• IMMUNE NETWORK
• /
• v.9 no.5
• /
• pp.192-202
• /
• 2009

Background: Little information is available the role of Nitric Oxide (NO) in host defenses during human tuberculosis (TB) infection. We investigated the modulating factor(s) affecting NO synthase (iNOS) induction in human macrophages. Methods: Both iNOS mRNA and protein that regulate the growth of mycobacteria were determined using reverase transcriptase-polymerase chain reaction and western blot analysis. The upstream signaling pathways were further investigated using iNOS specific inhibitors. Results: Here we show that combined treatment with 1,25-dihydroxyvitamin D3 (1,25-D3) and Interferon (IFN)-${\gamma}$ synergistically enhanced NO synthesis and iNOS expression induced by Mycobacterium tuberculosis (MTB) or by its purified protein derivatives in human monocyte-derived macrophages. Both the nuclear factor-${\kappa}B$ and MEK1-ERK1/2 pathways were indispensable in the induction of iNOS expression, as shown in toll like receptor 2 stimulation. Further, the combined treatment with 1,25-D3 and IFN-${\gamma}$ was more potent than either agent alone in the inhibition of intracellular MTB growth. Notably, this enhanced effect was not explained by increased expression of cathelicidin, a known antimycobacterial effector of 1,25-D3. Conclusion: These data support a key role of NO in host defenses against TB and identify novel modulating factors for iNOS induction in human macrophages.

• Journal of Microbiology and Biotechnology
• /
• v.27 no.1
• /
• pp.171-178
• /
• 2017

A series of novel effector molecules secreted by the type three secretion system (T3SS) of Shigella spp. have been reported in recent years. In this study, a proteomic approach was applied to study T3SS effectors systematically. First, proteins secreted by the S. flexneri wild-type strain after Congo Red induction were separated and identified using two-dimensional electrophoresis to display the relative abundance of all kinds of early effectors for the first time. Then, a gene deletion mutant of known virulence repressor (OspD1) and a gene overexpressed mutant of two known virulence activators (MxiE and IpgC) were constructed and analyzed to discover potential late effectors. Furthermore, the supernatant proteins of gene deletion mutants of two known translocators (IpaB and IpaD), which would constantly secrete effectors, were also analyzed. Among all of the secreted proteins identified in our study, IpaH1.4, IpaH_5, and IpaH_7 have not been reported before. These proteomics data of the secreted effectors will be valuable to understand the pathogenesis of S. flexneri.

• The Korean Journal of Physiology and Pharmacology
• /
• v.18 no.1
• /
• pp.61-66
• /
• 2014

Lung cancer is still the number one cause of death from cancer worldwide. The clinical effect of platinum-based chemotherapy for non-small cell lung cancer is constrained by the resistance to drug. To overcome chemo-resistance, various modified treatment including combination therapy has been used, but overall survival has not been improved yet. In this study, chemo-resistant lung cancer cells, A549/Cis and H460/Cis, were developed by long-term exposure of cells to cisplatin and the proliferative capability of these resistant cells was verified to be reduced. We found cytotoxic effect of epigallocatechin gallate (EGCG), a major catechin derived from green tea, on both the parental lung cancer cells, A549 and H460, and their cisplatin resistant cells, A549/Cis and H460/Cis. ELISA and Western blot analysis revealed that EGCG was able to increase interlukine-6 (IL-6) production per cell, whereas its downstream effector Signal transducers and activators of transcription 3 (STAT3) phosphorylation was not changed by EGCG, indicating that IL-6/STAT3 axis is not the critical signaling to be inhibited by EGCG. We next found that EGCG suppresses the expression of both Axl and Tyro 3 receptor tyrosine kinases at mRNA and protein level, explaining the cytotoxic effect of EGCG on lung cancer cells, especially, regardless of cisplatin resistance. Taken together, these data suggest that EGCG impedes proliferation of lung cancer cells including their chemo-resistant variants through downregulation of Axl and Tyro 3 expression.

• The Journal of the Korean dental association
• /
• v.54 no.4
• /
• pp.284-295
• /
• 2016

The aim of this study was to evaluate the effect of Epigallocatechin-3-Gallate (EGCG) on the alveolar bone metabolism in a collagen-induced arthritis (CIA) model in mice to enhance the understanding of rheumatoid arthritis (RA)-associated alveolar bone loss. Following the induction of CIA in animals (mice, n=16), mandibles were retrieved for micro-computed tomography (micro-CT) and isolation of alveolar bone cells (ABCs). In vitro osteogenic potentials of ABCs were evaluated and the mRNA expression of downstream effector genes was assessed. CIA was successfully induced in all animals, and micro-CT data showed that alveolar bone loss was significantly increased in the CIA group while the treatment of EGCG prevented the alveolar bone resorption. Osteogenesis by ABCs was significantly increased in the CIA+EGCG group in vitro. The analysis of mRNA expressions showed that osteoclastogenesis-associated genes were increased in CIA group while bone protecting genes were upregulated in EGCG treated group. The results demonstrate that EGCG downregulated the alveolar bone resorption in a CIA model in mice, and upregulation of bone protecting genes appear to be involved. Further studies are warranted.

• Biomedical Science Letters
• /
• v.13 no.3
• /
• pp.239-244
• /
• 2007

Hoxc8 is one of the homeotic developmental control genes regulating the expression of many downstream target genes, through which animal body pattern is established during embryonic development. In previous proteomics analysis, proliferating cell nuclear antigen (PCNA) which is also known as cyclin, has been implied to be regulated by Hoxc8 in F9 murine embryonic teratocarcinoma cell. When the 5' upstream region of PCNA was analyzed, it turned out to contain 20 Hox core binding sites (ATTA) in about 1.17 kbp (kilo base pairs) region ($-520{\sim}-1690$). In order to test whether this region is responsible for Hoxc8 regulation, the upstream 2.3 kbp fragment of PCNA was amplified through PCR and then cloned into the pGL3 basic vector containing a luciferase gene as a reporter. When the luciferase activity was measured in the presence of effector plasmid (pcDNA : c8) expressing murine Hoxc8, the PCNA promoter driven reporter activity was reduced. To confirm whether this reduction is due to the Hoxc8 protein, the siRNA against Hoxc8 (5'-GUA UCA GAC CUU GGA ACU A-3' and 5'-UAG UUC CAA GGU CUG AUA C-3') was prepared. Interestingly enough, siRNA treatment up regulated the luciferase activity which was down regulated by Hoxc8, indicating that Hoxc8 indeed regulates the expression of PCNA, in particular, down regulation in NIN3T3 cells. These results altogether indicate that Hoxc8 might orchestrate the pattern formation by regulating PCNA which is one of the important proteins involved in several processes such as DNA replication and methylation, chromatin remodeling, cell cycle regulation, differentiation, as well as programmed cell death.

• BMB Reports
• /
• v.41 no.6
• /
• pp.415-434
• /
• 2008

Phosphoinositide-specific phospholipase C is an effector molecule in the signal transduction process. It generates two second messengers, inositol-1,4,5-trisphosphate and diacylglycerol from phosphatidylinositol 4,5-bisphosphate. Currently, thirteen mammal PLC isozymes have been identified, and they are divided into six groups: PLC-$\beta$, -$\gamma$, -$\delta$, -$\varepsilon$, -$\zeta$ and -$\eta$. Sequence analysis studies demonstrated that each isozyme has more than one alternative splicing variant. PLC isozymes contain the X and Y domains that are responsible for catalytic activity. Several other domains including the PH domain, the C2 domain and EF hand motifs are involved in various biological functions of PLC isozymes as signaling proteins. The distribution of PLC isozymes is tissue and organ specific. Recent studies on isolated cells and knockout mice depleted of PLC isozymes have revealed their distinct phenotypes. Given the specificity in distribution and cellular localization, it is clear that each PLC isozyme bears a unique function in the modulation of physiological responses. In this review, we discuss the structural organization, enzymatic properties and molecular diversity of PLC splicing variants and study functional and physiological roles of each isozyme.

• Asian Pacific Journal of Cancer Prevention
• /
• v.13 no.3
• /
• pp.867-872
• /
• 2012

Background: $FOXP3^+$ regulatory T cells (Tregs) inhibit effector T cell functions and are implicated in tumour progression. However, together with microvessel density (MVD) they remain controversial prognostic predictors for renal cell carcinoma (RCC), and potential associations have yet to be determined. The objective of this study was to determine the prognostic significance of Tregs and MVD and their potential relationship in RCCs. Design: Paraffin-embedded tissues from 62 RCC patients were analysed using immunohistochemistry to detect $FOXP3^+$ lymphocytes, and double immunohistochemistry to detect different microvessel types in the tumour interior, rim and normal kidney tissue, and their correlation with clinicopathological characteristics. Survival analysis was also performed. Results: The presence of $FOXP3^+$ cells in the tumour interior or the rim showed no correlation with death from RCC and other pathological characteristics. Negative correlations were noted between the immature MVD in the tumour interior or the rim and tumour size, tumour stage and overall survival; however, there was no correlation with the nuclear grade or pathological type. A positive correlation between $FOXP3^+$ Tregs and immature MVD (r=0.363, P=0.014) and mature MVD (r=0.383, P=0.009) was confirmed in the tumour interior. However, there was no correlation between $FOXP3^+$ Tregs and mature MVD (r=0.281, P=0.076) or immature MVD (r=0.064, P=0.692) in the tumour rim. Conclusions: In this study, a positive correlation between the presence of $FOXP3^+$ Tregs and immature and mature MVD in RCC was confirmed, which suggests a link between suppression of immunity, tumour angiogenesis and poor prognosis.

• Molecules and Cells
• /
• v.38 no.5
• /
• pp.452-456
• /
• 2015

Obesity is the fifth leading risk for death globally, and a significant challenge to global health. It is a common, complex, non-malignant disease and develops due to interactions between the genes and the environment. DNA methylation can act as a downstream effector of environmental signals; analysis of this process therefore holds substantial promise for identifying mechanisms through which genetic and environmental factors jointly contribute to disease risk. To assess the effects of excessive weight and obesity on gene-specific methylation levels of promoter regions, we determined the methylation status of four genes involved in inflammation and oxidative stress [interleukin 6 (IL6), tumor necrosis factor ${\alpha}$ ($TNF{\alpha}$), mitochondrial transcription factor A (TFAM), and glucose transport 4 (GLUT4)] in blood cell-derived DNA from healthy women volunteers with a range of body mass indices (BMIs) by methylation-specific PCR. Interestingly, the samples from obese individuals ($BMI{\geq}30kg/m^2$) showed significantly increased hypermethylation for IL6 gene compared to normal weight ($BMI<23kg/m^2$) and overweight sample ($23kg/m^2{\leq}BMI<30kg/m^2$) (P = 0.034 and P = 0.026). However there was no statistically significant difference in promoter methylation of the other 3 genes between each group. These findings suggest that aberrant DNA methylation of IL6 gene promoter may play an important role in the etiology and pathogenesis of obesity and IL6 methylation could be used as molecular biomarker for obesity risk assessment. Further studies are required to elucidate the potential mechanisms underlying this relationship.

• Molecules and Cells
• /
• v.37 no.6
• /
• pp.467-472
• /
• 2014

Obesity is known to be strongly associated with cardiovascular disease and cancer, the leading causes of mortality worldwide, and develops owing to interactions between genes and the environment. DNA methylation can act as a downstream effector of environmental signals, and analysis of this process therefore holds substantial promise for identifying mechanisms through which genetic and environmental factors jointly contribute to disease risk. Global DNA methylation of peripheral blood cells has recently been proposed as a potential biomarker for disease risk. Repetitive element DNA methylation has been shown to be associated with prominent obesity-related chronic diseases, but little is known about its relationship with weight status. In this study, we quantified the methylation of Alu elements in the peripheral blood DNA of 244 healthy women with a range of body mass indexes (BMIs) using pyrosequencing technology. Among the study participants, certain clinical laboratory parameters, including hemoglobin, serum glutamic oxaloacetic transaminase, serum glutamic- pyruvic transaminase, total cholesterol, and triglyceride levels were found to be strongly associated with BMI. Moreover, a U-shaped association between BMI and Alu methylation was observed, with the lowest methylation levels occurring at BMIs of between 23 and $30kg/m^2$. However, there was no significant association between Alu methylation and age, smoking status, or alcohol consumption. Overall, we identified a differential influence of BMI on global DNA methylation in healthy Korean women, indicating that BMI-related changes in Alu methylation might play a complex role in the etiology and pathogenesis of obesity. Further studies are required to elucidate the mechanisms underlying this relationship.

• Proceedings of the Korea Environmental Mutagen Society Conference
• /
• 2002.05a
• /
• pp.25-35
• /
• 2002

Transcription factor Nrf2 is essential for the antioxidant responsive element (ARE)-mediated induction of phase II detoxifying and oxidative stress enzyme genes. Detailed analysis of differential Nrf2 activity displayed in transfected cell lines ultimately led to the identification of a new protein, which we named Keap1, that suppresses Nrf2 transcriptional activity by specific binding to its evolutionarily conserved amino-terminal regulatory domain. The closest homolog of Keap1 is a Drosophila actin-binding protein called Kelch, implying that Keap1 might be a Nrf2 cytoplasmic effector. We then showed that electrophilic agents antagonize Keap1 inhibition of Nrf2 activity in vivo, allowing Nrf2 to traverse from the cytoplasm to the nucleus and potentiate the ARE response. We postulate that Keap1 and Nrf2 constitute a crucial cellular sensor for oxidative stress, and together mediate a key step in the signaling pathway that leads to transcriptional activation by this novel Nrf2 nuclear shuttling mechanism. The activation of Nrf2 leads in turn to the induction of phase II enzyme and antioxidative stress genes in response to electrophiles and reactive oxygen species.