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Killer‐cell immunoglobulin‐like receptors (KIR) control natural killer (NK) cell functions by recognizing HLA molecules and modulating the activity of NK cells. The KIR gene cluster contains polymorphic and highly homologous genes. Diversity of the KIR region is achieved through differences in gene content, allelic polymorphism, and gene copy number, which result in unrelated individuals having different KIR genotypes and individualized immune responses that are relevant to multiple aspects of human health and disease. Therefore, KIR genotyping is increasingly used in epidemiological studies. Here, we developed multiplex polymerase chain reaction with sequence‐specific primers (PCR‐SSP) to compensate for the shortcomings of the conventional PCR‐SSP method, which is most commonly used for KIR analysis. Multiplex PCR‐SSP method involves six multiplex reactions that detect 16 KIR genes and distinguish variant types of some KIR genes by adding two reactions. The assay was evaluated in a blind survey using a panel of 40 reference DNA standards from the UCLA KIR Exchange Program. The results are 100% concordant with the genotype determined using Luminex‐based reverse sequence‐specific oligonucleotide typing systems. Additionally, we investigated the currently known 16 KIR genes and their common variants in 120 unrelated Korean individuals. The results were consistent with the KIR genotype previously reported by Hwang et al. This multiplex PCR‐SSP is an efficient method for analyzing KIR genotypes in both small‐ and large‐scale studies with minimal labor, reagents, and DNA. Furthermore, by providing a better definition of KIR polymorphisms it can contribute to developments in immunogenetics. [ABSTRACT FROM AUTHOR]

HLA‐A*31:217 differs from HLA‐A*31:01:02:01 in codon 37 in exon 2. [ABSTRACT FROM AUTHOR]

HLA‐B*27:264 differs from HLA‐B*27:05:02:01 in codon 49 in exon 2. [ABSTRACT FROM AUTHOR]

The hypothesis of the present study was that nitro-fatty acids (NO 2 -FAs) would suppress inflammation associated with periodontal disease. To test this hypothesis, we investigated the influence of nitrooleic acid, a prototypical NO 2 -FA, on the inflammatory response of murine macrophages activated with lipopolysaccharide (LPS) from Prevotella intermedia , a pathogen associated the etiology of different types of periodontal diseases. LPS was prepared from P. intermedia cells by using phenol-water protocol. Culture supernatants were assayed for nitric oxide (NO), interleukin-1β (IL-1β), and IL-6. Real-time polymerase chain reaction and immunoblotting analyses were performed to quantify messenger RNA and protein expression, respectively. The secreted embryonic alkaline phosphatase reporter assay was performed to measure NF-κB activation. The transcription factor assay kit was used to measure DNA-binding of NF-κB subunits. Findings obtained from the present study revealed that nitrooleic acid suppresses the generation and messenger RNA expression of inducible NO synthase-derived NO, IL-1β, and IL-6 in RAW264.7 cells activated with P. intermedia LPS and promotes macrophage polarization toward anti-inflammatory M2 phenotype. We also found that nitrooleic acid exerts its effect via heme oxygenase-1 induction and suppression of NF-κB signaling. The inhibition of NO and proinflammatory cytokine production by nitrooleic acid was independent from PPAR-γ, JNK, p38, and STAT1/3. Nitrooleic acid may represent a novel class of agent as a host modulator which has therapeutic benefit in periodontal disease, though more work is needed to confirm this. Proposed cellular mechanisms involved in the inhibition of Prevotella intermedia lipopolysaccharide (LPS)-induced production of nitric oxide (NO), interleukin-1β (IL-1β), and IL-6 by nitrooleic acid (OA-NO 2) in RAW264.7 cells. The generation and messenger RNA expression of inducible NO synthase-derived NO, IL-1β, and IL-6 induced by P. intermedia LPS were suppressed by OA-NO 2. OA-NO 2 encouraged the switch from proinflammatory M1 macrophages to anti-inflammatory M2 macrophage phenotype in cells activated with LPS. OA-NO 2 exerts its effect via heme oxygenase-1 induction and suppression of NF-κB signaling. [Display omitted] [ABSTRACT FROM AUTHOR]

There are few data about Pneumocystis jirovecii pneumonia (PCP) in children, particularly in developed countries. This study investigated the clinical characteristics and prognosis of the clinical PCP in non-HIV-infected Korean children. Children with positive results for the staining and/or polymerase chain reaction (PCR) for P. jirovecii between 2001 and 2021 were identified. Patients were grouped into clinical PCP, which comprised proven and modified probable cases, and non-PCP groups. Modified probable PCP (mp-PCP) indicate the case which P. jirovecii was detected by conventional PCR rather than real-time PCR test. The differences in demographic and clinical characteristics were analyzed between the groups. A total of 110 pneumonia cases with positive results for P. jirovecii PCR and/or stain were identified from 107 children. Of these, 28.2% were classified as non-PCP, 12.7% of proven PCP, and 59.1% of mp-PCP. Compared with the non-PCP group, the mp-PCP group had a significantly higher rate of solid organ transplantation (3.2% vs. 24.6%), fever (58.1% vs. 76.9%), tachypnea (25.8% vs. 66.2%), dyspnea (48.4% vs. 83.1%), desaturation (48.4% vs. 80.0%), and bilateral ground-glass opacity on chest radiograph (19.4% vs. 73.8%). However, when the mp-PCP group was compared with the proven PCP group, there was no statistically significant difference. For children with clinical PCP, age under 5 years of age (odds ratio [OR] 10.7), hospital-onset (OR 6.9), and desaturation as initial symptom (OR 63.5) were significant risk factors for death in multivariable analysis. Modified probable PCP might reliably reflect true PCP in terms of patient's demographic, clinical features, treatment response, and prognosis. Immunocompromised children with hospital-onset pneumonia who are younger than 5 years of age and have desaturation would be more cautiously and aggressively managed for survival through the screening for P. jirovecii by conventional PCR on appropriate lower respiratory specimens. [ABSTRACT FROM AUTHOR]

Background: This study aimed to assess and compare the clinical value of aqueous humor polymerase chain reaction (PCR) and serologic tests in patients diagnosed with suspected infectious uveitis.Methods: In this retrospective observational study, data of 358 patients who were diagnosed with suspected infectious uveitis and who underwent aqueous humor PCR testing were analyzed. PCR and serologic test results were compared with the clinical features.Results: The rates of initial diagnoses for infectious uveitis were higher with PCR (99 patients, 28%) compared to those with serologic tests (38 pateints, 11%). The diagnostic positivity of PCR was 29% for anterior uveitis, 0% for intermediate uveitis, 5% for posterior uveitis, and 30% for panuveitis. In particular, PCR was useful in confirming the diagnosis of cytomegalovirus and varicella-zoster virus infections and Toxoplasma gondii-associated uveitis. For PCR test, the sensitivity was 0.431, specificity was 0.985, and the negative and positive predictive values were 0.506 and 0.980, respectively. For IgM test, the sensitivity was 0.151, specificity was 0.970, and the negative and positive predictive values were 0.403 and 0.895, respectively.Conclusion: Aqueous humor PCR can be a valuable diagnostic tool for confirming the infectious etiology in patients clinically diagnosed with uveitis. PCR had good predictive and diagnostic value for anterior uveitis and panuveitis compared with that for intermediate and posterior uveitis. [ABSTRACT FROM AUTHOR]

Malignant melanoma is considered to be a heterogeneous disease that arises from altered genes and transformed melanocytes. In this study, special softjet cold atmospheric plasma was used to treat three different human melanoma cells using air and N2 gases to check the anti-melanoma activity. The physical effects by plasma revealed an increase in the temperature with the gradual reduction in pH at 60 sec, 180 sec and 300 sec air and N2 plasma treatment. Cellular toxicity revealed a decreased in cell survival (~50% cell survival using air gas and <~60% cell survival using N2 gas at 60 sec plasma treatment in G-361 cells). Gene analysis by q-PCR revealed that 3 min and 5 min air and N2 plasma treatment activated apoptotic pathways by triggering apoptotic genes in all three melanoma cell lines. The apoptosis was confirmed by DAPI staining and its related pathways were further explored according to protein-protein docking, and their probable activation mechanism was revealed. The pathways highlighted that activation of apoptosis which leads to cellular cascades and hence stimulation ASK1 (docking method) revealed that softjet plasma can be an effective modality for human melanoma treatment. [ABSTRACT FROM AUTHOR]

The major histocompatibility complex region has been suggested to play an important role in the development of autoimmune thyroid disease (AITD). In this study, we investigated the associations of human leukocyte antigen (HLA) alleles and amino acid variants of HLA with early-onset AITD. HLA class I and class II genes were analyzed in 116 Korean children with AITDs (Graves’ disease [GD]: 71, Hashimoto’s disease [HD]: 45) and 142 healthy controls. HLA-B*46:01 (OR = 3.96, Pc = 0.008), -C*01:02 (OR = 2.51 Pc = 0.04), -DPB1*02:02 (OR = 3.99, Pc = 0.04), and -DPB1*05:01 (OR = 4.6, Pc = 0.003) were significantly associated with GD, and HLA-A*02:07 (OR = 4.68, Pc = 0.045) and -DPB1*02:02 (OR = 6.57, Pc = 0.0001) with HD. The frequency of HLA-DPB1*05:01 was significantly higher in GD patients than in HD patients (Pc = 0.0005). Furthermore, differences were found between patients with Thyroid associated ophthalmopathy (TAO) and those without TAO in the distribution of HLA-B*54:01 (8.6% vs. 30.6%, P = 0.04) and -C*03:03 (37.1% vs. 11.1%, P = 0.02). In the analysis of amino acid variants of HLA molecules, both Leu35 (OR = 23.38, P = 0.0002) and Glu55 (OR = 23.38, P = 0.0002) of HLA-DPB1 were strongly associated with GD and showed different distributions between GD and HD (P = 0.001). Our results suggest that HLA alleles, especially amino-acid signatures of the HLA-DP β chain, might contribute to the molecular pathogenesis of early-onset AITD. [ABSTRACT FROM AUTHOR]

HLA‐B*13:144 differs from HLA‐B*13:01:01:01 in codon 12 in exon 2. [ABSTRACT FROM AUTHOR]

HLA‐A*24:480 differs from HLA‐A*24:02:01:01 in codon 178 in exon 3. [ABSTRACT FROM AUTHOR]

HLA‐A*02:877 differs from HLA‐A*02:06:01:01 in codon 183 in exon 3. [ABSTRACT FROM AUTHOR]

HLA‐B*44:454 differs from HLA‐B*44:03:02:01 in codon 81 in exon 2. [ABSTRACT FROM AUTHOR]

HLA‐C*07:478 differs from HLA‐C*07:02:01:01 in codon 81 in exon 2. [ABSTRACT FROM AUTHOR]

Adrenergic α2C receptor (ADRA2C) is an inhibitory modulator of the sympathetic nervous system. Knockout mice for this gene show physiological and behavioural alterations that are associated with the fight-or-flight response. There is evidence of positive selection on the regulation of this gene during chicken domestication. Here, we find that the neuronal expression of ADRA2C is lower in human and chimpanzee than in other primates. On the basis of three-dimensional chromatin structure, we identified a cis-regulatory region whose DNA sequences have been significantly accelerated in human and chimpanzee. Active histone modification marks this region in rhesus macaque but not in human and chimpanzee; instead, repressive marks are enriched in various human brain samples. This region contains two neuron-restrictive silencer factor (NRSF) binding motifs, each of which harbours a polymorphism. Our genotyping and analysis of population genome data indicate that at both polymorphic sites, the derived allele has reached fixation in humans and chimpanzees but not in bonobos, whereas only the ancestral allele is present among macaques. Our CRISPR/Cas9 genome editing and reporter assays show that both derived nucleotides repress ADRA2C, most likely by increasing NRSF binding. In addition, we detected signatures of recent positive selection for lower neuronal ADRA2C expression in humans. Our findings indicate that there has been selective pressure for enhanced sympathetic nervous activity in the evolution of humans and chimpanzees. [ABSTRACT FROM AUTHOR]

Pneumococcal surface protein A (PspA) is an important virulence factor of pneumococci and has been investigated as a primary component of a capsular serotype-independent pneumococcal vaccine. Thus, we sought to determine the genetic diversity of PspA to explore its potential as a vaccine candidate. Among the 190 invasive pneumococcal isolates collected from Korean children between 1991 and 2016, two (1.1%) isolates were found to have no pspA by multiple polymerase chain reactions. The full length pspA genes from 185 pneumococcal isolates were sequenced. The length of pspA varied, ranging from 1,719 to 2,301 base pairs with 55.7–100% nucleotide identity. Based on the sequences of the clade-defining regions, 68.7% and 49.7% were in PspA family 2 and clade 3/family 2, respectively. PspA clade types were correlated with genotypes using multilocus sequence typing and divided into several subclades based on diversity analysis of the N-terminal α-helical regions, which showed nucleotide sequence identities of 45.7–100% and amino acid sequence identities of 23.1–100%. Putative antigenicity plots were also diverse among individual clades and subclades. The differences in antigenicity patterns were concentrated within the N-terminal 120 amino acids. In conclusion, the N-terminal α-helical domain, which is known to be the major immunogenic portion of PspA, is genetically variable and should be further evaluated for antigenic differences and cross-reactivity between various PspA types from pneumococcal isolates. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy is both the most common complication and the leading cause of mortality associated with diabetes. Prunella vulgaris, a well-known traditional medicinal plant, is used for the cure of abscess, scrofula, hypertension and urinary diseases. This study confirmed whether an aqueous extract of Prunella vulgaris (APV) suppresses renal inflammation and fibrosis. In human mesangial cell (HMC), pretreatment of APV attenuated 25mM HG-induced suppressed TGF- and Smad-2/4 expression; it increased the expression level of Smad-7. Connective tissue growth factor (CTGF) and collagen IV, fibrosis biomarkers, were significantly decreased by APV. APV suppressed inflammatory factors such as intracellular cell adhesion molecule-1 (ICAM-1) and monocyte chemoattractant protein-1 (MCP-1). APV inhibited activation and translocation of nuclear factor kappa-B (NF-B) in HG-stimulated HMCs. Moreover, APV significantly improved HG-induced ROS in a dose-dependent manner. In diabetic rat models, APV significantly decreased blood glucose, blood urea nitrogen (BUN) and ameliorated plasma creatinine (PCr). APV reduced the PAS positivity staining intensity and basement membrane thickening in glomeruli of diabetic rats. Fibrosis related proteins such as collagen IV and TGF-1 were also inhibited by APV. These results suggest that APV has a significant protective effect against diabetic renal dysfunction including inflammation and fibrosis through disruption of the TGF-/Smad signaling. Therefore, APV may be useful in potential therapies that target glomerulonephritis and glomerulosclerosis, which lead to diabetic nephropathy. [ABSTRACT FROM AUTHOR]

Prion diseases are infectious and fatal neurodegenerative diseases which require the cellular prion protein, PrPC, for development of diseases. The current study shows that the PrPC augments infectivity and plaque formation of a mouse endogenous retrovirus, MuLV. We have established four neuronal cell lines expressing mouse PrPC, PrP+/+; two express wild type PrPC (MoPrPwild) and the other two express mutant PrPC (MoPrPmut). Infection of neuronal cells from various PrP+/+ and PrP-/- (MoPrPKO) lines with MuLV yielded at least three times as many plaques in PrP+/+ than in PrP-/-. Furthermore, among the four PrP+/+ lines, one mutant line, P101L, had at least 2.5 times as many plaques as the other three PrP+/+ lines. Plaques in P101L were four times larger than those in other PrP+/+ lines. Colocalization of PrP and CAgag was seen in MuLV-infected PrP+/+ cells. In the PrP-MuLV interaction, the involvement of galectin-3 and -6 was observed by immunoprecipitation with antibody to PrPC. These results suggest that PrPC combined with galectin-3 and -6 can act as a receptor for MuLV. P101L, the disease form of mutant PrPC results suggest the genetic mutant form of PrPC may be more susceptible to viral infection. [ABSTRACT FROM AUTHOR]

Live attenuated vaccines are extensively used worldwide to control the outbreak of infectious laryngotracheitis. Virulent field strains showing close genetic relationship with the infectious laryngotracheitis virus (ILTV) vaccines of chicken embryo origin have been detected in the poultry industry. Polymerase chain reaction-restriction fragment length polymorphism (PCR- RFLP) analysis, a reliable molecular epidemiological method, of multiple genomic regions was performed. The PCR-RFLP is a time-consuming method that requires considerable amount of intact viral genomic DNA to amplify genomic regions greater than 4 kb. In this study, six variable genomic regions were selected and amplified for sequencing. The multi-allelic PCR- sequence genotyping showed better discrimination power than that of previous PCR- sequencing schemes using single or two target regions. The allelic variation patterns yielded 16 strains of ILTV classified into 14 different genotypes. Three Korean field strains, 550/05/Ko, 0010/05/Ko and 40032/08/Ko, were found to have the same genotype as the commercial vaccine strain, Laryngo Vac (Zoetis, Florham Park, NJ, USA). Three other Korean field strains, 40798/10/Ko, 12/07/Ko, and 30678/14/Ko, showed recombined allelic patterns. The multi- allelic PCR-sequencing method was proved to be an efficient and practical procedure to classify the different strains of ILTV. The method could serve as an alternate diagnostic and differentiating tool for the classification of ILTV, and contribute to understanding of the epidemiology of the disease at a global level. [ABSTRACT FROM AUTHOR]

Copepods, small aquatic crustaceans, are the most abundant metazoan zooplankton and outnumber every other group of multicellular animals on earth. In spite of ecological and biological importance in aquatic environment, their morphological plasticity, originated from their various lifestyles and their incomparable capacity to adapt to a variety of environments, has made the identification of species challenging, even for expert taxonomists. Molecular approaches to species identification have allowed rapid detection, discrimination, and identification of cryptic or sibling species based on DNA sequence data. We examined sequence variation of a partial mitochondrial cytochrome C oxidase I gene (COI) from 133 copepod individuals collected from the Korean Peninsula, in order to identify and discriminate 94 copepod species covering six copepod orders of Calanoida, Cyclopoida, Harpacticoida, Monstrilloida, Poecilostomatoida and Siphonostomatoida. The results showed that there exists a clear gap with ca. 20 fold difference between the averages of within-specific sequence divergence (2.42%) and that of between-specific sequence divergence (42.79%) in COI, suggesting the plausible utility of this gene in delimitating copepod species. The results showed, with the COI barcoding data among 94 copepod species, that a copepod species could be distinguished from the others very clearly, only with four exceptions as followings: Mesocyclops dissimilis–Mesocyclops pehpeiensis (0.26% K2P distance in percent) and Oithona davisae–Oithona similis (1.1%) in Cyclopoida, Ostrincola japonica–Pseudomyicola spinosus (1.5%) in Poecilostomatoida, and Hatschekia japonica–Caligus quadratus (5.2%) in Siphonostomatoida. Thus, it strongly indicated that COI may be a useful tool in identifying various copepod species and make an initial progress toward the construction of a comprehensive DNA barcode database for copepods inhabiting the Korean Peninsula. [ABSTRACT FROM AUTHOR]

The purpose of this research was to covalently graft fibroblast growth factor 2 (FGF-2) onto biphasic calcium phosphate (BCP) via a bifunctional cross-linker technique and to estimate the optimal dose of FGF-2 resulting in the best osteogenic differentiation of human mesenchymal stem cells (hMSCs). SEM observation revealed that the surface of the 100 ng FGF-2 coated BCP was completely covered with the nanoparticles expected to be from the silane coupling agent. XRD, FT-IR, and XPS analysis showed that silane treatment, bifunctional cross-linker coating, and FGF-2 covalent grafts were conducted successfully without deforming the crystalline structure of BCP. An MTT assay demonstrated that FGF-2 coated BCP had good biocompatibility, regardless of the concentration of FGF-2, after 24 or 48 h of incubation. An alkaline phosphatase (ALP) activity assay (14 days of incubation) and the ALP gene expression level of real-time PCR analysis (7 days of incubation) revealed that 50, 100, and 200 ng FGF-2 coated BCP induced the highest activities among all experimental groups and control group (P < 0.05). Thus, low concentrations of FGF-2 facilitated excellent osteogenesis and were effective at enhancing osteogenic potential. Also, the bifunctional cross-linker technique is expected to be a more feasible way to induce osteogenic differentiation while minimizing the risk of FGF-2 overdose. [ABSTRACT FROM AUTHOR]

Bovine tuberculosis caused by Mycobacterium bovis has a wide range of hosts including cattle and humans, but its incidence in otters is very rare. Our report describes a case of bovine tuberculosis in an Asian small-clawed otter (Aonyx cinerea). A deceased female otter ~2–3 years of age that was raised in an aquarium was submitted to the Animal and Plant Quarantine Agency (Anyang, Republic of Korea) for autopsy in June 2013. Following gross pathological examination, many white nodules were observed in the lungs and mesentery. The nodules showed central necrosis infiltrated with lymphocytes and macrophages and surrounded by fibrous tissue. Acid-fast bacteria were detected in the necrotic foci, but no fungi were observed. Molecular analysis led to the detection of M. bovis, which is identified in otters in some European countries such as Spain and France. [ABSTRACT FROM AUTHOR]

We conducted a study on the relationship between leukocyte telomere length (LTL) and quality of life (QoL) in 82 couples aged 55 and older. LTL was measured by quantitative real-time PCR. QoL was evaluated using the physical component score (PCS) and mental component score (MCS) on the Korean version of the Short-Form Health Survey (SF-36). LTL was found to be independently associated with the SF-36 PCS in males (β=0.014, p=0.03) and the SF-36 MCS in females (β=1.16, p<0.01). Thus LTL is associated with QoL in gender-specific ways. [ABSTRACT FROM AUTHOR]

Several reports have indicated that dietary intake of DHA is associated with lower prevalence of periodontitis. In the present study, we investigated the effect of DHA on the production of proinflammatory mediators in murine macrophage-like RAW264.7 cells stimulated with lipopolysaccharide (LPS) isolated from Prevotellaintermedia, a pathogen implicated in inflammatory periodontal disease, and its mechanisms of action. LPS was isolated from lyophilised P. intermedia ATCC 25 611 cells using the standard hot-phenol–water protocol. Culture supernatants were collected and assayed for NO, IL-1β and IL-6. Real-time PCR analysis was carried out to detect the expression of inducible NO synthase (iNOS), IL-1β, IL-6 and haeme oxygenase-1 (HO-1) mRNA. Immunoblot analysis was carried out to quantify the expression of iNOS and HO-1 protein and concentrations of signalling proteins. DNA-binding activities of NF-κB subunits were determined using an ELISA-based assay kit. DHA significantly attenuated the production of NO, IL-1β and IL-6 at both gene transcription and translation levels in P. intermedia LPS-activated RAW264.7 cells. DHA induced the expression of HO-1 in cells treated with P. intermedia LPS. Selective inhibition of HO-1 activity by tin protoporphyrin IX significantly mitigated the inhibitory effects of DHA on LPS-induced NO production. DHA significantly attenuated the phosphorylation of c-Jun N-terminal kinase induced by LPS. In addition, DHA suppressed the transcriptional activity of NF-κB by regulating the nuclear translocation and DNA-binding activity of NF-κB p50 subunit and inhibited the phosphorylation of signal transducer and activator of transcription 1. Further in vivo studies are needed to better evaluate the potential of DHA in humans as a therapeutic agent to treat periodontal disease. [ABSTRACT FROM PUBLISHER]

HLA‐A*11:384 differs from HLA‐A*11:01:01:01 in codon 151 in exon 3. [ABSTRACT FROM AUTHOR]

High levels of expression of Wilms' tumor gene 1 ( WT1), survivin, or telomerase reverse transcriptase ( TERT) genes are introduced as leukemia-associated targets predicting clinical outcome. We prospectively investigated the leukemia-associated gene transcripts by real-time quantitative polymerase chain reaction from 151 adult patients with AML associated with the patients' clinical characteristics. The maximum levels of each gene in bone marrow were 64.4-, 8.1-, and 3.9-fold higher than those in the normal control, respectively. In contrast to the WT1 and TERT levels, survivin showed comparatively higher expression in the unfavorable cytogenetic group of patients. We found a significant difference in s urvivin levels between the CR and non- CR groups ( P = 0.0237). TERT expression levels were higher in patients who had a greater number of peripheral blood leukemic blasts at diagnosis ( P = 0.0191). Non- MRC subtypes and patients without specific mutations were the most powerful predictive factors for a better CR rate, by multivariate analyses. The lower levels of both WT1 and survivin co-expression ( P = 0.0129) and both survivin + TERT co-expression ( P = 0.0115) were significant factors for better OS. Besides lower initial levels of serum ferritin ( P = 0.0401), lower levels of WT1 ( P = 0.0438) and survivin ( P = 0.0401), lower levels of both WT1 and survivin co-expression ( P = 0.0031), and the three-gene combination of lower WT1 + survivin + TERT ( P = 0.0454) were powerful predictive factors for better EFS. As our findings were based on a single disease entity, that is, adult AML, they suggest that the expression of these genes may be critical for the immunobiology of AML to influence the clinical outcome in various ways. [ABSTRACT FROM AUTHOR]

Target size effect on the sensory signaling intensity of polydiacetylene (PDA) liposome microarrays was systematically investigated. Influenza A virus M1 peptide and M1 antibody were selected as a probe-target pair. While red fluorescence from the PDA liposome microarrays was observed when the larger M1 antibody was used as a target, when the same M1 antibody was used as a probe to detect the smaller M1 peptide sensory signal did not appear. The results reveal that the intensity of the PDA sensory signal is mainly related to the steric repulsion between probe-target complexes not the strength of the probe-target binding force. Based on this finding, we devised a PDA sensory system that directly detects influenza A whole virus as a larger target, and confirmed the target size effect on the signaling efficiency of PDA. [ABSTRACT FROM AUTHOR]

ABSTRACT Human cytomegalovirus (HCMV) stimulates cellular synthesis of DNA and proteins and induces transition of the cell cycle from G1 to S and G2/M phase, in spite of increased amounts of p53 in the infected cells. The immediate early protein IE2-86 kDa (IE86) tethers a transcriptional repression domain to p53; however, its repression of p53 function is not enough to abrogate the G1 checkpoint function of p53. Other HCMV proteins that suppress the activity of p53 were investigated in this study. Of the HCMV proteins that bind to p53 when assessed by immunoprecipitation and immunoblot analysis, HCMV UL44 was chosen as a candidate protein. It was found that reporter gene containing p53 consensus sequence was activated by transfection with wild type p53, but when plasmids of p53 with IE86 or UL44 were co-transfected, p53 transcriptional activity was decreased to 3-7% of the p53 control in a dose-dependent manner. When the deletion mutant of UL44 was co-transected with p53, the carboxyl one-third portion of UL44 had little effect on inhibition of p53 transcriptional activity. The amount of mRNA p21 was measured in H1299 by real time PCR after transfection of the combination of p53 and UL44 vectors and it was found that p21 transcription by p53 was inhibited dose-dependently by UL44. Increased G0/G1 and decreased S phases in p53 wild type-transfected H1299 cells were recovered to the level of p53 mutant type-transfected ones by the additional transfection of UL44 in a dose-dependent manner. In conclusion, the transcriptional activity of p53 is suppressed by UL44 as well as by IE86. [ABSTRACT FROM AUTHOR]

Abstract: Aims: Eupatilin (5,7-dihydroxy-3,4,6-trimethoxyflavone) is a pharmacologically active ingredients in StillenTM, a drug for the gastric mucosal ulcers. Eupatilin has been known to possess anti-peptic, anti-cancer, and anti-allergy activity. In this report, we defined the effect of eupatilin on the endotoxin-induced inflammation in lipopolysaccharide (LPS)-stimulated macrophages. Main methods: Mouse J774A.1 cell line and mouse peritoneal macrophages were used. Gene expression and production of inflammatory mediators were determined by real-time PCR and Western blot. Key findings: Eupatilin dose-dependently suppressed LPS-induced expression of inducible nitric oxide synthase (iNOS) and production of nitric oxide (NO). Eupatilin decreased LPS-induced expression of inflammatory mediators and pro-inflammatory cytokines such as cyclooxygenase-2, monocyte chemoattractant protein-1, tumor necrosis factor-α, interleukin (IL)-1β and IL-6. In addition, this suppression of inflammatory mediators was nuclear factor (NF)-κB dependent. Significance: Our findings imply that eupatilin suppresses inflammatory responses by the inhibition of NF-κB signaling pathway, and downstream inflammatory mediators in endotoxin-stimulated macrophages. [Copyright &y& Elsevier]

Objective: We sought to test two concepts: that nanoparticles can be used for in vivo gene delivery and that canine granulocyte-macrophage colony-stimulating factor (GM-CSF)/nanoparticles can have possibility to be used to treat transient (acute) canine leukopenia. Materials and Methods: We have generated a novel fluorescent-silica nanoparticle binding of canine GM-CSF gene; canine GM-CSF gene was inserted between the cytomegalovirus promoter and poly-adenylation sequences of simian virus 40, and the gene construct was ligated to fluorescent silica nanoparticles functionalized with tertiary amine. Results: When the GM-CSF/nanoparticles were injected into normal dogs, the GM-CSF was expressed in peripheral blood mononuclear cells for at least 9 days and there were significant increases in white blood cell counts, as confirmed by complete blood count, differential count, and flow cytometry. Significant increases in expression of major histocompatibility complex class II on granulocytes and in serum GM-CSF were also observed. Readministration of the nanoparticles was also effective and expression in various tissues was confirmed by reverse transcriptase polymerase chain reaction. Conclusions: These GM-CSF/nanoparticles may be useful for correction of acute leukopenia, such as chemotherapy-induced myelosuppression without developing neutralizing antibodies. [Copyright &y& Elsevier]

HLA‐DRB1*04:05:21 differs from HLA‐DRB1*04:05:01:01 in codon 83 in exon 2. [ABSTRACT FROM AUTHOR]

HLA‐B*51:284 differs from HLA‐B*51:01:01:01 in codon 44 in exon 2. [ABSTRACT FROM AUTHOR]

HLA‐B*15:529 differs from HLA‐B*15:01:01:01 in codon 138 in exon 3. [ABSTRACT FROM AUTHOR]

HLA‐B*54:41 differs from HLA‐B*54:01:01:01 in codon 121 in exon 3. [ABSTRACT FROM AUTHOR]

Kim S-J, Ha M-S, Choi E-Y, Choi J-I, Choi I-S.Prevotella intermedialipopolysaccharide stimulates release of nitric oxide by inducing expression of inducible nitric oxide synthase. J Periodont Res 2004; doi: 10.1111/j.1600-0765.2004.00757.x© Blackwell Munksgaard 2004The purpose of this study was to examine the effects of lipopolysaccharide fromPrevotella intermedia, a major cause of inflammatory periodontal disease, on the production of nitric oxide (NO) and expression of inducible nitric oxide synthase (iNOS) in the murine macrophage cell line RAW264.7. We also attempted to throw light on the signaling mechanisms involved inP. intermedialipopolysaccharide-induced NO production.Lipopolysaccharide fromP. intermediaATCC 25611 was prepared by the standard hot phenol–water method. NO production was assayed by measuring the accumulation of nitrite in culture supernatants. Western blot analysis of iNOS and analysis of reverse transcription-polymerase chain reaction (RT-PCR) products were carried out.We found thatP. intermedialipopolysaccharide can induce iNOS expression and stimulate the release of NO without additional stimuli and demonstrated an important role of the transcription factor nuclear factor-κB (NF-κB) and microtubule polymerization in NO production. The production of NO requiredl-arginine but not activation of protein kinase C or protein tyrosine kinase.The present study clearly shows thatP. intermedialipopolysaccharide fully induced iNOS expression and NO production in RAW264.7 cells in the absence of other stimuli. The ability ofP. intermedialipopolysaccharide to promote the production of NO may be important in the pathogenesis of inflammatory periodontal disease. [ABSTRACT FROM AUTHOR]

In areas with high prevalence of macrolide-resistant Mycoplasma pneumoniae (MRMP) pneumonia, treatment in children has become challenging. This study aimed to analyze the efficacy of macrolides and doxycycline with regard to the presence of macrolide resistance. We analyzed children with MP pneumonia during the two recent epidemics of 2014–2015 and 2019–2020 from four hospitals in Korea. Nasopharyngeal samples were obtained from children with pneumonia for MP cultures and polymerase chain reaction (PCR). Macrolide resistance was determined by the analysis of 23S rRNA gene transition. Time to defervescence and to chest X-ray improvement were analyzed. Of 145 cases, the median age was 5.0 years and MRMP accounted for 59 (40.7%). Among macrolide-susceptible MP (MSMP), 78 (90.7%) were treated with macrolides and 21 (35.6%) in the MRMP group with doxycycline. In MRMP pneumonia, shorter days to defervescence (2 vs. 5 days, p < 0.001) and to chest X-ray improvement (3 vs. 6 days, p < 0.001) in the doxycycline group than in the macrolide group was observed, whereas no differences were observed among children with MSMP pneumonia. Compared to macrolides, treatment with doxycycline resulted in better outcomes with a shorter time to defervescence and to chest X-ray improvement among children with MRMP pneumonia. [ABSTRACT FROM AUTHOR]

Abstract: Widespread emergence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has changed the epidemiology of S. aureus infections. We examined the molecular types and antibiotic susceptibility of CA-MRSA and health care–associated MRSA (HA-MRSA) among Korean children. MRSA isolates were obtained from patients admitted to university-affiliated tertiary hospitals in Korea, between 2006 and 2010. Molecular studies including multilocus sequence typing, SCCmec typing, and polymerase chain reaction amplification of PVL genes and antibiotic susceptibility tests were performed. SCCmec type IV was most frequently found for both CA-MRSA (80.0%) and HA-MRSA (56.4%). ST72-MRSA-SCCmec type IV and its single-locus variants were the most prevalent MRSA clones in the Korean pediatric population, both in community and in health care settings. The PVL genes were detected in 10% (4/40) of CA-MRSA isolates. Most of the clinical MRSA isolates showed vancomycin MIC ≥1.0 μg/mL. In conclusion, the molecular characteristics of HA-MRSA have been changing and CA-MRSA genotype overtook HA-MRSA genotype in health care settings. [Copyright &y& Elsevier]

Abstract: A microfluidic immunosensor utilizing Mie scattering immunoaggultination assay was developed for rapid and sensitive detection of porcine reproductive and respiratory syndrome virus (PRRSV) from lung tissue samples of domesticated pigs. Antibodies against PRRSV were conjugated to the surface of highly carboxylated polystyrene microparticles (diameter=920nm) and mixed with the diluted PRRSV tissue samples in a Y-shaped microchannel. Antibody–antigen binding induced microparticle immunoagglutination, which was detected by measuring the forward 45° light scattering of 380nm incident beam using microcallipered, proximity fiber optics. For comparison, multi-well experiments were also performed using the same optical detection setup. The detection limit was determined to be 10−3 TCID50 ml−1 for PRRSV dissolved in PBS, while those of previous RT-PCR studies for PRRSV were 101 TCID50 ml−1 (conventional assays) or <1TCID50 ml−1 (quantitative real-time assays). Mie scattering simulations were able to predict the shape of the PRRSV standard curve, indicating that any non-linearity of the standard curve can be interpreted purely as an optical phenomenon. Each assay took less than 5min. A strong correlation could be found between RT-PCR and this method for the lung tissue samples, even though their respective detection mechanisms are different fundamentally (nucleic acids for RT-PCR and virus antigens for light scattering immunoagglutination assay). Several different dilution factors were also tested for tissue samples, and 1/10 and 1/100 were found to be usable. If the microfluidic chips are used only once (i.e. without re-using them), both superior sensitivity and satisfactory specificity can be demonstrated. Specificity studies revealed the presence of Type II PRRSV and non-presence of Type I PRRSV and that the microfluidic chip assay could detect Type II North American strain of PRRSV for the animals tested. This work demonstrates the potential of the Mie scattering immunoassay on a microfluidic chip towards real-time detection system for viral pathogens in domesticated animals. [Copyright &y& Elsevier]

Abstract: Background: Phenylketonuria (PKU) is an autosomal recessive disorder caused by a deficiency of phenylalanine hydroxylase (PAH), which catalyzes the conversion of phenylalanine to tyrosine. The resultant hyperphenylalaninemia causes mental retardation, seizure, and abnormalities in behavior and movement. Methods: We analyzed gene expression profiles in brain tissues of Pah enu2 mice to elucidate the mechanisms involved in phenylalanine-induced neurological damage. The altered gene expression was confirmed by real-time PCR and Western blotting. To identify markers associated with neurological damage, we examined TTR expression in serum by Western blotting. Results: Gene expression profiling of brain tissue from a mouse model of PKU revealed overexpression of transthyretin (Ttr), sclerostin domain containing 1 (Sostdc1), α-Klotho (Kl), prolactin receptor (Prlr), and early growth response 2 (Egr2). In contrast to its overexpression in the brain, TTR expression was low in the sera of PKU mice offered unrestricted access to a diet containing phenylalanine. Expression of TTR decreased in a time-dependent manner in phenylalanine-treated HepG2 cells. Conclusions: These findings indicate that Ttr, Sostdc1, Kl, Prlr, and Egr2 can be involved in the pathogenesis of PKU and that phenylalanine might have a direct effect on the level of TTR in serum. [Copyright &y& Elsevier]

This study investigated the roles of β-lactamase and penicillin-binding protein 3 (PBP3) alterations in the development of recent antimicrobial resistance in nontypeable Haemophilus influenzae (NTHi) isolated from Korean children. Nasopharyngeal NTHi isolates from children at a tertiary children's hospital were tested for antimicrobial susceptibility using E-test. β-lactamase production was screened by the paper disc test, and polymerase chain reaction amplification of blaTEM and blaROB-1 was performed. The ftsI gene was amplified to identify PBP3 alteration. Of the 53 NTHi isolates, 69.8% were ampicillin nonsusceptible. The nonsusceptibility rates for cefaclor were 81.1%, cefpodoxime 69.8%, and amoxicillin/clavulanate 32.1%. About 60.3% and 32.1% of the isolates were genetically β-lactamase-nonproducing ampicillin-resistant (gBLNAR) and genetically β-lactamase-producing amoxicillin/clavulanate-resistant (gBLPACR) strains, respectively. Group III amino acid substitutions comprised 65.6% of the gBLNAR strains and 70.6% of the gBLPACR strains. MIC50 for amoxicillin/clavulanate, cefaclor, cefuroxime, cefpodoxime, and cefixime were more than 2–80 times higher in the gBLNAR and gBLPACR strains compared with gBLPAR strains. Group III gBLNAR strains had significantly higher ampicillin, amoxicillin/clavulanate, cefpodoxime, and cefixime minimum inhibitory concentrations than group II strains. Group III gBLNAR and gBLPACR NTHi strains are highly prevalent in Korea, raising the alarm about increasing β-lactam resistance in NTHi. [ABSTRACT FROM AUTHOR]

Purpose: The ability to predict individual responsiveness to cancer therapy is urgently needed. This is particularly true for patients with locally advanced rectal cancer (LARC) because a large proportion are resistant to preoperative chemoradiation therapy (CRT). In this study, we sought to identify markers that could predict response by comparing the gene expression profiles of the tumors of patients who received preoperative CRT.Methods and Materials: The basal gene expression profiles of tumors from 22 LARC patients who were responders (n=9) and nonresponders (n=13) to preoperative CRT were analyzed using RNA sequencing (RNA-Seq). To validate the RNA-Seq findings, real-time reverse transcriptase polymerase chain reaction (RT-PCR) was performed on tumor samples from an additional 40 LARC patients (n=20 responders; n=20 nonresponders). Candidate genes were stably overexpressed or knocked down in colorectal cancer (CRC) cell lines, and the effect on response to radiation was tested in vitro and also in vivo in a mouse xenograft model.Results: Eight differentially expressed (>16-fold) genes (B3GALT4, HSPA1B, KRBOX1, PPBP, PPP1R18, PSMB8, SLC39A7, and TAP2) associated with the preoperative CRT response were identified (P<.0005). Among these genes, real-time RT-PCR showed that PSMB8 and SLC39A7 were upregulated in the responsive group of the additional 40 LARC patients. In CRC cell lines, PSMB8 overexpression significantly reduced colony formation and increased the apoptosis-inducing molecules cleaved caspase-3 and cleaved PARP after 6-Gy irradiation. PSMB8 knockdown increased colony formation and decreased caspase-3 activation and cleaved PARP levels after irradiation. SLC39A7 overexpression had no significant effects on irradiated CSC cells. After irradiation of the xenografted mice, tumors that arose from CRC cell line HCT116 overexpressing PSMB8 grew more slowly than did those from HCT116 with vector alone.Conclusion: These results suggest that PSMB8 is a predictive marker of preoperative radiosensitivity in LARC patients. Clinical validation in a larger cohort is now required. [ABSTRACT FROM AUTHOR]

Due to significant antigenic variations between field isolates of porcine reproductive and respiratory syndrome virus (PRRSV), suboptimal cross-protection between different viruses impedes the effective control of PRRS via vaccination. Our previous study showed that chimeric viruses containing mixed structural genes from two distinct strains (VR2332 and JA142) of PRRSV were highly susceptible to the viral neutralizing activity of antisera generated against both parental strains. In this study, three chimeric viruses (JAP5, JAP56 and JAP2–6) were constructed by replacing ORF5, ORFs 5 and 6, and ORFs 2-6 of VR2332 with the corresponding genes of JA142, respectively, and their ability to confer cross-protection against challenge with the VR2332 and JA142 strains was evaluated in vivo . A total of 114 pigs were divided into 6 groups, and each group was intramuscularly injected with one of the 3 chimeric viruses ( n = 16 pigs per group), VR2332 ( n = 24), JA142 ( n = 24), or sham inoculum ( n = 18). At 44 days post-inoculation (dpi), these pigs were further divided into 15 groups ( n = 6 or 8 pigs per group) and intranasally challenged with VR2332, JA142, or sham inoculum. All pigs inoculated with one of the chimeric viruses prior to challenge had lower viremia levels than the challenge control pigs. Prior inoculation with JAP56 markedly decreased viremia to nearly undetectable levels in pigs challenged with either VR2332 or JA142. These results suggest that chimeric viruses harboring mixed structural genes from two distinct PRRSV strains can provide protection against both donor viruses. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this study was to develop a single-nucleotide polymorphism (SNP) PCR assay to be performed directly on respiratory samples for the simultaneous detection of Mycoplasma pneumoniae and its 23S rRNA gene mutations, which are responsible for macrolide resistance. For multiplex SNP PCR, two outer primers for amplification of the 23S rRNA gene and two mutant-specific primers for the discrimination of single base changes were designed. A total of 73 M. pneumoniae-positive samples and 100 M. pneumoniae-negative samples were analyzed using this assay. By SNP PCR, we detected two mutations conferring high-level macrolide resistance in 22 samples (A2063G from 20 and A2064G from 2 samples); these results are identical to those produced by the 23S rRNA gene sequencing of M. pneumoniae-positive samples. Thus, this assay can be used as a practical method for the simultaneous detection of M. pneumoniae and mutations associated with macrolide resistance directly from respiratory samples. [Copyright &y& Elsevier]

Abstract: To evaluate the diagnostic value of immunochromatographic test (ICT) on pleural fluid in diagnosing pneumococcal empyema in children and to determine pneumococcal serotypes, 62 exudative parapneumonic effusions from Korean children were tested with culture, ICT for S. pneumoniae, pneumococcal autolysin polymerase chain reaction (PCR), and subsequent sequencing. Of the 62 patients, culture was positive in 3 patients only (4.8%). Pneumococci were identified in 13 samples (21.0%) by sequencing-confirmed PCR and ICT, respectively. When pneumococcal empyema was defined by either positive culture or sequence confirmation, the sensitivity of ICT was 76.9% (10/13) and the specificity of ICT was 93.9%. Eight of 10 patients with positive ICT and culture-negative results had a history of prior antibiotics use, whereas none of the culture-proven cases had. Serotypes of PCR-positive samples were determined by multiplex PCR assays. Multiplex PCR detected serotypes 19A (6), 1 (1), 14 (1), 34 (1), and untypable (4). ICT on pleural fluid is a relatively sensitive and highly specific method for diagnosis of pneumococcal empyema, especially in children given prior antibiotics. [Copyright &y& Elsevier]

Summary: The Ku protein is essential for the repair of a majority of DNA double-strand breaks in mammalian cells. The purpose of this study was to investigate the relationship between the expression of Ku70/80 and sensitivity to radiation in cancer cell lines of the head and neck. The sensitivity to radiation in various head and neck cancer cell lines (AMC-HN-1 to -9) was analyzed by colony forming assay. Of the nine cell lines examined, the most radiosensitive cell line (AMC-HN-3) and the most radioresistant cell line (AMC-HN-9) were selected for this experiments. The expression of Ku70/80 was examined after irradiation using real time PCR, Western blotting and immunofluorescence in two different cell lines. Cell cycle distribution after irradiation were analysed. A differential radioresponse was demonstrated by expression of Ku70/80 in AMC-HN-3 and AMC-HN-9 cells. While the expression of Ku70 was slightly increased in the radioresistant AMC-HN-9 cell line, the expression of Ku80 was remarkably increased, suggesting a correlation between Ku80 expression and radiation resistance. Overexpression of Ku80 plays an important role in the repair of DNA damage induced by radiation. Ku80 expression may provide an effective predictive assay of radiosensitivity in head and neck cancers. [Copyright &y& Elsevier]

Abstract: Tumor endothelial marker7 (TEM7) is a transmembrane protein that is highly expressed in the tumor endothelium and neurons. In the present study, the expression profile of TEM7 mRNA and protein was investigated in the dorsal root ganglion (DRG) of the rats. In situ hybridization and reverse transcription polymerase chain reaction studies revealed that TEM7 mRNA expression was localized to the neuronal population of the sensory ganglion. Immunohistochemical analysis on TEM protein with specific antibodies further supported the spatial expression patterns of TEM7 mRNA in the DRG. The temporal expression of TEM7 mRNA in the DRG demonstrated a postnatal increase of TEM7 mRNA expression. Our results indicate that TEM7 may play a role in the peripheral sensory neurons of the vertebrate nervous systems. [Copyright &y& Elsevier]