This study was to examine whether the vitrified, one-step diluted and direct transferred Hanwoo IVM/IVF/IVC blastocysts can be successfully survived in vivo and they were succeeded into the live birth. For vitrification, blastocysts were serially exposed in glycerol (G) or/and ethylene glycol (EG) mixtures [10% (v/v) G for 5 min, 10% G plus 20% EG (v/v) for 5 min, and 25% G plus 25% EG (v/v) for 30 sect] which is diluted in 10% FBS added D-PBS. Thawing of straw was carried out in air for 10 sec and then in water bath of $25^{\circ}C$ for 20 sec. One-step dilution within the straw was done in water bath of $25^{\circ}C$ for 1 min. Vitrified and one-step diluted embryos were directly transferred into 36 (natural or hormone induced synchronized) recipient cows in 6 areas of Kyungsang Buk-Do. Pregnancies were confirmed at first when recipient cows did not return to the subsequent estrus cycle, and later by manual palpation per rectum on day 45, 90 and then living calves were derived into parturition. Overall pregnancy was 33.3%(12/36), However, higher pregnancy was obtained when the recipients exhibited estrus one day earlier than the age of transferred embryos (53.3 vs 25.0-27.3%), irrespective of synchronization methods. Also, parous recipients became pregnant higher than nulliparous heifers, And, there were not different in pregnancy rates by the aspect of corpus luteum (CL) quality of recipients (good, 29.4; fair, 37.5; poor, 33.3%). One hundred eight of frozen-thawed Hanwoo blastocysts were directly transferred into 36 recipient cows. In 12 of pregnant cows, 3 cows were aborted and 9 cows were calved [single, 66.7% (6/9): twin, 33.3% (3/9)]. Total embryo implantation rate was 11.1% (12/108). However, 9 Hanwoo calves were lived. Therefore, these results demonstrate that direct transfer technique of vitrified and one-step diluted bovine blastocysts can be applied easily and effectively with the higher pregnancy rate on field trial without the equipment and embryological skills.
Radioiodide uptake in thyroid follicular epithelial cells, mediated by a plasma membrane transporter, sodium iodide symporter (NIS), provides a first step mechanism for thyroid cancer detection by radioiodide injection and effective radioiodide treatment for patients with invasive, recurrent, and/or metastatic thyroid cancers after total thyroidectomy. NIS gene transfer to tumor cells may significantly and specifically enhance internal radioactive accumulation of tumors following radioiodide administration, and result in better tumor control. NIS gene transfers have been successfully performed in a variety of tumor animal models by either plasmid-mediated transfection or virus (adenovirus or retrovirus)-mediated gene delivery. These animal models include nude mice xenografted with human melanoma, glioma, breast cancer or prostate cancer, rats with subcutaneous thyroid tumor implantation, as well as the rat intracranial glioma model. In these animal models, non-invasive imaging of in vivo tumors by gamma camera scintigraphy after radioiodide or technetium injection has been performed successfully, suggesting that the NIS can serve as an imaging reporter gene for gene therapy trials. In addition, the tumor killing effects of I-131, ReO4-188 and At-211 after NIS gene transfer have been demonstrated in in vitro clonogenic assays and in vivo radioiodide therapy studies, suggesting that NIS gene can also serve as a therapeutic agent when combined with radioiodide injection. Better NIS-mediated imaging and tumor treatment by radioiodide requires a more efficient and specific system of gene delivery with better retention of radioiodide in tumor. Results thus far are, however, promising, and suggest that NIS gene transfer followed by radioiodide treatment will allow non-invasive in vivo imaging to assess the outcome of gene therapy and provide a therapeutic strategy for a variety of human diseases.
Han, E Jung;Kim, Seul Ki;Lee, Jung Ryeol;Jee, Byung Chul;Suh, Chang Suk;Kim, Seok Hyun
Clinical and Experimental Reproductive Medicine
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제42권4호
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pp.169-174
/
2015
Objective: To assess compliance with Korean guidelines for embryo transfer, the possible reasons for non-compliance, and multiple pregnancies according to each specific condition in compliant cycles. Methods: A single-institution, retrospective study was conducted of 256 fresh in vitro fertilization cycles during 2012-2014. To assess compliance with Korean guidelines, the maximum recommended number of embryos transferred (according to criteria of age, transfer day, and presence of favorable conditions) was compared with the actual number of embryos transferred. Clinical pregnancy rate (PR) was assessed as the percentage of pregnant women resulting from each set of transfer conditions, including the number of embryos transferred. The multiple pregnancy rate (MPR) was calculated as the percentage of pregnant women with a multifetal pregnancy. Results: The compliance rate with the Korean guidelines was 96.5% (247/256). Non-compliance occurred in nine cycles owing to poor embryo quality, repeated implantation failure, or hostile endometrium. In compliant cycles, the PR was 31.2% (77/247), and the MPR was 27.3% (21/77; 20 twins and one triplet). Higher MPR was noted in two types of transfer conditions: transfer of three cleavage embryos in women aged 35-39 years with favorable conditions (66.7%; primarily from those aged 35-37 years) and transfer of two blastocysts in women aged ${\geq}40$ years with favorable conditions (50%). Conclusion: Under the Korean guidelines, compliance rate was high in our center. Multiple pregnancies occurred primarily in group with favorable conditions. In high-risk groups for multiple pregnancies, reducing number of embryos transferred should be considered than suggested in the guideline.
Objective: This study evaluated the effect of cyclic pre-calcification treatment on the improvement of bioactivity and osseointegration of Ti-6Al-4V mini-screws. Methods: The experimental groups were: an untreated group (UT), an anodized and heat-treated group (AH), and an anodized treatment followed by cyclic pre-calcification treatment group (ASPH). A bioactive material with calcium phosphate was coated on the mini-screws, and its effects on bioactivity and osseointegration were evaluated in in vitro and in vivo tests of following implantation in the rat tibia. Results: As a result of immersing the ASPH group in simulated body fluid for 2 days, protrusions appearing in the initial stage of hydroxyapatite precipitation were observed. On the 3rd day, the protrusions became denser, other protrusions overlapped and grew on it, and the calcium and phosphorus concentrations increased. The removal torque values increased significantly in the following order: UT group (2.08 ± 0.67 N·cm), AH group (4.10 ± 0.72 N·cm), and ASPH group (6.58 ± 0.66 N·cm) with the ASPH group showing the highest value (p < 0.05). In the ASPH group, new bone was observed that was connected to the threads, and it was confirmed that a bony bridge connected to the adjacent bone was formed. Conclusions: In conclusion, it was found that the surface treatment method used in the ASPH group improved the bioactivity and osseointegration of Ti-6Al-4V orthodontic mini-screws.
A heart supplies blood of about 15, 000 liters to each human organ in a day. A normal function of heart valves is necessary to accomplish these enormous work of heart. The disease of heart valve develops to a narrowness of a closure, resulting in an abnormal circulation of blood. In an attempt to eliminate the affliction of heart valves, the operative method to replace with artificial heart valves has developed and saved numerous patients over past 30 years. This replacement operation has been performed since early 1960`s in Korea, but all the artificial heart valves used are imported from abroad with very high costs until recent years. New artificial heart valves have been developed in Korea Advanced Institute of Science and Technology since early 1980`s. The first developed valve was designed with a free-floating pyrolytic carbon disk that is suspended in a titanium cage. The design of the valve was tested in vitro, and in animals in 1987. The results from this study was that the eccentrically placed struts creates a major and minor orifice when the disc opens and stagnation of flow in the area of the minor orifice has led to valve thrombosis. In this work, the design of the valve was changed from a single - leaflet valve to double - leaflet one in order to resolve the problems observed in the first - year tests. Morphological and hemodynamic studies were made for the newly designed valves through the in vitro and in vivo tests. The design and partial materials of the artificial heart valve was improved comparing with first - year`s model. The disc in the valve was modified from single - leaflet to bi - leaflet, and the material of the cage was changed from titanium metal to silicon - alloyed pyrolytic carbon. A test was made for the valve in order to examine its mechanical performance and stability. Morphological and hemodynamic studies were made for the valve that had been implanted in tricuspid position of mongrel dogs. All the test animals were observed just before the deaths. A new artificial heart valve was designed and fabricated in order to resolve the problems observed in the old model. The new valve was verified to have good stability and high resistance to wear through the performance tests. The hemodynamic properties of the valve after implantation were also estimated to be good in animal tests. Therefore, the results suggest that the newly designed valve in this work has a good quality in view of the biocompatibility. However, valve thrombosis on valve leaflets and annulus were found. This morphological findings were in accordance with results of surface polishing status studies, indicating that a technique of fine polishing of the surface is necessary to develop a valve with higher quality and performance.
We had presented an inductive type intraocular pressure sensor (L-sensor) in previous work. The distance between a micro coil and a ferrite on the membrane was modulated by pressure, and as a result the inductance and resonant frequency were changed. However, L-sensor has some problems to implant in eyes. First problem is low sensitivity. When L-sensor was implanted in rabbit's eyes, resonant frequency of L-sensor was very hard to detect. Second problem is biocompatibility. Size of L-sensor is $6{\times}7{\times}1.2mm$. When L-sensor was implanted in the eyes, it caused the inflammation. Therefore, this study suggests an inductive and capacitive type IOP sensor (LCsensor). The sensitivity of the LC-sensor 27.3 kHz/mmHg under 60mmHg. It is much larger than 14 kHz/mmHg of the L-sensor. And the size of LC-sensor is 47% smaller than L-sensor. After 2 weeks from the implantation of LC-sensor into rabbit eyes, we measured the changes of resonant frequency of LC-sensor according to increased IOP by Balanced Salt Solution (BSS) injection. As a result, the sensitivity of LC-sensor in in vivo test is 25 kHz/mmHg. That is similar to the sensitivity of in vitro test.
This experiment was carried out to develop a new technique of identifying XX of XY-bearing bisected embryos prior to implantation by immunological method. H-Y antiserum prepared in inbred Wastar female rats by repeated immunization with spleen cells from males of the same strain. The reactivity of H-Y antibody was confirmed by culturing mouse embryos in the medium containing H-Y antiserum and complement obtained from the guinea pig. The optimal condition for the activity of H-Y antibody was also investigated by culturing embryos under the concentraton or affected H-Y antibody was also investigated by culturing embryos under the concentration or affected H-Y antibody and culture rate. However, production of live young or sex rates of male and female from embryos transferred with psudopregnant. The biological test with the morula stage embryos showed that H-Y antibody was formed in all female rats immunized with spleen cell, but it was formed only in 80% female rats immunized with the antigen. When the bisected mouse embryos were cultured in vitro for 5~6 hours in morula stage, of 457 bisected embryos 81.4% of then were developed to the blastocyst stage. When the concentration rate of complement to H-Y antiserum varied from 1.0~5.0${mu}ell$, the lysis-rate of embryo was 19.5 to 67.3%. The concentration rate of complement did not influence the lysis-rate of embryos(P<0.05). The morphology embryos of bisected, zona-free and intact embryos showed the embryos lysis rate of 58.6, 42.7 and 48.5% respectively(P<0.05). Pregnancy rate were 50.0, 45.5 and 57.1% in psudopregnant recipient transferred with bisected, zona-free and intact blastocyst embryos. However, production of live youngs, sexual rate of male or female was 24(50.0:50.0), 22(45.5:55.5) and 36(58.3:41.7)mice, but affected and non affected half embryos with H-Y antiserum treatment was 23.1 and 26.7%. Also production of live youngs and sexual rate was 14(92.9:7.1) and 17(17.6:82.4)mice in affected and non affected half embryos in H-Y antiserum treatment(P<0.05).
K. S. Chung;Ko, S. A;S. J. Song;J. T. Do;Park, Y. S.;Lee, H. T.
한국가축번식학회지
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제26권4호
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pp.385-394
/
2002
This study was constructed the correlations of the embryonic developmental rates and the frequency of chromosome aberration using ear-skin-fibroblast cell in nuclear transfer (NT) derived embryos. Karyoplast-oocyte complexes were fused and activated simultaneously, then cultured for seven days to assess development. The developmental rates of NT and in vitro fertilization (IVF) embryos were 55.4% vs 63.5%, 31.7% vs 33% and 13.4% vs 16.8% in 2 cell, 8 cell and blastocyst, respectively. Firstly, the frequency of chromosome aberrations were evaluated using fluorescent in situ hybridization (FISH) technique with porcine chromosome 1 submetacentric specific probe. Chromosome aberration was detected at day 3 on the embryo culture, the percentages of chromosomal aneuploidy in NT and IVF embryos at 4-cell stage were 40%, 31.3%, respectively. Secondly, embryonic fragmentation was evaluated at 4-cell stage embryo. Frequency of embryonic fragmentations was in 51.3% of NT, 61.3% of IVF, 28.9% of parthenogenetic activation at 4-cell stage. The proportion of fragmentation in NT embryos was higher than activation embryos. This result indicates that chromosomal abnormalities and embryonic fragments are associated with low developmental rate in porcine NT embryo. It is also suggest that abnormal porcine embryos produced by NT related with lower implantation rate, increased abortion rate and production of abnormal fetuses.
The trophectoderm is one of the earliest cell types to differentiate in the forming placenta. It is an important for the initial implantation and placentation during pregnancy. Trophoblast stem cells (TBSCs) develop from the blastocyst and are maintained by signals emanating from the inner cell mass. However, several limitations including rarity and difficulty in isolation of trophoblast stem cells derived from blastocyst still exist. To establish a model for trophoblast differentiation, we isolated TBSCs from human term placenta ($\geq$38 weeks) and characterized. Cell cycle was analyzed by measuring DNA content by FACS analysis and phenotype of TBSCs was characterized by RT-PCR and FACS analysis. TBSCs have expressed various markers such as self-renewal markers (Nanog, Sox2), three germ layer markers (hNF68, alpha-cardiac actin, hAFP), trophoblast specific markers (CDX-2, CK7, HLA-G), and TERT gene. In FACS analysis, TBSCs isolated from term placenta showed that the majority of cells expressed CD13, CD44, CD90, CD95, CD105, HLA-ABC, cytokeratin 7, and HLA-G. Testing for CD31, CD34, CD45, CD71, vimentin and HLA-DR were negative. TBSCs were shown to decrease the growth rate when cultured in conditioned medium without FGF4/heparin as well as the morphology was changed to a characteristic giant cell with a large cytoplasm and nucleus. In invasion assay, TBSCs isolated from term placenta showed invasion activities in in vivo using nude mice and in vitro Matrigel system. Taken together, these results support that an isolation potential of TBSCs from term placenta as well as a good source for understanding of the infertility mechanism.
Yi, Hyun Jeong;Koo, Hwa Seon;Cha, Sun Hwa;Kim, Hye Ok;Park, Chan Woo;Song, In Ok
Clinical and Experimental Reproductive Medicine
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제43권2호
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pp.133-138
/
2016
Objective: To determine the incidence of embryo retention (ER) in the transfer catheter following embryo transfer (ET) in blastocyst transfer and investigate whether retransferring retained embryos has an impact on reproductive outcomes in patients undergoing in vitro fertilization-ET. Methods: We retrospectively analyzed the records of 1,131 blastocyst transfers, which comprised 223 single blastocyst transfer (SBT) and 908 double blastocyst transfer (DBT) cycles. Each SBT and DBT group was classified depending on whether ET was performed without retained embryos in the catheter during the first attempt (without-ER group) or whether any retained embryos were found following ET (ER group) for the purpose of comparing reproductive outcomes in a homogenous population. Results: The overall incidence of finding retained embryos was 2.8% (32/1,131). There were no retained embryos in SBT cycles. In DBT cycles, implantation rates (30.0% vs. 26.6%), positive ${\beta}-hCG$ rates (57.2% vs. 56.2%), clinical pregnancy rates (45.3% vs. 46.9%), and live birth rates (38.9% vs. 43.8%) were not significantly different between the without-ER and ER groups. There were no significant differences in the mean birth weight (g) $2,928.4{\pm}631.8$ vs. $2,948.7{\pm}497.8$ and the mean gestational age at birth ($269.3{\pm}17.2days$ vs. $264.2{\pm}25.7days$). A total of nine cases of congenital birth defects were found in this study population. Eight were observed in the without-ER group and one in the ER group. Conclusion: Our results suggest that retransfer of retained embryos does not have any adverse impact on reproductive outcomes in blastocyst transfer cycles. Furthermore, our results support finding that SBT might be advantageous for decreasing the incidence of retained embryos in catheters.
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