• Journal of Biomedical Engineering Research
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• v.13 no.2
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• pp.133-140
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• 1992

Bioelectrical Impedance method for measuring human body composition is based upon the principle that the electrical conductivity of the fat-free mass (FFM) is far greater than that of the (at. Deter- mination o( impedance was nlade in 30 healthy and obese men aged $40.9{\pm}14.7$yr (mean${\pm}$SD) and 25 healthy and obese women aged $44.0{\pm}10.2$ yr. A commercial impedance body fat analyzer was used with a four electrode arrangement that introduced a painless constant current(ImA at 50 kHz) into the body. Linear relationships were found between impudence values and FFM(r=-0.786) and between weight and FFM(r=0.837). Signigicant increases in the correlation coefficients were ob served when the predicator Hta/z was regressed against FFM(r=0.912) where Ht Is height and Z Is impedance. A linear regression equation, FFM=0.586(Htf/z) +0.317(Weight) -1.674(r=0.977), was found. The correlation coefficient of % body fat between the impedance and infrared methods was 0.898. These data Indicate that the bioelectrical impedance technique is a reliable and valld approach for the estimation of human body composition. This method Is safe, noninvasivr, prouides rapld measure menu, requires little operator skill and subject cooperation, and Os portable. Finally measurement of percent body fat was made possible using both the regression equation and a developed impedance measuring device which measures impedance between wrist and ankle.

• Polymer(Korea)
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• v.36 no.2
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• pp.196-201
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• 2012

Electrospinning is a versatile process used to prepare micro or nano sized fibers from various materials dissolved in volatile solvents. This study reports electrospun pullulan fibrous webs fabricated through electrospinning using water as a solvent. The electrospinning conditions such as pullulan (PUL) concentration and applied voltage were optimized in order to obtain smooth electrospun fibers. The concentration of PUL greatly influenced the viscosity and surface tension of PUL solution. PUL beaded electrospun fibers were obtained from PUL solutions with concentrations lower than 5 wt%, while homogenous electrospun fibers were prepared from solutions with high concentration and high viscosity. The average diameters of PUL fibers were decreased to 200 nm when the polymer concentration was kept at 10 wt% and the applied voltage was fixed at 15 kV during electrospinning. PUL electrospun fiber exhibited higher solubility, flexibility, softness and adhesive strength.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.2
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• pp.177-185
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• 2005

Objective: This study was conducted to find an optimal condition for the vitrification of mouse morulae and expanded blastocysts. Materials and Methods: Mouse embryos were obtained at 2-cell stage and cultured to morula and expanded blastocyst stage in Human Tubal Fluid (HTF) medium supplemented with 10% Serum Substitute Supplement (SSS). The vitrification solutions used were EFS30, EFS35 and EFS40 that contains 30%, 35% and 40% ethylene glycol, respectively, with 18% ficoll and 0.5 M sucrose diluted in Dulbecco's phosphate-buffered saline (DPBS) medium supplemented with 10% SSS. The vitrification procedure was performed in EFS solution with three steps, followed by thawing in 6 steps with 0.5 M sucrose, and then survival and hatching-hatched rate per embryos recovered were compared among six groups. Results: After 24 h culture in different vitrification and thawing solution, the survival rate of morula embryos was 94.1%, 85.4% and 59.7% for EFS30, EFS35 and EFS40 group, respectively. Hatching rate of morula embryos after 72 h culture was 30.6%, 25% and 11.3% for EFS30, EFS35 and EFS40 group, respectively. The survival rate of expanded blastocyst embryos after 24 h culture was 90.4%, 98.5% and 100% for EFS30, EFS35 and EFS40 group, respectively. Hatching rate of expanded blastocyst embryos after 48 h culture was 46.2%, 57.6% and 64.3% for EFS30, EFS35 and EFS40 group, respectively. Conclusion: The EFS30 solution was the best for vitrification of mouse morulae. The EFS40 solution was the best for vitrification of mouse expanded blastocysts. The mouse expanded blastocyst was better than mouse morula for vitrification of mouse embryos.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.2
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• pp.137-148
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• 1999

Purpose of the present study was to find the optimal ovulation induction medicine for the maturation and development of immature oocytes and culture media for 2-cell embryos in the mouse model. ICR female mouse aged 6 to 8 weeks, were stimulated with 5 IU PMSG injection. At 47 to 50 hour post-PMSG injection, ovaries were dissected out and oocytes-cumulus complexes were punctured. The oocyte-cumulus complexes were cultured in media containing various ovulation induction medicine, CC, HMG and Metrodin for 18 hours. Female ICR mice were stimulated with 5 IU PMSG and 48 hours later were injected 5 IU of hCG, then female and male mice were mated. At 48 hour post-hCG injection, oviducts were dissected out and 2-cell embryos were flushed. The 2-cell embryos were cultured in various media, Ham's F-10 media of milli-Q water $(3^{\circ})$, Ham's F-10 media of HPLC (high performance liquid chromatography, Baxter) water, Medicult media, HTF (human tubal fluid) media for 96 hours. The results were as follows. 1. When the oocytes-cumulus complexes were cultured in $10^{-9}{\mu}g/ml{\sim}10^{-8}{\mu}g/ml$ of CC, those were suppressed in meiotic maturation $(28.2{\sim}33.7%)$. Whereas the oocytes-cumulus complexes were cultured in $10^{-7}{\mu}g/ml{\sim}10^{-4}{\mu}g/ml$, these were not effected in meiotic maturation $(54.5{\sim}72.7%)$. 2. When the oocytes-cumulus complexes were cultured in $10^{-4}{\mu}g/ml{\sim}10^{-1}{\mu}g/ml$ of Metrodin, those were suppressed in meiotic maturation $(35.7{\sim}41.5%)$. Meanwhile the oocytes-cumulus complexes were cultured in $10^{-7}{\mu}g/ml{\sim}10^{-5}{\mu}g/ml$, those were not effected in meiotic maturation $(54.2{\sim}70.3%)$. 3. When the oocytes-cumulus complexes were cultured in $10^{-5}{\mu}g/ml{\sim}10^{-4}{\mu}g/ml$ of HMG, those were suppressed in meiotic maturation $(48.2{\sim}50.4%)$. As being cultured in $10^{-7}{\mu}g/ml{\sim}10^{-6}{\mu}g/ml$, increased in meiotic maturation $(75.8{\sim}80.7%)$. 4. When the 2-cell embryos were cultured in Ham's F-10 media of milli-Q water $(3^{\circ})$, Ham's F-10 media of HPLC (high performance liquid chromatograpy, Baxter) water, Medicult media, HTF (human tubal fluid) media, developmental rates to blastocyst and hatching for 96 hour were 50.0%, 45.2%, 71.5% and 95.6%, respectively.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.2
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• pp.171-178
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• 1999

The follicular fluid (FF) of ovary contains various biological active products which affected on the growth of follicles and the fertilization of oocyte in physiological reproductive process of mammals. This study was designed to determine the effects of human FF on fertilization of oocyte and embryonal development in vitro culture. The FF was prepared as clear without blood contamination by needle aspiration from mature follicles of human at the time of oocytes retrieval for in vitro fertilization (IVF). As the medium for culture in vitro of embryonal cells, human tubal fluid (HTF) supplemented with follicular fluids at concentrations of 10%, 40% and pure FF were used. These effects were compared to control group of cultured embryos in HTF supplemented with 0.4% BSA (bovine serum albumin). For IVF, 64 eggs in control group, 67 eggs in 10% FF, 57 eggs in 40% FF and 64 eggs in pure FF were respectively allocated. And the rates of fertilization were almost similar in all groups as resulting 82.81% in control, 85.07% in 10% FF, 87.71% in 40% FF and 81.25% in pure FF. On the examination for embryonal cleavage from fertilized eggs, the rates of developing to 4 cell stage was similar in all groups, as results 98.11% in control, 98.27% in 10% FF and 98% in 40% FF but 78.84% in pure FF. And the rates of developing to 8-16 cell stage were significantly reduced as 44% in 40% FF and 44.23% in pure FF (p<0.05) compare to 71.69% in control media. As likewise, the rates of developing to morular stage were also significantly reduced to 36% (p<0.05) and 21.15% (p<0.01) respectively in 40% FF and pure FF. And the rates to blastocystic stage of embryo was lowest as 7.69% in pure FF (Table 1). The quality of embryonal cells on cleavage to the 8-16 cell stage was poorer, higher concentrations of FF. The rates of grade 1 in pure FF, as 23.07%, was lowest compare to those of other groups, in which the rates of grade 1 in control, 10% FF and 40% FF were 58.49%, 47.36% and 34% respectively. And on the contrary, the rate of grade 4 in pure FF was highest as 23.07%, while those were 5.66% in control, 8.77% in 10% FF and 20% in 40% FF (Table 2). On the viability of embryos, the rate of embryonal cell death was more rise, at the higher concentrations as well as longer exposure in the follicular fluid. At 48 hours after in vitro culture of embryos, the rate of survival embryos in pure FF was markedly lowered as 44.23%, compare to that of control (p<0.05). But there was not significant difference between the rates of survival embryos in each group beside the pure FF, which the rates were 77.35% in control, 70.17% in 10% FF and 60% in 40% FF respectively. And at 72 hours after in vitro culture, the rates of survival embryos were also significantly dropped to 21.15% in pure and 36% in 40% at concentration of FF compare to 62.26% in control (p<0.05, p<0.01). Finally, the rate of embryonal death at 96 hours after in vitro culture was highest as 82.69% in pure FF among all groups which those were 35.84 in control, 56.14% in 10% FF and 64% in 40% FF respectively (Fig. 1, 2, 3). In conclusion, this study suggests that the FF has no effects, in particular, to the in vitro fertilization of oocytes but exerted a bad effect to the cleavage, quality and viability of the embryonal cells during in vitro culture. However, the FF is harmful on embryonal development at conditions in higher concentration and especially on the embryos after $8{\sim}16$ cell stage.

• Korean Journal of Animal Reproduction
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• v.23 no.1
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• pp.85-92
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• 1999

This study was designed to determine effect of cryoprotectant kinds and cell stages on OPP vitrification method in mouse embryos. The freezing speed, cryoprotectants and cell stage could affect of embryo viability following various vitrification methods. The vitrification solution used were consisting of 40% (v/v) ethylene glycol, 18% (w/v) Ficoll, 0.3 M sucrose solution in holding medium (D-PBS supplemented with 5% FCS: HM) (EFS) or 16.5% ethylene glycol , 16.5% dimethyl sulfoxide, 0.5 M sucrose in HM (EDS). The embryos were collected from oviduct at 18 h after hCG injection and then washed and cultured in mHTF medium until use. In experiment 1, the blastocysts were vitrified by OPP straw to determine the optimal vitrification solution of EFS or EDS. The post-thaw survival rates at re-expanded stage rates were significantly different between EFS and EDS (95.0 vs 100%), but at hatching stage was not different between EFS and EDS (90.0 vs 95.0%). respectively. In experiment 2, zygotes, 2-, 4-cell, morula and blastocysts were vitrified by OPP method to determine the acceptable of early stage embryos. The development rates to expanded blastocyst in zygote (70.0%) were significantly lower rather than those in 2-, 4- 8-cell, compacted morula or blastocyst (89.7, 90.0, 92.8, 97.6 or 97.5%), respectively. However, the cell number of post-thaw developed to expanded blastocyst in blastocyst and control blastocyst stage (39.6$\pm$2.81, 35.7$\pm$2.98) were significanty higher than those in zygote, 2-, 4-, 8-cell, compacted morula (29.8$\pm$3.21, 31.3$\pm$3.83, 29.3$\pm$3.58, 28.9$\pm$3.21 or 30.8$\pm$2.93). In experiment 3, the zygotes were exposed in VSl for 1, 2, and 3 min to the optimal exposed time. The cleavage rates (91.6, 88.5, 88.9%) and develop mental rates to blastocyst (83.3, 74.3 and 69.4%) depends on the exposed time in VSl were not significantly different among 1, 2, or 3 min, respectively. The cell number also were not significantly different among exposed time in VS1. respectively. These results indicate that OPP method could be useful for vitrification either EFS or EDS vitrification solution. The post-thaw survival rates at zygote were significantly lower than those at 2-, 4-, 8-cell, morula or blastocyst, respectively. The zygote stage were more sensitive rather than late stage embryos. The exposing time in VS1 for 1 min was better than that for 2 or 3 min, even it was not significantly different. The OPP vitrification method could be useful of mouse embryos either with EFS or EDS vitrification solution.

• Clinical and Experimental Reproductive Medicine
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• v.34 no.2
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• pp.117-124
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• 2007

Objective: The aim of this study was to compare the efficacy of slow freezing with vitrification method for cryopreservation of mouse pronuclear stage embryos. Methods: Mouse pronuclear embryos obtained from superovulated mice and classified into 2 groups of slow freezing and vitrification. Slow freezing solution consisted of 1.5 M PROH, 0.1 M sucrose, while vitrification solution consisted of 40% ethylene glycol, 18% Ficoll and 0.5 M sucrose diluted in Dulbecco's phosphate-buffered saline supplemented with 10% SSS. Recovery and survival rates after thawing and development rates to hatching balstocyst stage were compared between two groups. Results: After freezing and thawing, recovery rate of slow freezing group was 93.8%, whereas vitrification group was 66.5% (p<0.01). Survival rate of recovered embryos were similar between two groups as 83.2% in slow freezing and 87.6% in vitrification. Embryo development rates to 2-cell stage after 24 hrs (77.0% vs 59.1%), 4-cell after 48 hrs (72.6% vs 53.3%), blastocyst after 96 hrs (53.1% vs 40.1%) of thawing were significantly higher in vitrification group than those of slow freezing group, respectively. Conclusion: The vitrification method may provide better developmental competence of frozen-thawed embryos than that of slow freezing method for cryopreservation of mouse pronuclear stage embryos.

• Journal of Embryo Transfer
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• v.29 no.3
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• pp.235-240
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• 2014

Ethylene glycol (EG) has been successfully used as a cryoprotectant for vitrification of mammalian embryos (including human embryos) due to its low formula weight and high permeation into cells compared with other cryoprotectants, including propylene glycol (PROH). Cryopreservation is able to store the surplus pre-embryos for freezing and furthermore thawing and transfer in a subsequent cycle. This study was carried out to evaluate the effects of embryonic stage, cryoprotectant, and freezing-thawing method on the rates of survival and development of the cryopreserved mouse early embryo and finally to establish the cryopreservation method of surplus embryos obtained during assisted reproductive technology (ART). Female ICR mice (6~8 weeks old) were induced to superovulate by sequential intraperitoneal injection of 5 IU PMSG and 5 IU hCG 48 h apart. Mouse embryos were collected according to its developmental stage after the injection of hCG. Embryos were cryopreserved not only during cryoprotectant step (1~4 step) but also in a variety of media (HTF, IVF medium, D-PBS) and cell stage. The results were as follows : There is no clear advantage in these freezing media of rapid method, but 4 cell and 8 cell of slow method (2, 3 and 4 step) have advantage in D-PBS. The development of embryos according to cell stage become greater in 8 cell stage. In the treatment steps of cryopreservation, the development of embryo to blastocyst was similar among rapid method, but the development of 4 cell and 8 cell embryos to blastocyst according to slow method was better than rapid method.

• Development and Reproduction
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• v.10 no.2
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• pp.105-113
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• 2006

Reactive oxygen species(ROS) generated in cellular metabolism have an effect on cell maturation and development. In human reproductive tract, oxidative injury by ROS may induce female infertility. Also, oxidative injury may be responsible for developmental retardation and arrest of mammalian preimplantation embryos. Activating transcription factor 4(ATF4) is a member of the cyclic-AMP response element-binding(CREB) familiy of basic region- leucine zipper(bZip). ATF4 is known to regulate stress response to protect cell from various stress factors and inducer of apoptisis. The purpose of this study was to investigate whether ATF4 is involved in the defensive mechanism in oxidative stress condition during the development of mouse preimplantation embryos. To verify the expression of ATF4 in oxidative stress condition, 2-cell stage embryos were cultured in HTF media containing 0.1mM, 0.5mM or 1mM hydrogen peroxide($H_2O_2$) for 1hr(2-cell), 8hr(4-cell), 17hr(8-cell), 24hr(morula), 48hr(early blastocyst) or 64hr(late blastocyst). The developmental rate decreased in the 0.1mM $H_2O_2$ treated group compared with control group. In embryos treated with 0.5mM and 1mM $H_2O_2$ showed 2-cell block. As a results of the semi-quantitative RT-PCR analysis of SOD1, ATF4 and Bax gene expression, SOD1, ATF4 and Bax genes were increased in 0.1mM, 0.5mM, 1mM $H_2O_2$ treated groups compared with control group. In 2-cell embryos, expression of SOD1, ATF4 and Bax genes were notably increased in 0.1mM, 0.5mM, 1mM $H_2O_2$ treated groups compared with control group. Immunofluorescence analysis showed that ATF4 protein was localized at the cytoplasm of preimplantation embryos. The increase in ATF4 immunoreactivety was observed in the 0.1mM, 0.5mM, 1mM $H_2O_2$ treated groups compared with control group. It suggests that oxidative stress by $H_2O_2$ induces expression of ATF4 and may be involved in protection mechanism in preimplantation embryos from oxidative injury.