Neuroprotection in a Novel Mouse Model of Multiple Sclerosis

The authors acknowledge the support of the Barts and the London Charity, the Multiple Sclerosis Society of Great Britain and Northern Ireland, the National Multiple Sclerosis Society, USA, notably the National Centre for the Replacement, Refinement & Reduction of Animals in Research, and the Wellcome Trust (grant no. 092539 to ZA). The siRNA was provided by Quark Pharmaceuticals. The funders and Quark Pharmaceuticals had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Thiram inhibits angiogenesis and slows the development of experimental tumours in mice

Thiram-tetramethylthiuram disulphide – a chelator of heavy metals, inhibited DNA synthesis and induced apoptosis in cultured bovine capillary endothelial cells. Bovine capillary endothelial cells were 10–60-fold more sensitive to thiram than other cell types. These effects were prevented by addition of antioxidants, indicating involvement of reactive oxygen species. Exogenously added Cu2+ impeded specifically and almost completely the inhibitory effect of thiram for bovine capillary endothelial cells. Moreover, thiram had markedly inhibited human recombinant Cu/Zn superoxide dismutase enzymatic activity (85%) in vitro. Moreover, PC12-SOD cells with elevated Cu/Zn superoxide dismutase were less sensitive to thiram treatment than control cells. These data indicate that the effects of thiram are mediated by inhibition of Cu/Zn superoxide dismutase activity. Oral administration of thiram (13–30 μg mouse−1), inhibited angiogenesis in CD1 nude mice. Tumour development is known to largely depend on angiogenesis. We found that oral administration of thiram (30 μg) to mice caused significant inhibition of C6 glioma tumour development (60%) and marked reduction (by 3–5-fold) in metastatic growth of Lewis lung carcinoma. The data establish thiram as a potential inhibitor of angiogenesis and raise the possibility for its use as therapy in pathologies in which neovascularisation is involved, including neoplasia

Multiple Aggregates and Aggresomes of C-Terminal Truncated Human αA-Crystallins in Mammalian Cells and Protection by αB-Crystallin

Cleavage of 11 (αA162), 5 (αA168) and 1 (αA172) residues from the C-terminus of αA-crystallin creates structurally and functionally different proteins. The formation of these post-translationally modified αA-crystallins is enhanced in diabetes. In the present study, the fate of the truncated αA-crystallins expressed in living mammalian cells in the presence and absence of native αA- or αB-crystallin has been studied by laser scanning confocal microscopy (LSM).YFP tagged αAwt, αA162, αA168 and αA172, were individually transfected or co-transfected with CFP tagged αAwt or αBwt, expressed in HeLa cells and studied by LSM. Difference in protein aggregation was not caused by different level of α-crystallin expression because Western blotting results showed nearly same level of expression of the various α-crystallins. The FRET-acceptor photo-bleaching protocol was followed to study in situ protein-protein interaction. αA172 interacted with αAwt and αBwt better than αA168 and αA162, interaction of αBwt being two-fold stronger than that of αAwt. Furthermore, aggresomes were detected in cells individually expressing αA162 and αA168 constructs and co-expression with αBwt significantly sequestered the aggresomes. There was no sequestration of aggresomes with αAwt co-expression with the truncated constructs, αA162 and αA168. Double immunocytochemistry technique was used for co-localization of γ-tubulin with αA-crystallin to demonstrate the perinuclear aggregates were aggresomes.αA172 showed the strongest interaction with both αAwt and αBwt. Native αB-crystallin provided protection to partially unfolded truncated αA-crystallins whereas native αA-crystallin did not. Aggresomes were detected in cells expressing αA162 and αA168 and αBwt co-expression with these constructs diminished the aggresome formation. Co-localization of γ-tubulin in perinuclear aggregates validates for aggresomes