• Clinical and Experimental Reproductive Medicine
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• v.25 no.2
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• pp.109-113
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• 1998

Objective: To ananlyze the direct effect of nitric oxide (NO), generated from sodium prusside (SNP) on the embryo developments in reproductive process. Design: Ova from mouse were treated to allow fertilization in in vitro culture. And the samples of fertilized ova were alloted into five alliqutos. Each alliquot was cultured in media treated with either concentration at 0 (n=92), $25{\mu}M$ (n=84), $50{\mu}M$ (n=80), $100{\mu}M$ (n=77), $500{\mu}M$ (n=54) of SNP. Main Outcome Measure: Rates of embryonal cell cleavages, viability and cell morphology were assessed during in vitro fertilization and culture. Results: As analyse the cell cleavage at 24 hours after in vitro culture of fertilised egg in variuos NO concentration, all of egg cells of each alliquot were developed to $2\sim4$ cell stage. But the alliquot of egg cells treated with $50{\mu}M$, which were totally degenerated. And also all embryonal cells of each alliquot were developed to 8 cell stage and morula stage on culture continuosly. And the embryonal cells of each alliquot were analysed at 24 and 48 hours following the in vitro culture. The rates of cell fragmentation and fusion were $4.2{\pm}3.4%$ in control group which is not treated with NO, while experimental groups was high, as rated $23.4{\pm}6.2%$ in $25{\mu}M$, $28.2{\pm}5.7%$ in $50{\mu}M$ and $32.1{\pm}6.4%$ in $100{\mu}M$ concentration of NO. Accordingly the rate of abnormal morphology of embryonal cell in control was lower significantly than that in each alliquot of experimental groups (p<0.05). And the degenerated rates of embryonal cells were 0% in control, $17.8{\pm}6.7%$ in $25{\mu}M$, $23.6{\pm}4.7%$ in $50{\mu}M$ and $26.8{\pm}11.2%$ in $100{\mu}M$ at 8 cells and morula on culture of 48 and 72 hours. On the examination of embryonal cells developed to blastocyst through in vitro culture, the rates of degenerated cells were $16.8{\pm}7.2%$ in control, $37.5{\pm}6.2%$ in $25{\mu}M$, $73.4{\pm}4.6%$ in $50{\mu}M$, 100% in $100{\mu}M$. Conclusion: This results suggeted that the NO in any concentrations is harmful on embryos in view of morphology as well as viability of cell, and the toxicity of NO on embryo is stronger at condition in higher concentration of NO.

• The Korea Journal of Herbology
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• v.29 no.6
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• pp.27-33
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• 2014

Objectives : The aims of this study were to exploring the therapeutic effect of Imperatae Rhizoma Extract(IRE) on Asthma. Methods : To investigate biological modulation activities of IRE, we conducted the cell-based assay whether IRE could regulate T helper 2 cells activity with EL4 T cells and mouse splenocytes, and followed animal study to conform the efficacy of their therapeutic potential on OVA-induced asthmatic mouse. Results : In cell study, IRE suppress the nuclear translocation of GATA binding protein-3 protein in phorbol 12-myristate 13-acetate/Ionomycin-stimulated EL4 T cells and Interleukine(IL)-4, IL-5 and IL-13 production in splenocytes at concentration dependent manner. In animal study, IRE-treated groups both 100mg/ml and 200mg/ml improve airway hypersensitibility reaction(AHR) response to methacholine about 30% and 40% with positive control group. Peritoneal blood analysis reveal that eosinophil number and ovalbumin-specific IgE is reduced by IRE treatment. Cell number of eosinophil is also reduced in bronchoalveolar lavage of IRE group like to peritoneal cell and real time-polymerase chain reaction data show that expression levels of IL-4, IL-5 and IL-13 were down regulated in lung tissue. Finally, histological analysis indicate that IRE protect the bronchial tissue damages through the accumulation of inflammatory cells and collagen, and these effect may be cause by interfering Th2 cells activity. Conclusions : Our data represent that IRE potentiates therapeutic activities to the allergic diseases such as asthma by regulating Th2 cells differentiation.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.62-62
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• 2001

Recently, production of transgenic animal by nuclear transfer has been known as a useful method. The production of cloned offspring derived from nuclear transfer depends upon a variety of factors such as species, donor cells type and cell cycle, and source of recipient ova. Therefore, we attempted a different transgenic methods using follicular granulosa cells (GCs). In general, ovulated GCs undergoes lutenization and transformation in vitro which might defective effects on developmental potential. In order to avoid the GCs transformation in vitro culture system, we introduced a direct injection of retrovirus into the follicles and then collected them mechanically from ovaries of 6-8 week-old ICR mice. Retrovirus vector constructed with pLN $\beta$ EGFP was injected into the follicles. The follicles are cultured in $\alpha$ -MEM supplemented with human FSH, LH and ITS in Costar Transwell dish for 4 days. Survival rate of virus injected follicles was 52.1% (12/23) and expression rate of EGPP gene was 33.3% (4/12). In this study, we found GCs performed transgenesis in our culture system. In addition, the GCs in follicle may be developed in vivo like environment rather than in vitro environment. Thus, the use of GCs as donor cells may be useful in the nuclear transfer for cloning of genetic modification. Therefore, these results suggest that follicular GCs can be transfected by viral vector during folliculogenesis in vitro.

• IMMUNE NETWORK
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• v.12 no.6
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• pp.253-260
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• 2012

${\alpha}$-Mangostin is a xanthon derivative contained in the fruit hull of mangosteen (Garcinia mangostana L.), and the administration of ${\alpha}$-Mangostin inhibited the growth of transplanted colon cancer, Her/CT26 cells which expressed Her-2/neu as tumor antigen. Although ${\alpha}$-Mangostin was reported to have inhibitory activity against sarco/endoplasmic reticulum $Ca^{2+}$ ATPase like thapsigargin, it showed different activity for autophagy regulation. In the current study, we found that ${\alpha}$-Mangostin induced autophagy activation in mouse intestinal epithelial cells, as GFP-LC3 transgenic mice were orally administered with 20 mg/kg of ${\alpha}$-Mangostin daily for three days. However, the activation of autophagy by ${\alpha}$-Mangostin did not significantly increase OVA-specific T cell proliferation. As we assessed ER stress by using XBP-1 reporter system and phosphorylation of $eIF2{\alpha}$, thapsigargin-induced ER stress was significantly reduced by ${\alpha}$-Mangostin. However, coadministration of thapsigargin with ${\alpha}$-Mangostin completely blocked the antitumor activity of ${\alpha}$-Mangostin, suggesting ER stress with autophagy blockade accelerated tumor growth in mouse colon cancer model. Thus the antitumor activity of ${\alpha}$-Mangostin can be ascribable to the autophagy activation rather than ER stress induction.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.1
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• pp.13-24
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• 2001

Objective: In muscle and neuronal cells, calcium channels have been classified by electrophysiological and pharmacological properties into (1) voltage-dependent $Ca^{2+}$-channel (1) P/Q-type $Ca^{2+}$-channel (2) N-type $Ca^{2+}$-channel (3) L-type $Ca^{2+}$-channel (4) T-type $Ca^{2+}$-channel (5) R-type $Ca^{2+}$-channel. The present study was done in order to investigate whether there is any difference in $Ca^{2+}$-channel distribution between activated and normally fertilized embryos. Methods: The immunocytochemical method was used to identify the existence of voltage-dependent $Ca^{2+}$-channels in parthenogenetically activated 2-cell embryos by ethanol and $SrCl_2$ treatment. These 2-cell embryos were obtained by exposure to 6% ethanol for 6 min and to 10 mM $SrCl_2$ for 2h. Results: P/Q-type $Ca^{2+}$-channels and L-type $Ca^{2+}$-channels have been identified. Whereas, three type of $Ca^{2+}$-channel P/Q-type, N-type, L-type have been identified in 2-cell embryos fertilized in vivo. Conclusion: Activation by ethanol was faster than those by $SrCl_2$. However, there was difference in DAB staining of the embryos between ethanol and $SrCl_2$ treatment (87.7% and 54.1 %). Intensity of staining was also different between ethanol- and $SrCl_2$-treated group. However, it has not been known why there was some difference in DAB staining and staining intensity in the present study.

• IMMUNE NETWORK
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• v.11 no.6
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• pp.383-389
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• 2011

Background: EY-6 is one of the newly synthesized indoledione derivatives to induce tumor cell-specific cell death. In this study, we investigated the mechanism of immunological death induced by EY-6 at mouse colon cancer cell as well as at the normal immune cell represented by dendritic cell. Methods: C57BL/6 mouse syngeneic colon cancer cell MC38 was treated with EY-6, and analyzed by MTT for viability test, flow cytometry for confirming surface expressing molecules and ELISA for detection of cytokine secretion. Normal myeloid-dendritic cell (DC) was ex vivo cultured from bone marrow hematopoietic stem cells of C57BL/6 mice with GM-CSF and IL-4 to analyze the DC uptake of dead tumor cells and to observe the effect of EY-6 on the normal DC. Results: EY-6 killed the MC38 tumor cells in a dose dependent manner (25, 50 and $100{\mu}M$) with carleticulin induction. And EY-6 induced the secretion of IFN-${\gamma}$ but not of TNF-${\alpha}$ from the MC38 tumor cells. EY-6 did not kill the ex-vivo cultured DCs at the dose killing tumor cells and did slightly but not significantly induced the DC maturation. The OVA-specific cross-presentation ability of DC was not induced by chemical treatment (both MHC II and MHC I-restricted antigen presentation). Conclusion: Data indicate that the EY-6 induced tumor cell specific and immunological cell death by modulation of tumor cell phenotype and cytokine secretion favoring induction of specific immunity eliminating tumor cells.

• Journal of Haehwa Medicine
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• v.12 no.2
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• pp.145-156
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• 2004

The purpose of this study was to investigate the inhibitory effect of the aroma therapy of three kinds of aroma oil mixtures on asthma. 1. The percentage of granulocytes/lymphocytes population in mouse OVA-induced asthma lung cells was decreased significantly compared with those of control group. 2. The number of CCR3+ cells, CD4+ cells, CD8+ cells, CD23 and CD3e+/CD69+ in lungs of the mice group treated with M1 were decreased significantly compared with those of control group. 3. The number of IgE+/B220+ cells in the lungs of the mice group treated with M1 decreased significantly compared with those of control group. But the number of B220+ cells in the lunes of the mice group treated with M1 didn't show significant difference compared with those of control group. 4. The number of Gr-1+/CD11b+ cells in lungs of the mice group treated with M1 didn't show significant difference compared with those of control group. But the number of CD11b+ cells in lungs of the mice group treated with M1 decreased significantly compared with those of control group.

• Biomedical Science Letters
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• v.16 no.2
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• pp.83-90
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• 2010

Hesperidin, a member of the flavanone group of flavonoids, can be isolated in large amounts from the rinds of some citrus species [e.g., Citrus aurantium L. (bitter orange), Citrus sinensis L. (sweet orange) and Citrus unshiu Marcov. (satsuma mandarin)], and has been reported to have anticarcinogenic, antihypotensive and antimicrobial properties. Despite the efficacy of these polyphenolic compounds as immune modulators, the effects of the flavonoids are poorly understood about allergic effect. In this study, we investigated whether hesperidin could influence on Th1 and Th2 balance. Allergic reactions included an increase in the number of eosinophils in bronchoalveolar lavage (BAL) fluid, an increase in inflammatory cell infiltration into the lung tissue around blood vessels and airways, airway luminal narrowing, the development of airway hyper-responsiveness (AHR). The administration of hesperidin before the last airway OVA challenge resulted in a significant inhibition of all asthmatic reactions. Accordingly, this study may provide evidence that hesperidin plays a critical role in the amelioration of the pathogenetic process of asthma in mice. These findings provide new insight into the immunopharmacological role of hesperidin in terms of its effects in a murine model of asthma, and also broaden current perspectives in our understanding of the immunopharmacological functions of hesperidin.

• Korean Journal of Animal Reproduction
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• v.26 no.2
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• pp.133-142
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• 2002

The objective of this study was to examine maturation, fertilization and developmental rate of the in vitro-grown mouse oocytes, and to compare these results with those of oocytes grown and matured in vivo. The preantral follicles isolated from 12-day-old mice were cultured on Transwell-COL membrane inserts. After in vitro growth and maturation, 72.5 % of oocytes grown in vitro produced polar body which can be comparable to in vivo growth (70.5 %). However, the mean oocyte diameter of the in vitro group (69.6$\pm$2.1$\mu$m) was smaller than that of the in vivo group (73.3$\pm$3.0$\mu$m). The fertilization rate was significantly lower (p<0.05) in the in vitro group (76.5%) than in the in vivo group (90.2%), however, there was no difference in the percentage of monospermic and polyspermic oocytes between two groups. The capacities of in vitro grown ova to cleave and develop to blastocyst were (57.8 and 14.4%, respectively) significantly lower (p<0.001) than those of the in vivo counterpart (84.4 and 56.6%, respectively). Moreover, the mean number of cells per blastocyst was significantly lower (p<0.05) in the in vitro group (39.0$\pm$10.8) than in the in vivo group (60.5$\pm$12.5). Live young were produced from transferred 2-cell embryos derived from in vitro-grown and matured oocytes. In conclusion, the results show that in vitro-grown oocytes did not achieve the developmental capacity of in vitro-grown oocytes.

• Korean Journal of Animal Reproduction
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• v.13 no.2
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• pp.113-120
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• 1989

In order to determine the sex of preimplantation embryos prior to transfer in cattle, a series of experiments were carried out using 45 Holstein donor cows to examine the ovarian response on the gonadotropin and PGF2${\alpha}$, and the morphology of fresh embryos or frozen/thawed embryos after deep freezing at -196$^{\circ}C$. The sexing of embryos treated with the medium containing H-Y antiserum(10%, v/v) and FITC anti-mouse IgG(10%, v/v) were analysed by chromosomal analysis, and the sex of the embryos which survived were ascertain after delivering the pups. The results obtained were summarized as follows ; 1. The average number of developed follicle and corpus luteum per cow were 13.5 and 8.1, and the ovalation rate was 60.1%. 2. Of 220-ova recovered, 75(34.1%) were morula and 91(41.4%) were blastocyst, and the morphological normal and abnormal rate of ova recovered were 75.5% and 24.5%, respectively. 3. Of 39 frozen/thawed embryos, the scores of normal morula and blastocyst, after thawing were 79.2%(19/24) and 73.3%(11/15). The average rate of frozen/thawed embryos which appeared morphologically normal post thawing was 76.9%(30/39). 4. The sex ratio was measured using the embryos treated with immunofluorescence assay to examine the relationship between embryo developmental stage, sex ratio of morula stage embryo was 42.2%(19/45) fluorescing and 57.8%(26/45) non-fluorescing, on the other hand, the ratio switched to 46.8%(29/62) fluorescing and 53.2%(33/62) non-fluorescing embryo in blastocyst stage. The sex ratio was also measured between fresh and frozen/thawed embryos, fresh and frozen/thawed treated embryos were indicated 45.8%(38/83) fluorescing, 54.2%(45/83) non-fluorescing and 41.7%(10/24) fluorescing, 58.3%(14/24) non-fluorescing. This trend indicated the approximal sex ratio was 1 : 1. 5. The result of karyotype test showed the successful rate of sexing embryo is fluorescing and non-fluorescing was 21.2%(7/33) and 29.6%(8/27). The female to male ratio within 33 fluorescing was 28.6 : 71.4, and the ratio of 27 non-fluorescing embryos was 87.7 : 12.5. 6. Of the embryo transferred after assignment of H-Y phenotype, five of the fluorescing embryos survived to term, all was males. Whereas six non-fluorescing embryos also survived to term and the sexes of the calves were 1 male 5 female.