Evaluation of acetylcholinesterase in an animal model of mania induced by d-amphetamine

The present study aims to investigate the effects of mood stabilizers, lithium (Li) and valproate (VPA), on acetylcholinesterase (AChE) activity in the brains of rats subjected to an animal model of mania induced by d-amphetamine (d-AMPH). In the reversal treatment, Wistar rats were first given d-AMPH or saline (Sal) for 14 days. Between days 8 and 14, the rats were treated with Li, VPA, or Sal. In the prevention treatment, rats were pretreated with Li, VPA, or Sal. AChE activity was measured in the brain structures (prefrontal cortex, hippocampus, and striatum). Li, alone in reversion and prevention treatments, increased AChE activity in the brains of rats. VPA, alone in prevention treatment, increased AChE activity in all brain regions evaluated; in the reversion, only in the prefrontal. However, d-AMPH decreased activity of AChE in the striatum of rats in both the reversion and prevention treatments. VPA was able to revert and prevent this AChE activity alteration in the rat striatum. Our findings further support the notion that the mechanisms of mood stabilizers also involve changes in AChE activity, thus reinforcing the need for more studies to better characterize the role of acetylcholine in bipolar disorder.     

Real-time quantitative PCR for assessment of antiviral drug effects against Epstein-Barr virus replication and EBV late mRNA expression

This study assesses the ability of quantitative real-time PCR to measure the effects of virus DNA polymerase inhibitors on EBV DNA and late mRNAs syntheses in EBV-producing cell lines. In-house real-time quantitative PCRs were used to measure EBV DNA (thymidine kinase) and mRNAs (BLLF1 gene/gp350/220, BVRF2 gene/protease) in P3HR-1 and B95-8 cells induced for EBV production by PMA and exposed to ganciclovir, cidofovir and foscarnet. The calculated 50% effective concentrations (EC(50)) for viral DNA replication inhibition in P3HR-1 cells after 7 days of drug exposure were 0.28+/-0.06, 0.29+/-0.01 and 13.6+/-0.17 microg/mL for ganciclovir, cidofovir and foscarnet, respectively. The EC(50) for B95-8 cells were 0.44+/-0.02, 0.70+/-0.06 and 46.8+/-0.5 microg/mL, respectively. The quantitation of the late viral mRNAs showed a decrease of 79-89% in the mRNA amount after 4 days of antiviral treatment. Nevertheless, a substantial amount of mRNA still remained detectable after drug exposure. The real-time PCR is an improvement in the attempt to simplify EBV DNA-quantitation for antiviral assays. The quantitation of late mRNA does not appear as more informative than DNA quantitation for the assessment of the DNA polymerase inhibitor activity, but it may be useful to assess the antiviral activity of drugs acting by another mechanism.     

Antioxidant treatment prevented late memory impairment in an animal model of sepsis

OBJECTIVE: Assess the effect of antioxidant treatment on late memory impairment and early hippocampus oxidative stress after cecal ligation and perforation. SUBJECTS: Male Wistar rats. INTERVENTIONS: Rats underwent sham operation or cecal ligation and perforation. Animals that underwent cecal ligation and perforation were divided into groups: 1) treated with basic support (50 mL/kg saline, 30 mg/kg ceftriaxone, and 25 mg/kg clindamycin every 6 hrs), 2) treated with basic support plus N-acetylcysteine (20 mg/kg N-acetylcysteine at 3, 6, 12, 18, and 24 hrs after cecal ligation and perforation), 3) treated with basic support plus deferoxamine (20 mg/kg deferoxamine at 3 and 24 hrs after cecal ligation and perforation), 4) treated with basic support plus N-acetylcysteine and deferoxamine, or 5) treated with N-acetylcysteine plus deferoxamine. MEASUREMENTS AND MAIN RESULTS: On days 10 and 30 after surgery, the animals underwent behavioral tasks: inhibitory avoidance task, habituation to an open field, and continuous multiple-trials step-down inhibitory avoidance task. The sepsis group showed significantly decreased performance in latency retention compared with the sham group in the inhibitory avoidance task. In the open-field task, the sepsis group presented memory impairment after sepsis. In the continuous multiple-trials step-down inhibitory avoidance task, the sepsis group showed a significant increase in the number of training trials required to reach the acquisition criterion. All these memory impairments were prevented by N-acetylcysteine plus deferoxamine treatment, but not its isolate use. In addition, the combined use of antioxidants attenuated oxidative damage in hippocampus 6 hrs after sepsis induction. CONCLUSIONS: Antioxidant treatment prevented the development of late cognitive deficits in an animal model of sepsis.     

Factors modulating the delivery and effect of enzymatic cargo conjugated with antibodies targeted to the pulmonary endothelium.

Vascular drug targeting may improve therapies, yet a thorough understanding of the factors that regulate effects of drugs directed to the endothelium is needed to translate this approach into the clinical domain. To define factors modulating the efficacy and effects of endothelial targeting, we used a model enzyme (glucose oxidase, GOX) coupled with monoclonal antibodies (anti-TM(34) or anti-TM(201)) to distinct epitopes of thrombomodulin, a surface determinant enriched in the pulmonary endothelium. GOX delivery results in conversion of glucose and oxygen into H(2)O(2) leading to lung damage, a clear physiologic endpoint. Results of in vivo studies in mice showed that the efficiency of cargo delivery and its effect are influenced by a number of factors including: 1) The level of pulmonary uptake of the targeting antibody (anti-TM(201) was more efficient than anti-TM(34)); 2) The amount of an active drug delivered to the target; 3) The amount of target antigen on the endothelium (animals with suppressed TM levels showed less targeting); and, 4) The substrate availability for the enzyme cargo in the target tissue (hyperoxia augmented GOX-induced injury). Therefore, both activities of the conjugates and biological factors control targeting and effects of enzymatic cargo. Understanding the nature of such "modulating biological factors" will hopefully allow optimization and ultimately applications of drug targeting for "individualized" pharmacotherapy.     

Risk of AKI with Gentamicin as Surgical Prophylaxis

In 2009, the Scottish government issued a target to reduce Clostridium difficile infection by 30% in 2 years. Consequently, Scottish hospitals changed from cephalosporins to gentamicin for surgical antibiotic prophylaxis. This study examined rates of postoperative AKI before and after this policy change. The study population comprised 12,482 adults undergoing surgery (orthopedic, urology, vascular, gastrointestinal, and gynecology) with antibiotic prophylaxis between October 1, 2006, and September 30, 2010 in the Tayside region of Scotland. Postoperative AKI was defined by the Kidney Disease Improving Global Outcomes criteria. The study design was an interrupted time series with segmented regression analysis. In orthopedic patients, change in policy from cefuroxime to flucloxacillin (two doses of 1 g) and single-dose gentamicin (4 mg/kg) was associated with a 94% increase in AKI (P=0.04; 95% confidence interval, 93.8% to 94.3%). Most patients who developed AKI after prophylactic gentamicin had stage 1 AKI, but some patients developed persistent stage 2 or stage 3 AKI. The antibiotic policy change was not associated with a significant increase in AKI in the other groups. Regardless of antibiotic regimen, however, rates of AKI were high (24%) after vascular surgery, and increased steadily after gastrointestinal surgery. Rates could only be ascertained in 52% of urology patients and 47% of gynecology patients because of a lack of creatinine testing. These results suggest that gentamicin should be avoided in orthopedic patients in the perioperative period. Our findings also raise concerns about the increasing prevalence of postoperative AKI and failures to consistently measure postoperative renal function.Reported rates of postoperative AKI vary because of the heterogeneity of the populations studied. Uncomplicated AKI is associated with a mortality of 10%, rising to 50% in the context of multiorgan failure and up to 80% if RRT is required.^1,2 It was thought that the presence of AKI was a marker of coexisting pathology that increased mortality risk, but recent reports demonstrate AKI as an independent risk factor for mortality.^3,4 The increasing incidence of AKI and its long-term consequences have significant socioeconomic and public health effects globally.⁵Clostridium difficile infection (CDI) is an important healthcare-associated infection. Antibiotic use increases the risk of CDI for at least 3 months⁶ and short courses of perioperative antibiotic prophylaxis have also been associated with an increased risk of CDI, particularly in the context of an established outbreak.⁷In 2009, the Scottish government issued a new target for all health boards to reduce CDI by at least 30% over 2 years.⁸ The Scottish Antimicrobial Prescribing Group also produced recommendations for all National Health Service (NHS) boards to restrict the use of antibiotics associated with a high risk of CDI.⁹ As part of a widespread antibiotic policy change at NHS Tayside, orthopedic antibiotic prophylaxis was changed from cefuroxime to gentamicin and flucloxacillin. After concerns raised by nephrologists and a small uncontrolled study in the Dumfries and Galloway region of Scotland that described an increased rate of AKI in patients after orthopedic surgery after this policy change,¹⁰ it was felt that further investigation was required.This study aimed to use robust methodology, in a larger, population-based study of adult patients undergoing orthopedic implant surgery, to evaluate the effect of the policy change on postoperative AKI. It is noteworthy that patients who underwent repair of a neck of femur (NOF) fracture received coamoxiclav as antibiotic prophylaxis after the policy change because of concerns raised by orthopedic surgeons with regard to administering gentamicin in this particular patient group. This analysis was then extended to evaluate postoperative AKI in other surgical specialties (urology, vascular, gastrointestinal, and gynecology) that had changed to a gentamicin-based regimen.     

ASIC proteins regulate smooth muscle cell migration

The purpose of the present study was to investigate Acid Sensing Ion Channel (ASIC) protein expression and importance in cellular migration. We recently demonstrated that Epithelial Na(+)Channel (ENaC) proteins are required for vascular smooth muscle cell (VSMC) migration; however, the role of the closely related ASIC proteins has not been addressed. We used RT-PCR and immunolabeling to determine expression of ASIC1, ASIC2, ASIC3 and ASIC4 in A10 cells. We used small interference RNA to silence individual ASIC expression and determine the importance of ASIC proteins in wound healing and chemotaxis (PDGF-bb)-initiated migration. We found ASIC1, ASIC2, and ASIC3, but not ASIC4, expression in A10 cells. ASIC1, ASIC2, and ASIC3 siRNA molecules significantly suppressed expression of their respective proteins compared to non-targeting siRNA (RISC) transfected controls by 63%, 44%, and 55%, respectively. Wound healing was inhibited by 10, 20, and 26% compared to RISC controls following suppression of ASIC1, ASIC2, and ASIC3, respectively. Chemotactic migration was inhibited by 30% and 45%, respectively, following suppression of ASIC1 and ASIC3. ASIC2 suppression produced a small, but significant, increase in chemotactic migration (4%). Our data indicate that ASIC expression is required for normal migration and may suggest a novel role for ASIC proteins in cellular migration.     

Fabrication and performance evaluation of new nanocomposite membranes based on sulfonated poly(phthalazinone ether ketone) for PEM fuel cells

The purpose of this work is to enhance the proton conductivity and fuel cell performance of sulfonated poly(phthalazinone ether ketone) (SPPEK) as a proton exchange membrane through the application of SrTiO₃ perovskite nanoparticles. Nanocomposite membranes based on SPPEK and SrTiO₃ perovskite nanoparticles were prepared via a casting method. The highest proton conductivity of nanocomposite membranes obtained was 120 mS cm⁻¹ at 90 °C and 95% RH. These enhancements could be related to the hygroscopic structure of SrTiO₃ perovskite nanoparticles and the formation of hydrogen bonds between nanoparticles and water molecules. The satisfactory power density, 0.41 W cm⁻² at 0.5 V and 85 °C, of the nanocomposite membrane (5 wt% content of nanoparticles) confirms their potential for application in the PEM fuel cells.

The purpose of this work is to enhance the proton conductivity and fuel cell performance of sulfonated poly(phthalazinone ether ketone) (SPPEK) as a proton exchange membrane through the application of SrTiO₃ perovskite nanoparticles.     

The Thiazole-5-Carboxamide GPS491 Inhibits HIV-1, Adenovirus,and Coronavirus Replication by Altering RNA Processing/Accumulation

Medicinal chemistry optimization of a previously described stilbene inhibitor of HIV-1, 5350150 (2-(2-(5-nitro-2-thienyl)vinyl)quinoline), led to the identification of the thiazole-5-carboxamide derivative (GPS491), which retained potent anti-HIV-1 activity with reduced toxicity. In this report, we demonstrate that the block of HIV-1 replication by GPS491 is accompanied by a drastic inhibition of viral gene expression (IC₅₀ ~ 0.25 µM), and alterations in the production of unspliced, singly spliced, and multiply spliced HIV-1 RNAs. GPS491 also inhibited the replication of adenovirus and multiple coronaviruses. Low µM doses of GPS491 reduced adenovirus infectious yield ~1000 fold, altered virus early gene expression/viral E1A RNA processing, blocked viral DNA amplification, and inhibited late (hexon) gene expression. Loss of replication of multiple coronaviruses (229E, OC43, SARS-CoV2) upon GPS491 addition was associated with the inhibition of viral structural protein expression and the formation of virus particles. Consistent with the observed changes in viral RNA processing, GPS491 treatment induced selective alterations in the accumulation/phosphorylation/function of splicing regulatory SR proteins. Our study establishes that a compound that impacts the activity of cellular factors involved in RNA processing can prevent the replication of several viruses with minimal effect on cell viability.