• Journal of Life Science
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• v.33 no.10
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• pp.797-807
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• 2023

To improve the functionality of mulberry, samples were fermented with Lactobacillus plantarum JCM 1149 (LP) or Pichia kudriavzevii Atz-EN-01 (PK), and their antioxidant and anti-obesity activities were compared to those of unfermented mulberry. After fermenting for 60 hr, the total polyphenol and flavonoid content of the PK-fermented mulberry (PKFM) and LP-fermented mulberry (LPFM) was 1.5-fold and 2-fold higher, respectively, while the total anthocyanin content was 1.3-fold and 1.5-fold higher in the PKFM and LPFM, respectively. DPPH radical scavenging activity was found to be 16.3% higher (86% vs. 100%) after PK fermentation and 8.1% higher (86% vs. 93%) after LP fermentation. The lipase inhibitory activity of the LPFM and PKFM was 62.9% and 52.5%, respectively. 3T3-L1 preadipocytes were treated with unfermented mulberry, LPFM, or PKFM at 200, 400, or 800 ㎍/ml and stained with oil-red-O. A slight difference in the staining was observed in samples treated with 400 ㎍/ml. However, treatment with 800 ㎍/ml significantly reduced staining compared to the control, and the LPFM exhibited relatively higher adipogenesis inhibitory activity than the PKFM. Blood triglyceride content increased by 9.5% in the high-fat diet group, but decreased by 17.1% in the control group, 37.1% in the LPFM group, and 41.6% in the PKFM group. The blood triglyceride content of the LPFM group decreased by 43.1% and 21.4% compared to the high-fat diet group and the control group, respectively, and that of the PKFM group decreased by 48.6% and 28.9% compared to the same groups. In conclusion, the results indicate that fermented mulberry has increased antioxidant activity, lipase inhibitory activity, and adipogenesis inhibition activity, and decreased blood triglyceride content compared to unfermented mulberry.

• Journal of Life Science
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• v.33 no.11
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• pp.905-914
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• 2023

To evaluate the effectiveness of the skin barrier improvement of lactic acid (LA) and gluconolactone (GL), the expression of filaggrin, loricrin, hyaluronic acid (HA), hyaluronan syhthase-2 (HAS2), and aquaporine-3 (AQP3) in keratinocytes, and the moisture content and transepidermal water loss (TEWL) by clinical trials were evaluated. The expression levels of filaggrin and locricrin, which are the main factors affecting the proper functioning of skin barrier function, and HA, HAS2, and AQP3, which are skin moisturizing-related proteins measured by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting. The results showed that the expression levels of the factors that decreased by H2O2 treatment were significantly increased by LA, GL, and a mixture of LA and GL at the mRNA and protein levels (p<0.05). The nanoemulsion containing a mixture of LA and GL was prepared using the emulsion inversion method, and the average particle size was 299.9 ± 0.287 nm. After measuring the TEWL of nanoemulsion using Vapometer, it was found that TEWL significantly decreased by 15.53% and 26.73% after two weeks and four weeks of product use, respectively, compared to TEWL before product use (p<0.001). Similarly, the skin moisture content of the nanoemulsion significantly increased by 15.40% and 26.59% after two weeks and four weeks of product use, respectively, compared to skin moisture content before product use (p<0.001). Therefore, the skin barrier function and moisturizing effect of a mixture of LA and GL are shown by increasing the moisture content and decreasing the TEWL by increasing the expression of filaggrin, loricrin, HA, HAS2, and AQP3. This suggests the possibility for the development of functional cosmetic ingredients in the future.

• Membrane Journal
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• v.34 no.1
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• pp.50-57
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• 2024

In this experiment, Pd-Ag-Cu membrane was manufactured using electroless plating on an α-Al₂O₃ support. Pd, Ag and Cu were each coated on the surface of the support through electroless plating and heat treatment was performed for 18 h at 500℃ in H₂ in the middle of electroless plating to form Pd alloy. The surface of the Pd-Ag-Cu membrane was observed through Scanning Electron Microscopy (SEM), and the thickness of the Pd membrane was measured to be 7.82 ㎛ and the thickness of the Pd-Ag-Cu membrane was measured to be 3.54 ㎛. Energy dispersive X-ray spectroscopy and X-ray diffraction analysis confirmed the formation of a Pd-Ag-Cu alloy with a composition of Pd-78wt%, Ag-8.81wt% and Cu-13.19wt%. The gas permeation experiment was conducted under the conditions of 350~450℃ and 1~4 bar in H₂ single gas and H₂/N₂ mixed gas. The maximum H₂ flux of the hydrogen separation membrane measured in H₂ single gas is 74.16 ml/cm²·min at 450℃ and 4 bar for the Pd membrane and 113.64 ml/cm²·min at 450℃ and 4 bar for the Pd-Ag-Cu membrane. In the case of the separation factor measured in H₂/N₂ mixed gas, separation factors of 2437 and 11032 were measured at 450℃ and 4 bar.

• Journal of Marine Life Science
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• v.9 no.1
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• pp.9-21
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• 2024

Paralytic shellfish poisoning (PSP) including Saxitoxin (STX) is caused by harmful algae, and poisoning occurs when the contaminated seafood is consumed. The mouse bioassay (MBA), a standard test method for detecting PSP, is being sanctioned in many countries due to its low detection limit and the animal concerns. An alternative to the MBA is the Neuro-2a cell-based assay. This study aimed to establish various test conditions for Neuro-2a assay, including cell density, culture conditions, and STX treatment conditions, to suit the domestic laboratory environment. As a result, the initial cell density was set to 40,000 cells/well and the incubation time to 24 hours. Additionally, the concentration of Ouabain and Veratridine (O/V) was set to 500/50 μM, at which most cells died. In this study, we identified eight concentrations of STX, ranging from 368 to 47,056 fg/μl, which produced an S-shaped dose-response curve when treated with O/V. Through inter-laboratory variability comparison of the Neuro-2a assay, we established five Quality Control Criteria to verify the appropriateness of the experiments and six Data Criteria (Top and Bottom OD, EC₅₀, EC₂₀, Hill slop, and R² of graph) to determine the reliability of the experimental data. The Neuro-2a assay conducted under the established conditions showed an EC₅₀ value of approximately 1,800~3,500 fg/μl. The intra- & inter-lab variability comparison results showed that the coefficients of variation (CVs) for the Quality Control and Data values ranged from 1.98% to 29.15%, confirming the reproducibility of the experiments. This study presented Quality Control Criteria and Data Criteria to assess the appropriateness of the experiments and confirmed the excellent repeatability and reproducibility of the Neuro-2a assay. To apply the Neuro-2a assay as an alternative method for detecting PSP in domestic seafood, it is essential to establish a toxin extraction method from seafood and toxin quantification methods, and perform correlation analysis with MBA and instrumental analysis methods.

• Tuberculosis and Respiratory Diseases
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• v.43 no.3
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• pp.308-322
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• 1996

Background : Neutrophils are considered to play critical roles in the development of acute respiratory distress syndrome. Histamine, which is distributed abundantly in lung tissue, increases the rolling of neutrophills via increase of P-selectin expression on the surface of endothelial cells and is known to have some interrelationships with IL-1, IL-8 and TNF-$\alpha$. We studied to investigate the effect of the histamine on the acute lung injury of the rats induced by intratracheal insufflation of TNF-$\alpha$ which has less potency to cause lung injury compared to IL-1 in rats. Methods : We intratracheally instilled saline or TNF(R&D, 500ng), IL-1(R&D, 50ng)or histamine of varius dose(1.1, 11 and $55\;{\mu}g/kg$) with and without TNF separately in Sprague-Dawley rats weighing 270-370 grams. We also intratracheally treated IL-1(50ng) along with histamine($55\;{\mu}g/kg$). In cases, there were synergistic effects induced by histamine on the parameters of TNF-induced acute lung injury, antihistamines(Sigma, mepyramine as a $H_1$ receptor blockade and ranitidine as a $H_2$ receptor blockade, 10 mg/kg in each)were co-administered intravenously to the rats treated TNF along with histamine($1.1\;{\mu}g/kg$) intratraeheally. Then after 5 h we measured lung lavage neutrophil numbers, lavage cytokine-induced neutrophil chemoattractants(CINC), lung myeloperoxidase activity(MPO) and lung leak. We also intratracheally insufflated TNF with/without histamine($11\;{\mu}g/kg$), then after 24 h measured lung leak in rats. Statistical analyses were done by Kruskal-Wallis nonparametric ANOVA test with Dunn's multiple comparison test or by Mann-Whitney U test. Results : We found that rats given TNF, histamine alone(11 and $55\;{\mu}g/kg$), and TNF with histamine(l.1, 11, and $55\;{\mu}g/kg$) intratracheally had increased (p<0.05) lung MPO activity compared with saline-treated control rats. TNF with histamine $11\;{\mu}g/kg$ had increased MPO activity (P=0.0251) compared with TNF-treated rats. TNF and TNF with histamine(1.1, 11, and $55\;{\mu}g/kg$) intratracheally had all increased (p<0.05) lung leak, lavage neutophil numbers and lavage CINC activities compared with saline. TNF with histamine $1.1\;{\mu}g/kg$ had increased (p=0.0367) lavage neutrophil numbers compared with TNF treated rats. But there were no additive effect of histamine with TNF compared with TNF alone in acute lung leak on 5 h and 24 h in rats. Treatment of rats with the $H_1$ and $H_2$ antagonists resulted in inhibitions of lavage neutrophil accumulations and lavage CINC activity elevations elicited by co-treated histamine in TNF-induced acute lung injury intratracheally in rats. We also found that rats given IL-1 along with histamine intratracheally did not have increase in lung leak compared with IL-1 treated rats. Conclusion : Histamine administered intratracheally did not have synergistic effects on TNF-induced acute lung leak inspite of additive effects on increase in MPO activity and lavage neutrophil numbers in rats. These observations suggest that instilling histamine intratracheally would not play synergistic roles in neutrophil-mediated acute lung injury in rats.

• Journal of the Korean association of regional geographers
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• v.3 no.2
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• pp.163-193
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• 1997

Medical services are a fundamental and essential service in all urban areas. The location and accessibility of medical service facilities and institutions are critical to the diagnosis, control and prevention of illness and disease. The purpose of this paper is to present the results of a study on the location of medical facilities in Kwangju and the utilization of these facilities by the inhabitants. The following information is a summary of the findings: (1) Korea, like many countries, is now witnessing an increase in the age of its population as a result of higher living standards and better medical services. Korea is also experiencing a rapid increase in health care costs. To ensure easy access to medical consultation, diagnosis and treatment by individuals, the hierarchical efficient location of medical facilities, low medical costs, equalized medical services, preventive medical care is important. (2) In Korea, the quality of medical services has improved significantly as evident by the increased number of medical facilities and medical personnel. However, there is still a need for not only quantitative improvements but also for a more equitable distribution of and location of medical services. (3) There are 503 medical facilities in Kwangju each with a need to service 2,556 people. This is below the national average of 1,498 inhabitants per facility. The higher locational quotient and satisfactory population per medical facility showed at the civic center. On the other hand, problem regions such as the traditional residential area in Buk-Gu, Moo-deung mountain area and the outer areas of west Kwangju still maintain rural characteristics. (4) In the study area there are 86 general medicine clinics which provide basic medical services. i. e. one clinic per every 14,949 residents. As a basic service, its higher locational quotient showed in the residential area. The lower population concentration per clinic was found in the civic center and in the former town center, Songjeong-dong. In recently build residential areas and in the civic center, the lack of general medicine clinics is not a serious medical services issue because of the surplus of medical specialists in Korea. People are inclined to seek a consultation with a specialist in specific fields rather than consult a general practitioner. As a result of this phenomenon, there are 81 internal medicine facilities. Of these, 32.1% provide services to people who are not referred by a primary care physician but who self-diagnose then choose a medical facility specializing in what they believe to be their health problem. Areas in the city, called dongs, without any internal facilities make up 50% of the total 101 dongs. (5) There are 78 surgical facilities within the area, and there is little difference at the locational appearance from internal medicine facilities. There are also 71 pediatric health clinics for people under 15 years of age in this area, represents one clinic per 5,063 people. On the quantitative aspect, this is a positive situation. Accessibility is the most important facility choice factor, so it should be evenly located in proportion to demander distribution. However, 61% of 102 dongs have no pediatric clinics because of the uneven location. (6) There are 43 obstetrical and gynecological clinics in Kwangju, and the number of residents being served per clinic is 15,063. These services need to be given regularly so it should increase the numbers. There are 37 ENT clinics in the study area with the lower concentration in Dong-gu (32.4%) making no locational differences by dong. There are 23 dermatology clinics with the largest concentration in Dong-Gu. There are 17 ophthalmic clinics concentrated in the residential area because of the primary function of this type of specialization. (7) The use of general medicine clinics, internal medicine clinics, pediatric clinics, ENT clinics by the inhabitants indicate a trend toward primary or routine medical services. Obstetrics and gynecology clinics are used on a regular basis. In choosing a general medicine clinic, internal medicine clinic, pediatric clinic, and a ENT clinic, accessibility is the key factor while choice of a general hospital, surgery clinic, or an obstetrics and gynecology clinic, thes faith and trust in the medical practitioner is the priority consideration. (8) I considered the efficient use of medical facilities in the aspect of locational and management and suggest the following: First, primary care facilities should be evenly distributed in every area. In Kwangju, the number of medical facilities is the lowest among the six largest cities in Korea. Moreover, they are concentrated in Dong-gu and in newly developed areas. The desired number of medical facilities should be within 30 minutes of each person's home. For regional development there is a need to develop a plan to balance, for example, taxes and funds supporting personnel, equipment and facilities. Secondly, medical services should be co-ordinated to ensure consistent, appropriate, quality services. Primary medical facilities should take charge of out-patient activities, and every effort should be made to standardize and equalize equipment and facility resources and to ensure ongoing development and training in the primary services field. A few specialty medical facilities and general hospitals should establish a priority service for incurable and terminally ill patients. (9) The management scheme for the inhabitants' efficient use of medical service is as follows: The first task is to efficiently manage medical facilities and related services. Higher quality of medical services can be accomplished within the rapidly changing medical environment. A network of social, administrative and medical organizations within an area should be established to promote information gathering and sharing strategies to better assist the community. Statistics and trends on the rate or occurrence of diseases, births, deaths, medical and environment conditions of the poor or estranged people should be maintained and monitored. The second task is to increase resources in the area of disease prevention and health promotion. Currently the focus is on the treatment and care of individuals with illness or disease. A strong emphasis should also be placed on promoting prevention of illness and injury within the community through not only public health offices but also via medical service facilities. Home medical care should be established and medical testing centers should be located as an ordinary service level. Also, reduced medical costs for the physically handicapped, cardiac patients, and mentally ill or handicapped patients should be considered.

• Tuberculosis and Respiratory Diseases
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• v.45 no.6
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• pp.1236-1251
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• 1998

Background : TNF-$\alpha$ appears to be a central mediator of the host response to sepsis. While TNF-$\alpha$ is mainly considered a proinflammatory cytokine, it can also act as a direct cytotoxic cytokine. However, there are not so many studies about the relationship bet ween TNF-$\alpha$ level and lung injury severity in ALI, particularly regarding the case of ALI caused by direct lung injury such as diffuse pulmonary infection. Recently, a natural defense mechanism, known as the stress response or the heat shock response, has been reported in cellular or tissue injury reaction. There are a number of reports examining the protective role of pre-induced heat stress proteins on subsequent LPS-induced TNF-$\alpha$ release from monocyte or macrophage and also on subsequent LPS-induced ALI in animals. However it is not well established whether the stress protein synthesis such as HSP can be induced from rat alveolar macrophages by in vitro or in vivo LPS stimulation. Methods : We measured the level of TNF-$\alpha$, the percentage of inflammatory cells in bronchoalveolar lavage fluid, protein synthesis in alveolar macrophages isolated from rats at 1, 2, 3, 4, 6, 12, and 24 hours after intratracheal LPS instillation. We performed histologic examination and also obtained histologic lung injury index score in lungs from other rats at 1, 2, 3, 4, 6, 12, 24 h after intratracheal LPS instillation. Isolated non-stimulated macrophages were incubated for 2 h with different concentration of LPS (0, 1, 10, 100 ng/ml, 1, or 10 ${\mu}g/ml$). Other non-stimulated macrophages were exposed at $43^{\circ}C$ for 15 min, then returned to at $37^{\circ}C$ in 5% CO2-95% for 1 hour, and then incubated for 2 h with LPS (0, 1, 10, 100ng/ml, 1, or 10 ${\mu}g/ml$). Results : TNF-$\alpha$ levels began to increase significantly at 1 h, reached a peak at 3 h (P<0.0001), began to decrease at 6 h, and returned to control level at 12 h after LPS instillation. The percentage of inflammatory cells (neutrophils and alveolar macrophages) began to change significantly at 2 h, reached a peak at 6 h, began to recover but still showed significant change at 12 h, and showed insignificant change at 24 h after LPS instillation compared with the normal control. After LPS instillation, the score of histologic lung injury index reached a maximum value at 6 h and remained steady for 24 hours. 35 kDa protein band was newly synthesized in alveolar macrophage from 1 hour on for 24 hours after LPS instillation. Inducible heat stress protein 72 was not found in any alveolar macrophages obtained from rats after LPS instillation. TNF-$\alpha$ levels in supernatants of LPS-stimulated macro phages were significantly higher than those of non-stimulated macrophages(p<0.05). Following LPS stimulation, TNF-$\alpha$ levels in supernatants were significantly lower after heat treatment than in those without heat treatment (p<0.05). The inducible heat stress protein 72 was not found at any concentrations of LPS stimulation. Whereas the 35 kDa protein band was exclusively found at dose of LPS of 10 ${\mu}g/ml$. Conclusion : TNF-$\alpha$ has a direct or indirect close relationship with lung injury severity in acute lung injury or acute respiratory distress syndrome. In vivo and in vitro LPS stimulation dose not induce heat stress protein 72 in alveolar macrophages. It is likely that 35 kDa protein, synthesized by alveolar macrophage after LPS instillation, does not have a defense role in acute lung injury.

• Tuberculosis and Respiratory Diseases
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• v.49 no.6
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• pp.691-702
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• 2000

Background : Acute lung injury (ALI) is a commonly encountered respiratory disease and its prognosis is poor when the treatment is not provided promptly and properly. However no specific pharmacologic treatment is currently available for ALI, although recently several supportive drugs have been under scrutiny. We studied anti-inflammatory effects of pentoxifylline (PF), a methylated xanthine, and ONO-5046, a synthetic neutrophil elastase inhibitor on lipopolysaccharide (LPS)-induced ALI in vitro. Methods : To establish an in vitro model of LPS-induced ALI, primary rat alveolar macrophages and peripheral neutrophils in various ratios (1:0, 5:1, 1:1, 1:5, 0:1) were co-cultured with transformed rat alveolar epithelial cells (L2 cell line) or vascular endothelial cells (IP2-E4 cell line) under LPS stimulation. Each experiment was divided into five groups-control, LPS, LPS+PF, LPS+ONO, and LPS+PF+ONO. We compared LPS-induced superoxide anion productions from primary rat alveolar macrophages and peripheral neutrophils in various ratios, and the resultant cytotoxicity on L2 cells or IP2-E4 cells between groups. In addition we also compared the productions of tumor necrosis factor (TNF)-$\alpha$ interleukin (IL)-$1{\beta}$, monocyte chemotactic protein(MCP)-1, IL-6, and IL-10 as well as mRNA expressions of TNF-$\alpha$ inducible nitric oxide synthetase(iNOS), and MCP-1 from LPS-stimulated primary rat alveolar macrophages between groups. Results : (1) PF and ONO-5046 in each or both showed a trend to suppress LPS-induced superoxide anion productions from primary rat alveolar macrophages and peripheral neutrophils regardless of their ratio, except for the LPS+PF+ONO group with the 1:5 ratio, although statistical significance was limited to a few selected experimental conditions. (2) PF and ONO-5046 in each or both showed a trend to prevent IP2-E4 cells from LPS-induced cytotoxicity by primary rat alveolar macrophages and peripheral neutrophils regardless their ratio, although statistical significance was limited to a few selected experimental conditions. the effects of PF and/or ONO-5046 on LPS-induced L2 cell cytotoxicity varied according to experimental conditions. (3) PF showed a trend to inhibit LPS-induced productions of INF-$\alpha$ MCP-1, and IL-10 from primary rat alveolar macrophages. ONO-5046 alone didnot affect the LPS-induced productions of proinflammatory cytokines from primary rat alveolar macrophages but the combination of PF and ONO-5046 showed a trend to suppress LPS-induced productions of INF-$\alpha$ and IL-10 PF and ONO-5046 in each or both showed a trend to increase LPS-induced IL-$\beta$ and IL-6 productions from primary rat alveolar macrophages. (4) PF and ONO-5046 in each or both showed a trend to attenuate LPS-induced mRNA expressions of TNF-$\alpha$ and MCP-1 from primary rat alveolar macrophages but at the same time showed a trend increase iNOS mRNA expression. Conclusion : These results suggest that PF and ONO-5046 may play a role in attenuating inflammation in LPS-induced ALI and that further study is needed to use these drugs as a new supportive therapeutic strategy for ALI.

• The Korean Journal of Nuclear Medicine Technology
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• v.15 no.2
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• pp.76-82
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• 2011

Purpose: Lymphoscintigraphy is absolutely being used standard examination in lymphatic diagnosis, evaluation after treatment, and it is useful for lymphedema to plan therapy. In case of lymphoscintigraphy of lower-extremity lymphedema, it had an effect on results if patients had not pose same position on the examination of 1 min, 1 hour and 2 hours after injection. So we'll study the methods to improve confidence with minimized quantitative analysis errors by influence factors. Materials and Methods: Being used the Infinia of GE Co. we injected $^{99m}Tc$-phytate 37 MBq (1.0 mCi) 4 sylinges into 40 people's feet hypodermically from June to August 2010 in Samsung Medical Center. After we acquired images of fixed and unfixed condition, we confirmed the count values change by attenuation of soft tissue and bone according to different feet position. And we estimated 5 times increasing 2 cm of distance between $^{99m}Tc$ point source and detector each time to check counts difference according to distance change by different feet position. Finally, we compared 1 and 6 min lymphoscintigraphy images with same position to check the effect of quantitative analysis results owing to difference of amounts of movement of the $^{99m}Tc$-phytate in the lymphatic duct. Results: Percentage difference regarding error values showed minimum 2.7% and maximum 25.8% when comparing fixed and unfixed feet position of lymphoscintigraphy examination at 1 min after injection. And count values according to distance were 173,661 (2 cm), 172,095 (4 cm), 170,996 (6 cm), 167,677 (8 cm), 169,208 counts (10 cm) which distance was increased interval of 2 cm and basal value was mean 176,587 counts, and percentage difference values were not over 2.5% such as 1.27, 1.79, 2.04, 2.42, 2.35%. Also, Assessment results about amounts of movement in lymphatic duct within 6 min until scanning after injection showed minimum 0.15%, and maximum 2.3% which were amounts of movement. We can recognize that error values represent over 20% due to only attenuation of soft tissue and bone except for distance difference (2.42%) and amounts of movement in lymphatic duct (2.3%). Conclusion: It was show that if same patients posed different feet position on the examination of 1 min, 1 hour and 2 hours after injection in the lymphoscintigraphy which is evaluating lymphatic flow of patients with lymphedema and analyzing amount of intake by lymphatic system, maximum error value represented 25.8% due to attenuation of soft tissue and bone, and PASW (Predictive Analytics Software) showed that fixed and unfixed feet position was different each other. And difference of distance between detector and feet and change of count values by difference of examination beginning time after injection influence on quantitative analysis results partially. Therefore, we'll make an effort to fix feet position and make the most of fixing board in lymphoscintigraphy with quantitative analysis.

• Korean Journal of Animal Reproduction
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• v.22 no.4
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• pp.425-435
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• 1998

To improve the efficiency of nuclear transplantation in bovine, in this study the development in vitro of nuclear transferred (NT) embryos was compared by different activation regimens of the enucleated oocytes. The effect of developmental stage and culture system of donor nuclei on fusion and development in vitro of NT embryos were also evaluated. Oocytes were collected from Hanwoo ovaries obtained from slaughterhouse and matured in Ham's F-10 supplemented with hormones. After 20~22 h maturation, the oocytes were vortexed to be free from cumulus cells and subsequently their nucleus and the first polar body were removed. Enucleated oocytes were divided into 3 groups for activation; the oocytes of group I were activated with ionomycin for 5 min and subsequently incubated in 6-dimetylarninopurine (DMAP) for 4 h, Those of group II were treated with DMAP for 4 h at 39 h after onset of in vitro maturation (IVM) and those of group III were kept in room temperature ($25^{\circ}C$) for 3 h at 39 h after onset of IVM. After in vitro fertilization (IVF) the embryos for muclear donor were cultured either by group culture (20 embryos /50 ${mu}ell$ drop) or individually (1 embryo /50 ${mu}ell$ drop) for 4 day and 5 day. At day 4 and 5 after IVF, blastomeres were separated in calcium-magnesium free medium, and then classified into small (day 5: $\leq$ 38 ${\mu}{\textrm}{m}$, day 4: $\leq$ 46 ${\mu}{\textrm}{m}$) and large (day 5 : $\geq$ 38 ${\mu}{\textrm}{m}$, day 4 ; $\geq$ 46 ${\mu}{\textrm}{m}$). The separated blastomeres were replaced into enucleated and activated recipient cytoplasm. The blastomere-oocyte complexes were fused by electrically. The NT embryos were cultured in TCM-199 containing 10% FCS in 39$^{\circ}C$, 5% $CO_2$ incubator for 7 day. The results obtained were summarized as follows; There were no differences in fusion and development to blastocyst between groups as group I (68%, 10%), group II (75%, 14%) and group III (73%, 9%), respectively. However, the cell number in blastocyst of NT embryos in group III were significantly fewer than in the other groups (P＜0.05). No differences in fusion and development to blastocyst were found between individual or group cultured and between small or large blastomeres of day 4 and day 5 donor embryos. From these results, it was concluded that the combination of ionomycin and DMAP, or treatment of DMAP at 39 h after onset of IVM were useful for the efficient of production of NT bovine embryos, and the individual cultured embryos could be simply used as donor nuclei for NT bovine embryo.