• Journal of Life Science
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• v.9 no.5
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• pp.556-563
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• 1999

Presence of antibody to the capsid protein p24 is the main diagnostic criterion, since this reflects reliable antibody response to HIV infection. However, it takes about 6-8 weeks for antibody production after infection and people who are infected but antibodies are not produced yet are classified as seronegative. Therefore, there is a strong need for an improved diagnostic method for better health security. As a first step for developing such an improved diagnostic system, gag protein of human immunodificiency virus type 1 was expressed in E. coli DH5$\alpha$. The gag fragment of HIV-1 (including a portion of p17 and whole p24) was amplified by polymerase chain reaction (PCR) and BamH I/EcoR I sites were created during PCR. The amplified DNA fragment was cleaved with BamH I/EcoR I and was subcloned into the GEX-2T vector which had been digested with BamHI/EcoRI, resulting gene fusion with gst gene of pGEX-2T. The recombinant DNA was transferred into E. coli DH5$\alpha$. The transformed bacteria were grown at 37$^{\circ}C$ for 3h and protein expression was induced with 0.1mM IPTG at $25^{\circ}C$ for 3h. Recombinant gag protein or GST-gag fusion protein was purified with glutathione-sepharose 4B bead and migrated as a single band when analyzed by 10% polyacrylamide gel. These proteins were confirmed by immunoblotting with anti-GST goat sera or Korean AIDS patients sera. The results of this study establish the expression and single step pulification of HIV-1 gag protein which can specifically bind with Korean AIDS patients sera.

• Clinical and Experimental Reproductive Medicine
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• v.32 no.1
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• pp.47-54
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• 2005

Objective: Evaluation of embryos using early cleavage to 2-cell stage has been proposed, but a critical time-point for selecting embryos is unclear. The aim of the present study is to provide a guideline including critical time-point in the selection of early cleaving embryo for the reduction of multiple pregnancies as well as the increase of pregnancy rate in human IVF. Methods: This prospective study was performed in 116 cycles from 85 patients who underwent conventional IVF or ICSI at the infertility clinic of Good Moonhwa Hospital from January 2002 to December 2003. Early cleavage (EC) of embryos to 2-cell stage was assessed at 25 h and 27 h postinsemination/microinjection. Embryos that had early cleaved at each time point were designated as EC-1 and EC-2, respectively, while others were designated as non-early cleavage (NEC). Results: At least one early cleavage embryo was observed in 54 (46.6%) for the EC-1 and 84 (72.4%) for the EC-2 of the 116 cycles assessed. Clinical pregnancy rates (PR) were significantly higher in the EC-1 group (66.7%) compared to the EC-2 group (53.6%) or the NEC group (31.2%) (p<0.05). Significant improvement of the pregnancy rate was found when at least two or more embryos were early cleaved at 25 h postinsemination or when the proportion of early cleavage embryo at 25 h postinsemination was higher than 20% (p<0.05). Conclusion: The critical time-point for the selection of early cleavage embryos with high implantation potential is more effective in 25 h postinsemination/microinjection compared to 27 h. The proportion as well as number of early cleavage embryos is also an important factor for the prediction of pregnancy outcome and the chance of multiple pregnancies. These results demonstrated that the evaluation of early cleavage embryos to 2-cell stage is an easy, simple, and objective method for the selection of good quality embryos suitable for embryo transfer.

• Pediatric Infection and Vaccine
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• v.17 no.1
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• pp.49-55
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• 2010

Purpose : The Purposes of this study are to identify the circulating etiologic viruses of acute lower respiratory tract infection in children and to understand the relation with clinical diagnosis. Methods : We obtained a total of 418 nasopharyngeal aspirates from children admitted for their acute lower respiratory tract infections at three tertiary hospitals in Seoul from September 2008 to March 2009. We performed multiplex RT-PCR to identify 14 etiologic viruses and analyzed their emerging patterns and clinical features. Results : Average age of patients was 16.4 months old and the ratio of male to female was 1.36. Viruses were detected in 56.2% of a total of 418 samples. Respiratory syncytial virus (35%) was the most frequently detected and followed by human rhinovirus (22%), human bocavirus, adenovirus, human metapneumovirus, parainfluenza virus, influenza virus and human coronavirus. Co-infection reached 21.9 % of positive patients. Conclusion : When we manage the patients with acute lower respiratory infectious diseases, we should remind the role of various viral pathogens, which might be circulating by seasons and by local areas.

• Biomedical Science Letters
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• v.29 no.4
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• pp.211-219
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• 2023

Caffeic acid phenethyl ester (CAPE) is an active component of propolis obtained from honeybee hives. CAPE possesses anti-mitogenic, anti-carcinogenic, anti-inflammatory, and immunomodulatory activities in diverse systems, which know as displays antioxidant activity and inhibits lipoxygenase activities, protein tyrosine kinase, and nuclear factor kappa B (NF-κB) activation. This study aimed to investigate the effect of CAPE on lipopolysaccharide (LPS)-induced human neutrophil phagocytosis. Human neutrophils were cultured with various concentrations of CAPE (1, 10, and 100 µM) with or without LPS. The pro-inflammatory proteins (tumor necrosis factor-alpha [TNF-α], interleukin [IL]-6 and IL-8) levels were measured after 4 h incubation. To investigate the intracellular signaling pathway, we measured the levels of mitogen-activated protein kinases (MAPK), including phosphorylation of p38, extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) and c-Jun N-terminal kinase (JNK). Next, to evaluate the potential phagocytosis, neutrophils were labeled with iron particles of superparamagnetic iron oxide nanoparticles (SPIONs, 40 nm) for 1 h in culture medium containing 5 mg/mL of iron. The labeling efficiency was determined by Prussian blue staining for intracellular iron and 3T-wighted magnetic resonance imaging. CAPE decreased the activation of intracellular signaling pathways, including ERK1/2 and c-Jun, and expression of pro-inflammatory cytokines, including TNF-α and IL-6, but had no effect on the signaling pathways of p38 and cytokine IL-8. Furthermore, images obtained after mannan-coated SPION treatment suggested that CAPE induced significantly higher signal intensities than the control or LPS group. Together, these results suggest that CAPE regulates LPS-mediated activation of human neutrophils to reduce phagocytosis.

• Clinical and Experimental Reproductive Medicine
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• v.22 no.3
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• pp.279-285
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• 1995

Despite the frequent incidence of embryo fragmentation in early human embryos, the reason of the embryo fragmentation has not been known yet. This study was conducted to investigate the histological difference(s) between fragmented (FR) and non-fragmented (NFR) human embryos focusing on comparison of mitochondrial distribution and protein synthesis. Multi-pronuclei zygotes (MPZ) such as three or more pronuclei containing in human in vitro fertilization and embryo transfer (IVF-ET) program were used for this study. MPZ were cultured in TCM-199 supplemented with 10% of human fetal cord serum (hFCS) in 5% $CO_2$ incubator at $37^{\circ}C$ for 24 hours. The cleaved embryos to 2-4 cells after 24 hours were grouped by their grade of fragmentation. Embryos were stained with Rhodamine123 (Rh123) and fluorescence was evaluated under the fluorescence microscope through PB 450-490 filter (Leitz). Regarding to protein synthesis during early human embryogenesis, there is no significant difference in the amount of synthetic proteins between FR and NFR embryos. Distribution of cytoplasmic organelles in embryos was evaluated by transmission electron microscope (TEM). The cytoplasmic distribution of mitochondria was different between FR and NFR embryos. The mitochondrial distribution was even in NFR, whereas severely aggregated in FR. It is not able to clarify in the present study whether this uneven mitochondrial distribution in FR embryo is the reason for embryo fragmentation or is the result from fragmentation. Physiological disparity related to the mitochondrial distribution may be one of the reasons for embryo fragmentation. Further studies should be addressed to investigate the physiological differences between FR and NFR embryos.

• Korean Journal of Animal Reproduction
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• v.18 no.2
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• pp.87-94
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• 1994

The objective of this study is to evaluate the developmental ability of mouse embryo in the presence of human amniotic fluid (hAF), The highest development rate was found in the culture media supplemented with 20% mid-term hAF but this rate was concomitantly reduced with more than 20% hAF. Furthermore, mouse two-cell embryos cultured in 20% mid-term hAF were developed more consistently to the expanded and hatched blastocyst stages compared to those cultured in simple medium. However, no significant differences in the embryo development rates were observed among the supplemented effects of 20% mid-term hAF, 0.3% bovine serum albumin (BSA), and 10% fetal calf serum (FCS), Development rates of two-ceiI mouse embryos cultured in 20% full-term hAF were declined compared to 20% mid-term hAF. Outgrowth of hatched blastocysts were observed when the embryos were cultured in medium containing 20% mid-term hAF or 10% FCS. But two-cell mouse embryos cultured in the presence of 20% full-term hAF or O.3% BSA was not observed their outgrowth. The kinetics of outgrowth processes in the presence of hAF were similar to those with 10% FCS. However, embryos with FCS showed a considerably greater extents of trophetodermal cell proliferation and outgrowth. Taken together, these data suggest that mid-term hAF may have a suitability for the mammalian embryos and induce embryonic outgrowth.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.16
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• pp.6627-6632
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• 2014

Purpose: The present study concerns molecular mechanisms involved in induction of apoptosis by a fungal taxol extracted from the fungus Cladosporium oxysporum in T47D human breast cancer cells. Materials and Methods: Apoptosis-induced by the fungal taxol was assessed by MTT assay, nuclear staining, DNA fragmentation, flow cytometry and pro- as well as anti-apoptotic protein expression by Western blotting. Results: Our results showed inhibition of T47D cell proliferation with an $IC_{50}$ value of $2.5{\mu}M/ml$ after 24 h incubation. It was suggested that the extract may exert its anti-proliferative effect on human breast cancer cell line by suppressing growth, arresting through the cell cycle, increase in DNA fragmentation as well as down-regulation of the expression of NF-${\kappa}B$, Bcl-2 and Bcl-XL and up-regulation of pro-apoptotic proteins like Bax, cyt-C and caspase-3. Conclusions: We propose that the fungal taxol contributes to growth inhibition in the human breast cancer cell through apoptosis induction via a mitochondrial mediated pathway, with possible potential as an anticancer therapeutic agent.

• Biomedical Science Letters
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• v.16 no.4
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• pp.341-347
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• 2010

In the present study, we investigated whether Houttuynia cordata Thumb (Saururaceae; HC) extracts have an anti-inflammatory effect in human monocytic THP-1 cells and human eosinophilic EoL-1 cells. The dried and powdered whole plants of HC were extracted with 80% EtOH. The combined extract (HC-1) was concentrated under reduced pressure. The residue was diluted with water, and then successively partitioned with n-hexane, EtOAc, and BuOH to produce the n-hexane (HC-2), EtOAc (HC-3), BuOH (HC-4), and the water-soluble fractions (HC-5), respectively. HC extracts have no cytotoxicity on THP-1 cells and EoL-1 cells at a high concentration of $10\;{\mu}g/ml$ for 24 h, except HC-2 extract ($10\;{\mu}g/ml$). Interleukin-6, Interleukin-8 and Monocyte chemoattractant protein-1 in THP-1 cells were increased after the treatment with the extract from house dust mite or LPS. The increase of cytokine production was strongly suppressed by HC-3 extract, in comparision with other extracts. HC-3 also had inhibitory effect on Interleukin-6 production increased by mite extract and LPS in EoL-1 cells. However, HC-3 extract increased Interleukin-8 production induced by mite extract and LPS in EoL-1 cells. These results suggest that HC extracts may be used as useful agents for treating allergic disorders such as asthma and atopic dermatitis.

• Reproductive and Developmental Biology
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• v.35 no.2
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• pp.185-190
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• 2011

To evaluate the effect of spermatozoa culture on glycosidase activity of frozen-thawed spermatozoa in human, the spermatozoa were treated experimentally and assayed for activities of ${\alpha}$-L-fucosidase, ${\alpha}$-D-mannosidase, ${\beta}$-D-galactosidase and N-acetyl-${\beta}$-D-glucosaminidase (${\beta}$-GlcNAc'ase). The ${\beta}$-GlcNAc'ase activity was at least two-folds higher than other glycosidases regardless of spermatozoa incubation (p<0.05). The spermatozoa motility was decreased with incubation periods, but no effects by different glycosidases on the changes of spermatozoa motility during the various periods of incubation. In all glycosidases, the spermatozoa-zona binding rates in spermatozoa without incubation were higher than in spermatozoa incubated for 2 h (p<0.05). ${\beta}$-GlcNAc'ase is present mainly in the plasma membrane of spermatozoa frozen-thawed in human. It was also shown that the glycosidase activity was increased in all glycosidases in spite of lower sperm-zona binding by spermatozoa incubation.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.7
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• pp.4095-4099
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• 2013

Oxidative stress induces apoptosis in many cellular systems including glioblastoma cells, with caspase-8 activation was regarded as a major contribution to $H_2O_2$-induced cell death. This study focused on the role of the autophagic protein p62 in $H_2O_2$-induced apoptosis in U87MG cells. Oxidative stress was applied with $H_2O_2$, and cell apoptosis and viability were measured with use of caspase inhibitors or autophagic mediators or siRNA p62, GFP-p62 and GFP-p62-UBA (del) transfection. We found that $H_2O_2$-induced U87MG cell death was correlated with caspase-8. To understand the role of p62 in MG132-induced cell death, the levels of p62/SQSTM1 or autophagy in U87MG cells were modulated with biochemical or genetic methods. The results showed that the over-expression of wild type p62/SQSTM1 significantly reduced $H_2O_2$ induced cell death, but knockdown of p62 aggravated the process. In addition, inhibition of autophagy promoted p62 and active caspase-8 increasing $H_2O_2$-induced apoptosis while induction of autophagy manifested the opposite effect. We further demonstrated that the function of p62/SQSTM1 required its C-terminus UBA domain to attenuate $H_2O_2$ cytotoxity by inhibition of caspase-8 activity. Our results indicated that p62/SQSTM1 was a potential contributor to mediate caspase-8 activation by autophagy in oxidative stress process.