Glycoprotein electrophoretic patterns have potential to monitor changes associated with neoplastic transformation in oral cancer

Alterations in glycoproteins, important cell surface constituents, have long been associated with various malignancies. The present investigation therefore explored the clinical significance of a glycoproteomics approach in patients with oral precancerous conditions (OPC) and patients with oral cancer. The study included 80 oral cancer patients, 50 patients with OPC, and 84 controls. Native polyacrylamide gel electrophoresis followed by Schiff’s staining was carried out to study the alterations in glycoproteins. The results showed significant elevation (p<0.0001) of 192 kDa, 170 kDa, 116 kDa and 44 kDa glycoproteins in oral cancer patients and patients with OPC compared with controls. The odds ratio indicated a significantly higher risk for oral cancer among users and especially chewers of tobacco. The levels of all the glycoprotein bands (192 kDa, 170 kDa, 116 kDa and 44 kDa) were higher in patients with a habit of tobacco use (WHT) than in patients with no habit of tobacco (NHT) and were also higher in WHT controls than in NHT controls. Moreover, a 230 kDa glycoprotein consistently appeared only in individuals with tobacco habits and an increasing trend was observed from WHT controls to patients with OPC to WHT oral cancer patients. In conclusion, the results indicated the potential utility of glycoprotein alterations in monitoring sequential changes occurring due to tobacco consumption during neoplastic transformation.

     

Impairment of reproduction in the red flour beetle, Tribolium castaneum (Herbst) (Col., Tenebrionidae) due to larval feeding on triflumuron-treated diet

The larvae of Tribolium castaneum (Herbst) of different ages were exposed to various concentrations viz., 0.0001, 0.001 and 0.01 ppm of triflumuron up to pupation. The adults were allowed to feed on untreated diet. The preoviposition period, fecundity, egg viability and incubation period of the eggs laid were studied. Triflumuron significantly lengthened the preoviposition and incubation periods (P < 0.001). Fecundity and egg viability were also reduced (P < 0.001) due to the treatment. Both the strains were affected by the treatment.     

Proteome changes associated with hippocampal MRI abnormalities in the lithium pilocarpine-induced model of convulsive status epilepticus

Convulsive status epilepticus is associated with subsequent hippocampal damage and development of mesial temporal sclerosis in a subset of individuals. The lithium pilocarpine model of status epilepticus (SE) in the rat provides a model in which to investigate the molecular and pathogenic process leading to hippocampal damage. In this study, a 2-DE-based approach was used to detect proteome changes in the hippocampus, at an early stage (2 days) after SE, when increased T2 values were detectable by magnetic resonance imaging. Gel image analysis was followed by LC-MS/MS identification of protein species that differed in abundance between pilocarpine-treated and control rats. The most significantly up-regulated species in the experimental animals was identified as heat shock 27-kDa protein, in line with findings in humans and in other experimental models of epilepsy. Additional up-regulated species included dihydropyrimidinase-related protein-2, cytoskeletal proteins (alpha-tubulin and ezrin) and dihydropteridine reductase. In summary, the hippocampus of rats subject to pilocarpine-induced SE exhibits specific changes in protein abundance, which likely relate to pathogenic, neuroprotective and neurogenic responses.     

Prediction of cyclin-dependent kinase phosphorylation substrates

Protein phosphorylation, mediated by a family of enzymes called cyclin-dependent kinases (Cdks), plays a central role in the cell-division cycle of eukaryotes. Phosphorylation by Cdks directs the cell cycle by modifying the function of regulators of key processes such as DNA replication and mitotic progression. Here, we present a novel computational procedure to predict substrates of the cyclin-dependent kinase Cdc28 (Cdk1) in the Saccharomyces cerevisiae. Currently, most computational phosphorylation site prediction procedures focus solely on local sequence characteristics. In the present procedure, we model Cdk substrates based on both local and global characteristics of the substrates. Thus, we define the local sequence motifs that represent the Cdc28 phosphorylation sites and subsequently model clustering of these motifs within the protein sequences. This restraint reflects the observation that many known Cdk substrates contain multiple clustered phosphorylation sites. The present strategy defines a subset of the proteome that is highly enriched for Cdk substrates, as validated by comparing it to a set of bona fide, published, experimentally characterized Cdk substrates which was to our knowledge, comprehensive at the time of writing. To corroborate our model, we compared its predictions with three experimentally independent Cdk proteomic datasets and found significant overlap. Finally, we directly detected in vivo phosphorylation at Cdk motifs for selected putative substrates using mass spectrometry.     

Immobilization of laccase onto spacer-arm attached non-porous poly(GMA/EGDMA) beads: application for textile dye degradation

Non-porous poly(glycidyl methacrylate/ethyleneglycol dimetacrylate) (poly(GMA/EGDMA)) beads were prepared by suspension polymerization. The enzyme (i.e. laccase) was covalently immobilized onto plain and spacer-arm attached poly(GMA/EGDMA) beads. The amount of immobilized enzyme on the plain and spacer-arm attached beads was determined as 5.6 and 4.9 mg/g, respectively. The maximum activity (V(max)) and Michaelis constant (K(m)) of laccase immobilized on the spacer-arm attached beads, were found to be 77.6 U/min and 0.47 mM, respectively. Finally, the immobilized laccase was operated in a batch system, and textile dye Reactive Red 120 was successfully decolorized in the enzyme reactor.     

Hypocretin-2 Saporin Lesions of the Ventrolateral Periaquaductal Gray (vlPAG) Increase REM Sleep in Hypocretin Knockout Mice

Ten years ago the sleep disorder narcolepsy was linked to the neuropeptide hypocretin (HCRT), also known as orexin. This disorder is characterized by excessive day time sleepiness, inappropriate triggering of rapid-eye movement (REM) sleep and cataplexy, which is a sudden loss of muscle tone during waking. It is still not known how HCRT regulates REM sleep or muscle tone since HCRT neurons are localized only in the lateral hypothalamus while REM sleep and muscle atonia are generated from the brainstem. To identify a potential neuronal circuit, the neurotoxin hypocretin-2-saporin (HCRT2-SAP) was used to lesion neurons in the ventral lateral periaquaductal gray (vlPAG). The first experiment utilized hypocretin knock-out (HCRT-ko) mice with the expectation that deletion of both HCRT and its target neurons would exacerbate narcoleptic symptoms. Indeed, HCRT-ko mice (n = 8) given the neurotoxin HCRT2-SAP (16.5 ng/23nl/sec each side) in the vlPAG had levels of REM sleep and sleep fragmentation that were considerably higher compared to HCRT-ko given saline (+39%; n = 7) or wildtype mice (+177%; n = 9). However, cataplexy attacks did not increase, nor were levels of wake or non-REM sleep changed. Experiment 2 determined the effects in mice where HCRT was present but the downstream target neurons in the vlPAG were deleted by the neurotoxin. This experiment utilized an FVB-transgenic strain of mice where eGFP identifies GABA neurons. We verified this and also determined that eGFP neurons were immunopositive for the HCRT-2 receptor. vlPAG lesions in these mice increased REM sleep (+79% versus saline controls) and it was significantly correlated (r = 0.89) with loss of eGFP neurons. These results identify the vlPAG as one site that loses its inhibitory control over REM sleep, but does not cause cataplexy, as a result of hypocretin deficiency.