Stabilization of the cyclin-dependent kinase 5 activator,p35, by paclitaxel decreases beta-amyloid toxicity in cortical neurons

One hallmark of Alzheimer's disease (AD) is the formation of neurofibrillary tangles, aggregated paired helical filaments composed of hyperphosphorylated tau. Amyloid-beta (Abeta) induces tau hyperphosphorylation, decreases microtubule (MT) stability and induces neuronal death. MT stabilizing agents have been proposed as potential therapeutics that may minimize Abeta toxicity and here we report that paclitaxel (taxol) prevents cell death induced by Abeta peptides, inhibits Abeta-induced activation of cyclin-dependent kinase 5 (cdk5) and decreases tau hyperphosphorylation. Taxol did not inhibit cdk5 directly but significantly blocked Abeta-induced calpain activation and decreased formation of the cdk5 activator, p25, from p35. Taxol specifically inhibited the Abeta-induced activation of the cytosolic cdk5-p25 complex, but not the membrane-associated cdk5-p35 complex. MT-stabilization was necessary for neuroprotection and inhibition of cdk5 but was not sufficient to prevent cell death induced by overexpression of p25. As taxol is not permeable to the blood-brain barrier, we assessed the potential of taxanes to attenuate Abeta toxicity in adult animals using a succinylated taxol analog (TX67) permeable to the blood-brain barrier. TX67, but not taxol, attenuated the magnitude of both basal and Abeta-induced cdk5 activation in acutely dissociated cortical cultures prepared from drug treated adult mice. These results suggest that MT-stabilizing agents may provide a therapeutic approach to decrease Abeta toxicity and neurofibrillary pathology in AD and other tauopathies.     

Effect of methyl alcohol on the morphology and conformational characteristics of silk sericin

Effects of methyl alcohol on the morphology and conformational characteristics of silk sericin (SS) were studied. Scanning electron microscope showed that morphology of SS lyophilized was dramatically changed from sponge-like structure to spherical fine particle type. X-ray diffraction method, infrared spectroscopy, and differential scanning calorimetry showed that the conformation of SS was random coil structure regardless of the addition of methyl alcohol. On the other hand, circular dichroism showed that the molecular states of SS were more densely packed.     

Synthesis and biological evaluation of thymine nucleosides fused with 3',4'-tetrahydrofuran ring

The pyrimidine nucleosides fused with 3',4'-tetrahydrofuran ring were successfully synthesized, starting from 1,2;5,6-di-O-isopropylidene-D-glucose and assayed for antiviral activities against HIV-1, HIV-2, EMCV, Cox. B3 and VSV. Thymine analogue (5) and its corresponding 2'-deoxy analogue (6) exhibited high cytotoxicity instead of giving antiviral activities.     

Vasopressin-SV40 T antigen expression in transgenic mice induces brain tumor and lymphoma

In order to understand the importance of various cis-acting elements in regulating VP gene expression, transgenic mice regulated by VP constructs were produced containing 3.8 kb of the 5' flanking region and all the exons and introns in the mouse VP gene, which was fused at the end of exon 3 to an SV40 T antigen (Tag). In the transgenic mice by the pVPSV.IGR3.6 construct, all the six transgenic mice died at the age of 2-6 weeks. In the transgenic mice by pVPSV.IGR2.1, 21% of them had brain tumors at 5 weeks and 100% of the mice had brain tumors after 24 weeks. Histological analysis of the transgenic mice revealed primitive neuroectodermal tumors (PNET) in the brain and lymphoma in the spleen and lymph nodes. The phenotype differences between the two transgenic mice suggest that tissue-specific expression might be regulated by cis-acting elements in the 1.5-kb of the 3(') flanking region, which are not contained in pVPSV.IGR2.1. In conclusion, pVPSV.IGR2.1 mice will be a valuable mouse model system for investigating PNET tumorigenesis in the brain and lymphoma in the lymph nodes and spleen.     

Synthesis of 5-azacytidine nucleosides with rigid sugar moiety as potential antitumor agents

The bicyclic 3'-O,5'-C-methylene-linked and 2'-O,5'-C-methylene-linked 5-azacytidine derivatives were readily synthesized from 1,2;5,6-di-O-isopropylidene-D-glucose and evaluated against several cancer cell lines.     

Design and synthesis of A3 adenosine receptor ligands, 2'-fluoro analogues of Cl-IB-MECA

Synthesis of 2'-deoxy-2'-fluoro-N6-substituted adenosines as bioisosteres of Cl-IB-MECA and their binding affinities to A3 adenosine receptor are described.     

Effect of doxycycline-regulated ERp57 expression on specific thrombopoietin productivity of recombinant CHO cells

In an attempt to increase the specific thrombopoietin (TPO) productivity (q(TPO)) of recombinant Chinese hamster ovary (rCHO) cells (TPO-33), the effect of expression level of ERp57, an isoform of protein disulfide isomerase, on q(TPO) was investigated. To regulate ERp57 expression level, the Tet-Off system was first introduced in TPO-33 cells and stable Tet-Off cells (TPO-33-Tet-Off) were screened by the luciferase assay. The rCHO cells with a doxycycline-regulated ERp57 expression system (TPO-33-ERp57) were obtained by cotransfection of pTRE-ERp57 and pTK-Hyg expression vectors into TPO-33-Tet-Off cells and subsequent screening by Western blot analysis of ERp57 and an enzyme-linked immunosorbent assay of secreted TPO. Western blot analysis showed that ERp57 expression level in TPO-33-ERp57 cells could be regulated tightly by the addition of different concentrations of doxycycline to a culture medium. A doxycycline concentration of 1 microg/mL, which did not influence cell growth and TPO production of TPO-33-Tet-Off cells, was high enough to suppress the ERp57 expression to a basal level. Compared with the basal level, a 1.7-fold increase in ERp57 expression level was obtained in the absence of doxycycline. This increased expression level of ERp57 resulted in a 2.1-fold increase in q(TPO) without growth inhibition, probably as a result of the chaperone-like activity of ERp57 in CHO cells. Taken together, the results obtained here demonstrate that q(TPO) of rCHO cells can be increased by elevating the expression level of ERp57.     

Osteopontin in kainic acid-induced microglial reactions in the rat brain

The present study was performed to investigate the spatial and temporal expression of osteopontin (OPN) mRNA in the rat brain after kainic acid-induced seizures, and to determine whether this phenomenon is associated spatiotemporally with the microglial reaction. The expression of OPN mRNA was detected using an in situ hybridization technique and Northern blot analysis. Following intraperitoneal injection of kainic acid (10 mg/kg), OPN mRNA was expressed in selective vulnerable areas, including the hippocampus, thalamus, hypothalamus, amygdala, and entorhinal cortex. Comparison of the morphology and localization with the established microglial marker OX-42 in the adjacent sections positively identified the OPN-expressed cells as microglia. Furthermore, double labeling experiments revealed that OPN mRNA expression was confined to ameboid-like cells among microglia stained with GSI-B4, an another microglial marker. These findings from a rat model of seizure support the notion that OPN can be synthesized in a subpopulation of reactive microglial cells. It can therefore be assumed that in the response of the brain to excitotoxic injury, synthesis of OPN occurs generally in a subset of activated microglia.