Rapid quantification of DNA methylation through dNMP analysis following bisulfite-PCR

We report a novel method for rapid quantification of the degree of DNA methylation of a specific gene. Our method combined bisulfite-mediated PCR and quantification of deoxyribonucleoside monophosphate (dNMP) contents in the PCR product through capillary electrophoresis. A specific bisulfite-PCR product was enzymatically hydrolyzed to dNMP monomers which were quantitatively analyzed through subsequent capillary electrophoresis. PCR following bisulfite treatment converts unmethylated cytosines to thymines while leaving methyl-cytosines unchanged. Then the ratio of cytosine to thymine determined by capillary electrophoresis represents the ratio of methyl-cytosine to cytosine in genomic locus of interest. Pure oligonucleotides with known sequences were processed in parallel as standards for normalization of dNMP peaks in capillary electrophoresis. Sources of quantification uncertainty such as carryovers of dNTPs or primers and incomplete hydrolysis were examined and ruled out. When the method was applied to samples with known methylation levels (by bisulfite-mediated sequencing) as a validation, deviations were within ±5%. After bisulfite-PCR, the analytical procedure can be completed within 1.5 h

Insulin Facilitates the Recovery of Myocardial Contractility and Conduction during Cardiac Compression in Rabbits with Bupivacaine-Induced Cardiovascular Collapse

Bupivacaine inhibits cardiac conduction and contractility. Insulin enhances cardiac repolarization and myocardial contractility. We hypothesizes that insulin therapy would be effective in resuscitating bupivacaine-induced cardiac toxicity in rabbits. Twelve rabbits were tracheally intubated and midline sternotomy was performed under general anesthesia. Cardiovascular collapse (CVC) was induced by an IV bolus injection of bupivacaine 10 mg/kg. The rabbits were treated with either saline (control) or insulin injection, administered as a 2 U/kg bolus. Internal cardiac massage was performed until the return of spontaneous circulation (ROSC) and the time to the return of sinus rhythm (ROSR) was also noted in both groups. Arterial blood pressure, and electrocardiography were continuously monitored for 30 min and plasma bupivacaine concentrations at every 5 min. The ROSC, ROSR and normalization of QRS duration were attained faster in the insulin-treated group than in the control group. At the ROSC, there was a significant difference in bupivacaine concentration between two groups. Insulin facilitates the return of myocardial contractility and conduction from bupivacaine-induced CVC in rabbits. However, recovery of cardiac conduction is dependent mainly on the change of plasma bupivacaine concentrations

Transfer Path Analysis of Output Noise Using Multi- dimensional Spectral Analysis Method For Vacuum Cleaner

ABSTRACT Noise reduction of vacuum cleaner is important, according as estimating quality of product. To reduce noise of vacuum cleaner, we need analysis of correct noise source and contribution grasping about Identified noise sources' output noise. Because noise sources' correlation exists in vacuum cleaner that is small and complicated system, and analysis is not easy. In this case, we need to apply Multi-dimensional spectral analysis (MDSA) method that can remove correlation among noise sources and grasp pure contribution degree of noise sources. In this study, we analyze transfer path analysis between output noise and input noise that measured in inside/outside of vacuum cleaner

Anti-inflammatory effects of Radix Gentianae Macrophyllae (Qinjiao), Rhizoma Coptidis (Huanglian) and Citri Unshiu Pericarpium (Wenzhou migan) in animal models

<p>Abstract</p> <p>Background</p> <p>KHU14, an ethanolic extract of <it>Radix Gentianae Macrophyllae </it>(<it>Qinjiao</it>), <it>Rhizoma Coptidis </it>(<it>Huanglian</it>) and <it>Citri Unshiu Pericarpium </it>(<it>Wenzhou migan</it>) was tested for its anti-inflammatory effects.</p> <p>Methods</p> <p>Three out of 20 herbs were found to have anti-inflammatory effects. The formulation of these herbs, i.e. KHU14 was tested for croton oil-induced ear edema, carrageenan-induced paw edema, acetic acid-induced capillary permeability, cotton pellet and delayed type hypersensitivity.</p> <p>Results</p> <p>KHU14 exhibited anti-inflammatory effects in animal models of acute and chronic inflammation. The anti-inflammatory activity of KHU14 observed was comparable to that of celecoxib. KHU14 inhibited the production of NO and PGE₂in LPS/IFN-gamma-stimulated peritoneal macrophages, and reduced edema and the amount of infiltrated cells in animal models.</p> <p>Conclusion</p> <p>KHU14 exhibited anti-inflammatory effects as demonstrated in typical immunological tests for anti-inflammation <it>in vitro </it>and <it>in vivo</it>.</p

Safe and high efficient method for application of liquid ethyl formate (FumateTM) to replace methyl bromide for treatment of imported nursery plants: Poster

There have been significantly increased reports of finding invasive quarantine pests with increasing import plants into Korea. Moreover, the efficacy and work safety issues have been reported regarding use of methyl bromide (MeBr) for fumigation of imported nursery plants. For replacement of MeBr use on imported plants, a new technology of using liquid ethyl formate has been registered in South Korea as FumateTM. The technology involved to mix ethyl format with nitrogen gas to form non-flamable ethyl format formulation. It has been evaluated on various imported plants. The FumateTM is recently developed and commercialized in Republic of Korea and Australia for quarantine treatments on fresh fruits, grains etc. Fumigation with FumateTM offers environmental-friendly and practically safe use of liquid ethyl formate. We have extended the use of liquid EF application technology to quarantine treatment of imported nursery plants.There have been significantly increased reports of finding invasive quarantine pests with increasing import plants into Korea. Moreover, the efficacy and work safety issues have been reported regarding use of methyl bromide (MeBr) for fumigation of imported nursery plants. For replacement of MeBr use on imported plants, a new technology of using liquid ethyl formate has been registered in South Korea as FumateTM. The technology involved to mix ethyl format with nitrogen gas to form non-flamable ethyl format formulation. It has been evaluated on various imported plants. The FumateTM is recently developed and commercialized in Republic of Korea and Australia for quarantine treatments on fresh fruits, grains etc. Fumigation with FumateTM offers environmental-friendly and practically safe use of liquid ethyl formate. We have extended the use of liquid EF application technology to quarantine treatment of imported nursery plants

Association of the programmed cell death 1 (PDCD1) gene polymorphism with ankylosing spondylitis in the Korean population

The PD-1 (programmed death 1) molecule is a negative regulator of T cells. PDCD1 (programmed cell death 1) has been reported to have a genetic association in systemic lupus erythematosus and rheumatoid arthritis in Caucasians. However, there are no reports on the association between this gene and ankylosing spondylitis (AS). The present study investigated the association of the PD-1 polymorphisms and the haplotypes with AS in a Korean population sample. In a case-control association study, two single-nucleotide polymorphisms, PD-1.5 C/T and PD-1.9 T/C, were genotyped in 95 AS patients and 130 healthy controls. The T allele of the PD-1.9 polymorphism was more frequent in the Korean male population with AS than in the Korean male controls (21.0% versus 6.9%, odds ratio 1.89, 95% confidence interval 1.483 to 2.408). The frequency of the CT haplotype (PD-1.5 C/T and PD-1.9 T/C) was higher in the AS patients (19%) than the controls (5.4%) (odds ratio 1.83, 95% confidence interval 1.559 to 2.521). The PD-1 polymorphism was demonstrated in Korean AS patients. The results suggest a genetic association between the PD-1 polymorphism and susceptibility to AS

Taurine chloramine differentially inhibits matrix metalloproteinase 1 and 13 synthesis in interleukin-1β stimulated fibroblast-like synoviocytes

It has been suggested that taurine chloramine (TauCl) plays an important role in the downregulation of proinflammatory mediators. However, little is known about its effect on the expression of matrix metalloproteinases (MMPs). In this study, we investigated the effects of TauCl on synovial expression of MMPs. The effects of TauCl on MMP expression in IL-1β stimulated fibroblast-like synoviocytes (FLSs) were studied using the following techniques. Real-time PCR and semi-quantitative PCR were employed to analyze the mRNA expression of MMPs. ELISA was used to determine protein levels of MMPs. Western blot analyses were performed to analyze the mitogen-activated protein kinase and inhibitor of nuclear factor-κB (IκB) kinase signalling pathways. Finally, electrophoretic mobility shift assay and immunohistochemistry were used to assess localization of transcription factors. IL-1β increased the transcriptional and translational levels of MMP-1 and MMP-13 in rheumatoid arthritis FLSs, whereas the levels of MMP-2 and MMP-9 were unaffected. TauCl at a concentration of 400 to 600 μmol/l greatly inhibited the transcriptional and translational expression of MMP-13, but the expression of MMP-1 was significantly inhibited at 800 μmol/l. At a concentration of 600 μmol/l, TauCl did not significantly inhibit phosphorylation of mitogen-activated protein kinase or IκB degradation in IL-1β stimulated rheumatoid arthritis FLSs. The degradation of IκB was significantly inhibited at a TauCl concentration of 800 μmol/l. The inhibitory effect of TauCl on IκB degradation was confirmed by electrophoretic mobility shift assay and immunochemical staining for localization of nuclear factor-κB. TauCl differentially inhibits the expression of MMP-1 and MMP-13, and inhibits expression of MMP-1 primarily through the inhibition of IκB degradation, whereas it inhibits expression of MMP-13 through signalling pathways other than the IκB pathway