• Asian-Australasian Journal of Animal Sciences
• /
• v.17 no.10
• /
• pp.1369-1373
• /
• 2004

The percentages of sperm motility and normal acrosome on the liquid boar semen diluted and preserved at $4^{\circ}C$ with lactose hydrate, egg yolk and N-acetyl-D-glucosamine (LEN) diluent were significant differences according to preservation day and incubation time, respectively. The sperm motility steadily declined from 96.9% at 0.5 h incubation to 78.8% at 6 h incubation at 1 day of preservation. However, the sperm motility rapidly declined after 4 day of preservation during incubation. The normal acrosome steadily declined from 93.3% at 0.5 h incubation to 73.8% at 6 h incubation at 1 day of preservation. However, the normal acrosome rapidly declined after 3 day of preservation during incubation. The rates of sperm penetration and polyspermy were higher in 5 and $10{\times}10^6$ sperm/ml than in 0.2 and $1{\times}10^6$ sperm/ml. Mean numbers of sperm in penetrated oocyte were highest in $10{\times}10^6$ sperm/ml compared with other sperm concentrations. The rates of blastocysts from the cleaved oocytes (2-4 cell stage) were highest in $1{\times}10^6$sperm/ml compared with other sperm concentrations. In conclusion, we found out that liquid boar sperm stored at $4^{\circ}C$ could be used for in vitro fertilization of pig oocytes matured in vitro. Also, we recommend $1{\times}10^6$sperm/ml concentration for in vitro fertilization of pig oocytes.

• Korean Journal of Animal Reproduction
• /
• v.21 no.1
• /
• pp.19-23
• /
• 1997

This present study was carried out to examine the effect of maturation media and liquid boar semen on in vitro maturation and feritilization of pig oocytes. The results obtained were as follows : When the oocytes were cultured for 36∼42 hours in mTCM-199, Waymouth MB 725/1 and mTLP-PVA medium, the maturation rates were 90%, 92% and 88%, respectively. The sperm penetration rates of pig oocyte matured in vitro were 87%(mTCM-199), 90%(Waymouth MB 725/1) and 86%(mTLP-PVA), respectively. The rates of nuclear maturation and fertilization of pig oocytes among three different media did not differ. However, the rate of male pronucleus formation of pig oocytes was significantly higher in pig oocytes matured in Waymouth MB 725/1(91%) than oocytes matured in mTCM-199(66%) and mTLP-PVA(62%) medium (P<0.05). When the collected sperm-rich fraction without diluent was used fro in vitro fertilization in mTCM-199 fertilization medium, the fertilization rate was 87.9%. However, when the liquid boar semen diluted with B tschwiler diluent was used at day 3 and 5 after dilution, the fertilization rate was 40.8% and 0.0%, respectively.

• Biomolecules & Therapeutics
• /
• v.17 no.3
• /
• pp.241-248
• /
• 2009

The purpose of this study was to evaluate the effect of methylsulfonylmethane (MSM) on hair growth promotion of magnesium ascorbyl phosphate (MAP) for the treatment of alopecia. Aqueous solutions of MAP 7.5% with or without MSM 1%, 5% or 10% were prepared and applied onto the depilated back skin of the male mice once a day for 20 days. The degree of hair growth was evaluated by visual scoring using hair growth quantification scale (0-5, 0 being initial state and 5 being complete hair growth). In vitro transdermal penetration and intradermal retention studies of MAP were performed with Franz diffusion cell using hairless mice skin. Hair growth in the group treated with the aqueous solution containing MAP 7.5% and MSM 10% was comparable to or better than the result in the group treated with minoxidil 5% solution. Hair growth promotion of MAP was dose-dependently increased by the presence of MSM used in combination with MAP 7.5% solution. The in vitro transdermal penetration of the MAP was decreased in proportion to the concentration of MSM. However, intradermal retention of MAP was profoundly and dose-proportionally increased as a function of MSM concentration, reaching 802 ${\mu}g/cm^2$ in the presence of MSM 10% (200-fold increase). The effect of MSM on hair growth promotion of MAP was dose-proportional to the concentration of MSM due to the enhanced intradermal retention of MAP in the presence of MSM. Therefore, topical application of MAP together with MSM appears to be useful for the treatment of alopecia.

• Journal of Korean Society of Forest Science
• /
• v.99 no.6
• /
• pp.871-877
• /
• 2010

This study was conducted to investigate the effects of host tree species, temperature and humidity on ex vitro for seed germination in endangered species of Loranthus tanakae. In addition, we compared seed shapes between Loranthus tanakae and Viscum album that we could have easily shown in ex vitro condition. Seeds were germinated after one week inoculation and followed to develop radicles. Seed germination rates of Loranthus tanakae were 80~95% in most of the experimental conditions. The highest rate of holdfast penetrated to host plants was 72% in Populus alba var. pyramidalis among 13 different species tested at $20^{\circ}C$. Also the rates of their penetration were 57% in Morus bombycis, 55% in Acer palmatum and 42% in Castanea crenata at $20^{\circ}C$. Seeds were germinated under condition without irrigation and followed to withered in 12 weeks later. Stages of seed germination of the Loranthus tanakae were followed by radicle induction, holdfast development, haustorium formation and penetration in order in total period of 14 weeks.

• Asian-Australasian Journal of Animal Sciences
• /
• v.17 no.2
• /
• pp.164-167
• /
• 2004

This study was carried out to investigate in vitro fertilization and development of in vitro matured pig oocytes inseminated with the Duroc boar sperm by different sperm washing media after thawing of the 5 ml frozen straws. Immature follicular oocytes (30-40) were transferred into each well of a Nunc 4-well multidish containing $500{\mu}l$ mTCM199 maturation medium. The sperm rich portion of ejaculates was collected into a 250 ml insulated vacuum bottle and gradually cooled 22 to $24^{\circ}C$ over a 2 h period. Semen was centrifuged at 800 g for 10 min and the seminal plasma discarded. Sperm were esuspended in a lactose-egg yolk and N-acetyl-Dglucosamine (LEN) diluent to contain $1{\times}10^{9}$ sperm/ml and cooled to $5^{\circ}C$ over a 2 h period. Immediately before freezing, semen was rediluted with an equal volume of LEN+4% glycerol and packed into 5 ml straws. After thawing of the 5 ml straw, the 5 ml semen was diluted with 20 ml Beltsville thawing solution (BTS) at room temperature. Oocytes were inseminated with untreated (unwashed and nonpreincubated) or treated sperm (washed two times in BTS, mTLP-PVA and mTBM media, respectively and nonpreincubated) with $2{\times}10^{7}$ sperm concentration. Oocytes were coincubated for 6 h in $500{\mu}l$ mTBM fertilization. At 6 h after IVF, oocytes were transferred into $500{\mu}l$ NCSU-23 culture medium for further culture of 6 h. Sperm penetration, polyspermy and male pronuclear formation of oocytes at 12 h after IVF and developmental ability of oocytes at 48 h after IVF were evaluated. Sperm penetration rate, male pronuclear formation and rate of cleaved embryos were higher in the BTS, mTLP-PVA and mTBM treatments than the unwashed treatment (p<0.05). The rate of blastocysts from the cleaved oocytes (2-4 cell stage) were higher in the mTLP-PVA treatment than in the unwashed, BTS and mTBM treatments. In conclusion, we recommend the washing of frozen-thawed sperm with mTLP-PVA medium before in vitro fertilization of oocytes in mTBM medium.

• Clinical and Experimental Reproductive Medicine
• /
• v.29 no.1
• /
• pp.57-66
• /
• 2002

Objective : This study was designed to investigate the interrelationship and clinical usefulness of sperm morphology by strict criteria (SM), acrosome reaction following ionophore challenge test (ARIC) and sperm penetration assay (SPA) using zona-free hamster ova as prognostic factors in in vitro fertilization. Materials and Methods: Semen samples were provided by 83 patients undergoing IVF. We first evaluated the differences between normal fertilization group and poor fertilization group on three andrologic tests. Secondly, we analyzed the relationship between the three andrologic tests and in vitro fertilization on IVF settings. Finally, we evaluated the effectiveness of the three andrologic tests as the prognostic indicators for fertilizing ability. Results: The fertilization rate of all men in the poor fertilization group was less than 30%; but there was no evidence that this poor fertilization was due to oocyte defects. The results of three andrologic tests were significatly higher in normal fertilization group. Fertilization rate (%) in vitro was highly correlated (p<0.001) with % normal sperm by SM, ARIC value (%), and SPA result. By using Receiver-Operator-Characteristic curve (ROC), we evaluated the effectiveness of these three tests. The sensitivity and specificity of SM, ARIC test and SPA in predicting fertilization potential in IVF setting were 76% and 75%, 84% and 90%, and 76% and 95%, respectively. Conclusion: Our data suggest that the three andrologic tests can be reliable tools as prognostic factors of sperm fertilizing ability. Among these test, ARIC test and SPA gave more accurate information on fertilizing capacity. ARIC test was shown to have a predictive value for fertilizing ability comparable to that of SPA that appears to be a simple and cost-effective addition to current andrology laboratory. Combined application of these three tests may give more information on predicting sperm fertilizing capacity.

• Journal of Environmental Health Sciences
• /
• v.42 no.1
• /
• pp.61-70
• /
• 2016

Objectives: The purpose of this study is to determine the exposure risk to tattoo components by analyzing skin absorption using the in vitro method. Tattoos are commonly used for cosmetic purposes, and the skin of not only the operator but of the people who are undergoing the cosmetic procedure is continuously exposed to hazardous chemicals. Methods: Skin permeation risk determination was conducted by the in vitro Franz diffusion cell method according to the ingredient types of tattoo dyes, such as volatile organic compounds (VOCs), non-volatile organic compounds and heavy metals, using hairless mouse full skin and human cadaver epidermis. Results: The major components with good skin penetration for each type of tattoo dye ingredient were clarified. Among the tatto dye ingredients, 1,2-Dichlorobenzene, Zn, Al, Pb and Ti showed good skin penetration. Most of the skin transmission rates were higher in hairless mouse full skin than in human cadaver epidermis. Conclusion: A possible exposure risk to hazardous substances in tattoo dyes was confirmed from this study. These results are expected to provide a positive contribution to the establishment of management regulations for tattoo dyes.

• Korean Journal of Animal Reproduction
• /
• v.25 no.4
• /
• pp.317-325
• /
• 2001

This study investigated the effect of ferrous sulfate (Fe$^{2+}$) and/or ascorbic acid (Asc) on fertilizing ability in vitro of frozen-thawed boar spermatozoa. Using chlortetracycline (CTC) fluorescence, the spermatozoa was treated in preincubation medium with control, Fe$^{2+}$(1 mM), Asc (0.5 mM) and Fe$^{2+}$Asc to assessed for acrosome reaction, and the oocyte penetration test to determine whether the Fe$^{2+}$ and/or Asc can promote the penetration ability in vitro. When frozen-thawed spermatozoa was washed with preincubation medium, there were significantly (P ＜ 0.05) more acrosome-reacted in medium with Fe$^{2+}$Asc (38%) than control (27%). The penetration rates were also significantly (P ＜ 0.05) higher in medium with Fe$^{2+}$Asc (76%) than control (55%). Next, the lipid peroxidation of sperm was evaluated on the basis of malondialdehyde production following same treatments. The addition of Fe$^{2+}$Asc to sperm suspension increases the formation of malondialdehyde. However, there were not significantly different under the all conditions. The sperm suspension were also treated with control, Fe$^{2+}$, Asc and Fe$^{2+}$/Asc and assayed for sulfhydry1(-SH) group content. In the Fe$^{2+}$/Asc group, sperm-SH group were higher than another groups. In spermatozoa treated with Fe$^{2+}$ and/or Asc, however, no changes in sperm -SH-groups were detected when compared to controls. In another experiment, the activity of sperm binding to zona pellucida was evaluated through binding to salt-stored porcine oocytes. In control and Asc treatment groups, sperm binding to zona pellucida were significantly (P ＜ 0.05) higher than in medium with Fe$^{2+}$. On the other hand, there is not a significant increase in binding to zona pellucida with spermatozoa treated by Fe$^{2+}$/Asc. In summary, the present study suggests that Fe$^{2+}$/Asc causes an enhancement in fertilizing ability that is associated with penetration rate increased without change of spermatozoa binding capacity to homologous zona pellucida.o homologous zona pellucida.

• Journal of the Society of Cosmetic Scientists of Korea
• /
• v.31 no.1 s.49
• /
• pp.43-49
• /
• 2005

Recently, according as people who have sensitive skin increase, we've been giving more importance to the safety of cosmetics. Especially, preservative is known to be one of the main stimuli which cause side-effects of cosmetics. However, there have been few reports describing cell cytotoxicity, skin penetration, oil-aqueous phase partition, anti-microbial activity of preservatives and their correlation with skin irritation. The study is aimed to develop low irritable preservative system with phenoxyethanol, one of the most commonly used preservatives in cosmetics, considering various factors mentioned above. According to our results of cell cytotoxicity against human normal fibroblasts by means of MTT assay, phenoxyethanol showed the lowest cytotoxicity when compared to other preservatives tested (cytotoxicity: pro-pylparaben > butylparaben > ethylparaben > methylparaben > triclosan > phenoxyethanol), but human patch test for assessing shin primary irritation revealed that phenoxyethanol has higher skin irritation than methylparaben and triclosan. We performed in vitro skin penetration test using horizontal Franz diffusion cells with skin membrane prepared from hairless mouse (5 ${\~}$ 8 weeks, male) to evaluate the rate of skin penetration of preservatives. From the results, we found that the higher irritable property of phenoxyethanol in human skin correlates with its predominant permeability (skin penetration: phenoxyethanol > methylparaben > ethylparaben > propylparaben > butylfaraben > triclosan). Therefore, we made an effort to reduce skin permeability of phenoxyethanol and found that not only the rate of skin penetration of phenoxyethanol but also its skin irritation is dramatically reduced in formulas containing oils with low polarity. In the experiments to investigate the effect of oil polarity on the oil-aqueous phase partition of phenoxyethanol, more than $70\%$ of phenoxyethanol was partitioned in aqueous phase in formulas containing oils with low polarity, while about $70 {\~} 90\%$ of phenoxyethanol was partitioned in oil phase in formulas containing oils with high polarity. Also, in aqueous phase phenoxyethanol showed greater anti-microbial activity. Conclusively, it appears that we can develop less toxic preservative system with reduced use dosage of phenox-yethanol and its skin penetration by changing oil composition in formulas.

• Clinical and Experimental Reproductive Medicine
• /
• v.47 no.3
• /
• pp.186-193
• /
• 2020

Objective: The effectiveness of Staphylococcus protein A (SPA) in improving the penetration ability of sperm and reducing antisperm antibody (ASA) titers in immunologically infertile males was evaluated. Methods: Seminal fluid samples were obtained from 15 infertile men, and ASA titers were assessed with the latex agglutination test. Identification of immunoglobulin (Ig) classes and characterization of the antigens involved in the immune response were performed using indirect immunofluorescence. Local ASAs typically present as a mixture of IgG and IgA classes. The capillary tube penetration method was used to assess the capability of spermatozoa to penetrate the cervical mucus (CM). Results: ASAs associated with the neck region of sperm showed a significantly lower migration distance in the CM of infertile females than ASAs associated with the head or tail segments. ASA-positive seminal fluid exhibited significant increases in the mean migration distance (2.6 ± 1.4 cm vs. 1.54 ± 1.1 cm, respectively; p< 0.001) and sperm concentration (174 ± 121.0 × 10³/mL vs. 101 ± 93.7 × 10³/mL, respectively; p= 0.033) after treatment with SPA compared to pre-treated samples. A significant reduction (p< 0.01) in the recorded ASA titer was detected. Conclusion: These results indicate that SPA can be used as a sorting regimen for insemination programs. However, further studies are warranted to assess its influence on pregnancy rate.