• Journal of Embryo Transfer
• /
• v.10 no.3
• /
• pp.203-208
• /
• 1995

This study was carried out to efficiently use the ultrarapid freezing method in the cryopreservation of mouse ova. For this, the effects of dehydration method, oval vigour and $0^{\circ}C$ controlling method on post-thawing viability were investigated. Fresh mouse ova were dehydrated in mPBS with 3.5M DMSO and /or 0.25M sucrose, and directly immersed in L$N_2$ for ultrarapidly freezing. The frozen ova were thawed at 37$^{\circ}C$, rehydrated in mPBS with 0.25M sucrose, and then repeatedly washed in HAM's Fl0 before evaluating the morphological normality of frozen-thawed ova. The results obtained showed that there was difference between treatments in a experiment. 1) The post-thawing viability of ova dehydrated in multi-step (48.4$\pm$13.8%) was higher than that of ova in two-step (40.9$\pm$14.0%). 2) The post-thawing viability of fertilized ova (87$\pm$14.0%) was significantly(p<0.0l) higher than that of unfertilized ova (5.4$\pm$5.4%). 3) The post-thawing viability of ova dehydrated and rehydrated using a cooling machine (95.8$\pm$4.2%) was significantly(p<0.05) higher than that on ice(84.1$\pm$9.9). In conclusion, in order to efficiently cryopreserve ova in vitro with ultrarapidly freezing method, highly viable embryos should be selected, heavy osmotic shock to the dehydrating ova should be avoided, and embryos in high osmotic condition were dehydrated and rehydrated in a constantly low temperature.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.20 no.3
• /
• pp.581-589
• /
• 2006

These studies were investigated the effects of Gamicheungpyehwadam-tang (CPHDT) on immune-cell regulation in association with bronchial asthma in OVA-induced mouse model. The administration of 400 mg/kg CPHDT significantly reduced the number of total cells in lung, peripheral lymph node and spleen in OVA-induced bronchial asthma mouse model. The administration of 400 mg/kg CPHDT significantly reduced $CD3^+,{\;}CD19^+$and $CD3^+,{\;}CD69^+$ cell numbers separated from lung, peripheral lymph node and spleen in OVA-induced bronchial asthma mouse model. CPHDT significantly reduced $CD3^+/CCR3^+,{\;}CD4^+,{\;}B220^+/IgE^+$, and $CD3^+/DX5^+$ cell numbers separated from lung, peripheral lymph node and spleen in OVA-induced bronchial asthma mouse model in a dose dependent manner, However, CPHDT significantly reduced $CD8^+$ cell numbers from only lung and spleen. The administration of CPHDT significantly reduced $NK^+$ cell numbers separated from lung of OVA-induced bronchial asthma mouse model in all concentrations, but 200 mg/kg CPHDT reduced $NK^+$ cell numbers separated from peripheral lymph node. These results suggest that CPHDT has anti-asthma and anti-allergy effects. In addition to, CPHDT may be useful treatment of asthma based on the further studies about the individual efficacy search of the components of CPHDT and the adding of variety drugs to CPHDT.

• Korean Journal of Animal Reproduction
• /
• v.16 no.3
• /
• pp.199-208
• /
• 1992

These studies were carried out to investigate optimal physological conditions for in vitro fertilization (IVF) of mouse ova. The unfertilized ova were obtained by superovulation from ICR mice of 4 to 6 weeks old. Tyrode's 280 solution was used as basal media, and pH and osmolality of basal media were adjusted with the supplementation of sodium bicarbonate and sodium chloride, respectively. The optimal pH, and osmolality of culture media and the optimum period of sperm preincubation were examined in fertilization in vitro of mouse ova and the subsequent culture in vitro of embryos. The pH range of media examined was designed from 6.5 to 7.5 with 0.2 interval and the range of osmolality from 250 to 370 mOsm with 20 interval, and the period of sperm preincubation examined was 30, 60, 120, and 180 minutes. The ova developed to 2-cell embryosafter 26hrs. of incubation with preincubated sperm were evaluated as in vitro fertilized ones. The results obtained were summarized as follows: 1. The percentage of in vitro fertilized ova was highest (64.7%) in media of pH 7.1 and lowest (38.0%) in pH 6.7. No significant difference in % fertilized ova was found from the media of pH 7.1 to 7.5. Compared with the result from pH 7.1 medium, the pollyspermy was increased signifciantly (p<0.05) in the media of pH over 7.5 and below 6.9;, and the % degenerated ova was significantly (p<0.05) increased in the media of pH below 6.9. 2. The percentage of in vitro fertilized ova was highest (69.4%) in media of osmolality 330 mOsm and lowest (47.9%) in osmolality 250 mOsm. No significant difference in % fertilized ova was found from the media of osmolality 310 to 350 mOsm. Compared with the result from osmolality 330 mOsm in medium, the polyspermy aws increased significantly(p<0.05) in the media of osmolality over 350 mosmol and blow 290 mOsm, and the % degenerated ova was significantly (P<0.05) increased in the media of osmolality below 290 mOsm. 3. The percentate of in vitro fertlilized ova was highest (62.7%) in media of period sperm preincubation 180 min. and lowest (40.4%) in sperm preincubation 30 minutes. No significant difference in % fertilized ova was found from the media of sperm preincubation 120 to 180 minutes. Compared with the result from sperm preincubation 180 minutes in medium, the polyspermy was low differ no significantly(P<0.05) in the media of period sperm preincubation, and the % degenerated ova was signifciantly(P<0.05) increased in the media of sperm presincubation below 60 minutes.

• Journal of Acupuncture Research
• /
• v.22 no.3
• /
• pp.185-200
• /
• 2005

Objective : The aim of this study was to investigate the asthma-suppressive and immune-regulatory effect of AHCR-HA(ASARI HERBA CUM RADICE Herbal-acupuncture) at Pyesu(BLl3) on OVA(ovalbumin)-induced asthma in mice. Methods : C57BL/6 mice out of all the experimental groups, except the Normal group and the AHCR-HA group, were sensitized and challenged with OVA The mice in the AHCR-HA group and the OVA-AHCR-HA group were treated with AHCR-HA(1%) at Pyesu(BL13). The mice in the OVA-Saline group were injected with saline at Pyesu(BL13). The mice in the OVA-Needle-Prick group were treated with a single prick with an injection needle at Pyesu(BL13). AHCR-HA saline injection and needle prick were administered for 8 weeks, three times a week. Result : 1. The populations of granulocytes, CD3e-/CCR3+ cells, CD69+/CD3e+ cells, CD4+ cells and CD23+/B220+ cells in the OVA-induced asthmatic mouse lungs decreased significantly by AHCR-HA. 2. The lung weight, total cells in lung, total leukocytes in BALF, eosinophils in BALF, collagen accumulation in the lung sections of the OVA-AHCR-HA group decreased significantly. 3.The concentrations of IL-4, IL-5, IL-13, IgE in BALF and serum of the OVA-AHCR-HA group decreased significantly. 4. The numbers of Gr-1+/CD11b+, CCR3+, CD3e+, CD19+, CD3e+/CD69+cells in the OVA-AHCR-HA group decreased significantly. 5. The mRAN expressions of $TNF-{\alpha}$, IL-5, IL-4 and IL-13 in lung of the OVA-AHCR-HA group decreased significantly. 6. The AHCR-HA group didn't show any considerable difference from the Normal group. The OVA-saline group and the OVA-Needle prick group showed suppressive effects on OVA-induced asthma however they were not statistically significant. Conclusion : These results suggest that AHCR-HA at Pyesu(BL13) is considered to be effective in treating asthma and to be put to practical use in the future asthma clinic.

• Asian-Australasian Journal of Animal Sciences
• /
• v.26 no.4
• /
• pp.545-551
• /
• 2013

This study aimed to present a mouse model of ovalbumin (OVA) induced allergic diarrhea under a sub-barrier system and investigate the development of gut microbiota in this model. Male BALB/c mice were systemically sensitized with OVA or sham-sensitized with saline, and followed by oral OVA intubation, leading to OVA-specific acute diarrhea. Compared with sham-sensitized mice, sera OVA-specific IgG1 and total IgE in OVA-sensitized mice were dramatically elevated, and the number of mast cells was greatly increased in the jejunum of the OVA-sensitized mice. Principle component analysis of the DGGE profile showed that samples from group of OVA-sensitized mice and group of sham-sensitized mice were scattered into two different regions. Real-time PCR analysis showed that the number of 16S rRNA gene copies of Lactobacillus in the colon of OVA-sensitized mice decreased significantly, while there was no significant difference in the number of Bifidobacterium and total bacteria. In conclusion, OVA-specific allergic diarrhea was successfully induced under a sub-barrier system, and changes of allergic reactions during induction was coupled with changes in gut microbiota, especially the number of colonic Lactobacillus, but the role of gut microbiota in the development of food allergic reactions needs to be further evaluated.

• The Journal of Pediatrics of Korean Medicine
• /
• v.20 no.1
• /
• pp.181-194
• /
• 2006

Objective : Author evaluate the effects of Citri Reticulatae Viride Pericarpium for the bronchial asthma using assesment of the vascular endothelial growth factor after Citri Reticulatae Viride Peicarpium was intravenously administered Ovalbumine(OVA)-sensitized and -challenged mice. Material and Methods : Twenty-four female mice, 8-10 weeks old and free of murine specific pathogens, were sensitized by intraperitoneal injection of OVA. Of the sensitized mice, ten mice didn't administrate Citri Reticulatae Viride Pericarpium and the other ten mice administrated Citri Reticulatae Viride Pericarpium. Mice were sensitized on the first and fourteen days introspectional injection of 20 ${\mu}g$ OVA. After$21^{st}\;22^{th}\;and\;23^{rd}$, the initial sensitization, the mice were challenged for 30 minutes with an aerosol of 1% OVA in saline. Citri Reticulatae Viride Pericarpium administered 200mg/kg in the tail of the mouse, one a day, for 7 days and beginning 14 days after first sensitization. Bronchoalveolar lavage was performed 72 hours after the last challenge, and total cell numbers in the BAL fluid were count. Also, lever of VEGF in the BAL fluid was measured by Enzyme immunoassays and Western blot analysis. Results: Total cell numbers in BAL fluid were significantly greater than from 72 hrs after OVA inhalation compared with cell numbers in the control group. However, there was no difference of the total cell numbers between OVA-challenge groups without Citri Reticulatae Viride Pericarpium, and OVA-challenge with Citri Reticulatae Viride Pericarpium. Enzyme immunoassay revealed that VEGF levels in the BAL fluids were significantly increased 72 hrs after OVA inhalation compared with levels in the control group. After administration of the Citri Reticulatae Viride Pericarpiu, the levels of the VEGF in BAL fluids 72 hrs after OVA inhalation reduced dramatically. Western blot analysis revealed that VEGF protein levels were increased in the all mice which were challenge with OVA, without administered Citri Reticulatae Viride Pericarpium, compared the normal mouse. However, in the groups of the administered Citri Reticulatae Viride Pericarpium, the VEGF protein level markedly decreased. Conclusion: Citri Reticulatae Viride Pericarpium might affect the treatment of the bronchial ashma as a inhibition of the VEGF.

• Clinical and Experimental Pediatrics
• /
• v.53 no.4
• /
• pp.481-487
• /
• 2010

Purpose: Recently many studies show early exposure during childhood growth to endotoxin (lipopolysaccharides, LPS) and/or early exposure to allergens exhibit important role in development of allergy including bronchial asthma. The aim of this study was to evaluate the role of endotoxin and allergen exposure in early life via the airways in the pathogenesis of allergic airways inflammation and airway hyperresposiveness (AHR) in mouse model of asthma. Methods: Less than one week-old Balb/c mice was used. Groups of mice were received either a single intranasal instillation of sterile physiologic saline, 1% ovalbumin (OVA), LPS or $1.0{\mu}g$ LPS in 1% OVA. On 35th day, these animals were sensitized with 1% OVA for 10 consecutive days via the airways. Animals were challenged with ovalbumin for 3 days on 55th days, and airway inflammation, hyperresponsiveness, and cytokine expression were assessed. Measurements of airway function were obtained in unrestrained animals, using whole-body plethysmography. Airway responsiveness was expressed in terms of % enhanced pause (Penh) increase from baseline to aerosolized methacholine. Lung eosinophilia, serum OVA-IgE and bronchoalveolar lavage (BAL) fluid cytokine levels were also assessed. ANOVA was used to determine the levels of difference between all groups. Comparisons for all pairs were performed by Tukey-Kramer honest significant difference test; $P$ values for significance were set to 0.05. Results: Sensitized and challenged mice with OVA showed significant airway eosinophilia and heightened responsiveness to methacholine. Early life exposure of OVA and/or LPS via the airway prevented both development of AHR as well as bronchoalveolar lavage fluid eosinophilia. Exposure with OVA or LPS also resulted in suppression of interleukin (IL)-4, 5 production in BAL fluid and OVA specific IgE in blood. Conclusion: These results indicate that antigen and/or LPS exposure in the early life results in inhibition of allergic responses to OVA in this mouse model of astham. Our data show that early life exposure with OVA and/or LPS may have a protective role in the development of allergic airway inflammation and development of allergen-induced airway responses in mouse model of asthma.

• Journal of the Korea Academia-Industrial cooperation Society
• /
• v.13 no.12
• /
• pp.6208-6214
• /
• 2012

The most essential but missing components to understand and use toxic substances from marine microalgae are developing the fast, easy and economical determining technology for detecting it. In this paper we produced the antibodies against saxitoxin (STX). Mariculture keyhole limpet hemocyanin (mcKLH) and ovalbumin (OVA) were used as carrier proteins. mcKLH-STX conjugates were injected into the peritonial cavity of BALB/c mouse for immunization. After bleeding from mouse, anti-STX antiserum was isolated. Indirect enzyme-linked immunosorbent assays (ELISA) was performed to determine antiserum titer using the microtiter plate coated with free STX and OVA-STX. A goat anti-mouse IgG-phosphatase conjugate was used as secondary antibody to enable chromogenic reaction. Reactions of anti-STX antiserum were very specific on the OVA-STX and free STX. Sensitivity of anti-STX antiserum on STX was very high and STX detection limit was to be 64.9 ng/kg for indirect ELISA.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.18 no.6
• /
• pp.1694-1698
• /
• 2004

We hope to evaluate the effects of Sabaek-san for the bronchial asthma using assesment on the metrix metalloproteinase-9(MMP-9) after Sabaek-san was intravenously administered OVA-sensitized and -challenged mice. Seventy-two female mice, 8-10 weeks of age and free of murine specific pathogens, were used. Of the seventy-two mice, twenty-four mice were not sensitized and forty-eight mice were sensitized by intraperitoneal injection of OVA. Of the sensitized mice, twenty-four mice didn't administrate Sabaek-san and twenty-four administrated Sabaek-san. Mice were sensitized on days 1 and 14 by intraperitoneal injection of 20 fig OVA. On days 21, 22 and 23 after the initial sensitization, the mice were challenged for 30 minutes with an aerosol of 1% OVA in saline. Sabaek-san administered 200㎎/㎏ in the tail of the mouse, one time per day, for 7 days, beginning 14 days after first sensitization. Bronchoalveolar lavage was performed 72 hours after the last challenge, and total cell numbers in the BAL fluid were count. Also, level of MMP-9 in the BAL fluid were measured by Enzyme immunoassays and Western blot analysis. Enzyme immunoassay revealed that MMP-9 levels in the BAL fluids significantly increased 72 h after OVA inhalation compared with levels in the control group. After administration of the Sabaek-san, the levels of the MMP-9 in BAL fluids 72 h after OVA inhalation reduced dramatically. Western blot analysis revealed that MMP-9 levels increased in the all mice which were challenge with OVA without administered Sabaek-san compared the normal mouse. However, in the groups of the administered Sabaek-san, the MMP-9 level markedly decreased. Sabaek-san might be effect the treatment of the bronchial asthma as a inhibition of the MMP-9.

• Clinical and Experimental Reproductive Medicine
• /
• v.20 no.1
• /
• pp.31-36
• /
• 1993

The effects of freezing and 1,2-propanediol on early and late pronucleate stage mouse ova were investigated in terms of survival after thawing and development in vitro. The samples were divided into two groups according to different age in pronucleate ova: ova in(1) early pronuclear stage with two distant pronuclei at 18h after hCG injection, and (2) late pronuclear stage with adjacent pronuclei at 30h. Zygotes in the late pronuclear stage have been proven to be more resistant to 1,2-propanediol, showing a significantly higher developmental rate than zygotes in early stage (80.3 versus 66.3%, <0.05), but survival rate was similar in the two groups (91.0 versus 93.5%). After freezing and thawing, survival and developmental rates were decreased in both groups when compared to the control group (54.3 versus 92.3%, 47.7 versus 73.3%. respectively). And developmental rate in the late pronuclear stage zygotes showed significantly higher than in early (55.4 versus 40.0%) after thawing. In conclusion, early pronucleate mouse ova have a lower developmental capacity in vitro and a lower survival rate after freezing and thawing than late ova. These findings suggest that the timing of freezing could be important for survival and further development in vitro in cryopreservation of human pronucleate ova.