=2.61; 95% CI=2.09-3.27; P=0.000), and TNM stage (1233 patients) (OR=6.85; 95% CI=2.04-23.04; P=0.002). Egger's tests suggested publication bias for depth of invasion, lymph node metastasis, lymph node metastasis and tumor size of heparanase mRNA and protein expression. Conclusions: This meta-analysis suggests that higher heparanase expression in gastric cancer is associated with clinicopathologic features of depth of invasion, lymph node metastasis and TNM stage at mRNA and protein levels, and of tumor size only at the protein level. Egger's tests suggested publication bias for these clinicopathologic features of heparanase mRNA and protein expression, and which may be caused by shortage of relevant studies. As a result, although abundant reports showed heparanase may be associated with clinicopathologic features in gastric cancer, this meta-analysis indicates that more strict studies were needed to evaluate its clinicopathologic significance.

Implant Isolation Characteristics for 1.25 Gbps Monolithic Integrated Bi-Directional Optoelectronic SoC (1.25 Gbps 단일집적 양방향 광전 SoC를 위한 임플란트 절연 특성 분석)

  • Kim, Sung-Il;Kang, Kwang-Yong;Lee, Hai-Young
    • Journal of the Institute of Electronics Engineers of Korea SD
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    • v.44 no.8
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    • pp.52-59
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    • 2007
  • In this paper, we analyzed and measured implant isolation characteristics for a 1.25 Gbps monolithic integrated hi-directional (M-BiDi) optoelectronic system-on-a-chip, which is a key component to constitute gigabit passive optical networks (PONs) for a fiber-to-the-home (FTTH). Also, we derived an equivalent circuit of the implant structure under various DC bias conditions. The 1.25 Gbps M-BiDi transmit-receive SoC consists of a laser diode with a monitor photodiode as a transmitter and a digital photodiode as a digital data receiver on the same InP wafer According to IEEE 802.3ah and ITU-T G.983.3 standards, a receiver sensitivity of the digital receiver has to satisfy under -24 dBm @ BER=10-12. Therefore, the electrical crosstalk levels have to maintain less than -86 dB from DC to 3 GHz. From analysed and measured results of the implant structure, the M-BiDi SoC with the implant area of 20 mm width and more than 200 mm distance between the laser diode and monitor photodiode, and between the monitor photodiode and digital photodiode, satisfies the electrical crosstalk level. These implant characteristics can be used for the design and fabrication of an optoelectronic SoC design, and expended to a mixed-mode SoC field.

Effect of Epigallocatechin Gallate on Phosphoinositide 3-kinase/Akt and Glycogen Synthase Kinase-3 Pathway in Oxidative-stressed N18D3 Cells Following $H_2O_2$ Exposure (산화성 손상을 받은 N18D3세포에서 Epigallocatechin gallate가 Phosphoinositide 3-kinase/Akt 및 Glycogen synthase kinase-3경로에 미치는 효과)

  • Koh, Seong Ho;Kwon, Hyug Sung;Oh, Hwa Soon;Oh, Jae Ho;Park, Ynun Joo;Kim, Jun Gyou;Kim, Ki Sok;Kim, Yang Soon;Yang, Ki Hwa;Kim, Seung U.;Kim, Seung H.;Jung, Hai Kwu
    • Korean Journal of Clinical Pharmacy
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    • v.13 no.1
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    • pp.29-39
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    • 2003
  • Neurodegenerative disorders are associated with apoptosis as a causing factor or an inducer. On the other hand, it has been reported that epigallocatechin gallate (EUG), one of antioxidants and flavonoids, and z-VAD-fmk, a nonselective caspase inhibitor, suppress oxidative-radical-stress-induced apoptosis. However, it is not yet known what is the effects of EGCG and z-VAD-fmk on the apoptotic pathway is through phosphoinositide 3-kinase (PI3K), Akt and glycogen synthase kinase-3 (GSK-3) as well as mitochondria, caspase-3 and poly (ADP-ribose) polymerase (PARP). We investigated the effects of EGCG by using $H_2O_2$ treated N18D3 cells, mouse DRG hybrid neurons. Methods: Following 30 min $100\;{\mu}m\;H_2O_2$ exposure, the viability of N18D3 cells (not pretreated vs. EGCG or z-VAD-fmk pretreated) was evaluated by using MTT assay. The effect of EGCG on immunoreactivity (IR) of cytochrome c, caspase-3, PARP, PI3K/Akt and GSK-3 was examined by using Western blot, and was compared with that of z-Y4D-fmk. Results: EGCG or z-VAD-fmk pretreated N18D3 cells showed increased viability. Dose-dependent inhibition of caspase-3 activation accompanied by PARP cleavage were demonstrated by pretreatment of both agents. However, inhibition of cytochrome c release was only detected in EGCG pretreated N18D3 cells. On the pathway through PI3K/Akt and GSK-3, however, the result of Western blot in EGCG pretreated N18D3 cells showed decreased IR of Akt and GSK-3 and increased IR of p85a PI3K, phosphorylated Akt and GSK-3, and contrasted with that in z-VAD-fmk pretreated N18D3 cells showing no changes on each molecule. Conclusion: These data show that EGCG affects apoptotic pathway through upstream signal including PI3K/Akt and GSK-3 pathway as well as downstream signal including cytochrome c and caspase-3 pathway. Therefore, these results suggest that EGCG mediated activation of PI3K/Akt and inhibition GSK-B could be new potential therapeutic strategy for neurodegenerative diseases associated with oxidative injury.

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Comparison of Various DNA Extraction Methods for Diagnosis of Tuberculosis Using a Polymerase Chain Reaction (중합효소연쇄반응을 이용한 결핵의 진단에 있어서 각종 DNA 추출방법의 비교)

  • Kim, Ju-Ock;Han, Pyo-Seong;Hong, Seok-Cheol;Lee, Jong-Jin;Cho, Hai-Jeong;Kim, Sun-Young
    • Tuberculosis and Respiratory Diseases
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    • v.40 no.1
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    • pp.43-51
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    • 1993
  • Background: The polymerase chain reaction (PCR) is a very sensitive method for the detecting of mycobacterial DNA. There are many reports revealing the efficacy of PCR for the diagnosis of M. tuberculosis, but there are many different methods for DNA extraction from Mycobacterium tuberculosis. Bead beater method is a very useful method for DNA extraction from clinical spectimens, but its procedures are relatively complicated and time-consuming. So we studied other methods for the DNA extraction from Mycobacterium tuberculosis $H_{37}Rv$ and some clinical specimens (5 smear positive sputa and 5 smear negative CSF). Method: We extracted the mycobacterial DNA with 6 different methods from H37Rv strain and clinical specimens. The methods included SDS-microwave oven method, NaOH lysis method, Triton X-100-Proteinase K method, Lysis buffer method, SDS-proteinase K method and bead beater method. The target DNA was 123bp of IS6110 and was detected by examination of ethidium bromide-stained agarose gels. Results: Among 6 methods, SDS-proteinase K method, bead beater method, lysis buffer method and triton X-100-proteinase K method were excellent, but SDS-proteinase K method was the best method in the aspect of simplicity and cost-effectiveness. Conclusion: We suggest that SDS-porteinase K method is a simple and convinient method and might be the best method for the extraction of mycobacterial DNA.

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Expression of Several Biologic Markers as Prognostic Markers in Non-Small Cell Lung Cancers (폐암조직에서 생물학적 지표들의 예후인자로서의 비교검토)

  • Kim, Sun-Young;Cho, Hai-Jeong;Suh, Ji-Won;Kim, Nam-Jae;Kim, Ju-Ock
    • Tuberculosis and Respiratory Diseases
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    • v.42 no.2
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    • pp.142-148
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    • 1995
  • Background: Despite modern diagnostic, staging, and therapeutic advances, esp. with molecular biologic techniques, the 5-year survival rate of all cases of lung cancer does not exceed 15%. Also, the incidence of lung cancer of both sex in Korea is increasing year by year and the lung cancer is one of the leading causes of cancer death. Therefore, it is strongly needed to develop the new combination of treatment modalities including neoadjuvant chemotherapy and to identify tumor specific characteristics with staging or prognostic markers. Here we present the clinical significance of several biologic tumor markers to use as a prognostic markers in patients with non-small cell lung cancers. Method: The survival has correlated with the expressibility of proliferative cell nuclear antigen (PCNA), epidermal growth factor receptor(EGFR), p53 and/or blood group antigen A(BGAA) using immunohistochemistry in 46 patients with non-small cell lung cancers. Results: 1) The expression rates of PCNA, EGFR, p53 and BGAA were 80.6%, 61.3%, 45.9% and 64.3%, respectively and those were not correlated to cell types or clinical stges. 2) The expression of BGAA was correlated with better survival in median survival and in 2-year survival rate and that of PCNA was correlated with worse survival in median survival and 2-year survival rate. 3) The expression of EGFR or p53 was not valuable to predict prognosis in non-small cell lung cancers. 4) With simultaneous applications of PCNA, EGFR and p53 immunostain, the patients with 2 or more negative expressions showed better prognosis than the patients with 2 or more positive expressions. Conclusion: It is suggested that the expression of blood group antigen may be a positive prognostic factor and that of PCNA may be a negative prognostic factor. Also, the combination of expressions of PCNA, EGFR and p53 may be used as a negative prognostic factor.

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A Study on the State of Health Functional Foods & Herbal Medicine Consumed by Elementary School Students (초등학생의 건강기능식품 및 한약 복용 실태에 대한 연구)

  • Kim, Mi-Ki;Jung, Ji-Ho;Ahn, Jae-Sun;Yim, Jung-Hoon;An, Min-Seop;Park, Jin-Su;Lee, Hai-Ja;Park, Eun-Jung
    • The Journal of Pediatrics of Korean Medicine
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    • v.23 no.3
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    • pp.143-153
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    • 2009
  • Objectives The purpose of this study is to get the basic information from patients how much they understood about their medication and also to know whether patients are making reasonable drug choice between Health Functional Foods and Herbal medicine. Methods 500 questionnaires were handed out to the parents of students in two elementary schools located in OO, Junlabukdo province. 421 questionnaires were completed to be evaluated. Results Among 421 subjects, 53.0% were female, and 47.0% were male. The percentages of the subjects consuming Health Functional Foods and herbal medicine were67.7% and 67.8%, respectively. Among those people who consumed Health Functional Food, 44.1% were using nutritional supplements, red ginseng or ginseng products (26.9%), chlorella products (11.5%), and plum extract products (7.7%). As for the reason to consume Health Functional Foods were varied, but 'in order to be healthy, although currently displaying no illness.'(43.0%) were the most responses among the given choices. On the other hand, the reason for consuming herbal medicine was 'In order to grow taller'(26.1%), 'In order to cure weak physical state frequently displaying common illnesses',(25.9%), and 'In order to cure diseases.'(23.3%). For the questions about effectiveness after consumption,the 69.9% subjects said that it seemed to be effected, and that % was slightly higher than that of subjects with consuming Health Functional Foods(64.4%). For question concerning preferences between Health Functional Foods and herbal medicine, 57.5% chose herbal medicine, and this percentage was higher than that of Health Functional Foods(42.5%).As for the reasons of additional consumption of the Health Functional Foods, subject answered as 'Easy to consume.'(41,6%), which was the most common among the subjects consuming Health Functional Foods. On other hand, the subjects of herbal medicine answered as that herbal medicine is 'more effective'(45.7%), and 'more trustworthy in preventing side-effects.'(40.3%). After consumption of the herbal medication, only 3.9% of the subjects consuming either Health Functional Foods or herbal medicine had side-effects. The most common side-effects were 'dermal reaction' which is normally caused by Health Functional Foods and 'indigestion' problems caused by herbal consumption. Conclusions According to the 421 subjects those involved in study, the percentages of consuming Health Functional Foods(67.7%) and herbal medicine(67.8%) were similar. The most commonly consumed products were a type of Health Functional Foods which were the nutrition-supplying products. Ginseng or red ginseng products were the next commonly used products. Health Functional Foods were commonly consumed for preventing illness and maintaining health rather than any other purpose. In contrary, herbal medicines were more commonly consumed for purposes such as for growth or treating certain type of disease. As a result of consumption, more than half of both subject replied as 'satisfied'. As for the side effects, dermal reaction was the most common problem for those with consuming Health Functional Foods, while indigestion was the most common side effect from the subjects with consuming herbal medicine.

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Neuroprotective Effects of Minocycline in Rat Brain Cortical Cell Culture Induced by Hypoxia (저산소 상태로 유도된 백서 뇌세포 배양에서 Minocycline의 뇌보호 효과)

  • Ha, Kyung A;Yang, Bum Seok;Kim, Jin Kyung;Kim, Hong Tae;Ha, Sung Jin;Lee, Jong Won;Chung, Hai Lee;Kim, Woo Taek
    • Clinical and Experimental Pediatrics
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    • v.46 no.11
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    • pp.1101-1106
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    • 2003
  • Purpose : In vivo, minocycline appears to be neuroprotective. Thus, the neuroprotective effects of minocycline were studied in a rat brain cortical cell culture induced by hypoxia. Methods : Cultured cells from the brains of Sprague-Dawley rats were divided into two sets of groups : normoxia groups treated with 5% $CO_2$ and hypoxia groups treated with 1% $CO_2$. After several days of incubation, the control groups were not treated with minocycline, while the sample groups were treated with either 1 or $10{\mu}g/mL$ of minocycline. The damaged cells were observed under a microscope, while apoptosis was detected using a TUNEL assay control-stained with DAPI. Results : Among the normoxia groups, the control and sample groups treated with 1 and $10{\mu}g/mL$ of minocycline were all statistically significantly different from each other. Meanwhile, among the hypoxia groups, although the control was significantly different from the sample groups, there was no statistically significant difference between the sample groups. When comparing the normoxia and hypoxia groups, there was a statistically significant difference between the control groups and sample groups treated with $1{\mu}g/mL$ of minocycline, yet no significant difference between the sample groups treated with $10{\mu}g/mL$ of minocycline. Conclusion : Minocycline was found to be neuroprotective in normoxia and hypoxia induced rat brain cortical cell cultures.

Growth Performance of Pangasiid Catfish, Silver carp and Catla in Polyculture (복합양식에서 pangasiid catfish, silver carp과 catla의 성장률)

  • Sarkar, Md. Reaz Uddin;Khan, Saleha;Haque, Md. Mahfuzul;Khan, Mohammed Nurul Absar;Luyen, Quoc-Hai;Choi, Jae-Suk
    • Journal of Life Science
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    • v.18 no.9
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    • pp.1186-1193
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    • 2008
  • The growth performance of pangasiid catfish (Pangasius hypophthalmus), silver carp (Hypophthalmichthys molitrix) and catla (Catla catla) was evaluated in polyculture at varied combinations and stocking ratios in earthen pond for a period of 16 weeks. Combinations of pangasiid catfish, silver carp and catla with the total density of 30,000 fish per hectare were evaluated. A commercial pelleted feed containing 28% crude protein and 6% lipid was fed only to pangasiid catfish at the rate of 8% of body weight during the first six weeks, 6% during the second six weeks and 4% thereafter twice daily. The growth and feed utilization efficiency of pangasiid catfish were found similar in all treatments. The growth of silver carp was markedly decreased upon the increase of its own stocking density. The highest growth of silver carp achieved in polyculture of pangasiid catfish, silver carp and catla at 2:1:1 ratio where density of silver carp was lowest. The growth of catla was found to be affected by the presence of silver carp. The growth rate of catla markedly declined with the increase of silver carp density. The highest growth of catla obtained in polyculture of pangasiid catfish only with catla stocked at 1:1 ratio. The highest yield and economic return achieved in polyculture of pangasiid catfish, silver carp and catla at the ratio of 2:1:1 due to higher growth of silver carp and higher market value of catla. The water quality conditions in all different treatment ponds were within optimum ranges throughout the culture period and the values did not show any significant variation (p>0.05) among the treatments.