• Asian Pacific Journal of Cancer Prevention
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• 제14권10호
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• pp.5645-5650
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• 2013

Src homology 2 domain containing (SHC) is a proto-oncogene which mediates cell proliferation and carcinogenesis in human carcinomas. Here, the SHC SH2-domain binding protein 1 (SHCBP1) was first established to be up-regulated in human hepatocellular carcinoma (HCC) tissues by array-base comparative genome hybridization (aCGH). Meanwhile, we examine and verify it by quantitative real-time PCR and western blot. Our current data show that SHCBP1 was up-regulated in HCC tissues. Overexpression of SHCBP1 could significantly promote HCC cell proliferation, survival and colony formation in HCC cell lines. Furthermore, knockdown of SHCBP1 induced cell cycle delay and suppressed cell proliferation. Furthermore, SHCBP1 could regulate the expression of activate extracellular signal-regulated kinase 1/2 (ERK1/2) and cyclin D1. Together, our findings indicate that SHCBP1 may contribute to human hepatocellular carcinoma by promoting cell proliferation and may serve as a molecular target of cancer therapy.

• Asian Pacific Journal of Cancer Prevention
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• 제13권3호
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• pp.761-766
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• 2012

Objective: Although various human cancer stem cells (CSCs) have been defined, their applications are restricted to immunocompromised models. Developing a novel CSC model which could be used in immunocompetent or transgenic mice is essential for further understanding of the biomolecular characteristics of tumor stem cells. Therefore, in this study, we analyzed murine lung cancer cells for the presence of CSCs. Methods: Side population (SP) cells were isolated by fluorescence activated cell sorting, followed by serum-free medium (SFM) culture, using Lewis lung carcinoma cell (LLC) line. The self-renewal, differentiated progeny, chemosensitivity, and tumorigenic properties in SP and non-SP cells were investigated through in vitro culture and in vivo serial transplantation. Differential expression profiles of stem cell markers were examined by RT-PCR. Results: The SP cell fraction comprised 1.1% of the total LLC population. SP cells were available to grow in SFM, and had significantly enhanced capacity for cell proliferation and colony formation. They were also more resistant to cisplatin in comparison to non-SP cells, and displayed increased tumorigenic ability. Moreover, SP cells showed higher mRNA expression of Oct-4, ABCG2, and CD44. Conclusion: We identified SP cells from a murine lung carcinoma, which possess well-known characteristics of CSCs. Our study established a useful model that should allow investigation of the biological features and pharmacosensitivity of lung CSCs, both in vitro and in syngeneic immunocompetent or transgenic/knockout mice.

• 대한한의학회지
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• 제22권3호
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• pp.156-168
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• 2001

Objectives : To evaluate the diverse actions of stimulation on the hematopoietic system, 4 formulas (KH I, KH 2, KH 3, KH 4) were studied. Method and Result : RT-PCR was performed to measure the gene expression of hematopoietic cytokines (TPO, GM-CSF, SCF, IL-3). When bone marrow cells were treated with KH 1, 2, 3, 4, the gene expressions of TPO, SCF, IL-3, and GM-CSF were increased. Flow cytometric analysis was performed to measure the expression of CD34+ cell activity. After 72 hrs culture supplemented with KH 1, 2, 3, 4, the percent of CD34+ cell of KH 2, 3, 4 were increased. To measure the expression of colony forming units - granulocyte erythrocytes, macrophages, megakaryocytes (CFU-GEMM) and burst forming unit-erythroid (BFU-E), semisolid clonogenic assay was performed. After 14 days of culture the number of CFU-GEMM and BFU-E of KH I, 2, 3, 4 were significantly increased compared to those of EPO groups (KH 1 P<0.0l, KH 2 P<0.05, KH 3 P<0.001, KH 4 P<0.0l). To determine the intracelluar TPO expression by KH 3, KH 4 in bone marrow cells, intracelluar staining and flow cytometric analysis were performed. After 24 hrs cultures, the TPO expression of the KH 3 and KH 4 treated groups were increased over those of the controlled groups (control : 50%, KH 3 : 87%, KH 4 : 78%). Conclusion : These results suggest that KH I, KH 2, KH 3, KH 4 have hematopoietic effects through increasing the production of hematopoietic cytokines and stimulating the activity of $CD34^{+}$ cells. This study also shows that KH 3 has a more effective hematopoietic effect than KH 1, 2, 4. These results suggest that the formulas (KH I, 2, 3, 4) can be applied to the patients with inappropriate hematopoietic system, and that KH 3 can be the most effective formula among these 4 in treating bone marrow disease in clinics.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5017-5021
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• 2013

Objective: MicroRNAs (miRNAs) are a small class of non-coding, single-stranded RNAs with a critical role in genesis and maintenance of renal cancer mainly through binding to 3'-untranslated regions (3'UTR) of target mRNAs, which causes a block of translation and/or mRNA degradation. The aim of the present study was to investigate the potential effects of miR-122 in human renal cell carcinomas. Methods: The expression level of miR-122 was quantified by qRT-PCR. MTT, colony formation, invasion and migration assays were used to explore the potential functions of miR-122 in human renal cell carcinoma cells. Results: Cellular growth, invasion and migration in two A498 and 786-O cells were significantly increased after miR-122 transfection. Further experiments demonstrated that overexpression of miR-122 resulted in the increase of phospho-Akt (Ser473) and phospho-mTOR (Ser2448), then activation of mTOR targets, p70S6K and 4E-BP1. Conclusions: The up-regulation of miR-122 may play an important role in the progress of renal cancer through activating PI3K/Akt signal pathway and could be a potential molecular target for anti-cancer therapeutics.

• 미생물학회지
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• 제38권4호
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• pp.230-230
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• 2002

Class Ⅲ chitin synthases in filamentous fungi are important for hyphal growth and differentiation of several filamentous fungi. A genomic clone containing the full gene encoding Chs4, a class Ⅲ chitin synthase in Penicillium chrysogenum, was cloned by PCR screening and colony hybridization from the genomic library. Nucleotide sequence analysis and transcript mapping of chs4 revealed an open reading frame (ORF) that consisted of 5 exons and 4 introns and encoded a putative protein of 915 amino acids. Nucleotide sequence analysis of the 5′flanking region of the ORF revealed a potential TATA box and several binding sites for transcription activators. The putative transcription initiation site at -716 position was identified by primer extension and the expression of the chs4 during the vegetative growth was confirmed by Northern blot analysis. Amino acid sequence analysis of the Chs4 revealed at least 5 transmembrane helices and several sites for past-transnational modifications. Comparison of the amino acid sequence of Chs4 with those of other fungi showed a close relationship between P chrysogenum and genus Aspergillus.

• Reproductive and Developmental Biology
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• 제30권4호
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• pp.287-291
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• 2006

We have developed a new passaging technique for the expansion of human embryonic stem cells (hESCs) that involves simply pipetting portions of hESCs acquired from colonies, reducing the laborious and time-consuming steps in the expansion of hESCs. Compared to general mechanical methods of passaging, our pipetting method allowed hESCs colonies to be broken into small fragments, which showed significantly higher attachment rates onto feeder cell layers. This technique produced three times the number of hESCs colonies than conventional mechanical methods. In addition, this pipetting method allowed us to distinguish differentiated hESCs from undifferentiated hESCs during hESCs colony pipetting. The hESCs cultured by pipetting method displayed normal human chromosomes for over 60 passages. According to RT-PCR and immunohistochemical analysis, the hESCs successfully maintained their undifferentiated state and pluripotency which was also confirmed by teratoma formation in viva Therefore, the pipetting method described in this study is a useful tool to efficiently and quickly expand hESCs on a large scale without enzyme treatment.

• 대한치과의사협회지
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• 제48권6호
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• pp.436-442
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• 2010

Objective: Transmission of S. mutans, a major dental caries pathogen, occurs mainly during the first 2.5 years of age. Children appear to acquire S. mutans mostly from their mothers, but few studies have investigated preventive effect of xylitol to S. mutans transmission from mother to child. The aim of this study was to perform a follow-up evaluation the preventive effect of xylitol chewing gum of the S. mutans of children's oral cavities, which included the characteristics of vertical transmission from mother to child. Methods: The mothers voluntarily participating in a women's oral health prevention program were divided into two groups (a control and a xylitol group). The subjects were 20 mother-child pairs, who were monitored for 30 months. Xylitol chewing gum group had consumed 2 gum pellets, 3 times a day for 24 months, and then they were followed until 30 months. At baseline, 24 and 30 months whole stimulated saliva samples were collected from the mothers. Children were also recruited from 6 months to 30 months after birth and were collected their dental plaque samples. After isolation and identification, the analysis of the colony count, transmission electron microscopy and real-time RT-PCR were performed to analyze the characteristics of S. mutans. Results: The S. mutans counts decreased steadily in the xylitol group at 24 months, but increased at 30 months. The similar results were showed at their children. While the glucan synthesis was decreased at xylitol group both mother and child. The expression of gtfB, gtfD and ftf were significantly reduced in the xylitol group both mother and child (p<0.05). Conclusions: These findings indicate that chewing xylitol gum over a long period may decrease the expression of the genes associated virulence and reduced the glucan synthesis of S. mutans, which can result the preventing the mother-to-child transmission of S. mutans.

• Journal of Microbiology and Biotechnology
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• 제24권11호
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• pp.1503-1509
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• 2014

With the purpose of facilitating the process of stable strain generation, a shuttle vector for integration of genes via a double recombination event into two ectopic sites on the Sulfolobus acidocaldarius chromosome was constructed. The novel chromosomal integration and expression vector pINEX contains a pyrE gene from S. solfataricus P2 ($pyrE_{sso}$) as an auxotrophic selection marker, a multiple cloning site with histidine tag, the internal sequences of malE and malG for homologous recombination, and the entire region of pGEM-T vector, except for the multiple cloning region, for propagation in E. coli. For stable expression of the target gene, an ${\alpha}$-glucosidase-producing strain of S. acidocaldarius was generated employing this vector. The malA gene (saci_1160) encoding an ${\alpha}$-glucosidase from S. acidocaldarius fused with the glutamate dehydrogenase ($gdhA_{saci}$) promoter and leader sequence was ligated to pINEX to generate pINEX_malA. Using the "pop-in" and "pop-out" method, the malA gene was inserted into the genome of MR31 and correct insertion was verified by colony PCR and sequencing. This strain was grown in YT medium without uracil and purified by His-tag affinity chromatography. The ${\alpha}$-glucosidase activity was confirmed by the hydrolysis of $pNP{\alpha}G$. The pINEX vector should be applicable in delineating gene functions in this organism.

• Journal of Microbiology and Biotechnology
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• 제27권2호
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• pp.226-233
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• 2017

Jeung-pyun, a fermented rice cake, is prepared by fermenting rice sourdough using makgeolli, a traditional Korean rice wine, in the presence of yeast and lactic acid bacteria (LAB). The goal of this study was to conduct biochemical and microbial analyses of five different rice sourdoughs, each fermented with a different commercial makgeolli, using culture-dependent and culture-independent approaches. All sourdough samples fermented with different makgeolli for 6.5 h showed different profiles in pH, total titratable acidity, organic acid concentration, and microbial growth. LAB belonging to different genera were identified based on colony morphology on modified MRS and sourdough bacteria agar medium. PCR-denaturing gradient gel electrophoresis analyses of the five sourdoughs showed different bands corresponding to LAB and yeast. 16S/26S rRNA gene sequence analyses of the samples confirmed that the predominant LAB in the five fermented rice doughs was Lactobacillus plantarum, Lb. pentosus, and Lb. brevis. Various other Lactobacillus spp. and Saccharomyces cerevisiae were common in all five fermented samples. This study provides comprehensive and comparative information on the microflora involved in fermentation of rice sourdough and signifies the need to develop effective starters to enrich the quality of jeung-pyun.

• Food Science and Biotechnology
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• 제16권1호
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• pp.99-103
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• 2007

The purpose of this study was to develop a simple detection method, possibly at the species-level, that allows for large-scale screening of bifidobacteria. Human fecal samples were plated on MRS-raffinose agar containing cysteine and neomycin sulfate, serving as selective pressure for bifidobacteria, and 0.003%(w/v) bromocresol green. All of the test strains grew well on this medium at $37{\pm}1^{\circ}C$, forming white colonies surrounded by yellow halos, which presented a sharp contrast against the green background. In this disc assay, the required incubation time to develop a yellowish zone varied with the species of Bifidobacterium that was tested, allowing for differential counts and easy identification at the species-level: 10-14 hr for B. bifidum, 20-22 hr for B. catenulatum and B. infantis. and 24-25 hr for B. longum and B. breve. No apparent color was observed for B. angulatum and B. adolescentis 28 hr after inoculation. To evaluate the results of pH indicator-based identification, individual isolates were subjected to a colony-PCR experiment with genus-specific primers. The amplified products from the isolates were in good accordance with those from the reference strains at a level of 95% agreement. These results suggest that the present method could be conveniently applied to cell counts, as well as to the preliminary identification of bifidobacteria from a variety of sample types including human feces, dairy products, and commercial probiotic supplements.