Ethanol-induced augmentation of annexin IV in cultured cells and the enhancement of cytotoxicity by overexpression of annexin IV by ethanol

The annexins are a family of highly homologous Ca(2+) and phospholipid binding proteins. The expressive amounts of several annexins have been shown to alter in certain pathological states such as brain ischemia and Alzheimer's disease. It has been demonstrated that ethanol induces cytotoxicity, which results in brain damage. In this study, we examined the relationship between ethanol-induced cytotoxicity and the intrinsic amount of annexins using cell lines (rat glioma C6 cells and human adenocarcinoma A549 cells). A decrease in the mitochondrial enzyme (dehydrogenase) activity, which is widely used to measure cytotoxicity, was observed with a high concentration of ethanol (200 mM or more) after a 24-h exposure in both C6 and A549 cells. Western blot analysis revealed that the amount of annexin IV was augmented in both cells by ethanol, whereas levels of annexins I and V were unchanged. The amount of annexin IV was augmented with increasing concentration of ethanol. The overexpression of annexin IV in C6 cells by transfection with annexin IV-DNA enhanced ethanol-induced cell lesion and was accompanied by NFkappaB activation. Thus, it might be indicated that the amount of annexin IV is selectively augmented and this augmentation facilitates the development of cell lesion by ethanol.     

Enhancement of tolerance to heavy metals and oxidative stress in Dunaliella tertiolecta by Zn-induced phytochelatin synthesis

The synthesis of phytochelatins (PCs) in a marine alga, Dunalliela tertiolecta, is strongly induced by Zn. Pretreatment of the cells with Zn enhances the tolerance toward toxic heavy metals such as Cd, Hg, Cu, Pb, and arsenate. Moreover, the pretreatment also increases the tolerance toward oxidative stress caused by hydrogen peroxide or paraquat. In vitro analysis shows that PC is a stronger scavenger of hydrogen peroxide and superoxide radical than glutathione. These results suggest that PCs inducibly synthesized by Zn treatment could play a role not only in detoxification of heavy metals but also in mitigation of oxidative stress.     

Amyloid beta protein activates PKC-delta and induces translocation of myristoylated alanine-rich C kinase substrate (MARCKS) in microglia

The increased accumulation of activated microglia containing amyloid beta protein (Abeta) around senile plaques is a common pathological feature in subjects with Alzheimer's disease (AD). Much less is known, however, of intracellular signal transduction pathways for microglial activation in response to Abeta. We investigated intracellular signaling in response to Abeta stimulation in primary cultured rat microglia. We found that the kinase activity of PKC-delta but not that of PKC-alpha or -epsilon is increased by stimulation of microglia with Abeta, with a striking tyrosine phosphorylation of PKC-delta. In microglia stimulated with Abeta, tyrosine phosphorylation of PKC-delta was evident at the membrane fraction without an overt translocation of PKC-delta. PKC-delta co-immunoprecipitated with MARCKS from microglia stimulated with Abeta. Abeta induced translocation of MARCKS from the membrane fraction to the cytosolic fraction. Immunocytochemical analysis revealed that phosphorylated MARCKS accumulated in the cytoplasm, particularly at the perinuclear region in microglia treated with Abeta. Taken together with our previous observations that Abeta-induced phosphorylation of MARCKS and chemotaxis of microglia are inhibited by either tyrosine kinase or PKC inhibitors, our results provide evidence that Abeta induces phosphorylation and translocation of MARCKS through the tyrosine kinase-PKC-delta signaling pathway in microglia.     

Duck and human pandemic influenza A viruses retain sialidase activity under low pH conditions

The majority of influenza A viruses isolated from wild birds, but not humans, can replicate in the duck intestinal tract. Here we demonstrate that all duck isolates tested universally retain sialidase activities under low pH conditions independent of their neuraminidase (NA) subtypes. In contrast, the sialidase activities of most isolates from humans and pigs practically disappear below pH 4.5, with the exception of four human pandemic viruses isolated in 1957 and 1968. Sequence comparisons among duck, human, and swine N2 NA subtypes indicate that amino acids at positions 153, 253, 307, 329, 344, 347, 356, 368, 390, and 431 may be associated with the low pH stability of duck and human pandemic N2 NAs. This finding suggests that the low pH stability of duck influenza A virus NA may be a critical factor for replication in the intestinal tract through the digestive tract of ducks, and that the properties of NAs are important for understanding the epidemiology of the influenza virus.