Inhibitory Effect of Inflexinol on Nitric Oxide Generation and iNOS Expression via Inhibition of NF-κB Activation

Inflexinol, an ent-kaurane diterpenoid, was isolated from the leaves of Isodon excisus. Many diterpenoids isolated from the genus Isodon (Labiatae) have antitumor and antiinflammatory activities. We investigated the antiinflammatory effect of inflexinol in RAW 264.7 cells and astrocytes. As a result, we found that inflexinol (1, 5, 10 μM) suppressed the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) as well as the production of nitric oxide (NO) in LPS-stimulated RAW 264.7 cells and astrocytes. Consistent with the inhibitory effect on iNOS and COX-2 expression, inflexinol also inhibited transcriptional and DNA binding activity of NF-κB via inhibition of IκB degradation as well as p50 and p65 translocation into nucleus. These results suggest that inflexinol inhibits iNOS and COX-2 expression through inhibition of NF-κB activation, thereby inhibits generation of inflammatory mediators in RAW 264.7 cells and astrocytes, and may be useful for treatment of inflammatory diseases

Akt regulates the expression of MafK, synaptotagmin I, and syntenin-1, which play roles in neuronal function

<p>Abstract</p> <p>Background</p> <p>Akt regulates various cellular processes, including cell growth, survival, and metabolism. Recently, Akt's role in neurite outgrowth has also emerged. We thus aimed to identify neuronal function-related genes that are regulated by Akt.</p> <p>Methods</p> <p>We performed suppression subtractive hybridization on two previously established PC12 sublines, one of which overexpresses the wild-type (WT) form and the other, the dominant-negative (DN) form of Akt. These sublines respond differently to NGF's neuronal differentiation effect.</p> <p>Results</p> <p>A variety of genes was identified and could be classified into several functional groups, one of which was developmental processes. Two genes involved in neuronal differentiation and function were found in this group. v-Maf musculoaponeurotic fibrosarcoma oncogene homolog K (MafK) induces the neuronal differentiation of PC12 cells and immature telencephalon neurons, and synaptotagmin I (SytI) is essential for neurotransmitter release. Another gene, <it>syntenin-1 </it>(<it>Syn-1</it>) was also recognized in the same functional group into which <it>MafK </it>and <it>SytI </it>were classified. Syn-1 has been reported to promote the formation of membrane varicosities in neurons. Quantitative reverse transcription polymerase chain reaction analyses show that the transcript levels of these three genes were lower in PC12 (WT-Akt) cells than in parental PC12 and PC12 (DN-Akt) cells. Furthermore, treatment of PC12 (WT-Akt) cells with an Akt inhibitor resulted in the increase of the expression of these genes and the improvement of neurite outgrowth. These results indicate that dominant-negative or pharmacological inhibition of Akt increases the expression of <it>MafK</it>, <it>SytI</it>, and <it>Syn-1 </it>genes. Using lentiviral shRNA to knock down endogenous Syn-1 expression, we demonstrated that Syn-1 promotes an increase in the numbers of neurites and branches.</p> <p>Conclusions</p> <p>Taken together, these results indicate that Akt negatively regulates the expression of <it>MafK</it>, <it>SytI</it>, and <it>Syn-1 </it>genes that all participate in regulating neuronal integrity in some way or another.</p