AAD-2004, a potent spin trapping molecule and microsomal prostaglandin E synthase-1 inhibitor, shows safety and efficacy in a mouse model of ALS

While free radicals and inflammation constitute major routes of neuronal injury occurring in neurodegenerative diseases, neither antioxidants nor nonsteroidal anti-inflammatory drugs (NSAIDs) have shown significant efficacy in human clinical trials. To explore the possibility that concurrent blockade of free radicals and PGE2-mediated inflammation might constitute a safe and effective therapeutic approach to certain neurodegenerative diseases, we have developed 2-hydroxy-5-[2-(4-trifluoromethylphenyl)-ethylaminobezoic acid (AAD-2004) as a derivative of aspirin. AAD-2004 completely removed free radicals at 50 nM as a potent spin trapping molecule and inhibited microsomal prostaglandin E synthase-1 (mPGES-1) with an IC50 of 230 nM. Oral administration of AAD-2004 blocked free radical formation, PGE2 formation, and microglial activation in the spinal motor neurons of SOD1G93A mice. As a consequence, AAD-2004 reduced autophagosome formation, axonopathy, and motor neuron degeneration, improving motor function and increasing life span. In these assays, AAD-2004 was superior to ibuprofen or riluzole. Gastric bleeding was not induced by AAD-2004 even at a dose 400-fold higher than that required to obtain maximal therapeutic efficacy in SOD1G93A mice. Targeting both mPGES-1 and free radicals may be a promising approach to reduce neurodegeneration in ALS and possibly other neurodegenerative diseases

Preparative Synthesis of dTDP-L-Rhamnose Through Combined Enzymatic Pathways

dTDP-L-rhamnose, an important precursor of O-antigen, was prepared on a large scale from dTMP by executing an one-pot reaction in which six enzymes are involved. Two enzymes, dTDP-4-keto-6-deoxy-D-glucose 3,5-epimerase and dTDP-4-keto-rhamnose reductase, responsible for the conversion of dTDP-4-keto-6-deoxy- D-glucose to dTDP-L-rhamnose, were isolated from their putative sequences in the genome of Mesorhizobium loti, functionally expressed in Escherichia coli, and their enzymatic activities were identified. The two enzymes were combined with an enzymatic process for dTDP-4- keto-6-deoxy-D-glucose involving TMP kinase, acetate kinase, dTDP-glucose synthase, and dTDP-glucose 4,6- dehydratase, which allowed us to achieve a preparative scale synthesis of dTDP-L-rhamnose using dTMP and glucose-1-phosphate as starting materials. About 82% yield of dTDP-L-rhamnose was obtained based on initial dTMP concentration at 20 mM dTMP, 1 mM ATP, 10 mM NADH, 60 mM acetyl phosphate, and 80 mM glucose-1- phosphate. From the reaction with 20 ml volume, approximately 180 mg of dTDP-L-rhamnose was obtained in an overall yield of 60% after two-step purification, that is, anion exchange chromatography and gel filtration for desalting. The purified product was identifiedbyHPLC, ESI-MS,andNMR,showingabout95%purity

Cloning, expression, purification, crystallization and X-ray crystallographic analysis of glucuronic acid dehy­drogenase from Chromohalobacter salexigens. Addendum

An addendum to the article by Ahn et al. [Acta Cryst. (2011) F67, 689–691]

Inhibition of T Cell Receptor Signal Transduction by Tyrosine Kinase–interacting Protein of Herpesvirus saimiri

T cells play a central role in orchestrating immunity against pathogens, particularly viruses. Thus, impairing T cell activation is an important strategy employed by viruses to escape host immune control. The tyrosine kinase–interacting protein (Tip) of the T lymphotropic Herpesvirus saimiri (HVS) is constitutively present in lipid rafts and interacts with cellular Lck tyrosine kinase and p80 endosomal protein. Here we demonstrate that, due to the sequestration of Lck by HVS Tip, T cell receptor (TCR) stimulation fails to activate ZAP70 tyrosine kinase and to initiate downstream signaling events. TCR ζ chains in Tip-expressing T cells were initially phosphorylated to recruit ZAP70 molecule upon TCR stimulation, but the recruited ZAP70 kinase was not subsequently phosphorylated, resulting in TCR complexes that were stably associated with inactive ZAP70 kinase. Consequently, Tip expression not only markedly inhibited TCR-mediated intracellular signal transduction but also blocked TCR engagement with major histocompatibility complexes on the antigen-presenting cells and immunological synapse formation. These results demonstrate that a lymphotropic herpesvirus has evolved a novel mechanism to deregulate T cell activation to disarm host immune surveillance. This process contributes to the establishment and maintenance of viral latency

Activation of PERK Signaling Attenuates Aβ-Mediated ER Stress

Alzheimer's disease (AD) is characterized by the deposition of aggregated beta-amyloid (Aβ), which triggers a cellular stress response called the unfolded protein response (UPR). The UPR signaling pathway is a cellular defense system for dealing with the accumulation of misfolded proteins but switches to apoptosis when endoplasmic reticulum (ER) stress is prolonged. ER stress is involved in neurodegenerative diseases including AD, but the molecular mechanisms of ER stress-mediated Aβ neurotoxicity still remain unknown. Here, we show that treatment of Aβ triggers the UPR in the SK-N-SH human neuroblastoma cells. Aβ mediated UPR pathway accompanies the activation of protective pathways such as Grp78/Bip and PERK-eIF2α pathway, as well as the apoptotic pathways of the UPR such as CHOP and caspase-4. Knockdown of PERK enhances Aβ neurotoxicity through reducing the activation of eIF2α and Grp8/Bip in neurons. Salubrinal, an activator of the eIF2α pathway, significantly increased the Grp78/Bip ER chaperone resulted in attenuating caspase-4 dependent apoptosis in Aβ treated neurons. These results indicate that PERK-eIF2α pathway is a potential target for therapeutic applications in neurodegenerative diseases including AD

predictive factors for longer operative times for thyroidectomy

Background/Objective: Conventional open thyroidectomy is considered as a safe surgery nowadays. However, surgeons sometimes encounter unexpected difficulty when performing thyroidectomies. The aim of this paper was to identify the predictors of a difficult thyroidectomy for the management of patients with papillary thyroid carcinoma. Methods: A database of patients who underwent open conventional thyroidectomy with cervical lymph node dissection after diagnosed papillary thyroid carcinoma between July 2008 and June 2013 was examined. In addition, the patients were subgrouped by difficult thyroidectomy (DT) and nondifficult thyroidectomy to determine the predictors of DT according to operation time. Clinicopathologic characteristics, surgical outcomes, and postoperative morbidities were investigated. Results: No between-group differences in clinicopathologic factors and postoperative complications, except for male sex (p < 0.001) and tumor size (p = 0.039), were noted. Male sex [odds ratio (OR) 4.158, 95% confidence interval (CI) 2.020–8.559, p = 0.043] and age < 45 years (OR 2.239, 95% CI 1.304–3.843, p = 0.003) were independent factors associated with DT in a multivariate logistic regression model. Elevated antithyroglobulin antibody (OR 1.004, 95% CI 1.000–1.008, p = 0.030) was a variable which is statistically significant, but not clinically significant. Discussion: Young age and male sex might be regarded as predictors of DT. Expecting DT before surgery might help surgeons, especially beginners, prevent troublesome situations

Molecular differentiation of common promoters in Salmonella class 1 integrons

The integron is a mobile gene element which harbors antibiotic-resistance gene cassettes capable of site-specific integration. Among the four known types of integrons, the class 1 integron has been associated with multidrug-resistance in pathogenic bacteria. These gene cassettes have been the focus of a series of studies. The gene cassettes share a common promoter, and their expression levels are affected not only by their proximity to the promoter, but also by the strength (weak, hybrid and strong) of the common promoter, P1, as well as the presence of the additional promoter, P2. In this study, we developed molecular methods for the differentiation of promoter structures using PCR, restriction enzyme analysis, and polyacrylamide gel electrophoresis, and have applied them to the characterization of class 1 integrons in 33 non-typhoidal Salmonella serotypes in Korea. Class 1 integrons were detected in four serotypes: S. Derby (SD), S. Istanbul (SI), S. Paratyphi B (SPB), and S. Livingstone (SL), and the amplicon sizes were 1.0 Kb (SD, SI and SPB) and 2.0 Kb (SL). All of the 1.0 kb amplicons harbored gene cassettes (aadA1 or aadA2), but the 2.0 kb amplicon harbored three (dhfrXII-orfF-aadA2) gene cassettes, which conferred streptomycin/spectinomycin (aadA) and trimethoprim (dhfr) resistances. Our promoter structure study revealed three types of promoters; strong P1 (SD), weak P1 (SPB and SL), and weak P1 + P2 (SI). In conclusion, the class 1 integrons were detected in Korean NTS, and their promoter structures were found to be variable. Therefore, our methods may prove helpful in terms of our understanding of molecular diversity, as well as the transmission of class 1 integrons and phenotype-genotype relationships in antibiotic-resistance.This work was supported by the Korea Research Foundation Grants KRF-005-E00076 and KRF-005-E00077