6-Mercaptopurine attenuates tumor necrosis factor-α production in microglia through Nur77-mediated transrepression and PI3K/Akt/mTOR signaling-mediated translational regulation

Physical interaction between Nur77 and p65. BV-2 cells were pretreated with 6-MP (50 μM) for 16 h followed by exposure to LPS (100 ng/ml) for 60 min. Nuclear extracts were harvested for immunoprecipitation (IP) experiments using anti-Nur77 and anti-p65 antibodies. Immunoblot (IB) analyses of the immunoprecipitates were performed using these antibodies. The immunoblots are representative of three independent experiments. (TIF 280 kb

Bioactive constituent characterization and antioxidant activity of ganoderma lucidum extract fractionated by supercritical carbon dioxide

Ganoderma lucidum has been recognized as a precious fungus in both Chinese and Japanese traditional medicine for centuries. It contains many bioactive ingredients such as triterpenoids and polysaccharides. The present study used supercritical carbon dioxide (SC-CO2) fractionation to fractionate Ganoderma lucidum extract into four fractions (R, F1, F2, & F3) and evaluate the correlation between the content of functional components and their antioxidant ability. Relatively high concentrations of the three types of bioactive constituents were simultaneously partitioned into different fractionation collecting vessels. The free radical scavenging ability was greatest in F1. The IC50 of DPPH scavenging ability was 0.90 mg/mL and that of ABTS radicals scavenging activity was 0.45 mg/mL. The correlation analysis of antioxidant ability with total triterpenoids and total polyphenols showed a positive relationship. In conclusion, this study showed that fractionation of Ganoderma lucidum extract using SC-CO2 fractionation technology was able to effectively partition its bioactive components including triterpenoids, polysaccharides and phenolic compounds and also to increase the antioxidant activities of the fractions

The glucocorticoid-Angptl4-ceramide axis induces insulin resistance through PP2A and PKCζ.

Chronic glucocorticoid exposure is associated with the development of insulin resistance. We showed that glucocorticoid-induced insulin resistance was attenuated upon ablation of Angptl4, a glucocorticoid target gene encoding the secreted protein angiopoietin-like 4, which mediates glucocorticoid-induced lipolysis in white adipose tissue. Through metabolomic profiling, we revealed that glucocorticoid treatment increased hepatic ceramide concentrations by inducing enzymes in the ceramide synthetic pathway in an Angptl4-dependent manner. Angptl4 was also required for glucocorticoids to stimulate the activities of the downstream effectors of ceramide, protein phosphatase 2A (PP2A) and protein kinase Cζ (PKCζ). We further showed that knockdown of PP2A or inhibition of PKCζ or ceramide synthesis prevented glucocorticoid-induced glucose intolerance in wild-type mice. Moreover, the inhibition of PKCζ or ceramide synthesis did not further improve glucose tolerance in Angptl4-/- mice, suggesting that these molecules were major downstream effectors of Angptl4. Overall, our study demonstrates the key role of Angptl4 in glucocorticoid-augmented hepatic ceramide production that induces whole-body insulin resistance

Ultrasonography-guided Percutaneous Interventional Procedures of the Spleen

Since the introduction of real-time ultrasonography (US) to the medicine in late 1970s, the unique benefit of the real-time cross-sectional imaging has made US one of the most widely used imaging modalities to guide interventional procedures. Among the intra-abdominal solid organs, the spleen is the least common solid organ considered for interventional procedures. Although splenic puncture for splenoportography was performed as early as the 1950s and has had a low complication rate, traditionally a direct splenic puncture is still avoided due to the risk of hemorrhage or laceration. US-guided percutaneous drainage of splenic abscesses has been used as a safe alternative procedure for more than 20 years, however, only a few series reporting such an interventional procedure have been published. This review describes briefly the usefulness, technique, safety, and the outcome of US-guided interventional procedures of the spleen

Glycogen synthase kinase-3β inactivation inhibits tumor necrosis factor-α production in microglia by modulating nuclear factor κB and MLK3/JNK signaling cascades

<p>Abstract</p> <p>Background</p> <p>Deciphering the mechanisms that modulate the inflammatory response induced by microglial activation not only improves our insight into neuroinflammation but also provides avenues for designing novel therapies that could halt inflammation-induced neuronal degeneration. Decreasing glycogen synthase kinase-3β (GSK-3β) activity has therapeutic benefits in inflammatory diseases. However, the exact molecular mechanisms underlying GSK-3β inactivation-mediated suppression of the inflammatory response induced by microglial activation have not been completely clarified. Tumor necrosis factor-α (TNF-α) plays a central role in injury caused by neuroinflammation. We investigated the regulatory effect of GSK-3β on TNF-α production by microglia to discern the molecular mechanisms of this modulation.</p> <p>Methods</p> <p>Lipopolysaccharide (LPS) was used to induce an inflammatory response in cultured primary microglia or murine BV-2 microglial cells. Release of TNF-α was measured by ELISA. Signaling molecules were analyzed by western blotting, and activation of NF-κB and AP-1 was measured by ELISA-based DNA binding analysis and luciferase reporter assay. Protein interaction was examined by coimmunoprecipitation.</p> <p>Results</p> <p>Inhibition of GSK-3β by selective GSK-3β inhibitors or by RNA interference attenuated LPS-induced TNF-α production in cultured microglia. Exploration of the mechanisms by which GSK-3β positively regulates inflammatory response showed that LPS-induced IκB-α degradation, NF-κBp65 nuclear translocation, and p65 DNA binding activity were not affected by inhibition of GSK-3β activity. However, GSK-3β inactivation inhibited transactivation activity of p65 by deacetylating p65 at lysine 310. Furthermore, we also demonstrated a functional interaction between mixed lineage kinase 3 (MLK3) and GSK-3β during LPS-induced TNF-α production in microglia. The phosphorylated levels of MLK3, MKK4, and JNK were increased upon LPS treatment. Decreasing GSK-3β activity blocked MLK3 signaling cascades through disruption of MLK3 dimerization-induced autophosphorylation, ultimately leading to a decrease in TNF-α secretion.</p> <p>Conclusion</p> <p>These results suggest that inactivation of GSK-3β might represent a potential strategy to downregulate microglia-mediated inflammatory processes.</p

Introduction of a strong temperature-sensitive phenotype into enterovirus 71 by altering an amino acid of virus 3D polymerase

AbstractIn 1998, an enterovirus 71 (EV71) epidemic in Taiwan resulted in 78 deaths; however, the molecular basis of EV71 pathogenicity remains poorly understood. Comparison of the deduced amino acid sequences in 3D polymerases of EV71clinical isolates showed the T251V or T251I substitution from 1986 and 1998 outbreaks. An EV71 replicon system showed that introducing an I251T mutation did not affect luciferase activities at 35 °C when compared with wild type; however, lower luciferase activities were observed when they were incubated at 39.5 °C. In addition, the I251T mutation in the EV71 infectious clone not only reduced viral replication at 39.5 °C in vitro but also decreased the virulence of the mouse adaptive strain MP4 in neonatal mice in an i.p. infection model. Therefore, these results suggested that the threonine at position 251 results in a temperature sensitivity phenotype of EV71 which may contribute to the attenuation of circulating strains