Mouse Anti Human Annexins II Hybridoma [FI8b2]

The capsids of viruses have a high degree of symmetry. Therefore, virus nanoparticles (VNPs) can be programmed to display many imaging agents precisely. Plant VNPs are biocompatible, biodegradable and non-infectious to mammals. We have carried out bioconjugation of sesbania mosaic virus (SeMV), a well characterized plant virus, with fluorophores using reactive lysine-N-hydroxysuccinimide ester and cysteine-maleimide chemistries. Monitoring of cellular internalization of labelled SeMV nanoparticles (NPs) by confocal microscopy and flow cytometry showed that the particles have a natural preference for entry into MDA-MB-231 (breast cancer) cells, although they could also enter various other cell lines. The fluorescence of SeMV NPs labelled via the cysteines with Cy5.5 dye was found to be more stable and was detectable with greater sensitivity than that of particles labelled via the lysines with Alexa Fluor. Live-cell imaging using SeMV internally labelled with Cy5.5 showed that it could bind to MDA-MB-231 cells in less than 5 minutes and enter the cells within 15 minutes. The particles undergo endolysosomal degradation by 6 h as evidenced by their co-localization with LAMP-1. Far-western blot analysis with a HeLa cell membrane protein fraction showed that SeMV interacts with 54-, 35- and 33-kDa proteins, which were identified by mass spectrometry as vimentin, voltage-dependent anion-selective channel protein (VDAC1), and annexin A2 isoform 2 (ANXA2), respectively, suggesting that the particles may bind and enter the cell through these proteins. The results presented here demonstrate that the SeMV NPs provide a new platform technology that could be used to develop in vivo imaging and targeted drug delivery agents for cancer diagnosis and therapy.

Previously, galectin-1 (GAL1) was found to be up-regulated in obesity-prone subjects, suggesting that use of a GAL1 inhibitor could be a novel therapeutic approach for treatment of obesity. We evaluated thiodigalactoside (TDG) as a potent inhibitor of GAL1 and identified target proteins of TDG by performing comparative proteome analysis of white adipose tissue (WAT) from control and TDG-treated rats fed a high fat diet (HFD) using two dimensional gel electrophoresis (2-DE) combined with MALDI-TOF-MS. Thirty-two spots from a total of 356 matched spots showed differential expression between control and TDG-treated rats, as identified by peptide mass fingerprinting. These proteins were categorized into groups such as carbohydrate metabolism, tricarboxylic acid (TCA) cycle, signal transduction, cytoskeletal, and mitochondrial proteins based on functional analysis using Protein Annotation Through Evolutionary Relationship (PANTHER) and Database for Annotation, Visualization, Integrated Discovery (DAVID) classification. One of the most striking findings of this study was significant changes in Carbonic anhydrase 3 (CA3), Voltage-dependent anion channel 1 (VDAC1), phosphatidylethanolamine-binding protein 1 (PEBP1), annexin A2 (ANXA2) and lactate dehydrogenase A chain (LDHA) protein levels between WAT from control and TDG-treated groups. In addition, we confirmed increased expression of thermogenic proteins as well as reduced expression of lipogenic proteins in response to TDG treatment. These results suggest that TDG may effectively prevent obesity, and TDG-responsive proteins can be used as novel target proteins for obesity treatment.