Differential distribution of TASK-1, TASK-2 and TASK-3 immunoreactivities in the rat and human cerebellum

In this work, the distributions of some acid-sensitive two-pore-domain K⁺ channels (TASK-1, TASK-2 and TASK-3) were investigated in the rat and human cerebellum. Astrocytes situated in rat cerebellar tissue sections were positive for TASK-2 channels. Purkinje cells were strongly stained and granule cells and astrocytes were moderately positive for TASK-3. Astrocytes isolated from the hippocampus, cerebellum and cochlear nucleus expressed TASK channels in a primary tissue culture. Our results suggest that TASK channel expression may be significant in the endoplasmic reticulum of the astrocytes. The human cerebellum showed weak TASK-2 immunolabelling. The pia mater, astrocytes, Purkinje and granule cells demonstrated strong TASK-1 and TASK-3 positivities. The TASK-3 labelling was stronger in general, but it was particularly intense in the Purkinje cells and pia mater.Received 25 February 2004; received after revision 19 April 2004; accepted 28 April 2004     

Diversity of structures and properties among catalases

More than 300 catalase sequences are now available, divided among monofunctional catalases (> 225), bifunctional catalase-peroxidases (> 50) and manganese-containing catalases (> 25). When combined with the recent appearance of crystal structures from at least two representatives from each of these groups (nine from the monofunctional catalases), valuable insights into the catalatic reaction mechanism in its various forms and into catalase evolution have been gained. The structures have revealed an unusually large number of modifications unique to catalases, a result of interacting with reactive oxygen species. Biochemical and physiological characterization of catalases from many different organisms has revealed a surprisingly wide range of catalatic efficiencies, despite similar sequences. Catalase gene expression in micro-organisms generally is controlled either by sensors of reactive oxygen species or by growth phase regulons, although the detailed mechanisms vary considerably.Received 2 June 2003; received after revision 24 June 2003; accepted 1 July 2003     

FK506 sensitizes mammalian cells to high osmolarity by modulating p38 MAP kinase activation

The immunosuppressants tacrolimus (FK506) and cyclosporin A (CsA) have increased the survival rates in organ transplantation. Both drugs inhibit the protein phosphatase calcineurin (CaN) in activated T cells, exhibiting similar side-effects. Diabetes is observed more often in FK506 than CsA therapy, probably due to inhibition of new molecular targets other than CaN. We studied FK506 toxicity in mammalian cells. FK506, but not CsA, regulated p38 activation by osmotic stress, and decreased viability in osmostressed cells. In addition, FK506 treatment strongly increased the phosphorylation of the eukaryotic initiation factor-2a (eIF-2a) subunit. eIF-2a phosphorylation, p38 inhibition and cell lethality were relieved by addition of excess amino acids to the medium, suggesting that amino acid availability mediated FK506 toxicity. Therefore, these FK506-dependent responses could be relevant to the non-therapeutic effects of FK506 therapy.Received 16 October 2003; received after revision 8 January 2004; accepted 14 January 2004     

Differential effects of monastrol in two human cell lines

The kinesin-related protein HsEg5 plays essential roles in mitotic spindle dynamics. Although inhibition of HsEg5 has been suggested as an aid in cancer treatment, the effects of such inhibition on human cells have not been characterized. Here we studied the effects of monastrol, an allosteric HsEg5 inhibitor, on AGS and HT29 cell lines and compared them to those of taxol. While both cell lines were similarly sensitive to taxol, AGS cells were more sensitive to monastrol. The differences in sensitivity were determined by the degree of inhibitory effect on cell proliferation, reversibility of monastrol-induced G2/M arrest, intracellular phenotypes and induction of apoptosis. In both cell lines, monastrol-induced apoptosis was accompanied by mitochondrial membrane depolarization and poly-ADP-ribose polymerase 1 cleavage. In AGS, but not HT29 cells, monastrol-induced apoptosis involved a prominent cleavage of procaspases 8 and 3. While in AGS cells, monastrol induced the formation of symmetric microtubule asters only, in HT29 cells, asymmetric asters were also formed, which may be related to specific HsEg5 functions in HT29 cells.Received 18 February 2004; received after revision 30 May 2004; accepted 16 June 2004     

Polarization of immunity induced by direct injection of naked sequence-stabilized mRNA vaccines

In the context of developing a safe genetic vaccination strategy we tested and studied globin-stabilized mRNA-based vaccination in mice. This vaccination strategy has the advantages of genetic vaccination (easy production, adaptability to any disease and inexpensive storage when lyophilized), but not the drawbacks of DNA vaccination (long-term uncontrolled expression of a transgene, possibility of integration into the host genome and possible induction of anti-DNA antibodies). We report here that injection of naked -globin untranslated region (UTR)-stabilized mRNA coding for -galactosidase is followed by detectable translation in vivo. In addition, we show that such a vaccination strategy primes a T helper 2 (Th2) type of response which can be enhanced and shifted to a Th1-type immune response by application of recombinant granulocyte/macrophage colony-stimulating factor 1 day after mRNA injection. Our data demonstrate that the administration of globin UTR-stabilized mRNA is a versatile vaccination strategy that can be manipulated to fit the requirement of antiviral, antibacterial or antitumor immunity.Received 14 June 2004; received after revision 19 July 2004; accepted 9 August 2004     

Dislocation-driven surface dynamics on solids

Dislocations are line defects that bound plastically deformed regions in crystalline solids. Dislocations terminating on the surface of materials can strongly influence nanostructural and interfacial stability, mechanical properties, chemical reactions, transport phenomena, and other surface processes. While most theoretical and experimental studies have focused on dislocation motion in bulk solids under applied stress and step formation due to dislocations at surfaces during crystal growth, very little is known about the effects of dislocations on surface dynamics and morphological evolution. Here we investigate the near-equilibrium dynamics of surface-terminated dislocations using low-energy electron microscopy. We observe, in real time, the thermally driven nucleation and shape-preserving growth of spiral steps rotating at constant temperature-dependent angular velocities around cores of dislocations terminating on the (111) surface of TiN in the absence of applied external stress or net mass change. We attribute this phenomenon to point-defect migration from the bulk to the surface along dislocation lines. Our results demonstrate that dislocation-mediated surface roughening can occur even in the absence of deposition or evaporation, and provide fundamental insights into mechanisms controlling nanostructural stability.