Role of N-terminal tau domain integrity on the survival of cerebellar granule neurons.

Although the role of the microtubule-binding domain of the tau protein in the modulation of microtubule assembly is widely established, other possible functions of this protein have been poorly investigated. We have analyzed the effect of adenovirally mediated expression of two fragments of the N-terminal portion - free of microtubule-binding domain - of the tau protein in cerebellar granule neurons (CGNs). We found that while the expression of the tau (1-230) fragment, as well as of full-length tau, inhibits the onset of apoptosis, the tau (1-44) fragment exerts a powerful toxic action on the same neurons. The antiapoptotic action of tau (1-230) is exerted at the level of Akt-mediated activation of the caspase cascade. On the other hand, the toxic action of the (1-44) fragment is not prevented by inhibitors of CGN apoptosis, but is fully inhibited by NMDA receptor antagonists. These findings point to a novel, physiological role of the N-terminal domain of tau, but also underlay that its possible proteolytic truncation mediated by apoptotic proteases may generate a highly toxic fragment that could contribute to neuronal death.

Interaction of cationic phospholipid vesicles with carbonic anhydrase.

The possibility of entrapping the enzyme carbonic anhydrase into liposomes, in order to obtain small, membrane-confined bioreactors for biotechnological or biomedical applications, was studied. Neutral liposomes (dipalmitoylphosphatidylcholine/cholesterol) or cationic liposomes (dipalmitoylphosphatidylcholine/cholesterol/stearylamine) with different dipalmitoylphosphatidylcholine/stearylamine ratios have been used to trap carbonic anhydrase. Kinetic experiments showed that carbonic anhydrase was being trapped into cationic liposomes, but not into neutral ones. A significant amount of carbonic anhydrase was sitting onto the external surface of liposomes when the ratio dipalmitoylphosphatidylcholine/cholesterol/stearylamine was 6:3:1, but not when it was 5:3:2. Morphological analysis by electron microscopy showed that the presence of carbonic anhydrase induced a significant swelling in the 6:3:1 cationic vesicles, related to the activity of the enzyme.

Cellular uptake and delivery monitoring of liposome/DNA complexes during in vitro transfection of CFTR gene.

The cellular uptake and distribution of cationic liposomes Dc-Chol/DOPECFTR gene complexes were assessed by electronic and confocal laser scanner microscopy (CLSM) for the CFTR gene transfer to human adenocarcinoma and tracheal epithelial cell lines. Cationic lipid forms unilamellar and multilamellar vesicles capable of rapid and efficient transport of gene into target cells. The number of fluorescent complexes was increasing with time in cells up to 6 hours showing a punctate and homogeneous DNA distribution in the cytoplasmatic and nuclear compartments, including the nucleolus. No significant difference in the biochemical and cellular behavior was observed between the investigated system and other systems previously tested. This study adds new insights into the CFTR cationic liposome-mediated gene delivery.

Three dimensional (3D) analysis of the morphological changes induced by 50 Hz magnetic field exposure on human lymphoblastoid cells (Raji).

Human Raji B lymphoid cells after exposure for 64 h to a 1 mT (rms) 50 Hz sinusoidal magnetic field showed a reorganization of membrane and cytoskeletal components. Atomic force microscopy in air revealed several modifications in 80% of the exposed cells, such as loss of microvilli-like structures followed by progressive appearance of membrane introflections. This change in plasma membrane morphology was also accompanied by a different actin distribution, as detected by phalloidin fluorescence. These observations support our previous hypothesis that electric and magnetic fields may modify the plasma membrane structure.