Among a myriad of pathogen-associated molecular pattern-sensing receptors, toll-like receptors (TLRs) are the principal core sensors of the host. Despite intensive studies for the expression of TLRs and their roles in the central nervous system, controversies remain regarding the expression and the function of TLR4 in human astrocytes. In order to clarify this issue, we attempted to verify functional expression of TLR4 in human astrocytes. Using Reverse transcription-polymerase chain reaction (RT-PCR), we confirmed that the human astrocytes express the TLR4 constitutively. To determine the function of TLR4, astrocytes were treated with TLR4 ligand or lipopolysaccharide (LPS), and then inflammatory cytokines expressions were checked using RT-PCR and enzyme-linked immunosorbent assay. Nuclear factor (NF)-κB activation was checked using electrophoretic mobility shift assay. Treatment of astrocytes with LPS increased tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-8 expression and induced NF-κB activation. Neutralizing anti-TLR4 antibody blocked the effect of LPS on cytokine production and NF-κB activation in astrocytes. The effect of LPS on cytokine production and NF-κB activation was shown in the presence of serum but not in the absence of serum. Therefore, we investigated the sensing mechanism of LPS in human astrocytes. Human astrocytes were treated with LPS following neutralizing anti-CD14 antibody treatment in the presence of serum. Neutralizing anti-CD14 antibody treatment abolished the effect of LPS on cytokine expression and NF-κB activation. Additionally, supplement of recombinant CD14 in serum-free media induced LPS effect on cytokine production and NF-κB activation. In these results, we showed that human astrocytes constitutively express functional TLR4 and require soluble CD14 to recognize LPS.
Astrocytes* ; Central Nervous System ; Culture Media, Serum-Free ; Cytokines ; Electrophoretic Mobility Shift Assay ; Enzyme-Linked Immunosorbent Assay ; Humans* ; Interleukin-8 ; Interleukins ; Toll-Like Receptors ; Tumor Necrosis Factor-alpha

With fast development and wide applications of next generation sequencing (NGS), genomic sequence information is within reach to various research fields. Three benchtop NGS instruments are now available. The 454 GS Junior (Roche), Ion PGM (Life Technologies) and MiSeq (Illumina) are laser-printer sized and offer modest set-up and running costs. By reviewing 2 studies that compared the performance of these instruments, the major characteristics of each benchtop platforms are compared to enable direct comparisons. The 454 GS Junior generated the longest reads and most contiguous assemblies but had the lowest throughput. The Ion Torrent PGM had the highest throughput and fastest run time. The MiSeq had the highest throughput per run and lowest error rates. The Ion Torrent PGM and 454 GS Junior both produced homopolymer-associated indel errors. Although all the platforms allow multiplexing of samples, details of experimental design, library preparation and data analysis may constrain the options. The features of the platforms provide opportunities both to conduct groundbreaking studies and to waste money. Thus, careful considerations should be made before purchasing or using any of them.
Research Design ; Running ; Statistics as Topic

Cyclic guanosine monophosphate adenosine monophosphate (cGAMP) synthase (cGAS) detects human immunodeficiency virus (HIV) and produces cGAMP to induce cytokines. Reverse transcribed DNA of HIV is critical for triggering innate immune responses as inhibitor of HIV reverse transcriptase blocked the induction of interferon-beta by the virus. Furthermore, knockout of cGAS in human or mouse cell lines abrogated the production of cytokines by HIV infection highlighting the essential role of cGAS in detection of HIV and other retroviruses.
Adenosine Monophosphate ; Animals ; Cell Line ; Cytokines ; DNA ; Guanosine Monophosphate ; HIV Infections ; HIV Reverse Transcriptase ; HIV* ; Humans ; Immunity, Innate ; Interferon-beta ; Mice ; Retroviridae

Group A rotaviruses are a major cause of acute gastroenteritis in young children worldwide. For the proper management of rotavirus infections, knowledge of the distribution of G and P genotypes including detection of emerging genotype is crucial. Therefore, the aim of this study is to describe epidemiological changes in rotavirus gastroenteritis in Gwangju metropolitan city, South Korea. Stool samples were collected from 14,314 patients with diarrhea, who visited hospitals in Gwangju from 2008 to 2012. Samples were screened for rotavirus with Enzyme-Linked Immunosorbent Assay (ELISA) method and rotavirus P (VP4), G (VP7) genotypes were determined by reverse-transcription polymerase chain reaction. And we performed nucleotide sequencing analysis. Among a total of 14,314 samples investigated 1,982 samples (13.8%) were ELISA positive. Genotyping of Rotavirus was performed using 526 rotavirus samples. The most prevalent circulating G genotype was G1 (40.5%), followed by G2 (27.6%), G3 (19.4%), G9 (9.7%), G4 (2.5%) and G12 (0.4%). The predominant type of P genotypes was P[8] (69.6%), followed by P[4] (27.8%) and P[6] (2.3%). In this study, 13 G-P combinations were detected. From 2008 to 2010, G1P[8] was the most prevalent, followed by G3P[8]. Whereas, 2011 and 2012, G2P[4] was the most common, followed by G1P[8]. Rotavirus gastroenteritis is a common disease associated with significant morbidity, mortality and economic burden. Ongoing rotavirus surveillance to understand the distribution of G and P genotypes will be critical for the development of effective prevention measurements.
Child ; Diarrhea ; Enzyme-Linked Immunosorbent Assay ; Gastroenteritis ; Genotype ; Gwangju ; Humans ; Korea ; Molecular Epidemiology* ; Mortality ; Polymerase Chain Reaction ; Rotavirus Infections ; Rotavirus*

Turnip yellow mosaic virus (TYMV) is a spherical plant virus that has a single 6.3 kb positive strand RNA as a genome. Previously, we have made the recombinant TYMV construct containing a 0.7 kb eGFP gene or a 1.8 kb GUS gene. The genomic RNAs from these constructs were efficiently encapsidated. To examine in more detail whether size constraint exists for replication and packaging of TYMV, we have inserted into the TY-GUS an extra sequence derived from either eGFP or GUS. We also made a recombinant containing RNA1 sequence of Flock house virus. These TYMV recombinants were introduced into Nicotiana benthamiana leaves by agroinfiltration. Northern blot analysis of the viral RNAs in the agroinfiltrated leaves showed that the genomic RNA band from the recombinant TYMV became weaker as longer sequence was inserted. The result also showed that the efficiency of genomic RNA encapsidation decreased sharply when an extra sequence of 2.2 kb or more was inserted. In contrast, the recombinant subgenomic RNA containing an extra sequence of up to 3.2 kb was efficiently encapsidated. Overall, these results show that size constraint exists for replication and encapsidation of TYMV RNA.
Blotting, Northern ; Genome ; Genome Size* ; Plant Viruses ; Product Packaging* ; RNA ; RNA, Viral ; Tobacco ; Tymovirus*

Helicobacter pylori, a gram-negative bacterium, is a causative agent of gastroduodenal diseases of human. Human immune system produces harmful reactive oxygen species to kill this bacterium that locates the microaerophilic mucous layer. H. pylori harbors various antioxidant enzymes including SodB, KatA and AhpC to protect the oxygen toxicity. We removed the catalase gene (katA) from H. pylori 26695 genome, and the change of profile of the gene expression of the mutant was analyzed by high resolution 2-DE followed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), tandem MS and microarray analysis. Eleven and 37 genes were upregulated and downregulated in the mutant respectively, either transcriptionally or translationally. Expression level of pfr and hp1588 that were decreased on protein level in the mutant was confirmed by RT-PCR analysis.
Catalase ; Gene Expression ; Genome ; Helicobacter pylori* ; Humans ; Immune System ; Mass Spectrometry ; Microarray Analysis ; Oxygen ; Proteome* ; Reactive Oxygen Species

Clostridium perfringens food poisoning ranks among the most common gastrointestinal diseases in developed countries. In Korea, C. perfringens food poisoning gradually increases. Using PCR, 72 strains of C. perfringens isolated in Seoul, 2013 were tested for the presence of toxin genes. Of the tested strains, 32 isolates carried the cpe gene, 37 isolates carried the cpb2 gene and 3 isolates carried the cpe and cpb2 genes, respectively. 32 cpe-positive strains were isolated from the food poisoning patient, whereas among 37 cpb2-positive strains, 22 strains were isolated from asymptomatic person. To investigate epidemiological relationship between the isolates, Pulsed-filed gel electrophoresis (PFGE) was performed. The genetic relatedness of the isolates ranged from 55.9% to 100% and 47 distinct PFGE profiles were observed. The results show that the cpe-positive outbreak strains showed close genetic relation, whereas the cpb2-positive isolates revealed a wide genetic diversity.
Clostridium perfringens* ; Developed Countries ; Electrophoresis ; Foodborne Diseases* ; Gastrointestinal Diseases ; Genetic Variation ; Humans ; Korea ; Molecular Epidemiology* ; Polymerase Chain Reaction ; Seoul

Gastric cancer is the third most common cancer and the third most frequent cause of cancer mortality in Asia. It is predicted that gastric cancer will remain an important cause of death at least during the next half century because of the increasing number of new cases in an aging population. However, little has been revealed about the role of gastric microbes and their reaction to gastric cancer. In this study, we identified differences in the microbial communities between gastric cancer and normal gastric mucosa by comparing the microbiomes of tissues from the same patients. The clustering analysis results showed different bacterial communities between normal gastric mucosa and gastric cancer. A comparison of bacterial communities at the species level revealed that Helicobacter pylori was significantly reduced in cancer tissue compared to that in normal gastric mucosa in the same patient. A comparison at the genus level showed that Propionibacterium spp., Staphylococcus spp., and Corynebacterium spp. had significantly reduced populations in cancer tissue, whereas Clostridium spp. and Prevotella spp. had significantly increased populations in cancer tissue.
Aging ; Asia ; Cause of Death ; Clostridium ; Corynebacterium ; Gastric Mucosa ; Helicobacter pylori ; Humans ; Microbiota ; Mortality ; Mucous Membrane* ; Prevotella ; Propionibacterium ; Staphylococcus ; Stomach Neoplasms ; Stomach*

Oral infection with Porphyromonas (P.) gingivalis causes periodontitis that is manifested by the destruction of gingival connective tissues. Although a few types of antibiotics are effective against the infection, its use induces the appearance of drug-resistant bacteria. The present study shows that the fermented product of Aspergillus (A.) oryzae S-03, cultivated on the fat-removed soybean, inhibits the cell growth of the P. gingivalis. Likewise, the fermented product of the S-03 strain cultured for 26~42 h displays an inhibitory activity to gingipain as a virulence factor of P. gingivalis. The activity is not lost even with heat treatment at 100degrees C for 15 min. We also demonstrate that the S-03 strain exhibits high protease activity. In addition, the strain does not produce aflatoxin because of the loss of a regulatory gene, aflR, necessary for the toxin biosynthesis.
Aflatoxins ; Anti-Bacterial Agents ; Aspergillus ; Aspergillus oryzae* ; Bacteria ; Connective Tissue ; Genes, Regulator ; Hot Temperature ; Oryza ; Periodontitis ; Porphyromonas ; Porphyromonas gingivalis* ; Soybeans ; Virulence*

Resistance to antibiotics is becoming a very serious problem, with so-called superbugs exhibiting resistance to nearly all conventional antibiotic drugs. Consequently, these organisms often cause severe illness and even death. Alternatives to conventional antibiotics are antimicrobial peptides (AMPs). These widely expressed short peptides, which have been isolated from insects, plants, marine organisms and mammals, including humans, show strong antimicrobial activity against both Gram-negative and Gram-positive bacteria. Most AMPs act by disrupting the bacterial membrane through "Barrel-stave", "Toroidal pore", "carpet" mechanism. In addition, AMPs may prevent septic shock through strongly binding lipopolysaccharides and lipoteichoic acid located on the bacterial membrane. The action mechanisms of AMP to minimize the likelihood developing resistance to the peptides would be particular advantage. For these reasons, we anticipate that AMPs will replace conventional antibiotic drugs in a variety of contexts.
Anti-Bacterial Agents ; Aquatic Organisms ; Gram-Positive Bacteria ; Humans ; Insects ; Lipopolysaccharides ; Mammals ; Membranes* ; Peptides* ; Shock, Septic