Sun Ju Choi

Double-stranded RNA mediates interferon regulatory factor 3 activation and interleukin-6 production by engaging Toll-like receptor 3 in human brain astrocytes

Toll‐like receptor 3 (TLR3) participates in the innate immune response by recognizing viral pathogens. In this study, human brain astrocytes were found to constitutively express TLR3, and this expression was increased by interferon‐γ (IFN‐γ) or double‐stranded RNA (dsRNA). Treatment employing dsRNA in astrocytes induced IFN regulatory factor 3 (IRF3) phosphorylation, dimer formation and nuclear translocation followed by STAT1 activation. This treatment also activated nuclear factor‐κB, p38 and c‐Jun N‐terminal kinase significantly, while activating extracellular signal‐regulated kinase to a lesser extent. Treatment with anti‐TLR3 antibody inhibited dsRNA‐mediated interleukin‐6 (IL‐6) production. In the presence of mitogen‐activated protein kinase inhibitors, astrocytes failed to secrete IL‐6 in response to dsRNA treatment. Therefore, dsRNA‐induced IL‐6 production is dependent on mitogen‐activated protein kinases and type I IFN production is dependent on IRF3 in brain astrocytes. These results suggest that brain inflammation, which produces inflammatory cytokines and type I IFNs, may enhance TLR3 expression in astrocytes. Additionally, upregulated TLR3 might modulate inflammatory processes by producing proinflammatory cytokines.

p38 Mitogen-activated protein kinase modulates expression of tumor necrosis factor-related apoptosis-inducing ligand induced by interferon-γ in fetal brain astrocytes

This study describes the involvement of the p38 mitogen‐activated protein kinase (MAPK) during interferon‐γ (IFN‐γ) signaling in fetal brain astrocytes. In some pathological conditions of brain, p38 MAPK transduces stress‐related signals, increases expression of proinflammatory cytokines, and induces cellular damage or apoptosis. In astrocytes, the tumor necrosis factor (TNF)‐related apoptosis‐inducing ligand (TRAIL) expression level was increased by IFN‐γ. AG490, a JAK inhibitor, blocked TRAIL expression induced by IFN‐γ. SB203580, a specific p38α and p38β2 MAPK inhibitor, decreased the TRAIL expression induced by IFN‐γ. The phosphorylation of the Ser727 site of STAT1, but not the Tyr701 site, was inhibited by SB203580. These results suggest that p38 MAPK modulates STAT1 phosphorylation in IFN‐γ signaling in fetal brain astrocytes.

Differential expression, shedding, cytokine regulation and function of TNFR1 and TNFR2 in human fetal astrocytes.

Tumor necrosis factor (TNF)-α induces pleiotropic cellular effects through a 55kDa, type 1 receptor (TNFR1) and a 75kDa type 2 receptor (TNFR2). Moreover, it participates in the pathogenesis of several CNS diseases, including demyelinating diseases. TNF-α receptors are differentially expressed and are regulated in many cell types. However, data regarding the TNF-α receptor expression and regulation in human astrocytes is limited to date. We investigated TNF-α receptor expression, its regulation by cytokines, and its functional role in primary cultured human fetal astrocytes, which are the most abundant cellular population in the central nervous system and are known to be immunologically active. In this study, astrocytes were found to constitutively and predominantly transcribe, translate and shed TNFR1 rather than TNFR2, but TNFR2 expression was increased by adding TNF-α, IL-1, and IFN-γ, but not by adding LPS. To determine the functional roles of TNFR1 and TNFR2 on TNF induction, we investigated NF-κB activation and TNF-α induction after neutralizing TNFR1 and TNFR2 by an antibody treatment. We found that NF-kB activation and TNF-α induction are blocked by TNFR1 neutralizing antibody treatments.

Effects of cytokines on VEGF expression and secretion by human first trimester trophoblast cell line

PROBLEM: The mechanism through which vascular endothelial growth factor (VEGF) regulation occurs at the feto‐maternal interface is poorly understood. The aim of this study was to investigate the effects of various cytokines on VEGF expression and secretion by trophoblast cells.

Influence of Bacterial Presence on Biofilm Formation of Candida albicans

Purpose Candida albicans is an opportunistic pathogen that is commonly found in human microflora. Biofilm formation (BF) is known as a major virulence factor of C. albicans. The aim of this study was to examine the influence of bacterial presence on biofilm formation of C. albicans. Materials and Methods The BF of Candida was investigated when it was co-cultured with C. albicans (C. albicans 53, a yeast with a low BF ability, and C. albicans 163, a yeast with high BF ability) and bacteria. BF was assessed with XTT reduction assay. A scanning electron microscope was used to determine the structure of the biofilm, and real-time reverse transcriptase polymerase chain reaction was used to amplify and quantify hyphae-associated genes. Results Co-culturing with two different types of bacteria increased the BF value. Co-culturing with C. albicans 53 and 163 also increased the BF value compared to the value that was obtained when the C. albicans was cultured individually. However, co-culturing with bacteria decreased the BF value of C. albicans, and the BF of C. albicans 163 was markedly inhibited. The expression of adherence and morphology transition related genes were significantly inhibited by co-culturing with live bacteria. Conclusion Bacteria have a negative effect on the formation of biofilm by C. albicans. This mechanism is the result of the suppression of genes associated with the hyphae transition of C. albicans, and bacteria particles physically affected the biofilm architecture and biofilm formation.

NF‐κB‐dependency and consequent regulation of IL‐8 in echinomycin‐induced apoptosis of HT‐29 colon cancer cells

The present study was to see whether echinomycin‐induced apoptosis would be NF‐κB‐dependent and if so, whether echinomycin would activate or inhibit NF‐κB as well as resultant chemokine IL‐8 expression. In HT‐29 cells echinomycin activated NF‐κB in time‐dependent manner. EMSA in the presence of antibodies specific for p50 and p65 subunits indicated that echinomycin‐induces the translocation of p50–p65 heterodimeric subunits of NF‐κB. Levels of IκB were detected at initial echinomycin treatment and thereafter decreased, faintly seen after a 6 h treatment. In contrast p‐IκB levels were clearly detected throughout 6–24 h of echinomycin treatment, albeit initially fainted. To clarify the role of NF‐κB on IL‐8 expression in echinomycin‐mediated apoptosis of HT‐29 cells, ELISA plus RT‐PCR clearly showed that IL‐8 production is inducible by echinomycin treatment. Using a specific inhibitor, IL‐8 regulation at echinomycin treatment in HT‐29 cells occurred via both caspase‐3 and NF‐κB‐dependent signal pathway. To confirm whether two different pathways (NF‐κB and caspase) would be coupled, only NF‐κB inhibitor (PDTC) and caspase‐3 specific inhibitor (Z‐DEVD‐FMK) together significantly attenuated echinomycin‐initiated apoptosis of HT‐29 cells, pretreatment of HT‐29 cells with PDTC rarely affected echinomycin‐induced caspase‐3 activation. So echinomycin‐induced apoptosis in HT‐29 cells occurs via NF‐κB activation independent of caspase‐3 activation modulating the resultant‐linked key chemokine IL‐8 expression and echinomycin‐induced apoptosis is NF‐κB‐dependant and directly related to NF‐κB activation, consequently regulating IL‐8 expression.

Proteus vulgaris and Proteus mirabilis Decrease Candida albicans Biofilm Formation by Suppressing Morphological Transition to Its Hyphal Form

Purpose Candida albicans (C. albicans) and Proteus species are causative agents in a variety of opportunistic nosocomial infections, and their ability to form biofilms is known to be a virulence factor. In this study, the influence of co-cultivation with Proteus vulgaris (P. vulgaris) and Proteus mirabilis (P. mirabilis) on C. albicans biofilm formation and its underlying mechanisms were examined. Materials and Methods XTT reduction assays were adopted to measure biofilm formation, and viable colony counts were performed to quantify yeast growth. Real-time reverse transcriptase polymerase chain reaction was used to evaluate the expression of yeast-specific genes (rhd1 and rbe1), filament formation inhibiting genes (tup1 and nrg1), and hyphae-related genes (als3, ece1, hwp1, and sap5). Results Candida biofilm formation was markedly inhibited by treatment with either living or heat-killed P. vulgaris and P. mirabilis. Proteus-cultured supernatant also inhibited Candida biofilm formation. Likewise, treatment with live P. vulgaris or P. mirabilis or with Proteus-cultured supernatant decreased expression of hyphae-related C. albicans genes, while the expression of yeast-specific genes and the filament formation inhibiting genes of C. albicans were increased. Heat-killed P. vulgaris and P. mirabilis treatment, however, did not affect the expression of C. albicans morphology-related genes. Conclusion These results suggest that secretory products from P. vulgaris and P. mirabilis regulate the expression of genes related to morphologic changes in C. albicans such that transition from the yeast form to the hyphal form can be inhibited.