Inhibition of inducible nitric oxide synthase and cyclooxygenase II by Platycodon grandiflorum saponins via suppression of nuclear factor-kappaB activation in RAW 264.7 cells

Saponins are glycosidic compounds present in many edible and inedible plants. They exhibit potent biological activities in mammalian systems, including several beneficial effects such as anti-inflammation and immunomodulation. In this study, we investigated the effects of seven platycodin saponins on the activities of inducible nitric oxide synthase (iNOS) and cyclooxygenase II (COX-2) in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. We found that 2"-O-acetyl polygalacin D (S1), platycodin A (S2), platycodin D (S3), and polygalacin D (S6) inhibited LPS-induced NO production in a concentration-dependent manner. Furthermore, these compounds inhibited the expression of LPS-induced iNOS and COX-2 protein and mRNA without an appreciable cytotoxic effect on RAW 264.7 macrophages, and could suppress induction by LPS of pro-inflammatory cytokines such as prostaglandin E2 (PGE2). Treatment with these compounds of RAW 264.7 cells transfected with a reporter construct indicated a reduced level of LPS-induced nuclear factor-kappaB (NF-kappaB) activity and effectively lowered NF-kappaB binding as measured by electrophoretic mobility shift assay (EMSA). The suppression of NF-kappaB activation appears to occur through the prevention of inhibitor kappaB (IkappaB) degradation. In vivo, platycodin saponin mixture (PS) and S3 protected mice from the lethal effects of LPS. The 89% lethality induced by LPS/galactosamine was reduced to 60% and 50% when PS and S3, respectively, were administered simultaneously with LPS. These results suggest that the main inhibitory mechanism of the platycodin saponins may be the reduction of iNOS and COX-2 gene expression through blocking of NF-kappaB activation.     

In vitro characterization of protein kinase CKII beta mutants defective in beta-beta dimerization

Protein kinase CKII is composed of two catalytic (alpha or alpha') subunits and two regulatory (beta) subunits. The beta subunit mediates tetramer formation through beta-beta homodimerization and alpha-beta heterodimerization. In a previous study R26 and R75, point mutants of CKIIb defective in beta-beta dimerization, were isolated. In the present work we characterized these CKIIbeta mutants in vitro. Purified R26 and R75 bound to CKIIalpha but were defective in binding to CKIIbeta. R75 stimulated the catalytic activity of CKII whereas R26 gave little stimulation, and poly-L-lysine increased the stimulation of catalytic activity by R26 or R75. Circular dichroism and intrinsic fluorescence data pointed to different conformational changes in R26 and R75. Molecular modeling of these mutants provides an explanation of the difference in their ability to interact with CKIIbeta and to activate CKIIalpha.     

Transition from Diffusion‐Controlled Intercalation into Extrinsically Pseudocapacitive Charge Storage of MoS2 by Nanoscale Heterostructuring

2D nanomaterials have been found to show surface‐dominant phenomena and understanding this behavior is crucial for establishing a relationship between a material's structure and its properties. Here, the transition of molybdenum disulfide (MoS₂) from a diffusion‐controlled intercalation to an emergent surface redox capacitive behavior is demonstrated. The ultrafast pseudocapacitive behavior of MoS₂ becomes more prominent when the layered MoS₂ is downscaled into nanometric sheets and hybridized with reduced graphene oxide (RGO). This extrinsic behavior of the 2D hybrid is promoted by the fast Faradaic charge‐transfer kinetics at the interface. The heterostructure of the 2D hybrid, as observed via high‐angle annular dark field–scanning transmission electron microscopy and Raman mapping, with a 1T MoS₂ phase at the interface and a 2H phase in the bulk is associated with the synergizing capacitive performance. This 1T phase is stabilized by the interactions with the RGO. These results provide fundamental insights into the surface effects of 2D hetero‐nanosheets on emergent electrochemical properties.     

Differences of urease activity and expression of associated genes according to gastric topography

Background: We hypothesize that pH difference between acid‐secreting corpus and non‐secreting antrum might influence the activity of H. pylori’s urease and/or related genes. We therefore measured urease activity and the expression of amiE whose encoded protein that hydrolyzes short‐chain amides to produce ammonia. Materials and Methods: Fifty‐four patients were recruited into this study. Each gastroscopic biopsy specimen collected from the antrum and body of each patient was immediately used to measure urease activity using serial changes of urease activity (ammonia levels) during 60 minutes. Probe specific for amiE was labeled with a biotin nick‐translation kit and was used to detect expression of these genes (mRNA) in fresh‐frozen gastroscopic biopsy specimens using fluorescent in situ hybridization (FISH). Results: Urease activity at 60 minutes from the gastric antrum and body of all patients infected with H. pylori was 399.5 ± 490.5 and 837.9 ± 1038.9 μg/dL, respectively (p = .004). Urease activity in the antrum was correlated with H. pylori density. Urease activity or H. pylori density in the antrum was significantly correlated with chronic active inflammation; in contrast, this correlation was not found in the gastric body. The expression level of amiE was 1.5 times higher (p < .05) in the gastric body compared with the antrum. Conclusion: Topographically, the urease activity in body was much higher than in antrum. The expression level of amiE was higher in the gastric body compared with the antrum.     

Insecticide susceptibility and resistance of Culex tritaeniorhynchus (Diptera: Culicidae) larvae collected from Gwangju,Republic of Korea

The susceptibility of Culex tritaeniorhynchus collected from Gwangju, Jeollabuk Province, Republic of Korea (ROK) to insecticides was evaluated under laboratory conditions using ten insecticides (7 pyrethroids and 3 organophosphates) that are currently applied by local public health centers in the ROK. Based on the values of median lethal concentration (LC₅₀), Cx. tritaeniorhynchus larvae were most susceptible to chlorpyrifos (0.006 ppm), fenitrothion (0.022 ppm), fenthion (0.035 ppm) and bifenthrin (0.038 ppm), and were least susceptible to esbiol (1.722 ppm). In comparative resistance tests, the resistance ratios (RRs) of seven insecticides were compared among each other using two strains of Cx. tritaeniorhynchus that were collected from the same locality during 1992 and 2010. Culex tritaeniorhynchus demonstrated significantly increased RRs to pyrethroids over time, while demonstrating decreased RRs among the organophosphates. Among the pyrethroids, permethrin had the highest RR values of 182.1‐ and 833.3‐fold differences, followed by etofenprox with RRs of 138.4‐ and 224.1‐fold differences in values of LC₅₀ and concentration that produced 90% mortality (LC₉₀), respectively. Culex tritaeniorhynchus strains demonstrated the least amount of change in susceptibility to the organophosphates, chlorpyrifos, fenitrothion and fenthion with 0.020‐, 0.019‐ and 0.001‐fold differences in resistance ratios (RRLC₅₀), respectively.     

Screening and characterization of a cellulase gene from the gut microflora of abalone using metagenomic library

A metagenomic fosmid library was constructed using genomic DNA isolated from abalone intestine. Screening of a library of 3,840 clones revealed a 36 kb insert of a cellulase positive clone (pAMHElO). A shotgun clone library was constructed using the positive clone (pAMHElO) and further screening of 3,840 shotgun clones with an approximately 5 kb insert size using a Congo red overlay revealed only one cellulase positive clone (pAMHL9). The pAMHL9 consisted of a 5,293-bp DNA sequence and three open reading frames (ORFs). Among the three ORFs, cellulase activity was only shown in the recombinant protein (CelAMll) coded by ORF3, which showed 100% identity with outer membrane protein A from Vibrio alginolyticus 12G01, but no significant sequence homology to known cellulases. The expressed protein (CelAMll) has a molecular weight of approximately 37 kDa and the highest CMC hydrolysis activity was observed at pH 7.0 and 37°C. The carboxymethyl cellulase activity was determined by zymogram active staining and different degraded product profiles for CelAMll were obtained when cellotetraose and cellopentaose were used as the substrates, while no substrate hydrolysis was observed on oligosaccharides such as cellobiose and cellotriose.