• Tuberculosis and Respiratory Diseases
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• v.56 no.5
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• pp.450-464
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• 2004

Background : The FHIT (fragile histidine triad) gene is a frequent target of deletions associated with abnormal RNA and protein expression in lung cancer. Previous studies have shown FHIT gene transfer into lung cancer cell line lacking FHIT protein expression resulted in inhibition of tumor cell growth attributable to the induction of apoptosis and reversion of tumorigenecity. However, the mechanism of the tumor suppressor activity of the FHIT gene and the cellular pathways associated with its function are not completely understood. Methods : To gain insight into the biological function of FHIT, we compared the NCI-H358 cell line with its stable FHIT transfectants after treatment with cisplatin or paclitaxel. We investigated the effects of FHIT gene expression on cell proliferation, apoptosis, and activation of caspase system and Bcl-2 family. The induction of apoptosis was evaluated by using DAPI staining and flow cytometry. Activation of caspases and Bcl-2 members was evaluated by Western blot analysis. Results : A significantly increased cell death was observed in FHIT transfectants after cisplatin or paclitaxel treatment and this was attributable to the induction of apoptosis. Remarkable changes in caspases and Bcl-2 family were observed in the transfected cells as compared with the control cells after treatment with paclitaxel. Activation of caspase-3 and caspase-7 was markedly increased in cells expressing FHIT. Expression level of Bcl-2 and Bcl-xL protein was significantly decreased and that of Bax and Bad protein was significantly increased in the transfected cells. Conclusion : FHIT gene delivery into lung cancer cells results in enhanced apoptosis induced by treatment with cisplatin or paclitaxel. The data suggest that apoptosis in FHIT-expressing cells could be related to activation of caspase pathway and Bcl-2 family.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.12
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• pp.5055-5061
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• 2014

In this study, an anti-oxidant and anti-tumor protein Latcripin-3 of Lentinula edodes C91-3 was expressed in Escherichia coli. for the first time. According to the cDNA library, the full-length gene of Latcripin-3 was cloned by the methods of 3'-full rapid amplification of cDNA Ends (RACE) and 5'-full RACE. The structural domain gene of Latcripin-3 was inserted into the pET32 a(+). The functional protein of Latcripin-3 was expressed in Rosetta-gami (DE3) E. coli, evaluated by Western blotting and mass spectrometry. DPPH testing showed that the protein Latcripin-3 can scavenge free radicals remarkably well. The activity of functional protein Latcripin-3 on A549 cells was studied with flow cytometry and the MTT method. The MTT assay results showed that there was a decreases in cell viability in a dose-dependent and time-dependent manner in protein Latcripin-3 treated groups. Flow cytometry demonstrated that Latcripin-3 can induce apoptosis and block S phase dramatically in human A549 lung cancer cells as compared to the control group. At the same time, the cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. This research offers new insights and advantages for identifying anti-oxidant and anti-tumor proteins.

• Reproductive and Developmental Biology
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• v.30 no.1
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• pp.27-34
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• 2006

This study was conducted to compare the reproduction ability of the wild type boar and recombinant human erythropoietin (hEPO) transgenic boar semen. Ejaculated boar semen was analyzed by flow cytometry, Elisa and IVF methods. In experiment 1, flow cytometric analysis showed that the live sperm ratio of transgenic boar sperm significantly lower (P<0.05) than that of wild type boar after incubation at 20, 22, 24 and 26 hr. In experiment 2, the presence and levels of various cytokines (IL-6, IL-10 and $TNF-{\alpha}$) to related animal reproduction in the seminal and blood plasma were examined using specific enzyme immunoassay. There was no significant difference between both groups. In experiment 3, the fertilizing capacity and developmental ability of both boar sperm were compared. The transgenic boar sperm had a significantly low capacity of penetration, sperm-zona binding, embryo development, and blastocyst formation compared to wild type sperm (P<0.05). These results suggest that transgenic boar sperm harboring hEPO gene has low sperm viability than wild type boar, and it is a reason to decrease of fertility and litter size.

• Journal of the korean academy of Pediatric Dentistry
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• v.31 no.2
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• pp.304-313
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• 2004

Xylitol, a five-carbon natural sugar alcohol, is widely used non-cariogenic sugar substitute. In present study, the effects of xylitol on the expression of mRNA for glucosyltransferase which synthesizes glucan from sucrose were detected by Fluorescent in situ hybridization (FISH) and flow cytometry. FITC fluorescences for mRNA of gtfB, gtfC and gtfD were decreased further with increasing concentration of xylitol from 1% to 10% when detected by FISH. Flow cytometric analysis also showed that the expression of gtfB, gtfC and gtfD was increased by the addition of sucrose and decreased by the addition of xylitol to BHI broth containing 1% sucrose. In conclusion, the expression of gtfB, gtfC and gtfD mRNA was decreased by the addition of xylitol.

• Journal of Microbiology and Biotechnology
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• v.29 no.3
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• pp.454-464
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• 2019

Salmonellosis is a highly contagious bacterial disease that threatens both human and poultry health. Tests that can detect Salmonella in the field are urgently required to facilitate disease control and for epidemiological investigations. Here, we combined loop-mediated isothermal amplification (LAMP) with a chromatographic lateral flow dipstick (LFD) to rapidly and accurately detect Salmonella. LAMP primers were designed to target the Salmonella invA gene. LAMP conditions were optimized by adjusting the ratio of inner to outer primers, $MgSO_4$ concentration, dNTP mix concentration, amplification temperature, and amplification time. We evaluated the specificity of our novel LAMP-LFD method using six Salmonella species and six related non-Salmonella strains. All six of the Salmonella strains, but none of the non-Salmonella strains, were amplified. LAMP-LFD was sensitive enough to detect concentrations of Salmonella enterica subsp. enterica serovar Pullorum genomic DNA as low as $89fg/{\mu}l$, which is 1,000 times more sensitive than conventional PCR. When artificially contaminated feed samples were analyzed, LAMP-LFD was also more sensitive than PCR. Finally, LAMP-LFD gave no false positives across 350 chicken anal swabs. Therefore, our novel LAMP-LFD assay was highly sensitive, specific, convenient, and fast, making it a valuable tool for the early diagnosis and monitoring of Salmonella infection in chickens.

• The Korea Journal of Herbology
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• v.26 no.2
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• pp.1-10
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• 2011

Objective ： Lonicera japonica contains anti complementary polysaccharides and polyphenolic compound. Among these polyphenolic substances, chlorogenic acid is the major active component of this plant. However, the immunological mechanisms for these activities, have not been elucidated, nor the active components. To clarify immunomodulatory effects of those we examined the relationship between the activity of CD8+ T cell-mediated lysis and the frequency of cytokine profiles in spleen, thymus (especially IFN-${\gamma}$, IL-4, GM-CSF etc.) expressing CD8+ T cells activated by IL-2. Methods : To study immunomodulatory effects ethyl acetate fraction from Lonicera japonica, chlorogenic acid on cytokine gene expression from spleen, thymus cells, RT-PCR was performed after quantitative normalization for each gene by a densitometry using ${\beta}$-actin gene expression. A modified standard $^{51}Cr$-release assay was used to measure cytotoxic activities of cytotoxic T cells. Spleen, thymus cells from NOD mice were stained with CD3, CD4, CD44, CD69 in staining buffer and analyzed by two color flow cytometry. Results : We showed that ethyl acetate fraction from Lonicera japonica in combination with IL-2 resulted in a significant enhancement of PCR products for IFN-${\gamma}$, IL-4, IL-10, GM-CSF, IL-6 and cytotoxtic CD8+ T cell proportion in spleen and thymus T cells in NOD mice. This suggests that IFN-${\gamma}$, IL-6 like IL-4 may be acting as a regulatory rather than proinflammatory cytokine. Conclusions : In conclusion, based on the results of the present study which showed that ethyl acetate fraction from Lonicera japonica and chlorogenic acid upregulating cytokine gene expression in spleen and thymus, we are tempted to speculate that some of the therapeutic efficacies such as anti-diabetic activity of Lonicera japonica are due to the immunomodulatory its ethylacetate fraction and chlorogenic acid.

• Parasites, Hosts and Diseases
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• v.56 no.2
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• pp.153-165
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• 2018

Development of an effective vaccine is critically needed for the prevention of malaria. One of the key antigens for malaria vaccines is the apical membrane antigen 1 (AMA-1) of the human malaria parasite Plasmodium falciparum, the surface protein for erythrocyte invasion of the parasite. The gene encoding AMA-1 has been sequenced from populations of P. falciparum worldwide, but the haplotype diversity of the gene in P. falciparum populations in the Greater Mekong Subregion (GMS), including Thailand, remains to be characterized. In the present study, the AMA-1 gene was PCR amplified and sequenced from the genomic DNA of 65 P. falciparum isolates from 5 endemic areas in Thailand. The nearly full-length 1,848 nucleotide sequence of AMA-1 was subjected to molecular analyses, including nucleotide sequence diversity, haplotype diversity and deduced amino acid sequence diversity and neutrality tests. Phylogenetic analysis and pair-wise population differentiation ($F_{st}$ indices) were performed to infer the population structure. The analyses identified 60 single nucleotide polymorphic loci, predominately located in domain I of AMA-1. A total of 31 unique AMA-1 haplotypes were identified, which included 11 novel ones. The phylogenetic tree of the AMA-1 haplotypes revealed multiple clades of AMA-1, each of which contained parasites of multiple geographical origins, consistent with the $F_{st}$ indices indicating genetic homogeneity or gene flow among geographically distinct populations of P. falciparum in Thailand's borders with Myanmar, Laos and Cambodia. In summary, the study revealed novel haplotypes and population structure needed for the further advancement of AMA-1-based malaria vaccines in the GMS.

• The Plant Pathology Journal
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• v.33 no.3
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• pp.296-306
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• 2017

Sugarcane mosaic virus (SCMV) is one of the most damaging viruses infecting sugarcane, maize and some other graminaceous species around the world. To investigate the genetic diversity of SCMV in Iran, the coat protein (CP) gene sequences of 23 SCMV isolates from different hosts were determined. The nucleotide sequence identity among Iranian isolates was more than 96%. They shared nucleotide identities of 75.5-99.9% with those of other SCMV isolates available in GenBank, the highest with the Egyptian isolate EGY7-1 (97.5-99.9%). The results of phylogenetic analysis suggested five divergent evolutionary lineages that did not completely reflect the geographical origin or host plant of the isolates. Population genetic analysis revealed greater between-group than within-group evolutionary divergence values, further supporting the results of the phylogenetic analysis. Our results indicated that natural selection might have contributed to the evolution of isolates belonging to the five identified SCMV groups, with infrequent genetic exchanges occurring between them. Phylogenetic analyses and the estimation of genetic distance indicated that Iranian isolates have low genetic diversity. No recombination was found in the CP cistron of Iranian isolates and the CP gene was under negative selection. These findings provide a comprehensive analysis of the population structure and driving forces for the evolution of SCMV with implications for global exchange of sugarcane germplasm. Gene flow, selection and somehow homologous recombination were found to be the important evolutionary factors shaping the genetic structure of SCMV populations.

• The Journal of Korean Obstetrics and Gynecology
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• v.19 no.2
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• pp.186-198
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• 2006

Purpose : The purpose of this study is to demonstrate the direct inhibitory effect of Hyulbuchukeotang on the proliferation of uterine leiomyoma cells through an experiment treating uterine leiomyoma cells cultivated by explantation with indicated concentrations of Hyulbuchukeotang and to research the gene expression related to cell cycle ill order to discover the connection with apoptosis and its mechanism by analyzing cell cycle. Methods : After primary culture of uterine leiomyoma cells, the cultivated uterine leiomyoma cells were treated with indicated concentrations of Hyulbuchukeotang for 24 hours. The inhibitory effect on the cell proliferation was determined by the cell count assay. The value of a cell count assay represent the percentage of cells in a phase of the cell cycle compared with total cells. In addition, a link between Hyulbuchukeotang and apoptosis was examined through flow cytometric analysis by FACS and DNA fragmentation analysis. Finally, the degree of gene expression related to cell cycle was evaluated by Western blot analysis. Results : The inhibitory effect of Hyulbuchukeotang increase of uterine leiomyoma cells treated with indicated concentrations of Hyulbuchkeotang increases. The result of gene expression related to G1 phase after treating with 100, 250, 500, 1,000 ${\mu}g/ml$ concentrations of Hyulbuchukeotang. on uterine leiomyoma cells is that the gene expression of p27 was increased but that of p53 an p21 remained unchanged and the gene of pRB, pro-caspase 3 was decreased. Conclusion Through the mentioned experiments, it is demonstrated that Hyulbuchkeotang is effective in inhibiting Proliferation of uterine leiomyoma cells by extending cell cycle G1. However it is not considered that the inhibitory effect results from the aptoposis.

• Journal of Society of Preventive Korean Medicine
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• v.22 no.1
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• pp.107-127
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• 2018

Objectives : The present study investigated the anti-obese and blood flow improvement activities of aqueous extracts of ginseng berry (GBe) on the mild diabetic obese mice as compared with metformin. Methods : After end of 56 days of continuous oral administrations of GBe 150, 100 and 50 mg/kg, or metformin 250 mg/kg, anti-obese and blood flow improvement effects - the changes of body weights, body and abdominal fat density by in live dual-energy x-ray absorptionmetry (DEXA), tail bleeding time, prothrombin time (PT), activated partial thromboplastin time (aPTT), serum total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL) and high density lipoprotein (HDL) levels, aorta and serum cyclic guanosine monophosphate (cGMP), nitric oxide (NO) and endothelin (ET)-1 levels, aorta phosphorylated PI3K (pPI3K), phosphorylated Akt (pAkt) and phosphorylated p38 MAPK (pp38 MAPK) levels were systemically analyzed. In addition, aorta vascular dilation and constriction related gene mRNA expressions - PI3K, Akt, eNOS, p38 MAPK and ET-1 were also analyzed by realtime RT-PCR. Results : The obesity and related blood flow impairment, induced by 84 days of continuous HFD supply, were significantly inhibited by 56 days of continuous oral treatment of GBe 150, 100 and 50mg/kg, dose-dependently, and they also dramatically normalized the changes of the aorta vascular dilation and constriction related gene mRNA expressions, also dose-dependently. Especially, GBe 150 mg/kg constantly showed favorable inhibitory activities against type II diabetes related obesity and vascular disorders through PI3K/Akt pathway and p38 MAPK mediated cGMP, NO and ET-1 expression modulatory activities, as comparable to those of metformin 250 mg/kg in HFD mice. Conclusion : By assessing the key parameters for anti-obese and blood flow improvement activities on the HFD-induced mild diabetic obese mice, the present work demonstrated that GBe 150, 100 and 50 mg/kg showed favorable anti-obese and blood flow improvement effects in HFD-induced type II diabetic mice, through PI3K/Akt pathway and p38 MAPK mediated cGMP, NO and ET-1 expression modulatory activities.