Objective: This study was carried out to investigate the effects of 3-dimensional co-culture of human endometrial cells decidualized with progesterone and TGF-${\beta}1$ on the development of 2-cell mouse embryos. Methods: Stromal and epithelial cells isolated from human endometrial tissue were immunostained for cytokeratin and vimentin. Expression of TGF-${\beta}1$, its receptor-1, -2, integrin-${\beta}3$ and prolactin in mono or co-culture according to three different hormone conditions was investigated by RT-PCR. Differential staining was used to investigate the number of ICM and trophectoderm of hatched mouse blastocysts in different three conditions. Results: The immunohistochemical study was positive for cytokeratin or vimentin and confirmed that epithelial and stromal cells were isolated from endometrial tissue successfully. In co-culture, TGF-${\beta}1$, its receptor-1, integrin-${\beta}3$ and prolactin except TGF-${\beta}1$-r2 were expressed in progesterone dominant condition. The hatching and attaching rate were higher in the co-culture with decidualized cells (p<0.05). However, we observed that lots of the incomplete hatched blactocysts attached on non-decidualized cells. The ICM number of hatched mouse blastocysts was higher in co-culture with decidualized and non decidualized cells than media only culture (p<0.05). The trophectoderm number of hatched blastocyst was higher in the co-culture with decidualized cells than non-decidualized cells or media only culture (p<0.05). Conclusion: The administration of progesterone, estrogen and TGF-$\beta$ could induce decidualization of stromal and epithelial cells isolated from human endometrial tissue using 3-dimensional co-culture, and the decidualization of human endometrial cells could increase the hatching and attaching rate of 2-cell mouse embryos.
Purpose : This study quantitatively evaluated the apoptosis In human peripheral blood lymphocytes using flow cytometry, and investigated the possibility of using this method, with a small amount of blood, and the time and dose dependence of radiation-induced apoptosls. Materials and Methods : Peripheral blood lymphocyes were isolated from the heparinized venous blood of 11 healthy volunteers, 8 men and 3 women, with each 10 ml of blood being divided Into IS samples. The blood lymphocytes were Irradiated using a linear accelerator at a dose rate of 2.4 Gy/min, to deliver doses of 0.5, 1, 2 and S Gy. The control samples, and Irradiated cells, were maintained in culture medium for 24, 48 and 72 hours fellowing the Irradiation. The number of apoptotic cells after the in vitro X-irradiation was measured by flow cytometry after Incubation periods of 24, 48 and 72 hours. We also observed the apoptotic cells using a DNA fragmentation assay and electron microscopy. Results : The rate oi spontaneous apoptosis increased in relation to the time interval following irradiation (1.761 ${\pm}$0.161, 3.563${\pm}$0.554, 11.098${\pm}$2.849, at 24, 48, and 72 hours). The apoptotli cells also increased In the samples irradiated with 0.5, 1, 2 and 5 Gy, In a radiation dose and time interval after Irradiation manner, with the apoptosls being too great at 72 hours after Irradiation. The dose-response curves were characterized by an Initial steep Increase In the number of apoptotic cells for Irradiation doses below 2 Gy, with a flattening of the curves as the dose approached towards 5 Gy. Conclusion :The flow cytometric assay technique yielded adequate data, and required less than 1 mL of blood. The time and dose dependence of the radiation-induced apoptosis, was also shown. It is suggested that the adequate time Interval required for the evaluation of apoptosis would be 24 to 48 hours after blood sampling.
This study was conducted for screening on antioxidant activity of 429 plants and selecting new potential antioxidant candidates. In vitro test models such as scavenging activity on DPPH radical and inhibitory activity on linoleic acid oxidation were used in the preliminary study. Flower of Sanguisorba officinalis, flower of Sedum kamtschaticum, flower of Rumex obtusifolius, and root of Sedum kamtschaticum showed very effective antioxidant activity on DPPH radical and linoleic acid oxidation. Those plants showed 8.1, 9.4, 9.9, $11{\mu}g/ml$ in DPPH radical scavenging activity as $SC_{50}$ and did 80.4, 80.1, 84.5, 88.0% in inhibition activity on linoleic acid oxidation, respectively. Root of Sedum middendorfianum M. showed positive effects in superoxide radical scavenging activity ($38.4{\mu}g/ml$) and inhibitory effect on $CuSO_4$-induced LDL oxidation (53.8% at final concentration of $1{\mu}g/ml$). Gleditsia japonica Mig. showed high antioxidant activity on LDL oxidation as 71.6% at final concentration of $1{\mu}g/ml$ and total phenol content of 958.5 mg% as tannic acid equivalent. In conclusion, we think that these plants having potent antioxidant activity might be studied further and could be used as new resources for many purposes including healthy food, functional cosmetics and drug development etc.
Recently, the importance of toxoplasmosis is raised as a complication in immunosuppressed or AIDS patients. Our study focused on the identification of a variety of Toxoplasma antigens by immunoblotting. Rabbits and BALB/c mice were immunized with Toxoplosmo Iysate (RH strain) , frozen tachyzoites (RH strain) or cysts (Beverly and Fukaya strain) . Blood were collected from ear vein, heart or orbital plexus for detecting the serum antibody levels. For excretory-secretory (E.S) antigens, T gondii (RH) tachyzoite were cultured in CHL (Chinese hamster lung) cells with MEM containing of 5% FCS. After 72hrs, culture supernatant was collected. BALB/c mice were inoculated with RH tachyzoite intraperitoneally and peritoneal fluids were extracted three days later. E.S antigens were detected in culture supernatant and infected mouse peritoneal fluid by EITB. Serum IgG levels in rabbit were 1 :512 of 10 days after primary immunization, 1 : 2,048 of 10 days after secondary immunization, 1: 1,024 of 20 days after secondary immunization by IFAT, respectively. Serum IgG levels of immunized mice were 1:128 after 7 weeks. Tachyzoite antigens of the RH strain were detected 25 protein bands ranging 10 kDa-220 kDa of molecular weights with Coomassie blue stain. Toxoplcsma major antigens corresponding to n of 24 kDa, 27 kDa,30 kDa, 35 kDa, 38 kDa were recognized by IgG and IgM antibodies. Excretory-secretory antigens present in culture supernatant with M. W. of 20, 30 kDa and in infected mouse peritoneal fluid with M.W. of 33 (P30), 45 kDa. When RH tachyzoite antigen was probed with different mice sera immunized with 2 strains of T gondii, the IgG antibody bud of Fukaya and Beverly strain (8 week-serum) is identical to those of RH strain. It is considered that the 30 kDa polypeptide detected in excretory- secretory materials and Iysate was important major antigen of T gondii (RH).
Hwang-Bo, Jeon;Jang, Kyung Ok;Chung, Hayoung;Park, Jong-Hwa;Lee, Tae Hoon;Kim, Jiyoung;Chung, In Sik
The Korean Journal of Food And Nutrition
/
v.29
no.6
/
pp.998-1007
/
2016
The objective of this study was to investigate the effects of a lettuce (Lactuca sativa L.) extract on the inflammation of human umbilical vein endothelial cell (HUVEC) and blood lipid improvement in hypercholesterolemic mice fed a high cholesterol diet. The lettuce extract (100% ethanol extract) inhibited the expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in HUVEC treated with tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$). The lettuce extract suppressed the adhesion of THP-1 to TNF-${\alpha}$-treated HUVEC. The lettuce extract decreased the TNF-${\alpha}$-stimulated production of proinflammatory cytokine interleukin-6, interleukin-8 and chemokine monocyte chemotactic protein 1. In hypercholesterolemic mice, the lettuce extract reduced serum total cholesterol, triglyceride, and low-density lipoprotein-cholesterol level, while the lettuce extract elevated high-density lipoprotein-cholesterol level, resulting in the decrease of atherogenic index and cardiac risk factor level. These results suggested that lettuce extract can be an useful resource to show an anti-inflammatory effect and improve lipid metabolism.
Purpose: The aim of this study was to investigate distribution of particle size in phytate kit and compare filtered method with non-filtered method using 200 nm filter for sentinel lymphoscintigraphy (SLS). Materials and Methods: Five phytate kit of having the same available period was measured by particle size analyzer. For in-vivo experiment, $^{99m}Tc$-phytate was injected intradermally at both foot to perform lymphoscintigraphy. Imaging was acquired at 1hour after injection. Region of interest (ROI) was drawn in inguinal and background area for analysis. RAW 264.7 cells (Murine macrophage cell) were prepared for measurement of celluar uptake as a representative of macrophages. Paired t-test was performed using SPSS (SPSS Inc, USA) for statistical analysis. Results: The size of most particle in Techne phytate kit was distributed in 130~650 nm(90.5 %). In-vivo study, the ROI analysis showed similar result between filtered and non-filtered sample, and the numerical value of count/pixel were $58.3{\pm}5.97$ and $60.2{\pm}4.88$. In-vitro study, cellular uptake study also showed no difference between filtered and non-filtered sample by gamma counting. Conclusion: The present study demonstrates that there was no meaning of 200 nm filtered method for SLS using $^{99m}Tc$-phytate.
In this study diverse dehydrogenative polymers (DHPs) were synthesized with three precursors of native lignin [p-coumaryl alcohol (PCA), coniferyl alcohol (CA), sinapyl alcohol (SA)] in the presence of horseradish peroxidase (HRP, EC. 1.11.1.7)/$H_2O_2$. To compare the structural features between DHPs and native lignin, the DHPs as well as pine/poplar milled wood lignins were simultaneously subjected to gel permeation chromatography (GPC) to determine average molecular weights and derivatization followed by reductive cleavage (DFRC) to investigate the frequency of ${\beta}$-O-4 linkage. The highest yield of DHP was measured to 71% when CA was solely injected (G-DHP) and the yield of H-DHP was 42%. However, single injection of SA could not form any polymer in this system. The average molecular weights of DHPs were ranged between 3,000~4,700, which were only 1/2 fold compared with that of pine MWL (G-type lignin: Mw 7,340) and 1/3 scale compared with that of poplar MWL (GS-type lignin: Mw 13,250). DFRC analysis revealed that the formation of ${\beta}$-O-4 linkage during dehydrogenative polymerization was the highest in the GS-DHP with ca. 502 ${\mu}mol$/g, which was, however, remained to only 50% compared to that in poplar MWL (1107 ${\mu}mol$/g). The ${\beta}$-O-4 linkage was estimated to ca. 286 ${\mu}mol$/g In the G-DHP, which was twice as much as that of H-DHP(127 ${\mu}mol$/g). Similar to GS-DHP, only half amount of ${\beta}$-O-4 linkage, compared to pine MWL, was formed during in vitro polymerization of CA by horseradish peroxidase/$H_2O_2$.
Tak, Byung-Yun;Chung, Myung-Sook;Joo, Kyoung-Hwan;Rim, Han-Jong
Journal of agricultural medicine and community health
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v.16
no.2
/
pp.134-140
/
1991
The amino acid constituents of Paragonimus westermani were very imperfectively known. Lee(1964) detected 9 amino acids in the tissue hydrolysates of P. westermani, and 13 amino acids were detected from the cyst content and body fluid constituents of P. ohirai, But, the quantity of amino acids in P. westermani is still unknown. In the present investigation 18 amino acids, the fundamental constituents of proteins, were quantitatively studied by high performance liquid chromatography. The results obtained were as follows : A total of 18 amino acids were recovered in protein hydrolysates of P. westermani obtained from cat. ; They were cystein, aspartic acid, glutamic acid, serine. glycine, histidine. arginine, threonine, alanine, proline, tyrosine, valine, methionine, isoleucine, leucine, phenylalanine, tryptophan and lysin. Among them, glutamic acid was the most abundant form and tryptophan, cystein, methionine, and histidine constitute minor portion of hydrolysates. Compared to the normal P. westermani, the volume of hydrolysates obtained from the praziquantel(PZQ) treated worm-0.lug PZQ/ml saline for 6 hours incubation, and $3{\times}25$mg/kg bwt${\times}$2days in vivo treatment was generally increased except tryptophan. A total of 17 free amino acids were identified and the volume was 174.18 umol/1 gm wet weight P. westermaini. Among them, glycine and alanine constitute 28% of total volume. No significant differences were observed in the material obtained from worm treated with PZQ. However, slight increase of serine, arginine and the slight diminution of glutamic acid and proline was observed.
Purpose : The purpose of this study was a phantom study to measure the diffusion properties of water molecules by steady-state free precession diffusion-weighted imaging (SSFP- DWI) with a low b-value and to determine if this sequence might be useful for application to the evaluation of bone marrow pathology. Materials and methods : 1. The phantom study: A phantom study using two diffusion weighted sequences for the evaluation of the diffusion coefficient was performed. Three water-containing cylinders at different temperatures were designed: phantom A was $3^{\circ}C$, B was $23^{\circ}C$ and C was $63^{\circ}C$. Both SSFP and echo planar imaging (EPI) sequences (b-value: $1000s/mm^2$) were performed for comparison of the diffusion properties. The Signal to noise ratios (SNR) and apparent diffusion coefficient (ADC) values of the three phantoms using each diffusion-weighted sequence were assessed. 2. The Clinical study: SSFP-DWI was performed in 28 patients [sacral insufficiency fractures (10), osteoporotic lumbar compression fractures (10), malignant compression fractures (8)]. To measure the ADC maps, a diffusion-weighted single shot stimulated echo-acquisition mode sequence ($650s/mm^2$) was obtained using the same 1.5-T MR imager Results : For the phantom study, the signal intensity on the SSFP as well as the classic EPI-based DWI was decreased as the temperature increased in phantom A to C. The ADC values of the phantoms on EPI-DWI were $0.13{\times}10^{-3}mm^2/s$ in phantom A, $0.22{\times}10^{-3}mm^2/s$ in B and $0.37{\times}10^{-3}mm^2/s$. in C. The SSFP can be regarded as a DWI sequence in view of the series of signal decreases. Conclusion : Bone marrow pathologies with different diffusion coefficients were evaluated by SSFP-DWI. All benign fractures were hypointense compared to the adjacent normal bone marrow where as the malignant fractures were hyperintense compared to the adjacent normal bone marrow.
We investigated the optimal concentration and exposure time of cycloheximide(CHX) on development of activated porcine oocytes following electrical pulse(EP). After 42~44 h maturation, oocytes were treated with 0.1% hyaluronidase, and denuded cumulus cells by pipetting. Oocytes were stimulated by electric pulse (1.2 kV/cm, 30 $\mu$sec, 1 pulse) or incubated for 3, 5 and 7 h in cycloheximide (1, 5 and 10 $\mu\textrm{g}$/$m\ell$, respectively) following electric pulse, and cultured for 8 days. Cleavage rate of oocytes activated with 10 $\mu\textrm{g}$/$m\ell$ CHX following EP was significantly (P<0.05) higher than those of 1 $\mu\textrm{g}$/$m\ell$ (86.8% vs. 74.4%). The developmental competence of oocytes incubated to 5 $\mu\textrm{g}$/$m\ell$ of CHX was significantly (P<0.05) higher development to blastocysts (13.3%), compared with 10 $\mu\textrm{g}$/$m\ell$ of CHX (5.6%). When the oocytes were activated with 5$\mu\textrm{g}$/$m\ell$ CHX for 3, 5, and 7 h following EP, the cleavage rate of oocytes in 5 h group(86.6%) was significantly (P<0.05) higher than that in 3 h group(73.2%). The developmental rate of oocytes to morula in 5 and 7 h groups(26.7% and 16.4%) were significantly (P<0.05) high than that in 3 h group(14.5%). Matured oocytes were activated with electric pulse (EP) or electric pulse combined with cycloheximide (EP + CHX) and cultured for 8 days. The rate of cleavage and development to blastocyst (80.1% and 11.6%) of activated with EP group were similar to EP combined with CHX group. When activated with EP or EP combined with CHX, the mean cell number of blastocysts were less in the activated with EP (18.67$\pm$5.53) than in the activated EP combined CHX (20.71$\pm$6.16), but not significantly different. This results suggest that, when the porcine oocytes were activated with CHX following EP, the developmental rate of activated oocytes can be improved by treated with a concentration of 5 $\mu\textrm{g}$/$m\ell$ CHX for 5 h exposure time.
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