• Asian Pacific Journal of Cancer Prevention
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• 제15권14호
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• pp.5577-5582
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• 2014

Objective: To observe the effects of metastasis-associated tumor gene family 2 (MTA2) depletion on human breast cancer cell proliferation and metastasis. Methods: A short-hairpin RNA targeting MTA2 was chemically synthesized and transfected into a lentivirus to construct Lv-shMTA2 for infection into the MDA-MB231 human breast cancer cell line. At 48 hours after infection cells were harvested and mRNA and protein levels of MTA2 were determined by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting, respectively. Cell viability and metastasis were assessed by CCK-8, wound-healing assay and Transwell assay, respectively. In addition, a xenograft model of human breast cancer was constructed to investigate cancerous cell growth and capacity for metastasis. Results: After infection with Lv-shMTA2, mRNA and protein levels of MTA2 was significantly reduced (p<0.05) and MDA-MB231 cell proliferation and metastasis were inhibited (p<0.05). In addition, mean tumor size was smaller than that in control group nude mice (p<0.05) and numbers of metastatic deposits in lung were lower than in control group mice (p<0.05). Depletion of MTA2 affected MMP-2 and apoptosis-related protein expression. Conclusions: For the first time to our knowledge we showed that MTA2 depletion could significantly inhibit human breast cancer cell growth and metastasis, implying that MTA2 might be involved in the progression of breast cancer. The role of MTA2 in breast cancer growth and metastasis might be linked with regulation of matrix metalloproteinase and apoptosis.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.5467-5472
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• 2013

Objective: To construct short hairpin RNA (shRNA) eukaryotic expression vectors targeting Livin and Survivin genes, and to explore the impact of co-transfection of Livin and Survivin shRNA expression vectors on the biological behavior of HepG2 cells. Methods: shRNA eukaryotic expression vectors pSD11-Livin and pSD11-Survivin were designed and constructed then transfected into HepG2 cells separately or in combination. mRNA and protein expression in transfected cells was assessed by quantitative fluorescence PCR and Western blotting, respectively. Cell proliferation was measured by MTT assay and cell apoptosis by TUNEL assay. Results: The Livin and Survivin shRNA eukaryotic expression vectors were successfully constructed and transfected into HepG2 cells. The relative mRNA expression levels of Livin and Survivin in HepG2 cells co-transfected with pSD11-Livin and pSD11-Survivin were $0.12{\pm}0.02$ and $0.33{\pm}0.13$, respectively, which was significantly lower than levels in cells transfected with either pSD11-Livin or pSD11-Survivin (P<0.05). The relative protein expression levels of Livin and Survivin in the co-transfected cells were also significantly decreased compared to single-transfection (P<0.05). The inhibition rate of cell growth in the co-transfection group was higher than that in the single-transfection groups at 48 h, 60 h, or 72 h after transfection (P<0.01). The apoptotic rate increased to the greatest extent in the co-transfection group relative to any other group (P<0.05). Conclusions: Co-transfection with pSD11-Livin and pSD11-Survivin was more efficient than transfection with either vector alone in reducing the mRNA and protein expression of Livin and Survivin genes in HepG2 cells. Co-transfection also inhibited the proliferation of transfected cells more than the other groups, and induced cellular apoptosis more effectively.

• Archives of Plastic Surgery
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• 제33권1호
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• pp.5-12
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• 2006

Protein tyrosine kinase(PTK), protein kinase C(PKC), oxidase, as a mediator, take a significant role in signal transduction pathway of angiogenesis. The authors utilized the inhibitors, targeting the formation of three co-enzyme in signal transduction pathway in order to quantify the suppression of abnormal vascular endothelial cell proliferation induced by DMH, to compare the level suppression in each up-regulated growth factors, CTGF, CYR61, $ITG{\beta}1$, FHL2, and to identify the relationship between abnormal cell proliferation and signal transduction pathway. Five groups were established; Control group, Group of DMH, Group of DMH-mixed Herbimycin, inhibitor of protein tyrosine kinase, Group of DMH-mixed Calphostin C, inhibitor of protein kinase C, Group Of Dmh-Mixed 10U Catalase, Inhibitor Of oxidase. The rise of vascular endothelial cell was compared by MTT assay, and four growth factors were analysed with RT-PCR method, at pre-administration, 4, 8, 12, and 24 hours after administration. In comparison of abnormal proliferation of vascular endothelial cell induced by DMH, suppression was noticed in Herbimycin and Calphostin C group, and Calphostin C group revealed higher suppression effect. Nevertheless, Catalase group did not have any suppression. In manifestation of four growth factors, Herbimycin and Calphostin C group presented similar manifestation with control group, except in $ITG{\beta}$. Catalse group had similar manifestation with DMH group in all four growth factors. Abnormal proliferation of vascular endothelial cell induced by DMH have a direct relationship with PTK and PKC, more specifically to PKC. Oxidase was confirmed not to have any relevance.

• Molecules and Cells
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• 제38권5호
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• pp.416-425
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• 2015

NF-E2-related factor 2 (Nrf2), a basic leucine zipper transcription factor, has recently received a great deal of attention as an important molecule that enhances antioxidative defenses and induces resistance to chemotherapy or radiotherapy. In this study, we investigated the apoptosis-inducing and Nrf2- upregulating effects of quercetin on malignant mesothelioma (MM) MSTO-211H and H2452 cells. Quercetin treatment inhibited cell growth and led to upregulation of Nrf2 at both the mRNA and protein levels without altering the ubiquitination and extending the half-life of the Nrf2 protein. Following treatment with quercetin, analyses of the nuclear level of Nrf2, Nrf2 antioxidant response element-binding assay, Nrf2 promoter-luc assay, and RT-PCR toward the Nrf2-regulated gene, heme oxygenase-1, demonstrated that the induced Nrf2 is transcriptionally active. Knockdown of Nrf2 expression with siRNA enhanced cytotoxicity due to the induction of apoptosis, as evidenced by an increase in the level of proapoptotic Bax, a decrease in the level of antiapoptotic Bcl-2 with enhanced cleavage of caspase-3 and PARP proteins, the appearance of a sub-$G_0/G_1$ peak in the flow cytometric assay, and increased percentage of apoptotic propensities in the annexin V binding assay. Effective reversal of apoptosis was observed following pretreatment with the pan-caspase inhibitor Z-VAD. Moreover, Nrf2 knockdown exhibited increased sensitivity to the anticancer drug, cisplatin, presumably by potentiating the oxidative stress induced by cisplatin. Collectively, our data demonstrate the importance of Nrf2 in cytoprotection, survival, and drug resistance with implications for the potential significance of targeting Nrf2 as a promising strategy for overcoming resistance to chemotherapeutics in MM.

• 대한미생물학회지
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• 제35권2호
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• pp.181-190
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• 2000

To investigate resistance to lamivudine (3TC), we examined the incidence of M184V in 20 HIV-1 patients treated with 3TC for $13.1{\pm}9$ months. Fourteen of 20 patients had been exposed to zidovudine (ZDV) or didanosine (ddI) prior to 3TC therapy. Nested PCR targeting to reverse transcriptase (RT) and direct sequencing were performed for peripheral blood mononuclear cells sampled serially. There were resistance mutations to ZDV in at least 9 patients at baseline, although there was no resistance mutation to 3TC. We could detect M184V in 6 (30%) out of 20 patients. The incidence of M184V increased as the duration of therapy prolongs (13% in samples <12 months; 47% in samples ${\ge}12$ months). The frequency of mutation M184V was higher in patients with previous mutation to ZDV than in patients with wild type. Resistance mutation was not detected in 7 patients. This study shows that resistance to 3TC tends to develop rapidly in patients with baseline mutations or two drugs combination therapy than in those treated simultaneously with triple drugs. This report is the first on resistance to 3TC in Korean AIDS patients.

• Parasites, Hosts and Diseases
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• 제56권3호
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• pp.281-285
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• 2018

Cryptosporidium is the most common protozoan that can infect a wide range of animals, including mammals and birds. Avian Cryptosporidium spp. can cause enteric and respiratory diseases which can be fatal in birds and some species are zoonotic. Companion birds have the potential as reservoir due to their close contact with humans. Pet shops are the major source of companion birds. However, few reports are available regarding Cryptosporidium spp. infection among companion birds kept in pet shops. The present study reports the prevalence and molecular characteristics of Cryptosporidium spp. among companion birds kept in pet shops in Japan. A total of 265 fresh fecal samples were obtained from birds kept in 4 pet shops; these birds belonged to 41 species in 3 bird orders. A nested polymerase chain reaction (PCR) assay targeting the small subunit rRNA gene was employed for the detection of Cryptosporidium spp. A total of 24 samples (9.1%) were positive, and Cryptosporidium spp. were detected from all pet shops. The prevalence of Cryptosporidium spp. in each of the bird orders was 6.5% (10/153) in Psittaciformes, 14.4% (13/90) in Passeriformes, and 4.5% (1/22) in Galliformes. Based on sequence analysis, 13 (54.2%) isolates were classified to C. galli, 8 (33.3%) were avian genotype III, and the remaining 3 (12.5%) were C. baileyi. No infection with zoonotic C. meleagridis and no coinfection with multiple Cryptosporidium spp. and/or genotypes were observed. The zoonotic potential of Cryptosporidium spp. infecting companion birds kept in pet shops in Japan is likely to be low.

• Journal of Microbiology and Biotechnology
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• 제27권7호
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• pp.1288-1299
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• 2017

Composting is widely used to transform waste into valuable agricultural organic fertilizer. Anaerobic ammonium-oxidizing (anammox) bacteria play an important role in the global nitrogen cycle, but their role in composting remains poorly understood. In the present study, the community structure, diversity, and abundance of anammox bacteria were analyzed using cloning and sequencing methods by targeting the 16S rRNA gene and the hydrazine oxidase gene (hzo) in samples isolated from compost produced from cow manure and rice straw. A total of 25 operational taxonomic units were classified based on 16S rRNA gene clone libraries, and 14 operational taxonomic units were classified based on hzo gene clone libraries. The phylogenetic tree analysis of the 16S rRNA gene and deduced HZO protein sequences from the corresponding encoding genes indicated that the majority of the obtained clones were related to the known anammox bacteria Candidatus "Brocadia," Candidatus "Kuenenia," and Candidatus "Scalindua." The abundances of anammox bacteria were determined by quantitative PCR, and between $2.13{\times}10^5$ and $1.15{\times}10^6$ 16S rRNA gene copies per gram of compost were found. This study provides the first demonstration of the existence of anammox bacteria with limited diversity in cow manure composting.

• 농업과학연구
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• 제41권3호
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• pp.157-161
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• 2014

Understanding the gene flow from genetically modified (GM) crops to conventional crops is important to prevent and mitigate seed contamination caused by pollen-mediated gene flow. We conducted a field test to investigate the gene flow from diamondback moth resistant GM cabbage (Brassica oleracea var. capitata) containing cry1Ac1 gene, to a non-GM control line AD126. GM and non-GM cabbage plants were cultivated in the field and pollinated using Bombus terrestris under the nets during the flowering periods. After seeds were collected from non-GM plants, hybrids between them and the GM cabbages were screened by multiplex PCR targeting cry1Ac1 gene. Out of 878 germinated seedlings, 168 hybrids were found and the average gene flow frequency was 19.7%. Because cabbage is mainly pollinated by insect pollinators, large-scale field tests are needed to study gene flow of GM cabbage.

• Molecules and Cells
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• 제38권10호
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• pp.895-903
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• 2015

Non-coding microRNAs (miRNAs) regulate the translation of target messenger RNAs (mRNAs) involved in the growth and development of a variety of cells, including primordial germ cells (PGCs) which play an essential role in germ cell development. However, the target mRNAs and the regulatory networks influenced by miRNAs in PGCs remain unclear. Here, we demonstrate a novel miRNAs control PGC development through targeting mRNAs involved in various cellular pathways. We reveal the PGC-enriched expression patterns of nine miRNAs, including miR-10b, -18a, -93, -106b, -126-3p, -127, -181a, -181b, and -301, using miRNA expression analysis along with mRNA microarray analysis in PGCs, embryonic gonads, and postnatal testes. These miRNAs are highly expressed in PGCs, as demonstrated by Northern blotting, miRNA in situ hybridization assay, and miRNA qPCR analysis. This integrative study utilizing mRNA microarray analysis and miRNA target prediction demonstrates the regulatory networks through which these miRNAs regulate their potential target genes during PGC development. The elucidated networks of miRNAs disclose a coordinated molecular mechanism by which these miRNAs regulate distinct cellular pathways in PGCs that determine germ cell development.

• Molecules and Cells
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• 제40권2호
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• pp.151-161
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• 2017

Proper synaptic function in neural circuits requires precise pairings between correct pre- and post-synaptic partners. Errors in this process may underlie development of neuropsychiatric disorders, such as autism spectrum disorder (ASD). Development of ASD can be influenced by genetic factors, including copy number variations (CNVs). In this study, we focused on a CNV occurring at the 16p11.2 locus in the human genome and investigated potential defects in synaptic connectivity caused by reduced activities of genes located in this region at Drosophila larval neuromuscular junctions, a well-established model synapse with stereotypic synaptic structures. A mutation of rolled, a Drosophila homolog of human mitogen-activated protein kinase 3 (MAPK3) at the 16p11.2 locus, caused ectopic innervation of axonal branches and their abnormal defasciculation. The specificity of these phenotypes was confirmed by expression of wild-type rolled in the mutant background. Albeit to a lesser extent, we also observed ectopic innervation patterns in mutants defective in Cdk2, Gq, and Gp93, all of which were expected to interact with Rolled MAPK3. A further genetic analysis in double heterozygous combinations revealed a synergistic interaction between rolled and Gp93. In addition, results from RT-qPCR analyses indicated consistently reduced rolled mRNA levels in Cdk2, Gq, and Gp93 mutants. Taken together, these data suggest a central role of MAPK3 in regulating the precise targeting of presynaptic axons to proper postsynaptic targets, a critical step that may be altered significantly in ASD.