Exaggerated Nighttime Sleep and Defective Sleep Homeostasis in a Drosophila Knock-In Model of Human Epilepsy

Despite an established link between epilepsy and sleep behavior, it remains unclear how specific epileptogenic mutations affect sleep and subsequently influence seizure susceptibility. Recently, Sun et al. (2012) created a fly knock-in model of human generalized epilepsy with febrile seizures plus (GEFS+), a wide-spectrum disorder characterized by fever-associated seizing in childhood and lifelong affliction. GEFS+ flies carry a disease-causing mutation in their voltage-gated sodium channel (VGSC) gene and display semidominant heat-induced seizing, likely due to reduced GABAergic inhibitory activity at high temperature. Here, we show that at room temperature the GEFS+ mutation dominantly modifies sleep, with mutants exhibiting rapid sleep onset at dusk and increased nighttime sleep as compared to controls. These characteristics of GEFS+ sleep were observed regardless of sex, mating status, and genetic background. GEFS+ mutant sleep phenotypes were more resistant to pharmacologic reduction of GABA transmission by carbamazepine (CBZ) than controls, and were mitigated by reducing GABA_A receptor expression specifically in wake-promoting pigment dispersing factor (PDF) neurons. These findings are consistent with increased GABAergic transmission to PDF neurons being mainly responsible for the enhanced nighttime sleep of GEFS+ mutants. Additionally, analyses under other light conditions suggested that the GEFS+ mutation led to reduced buffering of behavioral responses to light on and off stimuli, which contributed to characteristic GEFS+ sleep phenotypes. We further found that GEFS+ mutants had normal circadian rhythms in free-running dark conditions. Interestingly, the mutants lacked a homeostatic rebound following mechanical sleep deprivation, and whereas deprivation treatment increased heat-induced seizure susceptibility in control flies, it unexpectedly reduced seizure activity in GEFS+ mutants. Our study has revealed the sleep architecture of a Drosophila VGSC mutant that harbors a human GEFS+ mutation, and provided unique insight into the relationship between sleep and epilepsy.     

Gap Junctions Mediate Glucose Transport Between GLUT1-Positive and -Negative Cells in the Spiral Limbus of the Rat Cochlea

To elucidate the role of the spiral limbus in glucose transport in the cochlea, we analyzed the expression and localization of GLUT1, connexin26, connexin30, and occludin in the spiral limbus of the rat cochlea. GLUT1 and occludin were detected in blood vessels. GLUT1, connexin26, connexin30, and occludin were also expressed in fibrocytes just basal to the supralimbal lining cells. Connexin26 and connexin30 were present among not only these GLUT1-positive fibrocytes but also GLUT1-negative fibrocytes. In vivo glucose imaging using 6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-6-deoxyglucose (6-NBDG, MW 342) together with Evans Blue Albumin (EBA, MW 68,000) showed that 6-NBDG was rapidly distributed throughout the spiral limbus, whereas EBA was localized only in the vessels. Moreover, the gap junctional uncoupler heptanol inhibited the distribution of 6-NBDG. These findings suggest that gap junctions play an important role in glucose transport in the spiral limbus, i.e., that gap junctions mediate glucose transport from GLUT1-positive fibrocytes to GLUT1-negative fibrocytes in the spiral limbus.     

MiR-376c Down-Regulation Accelerates EGF-Dependent Migration by Targeting GRB2 in the HuCCT1 Human Intrahepatic Cholangiocarcinoma Cell Line

MicroRNA miR-376c was expressed in normal intrahepatic biliary epithelial cells (HIBEpiC), but was significantly suppressed in the HuCCT1 intrahepatic cholangiocarcinoma (ICC) cell line. The biological significance of the down-regulation of miR-376c in HuCCT1 cells is unknown. We hypothesized that miR-376c could function as a tumor suppressor in these cells. To test this hypothesis, we sought the targets of miR-376c, and characterized the effect of its down-regulation on HuCCT1 cells. We performed proteomic analysis of miR-376c-overexpressing HuCCT1 cells to identify candidate targets of miR-376c, and validated these targets by 3′-UTR reporter assay. Transwell migration assays were performed to study the migratory response of HuCCT1 cells to miR-376c overexpression. Furthermore, microarrays were used to identify the signaling that were potentially involved in the miR-376c-modulated migration of HuCCT1. Finally, we assessed epigenetic changes within the potential promoter region of the miR-376c gene in these cells. Proteomic analysis and subsequent validation assays showed that growth factor receptor-bound protein 2 (GRB2) was a direct target of miR-376c. The transwell migration assay revealed that miR-376c significantly reduced epidermal growth factor (EGF)-dependent cell migration in HuCCT1 cells. DNA microarray and subsequent pathway analysis showed that interleukin 1 beta and matrix metallopeptidase 9 were possible participants in EGF-dependent migration of HuCCT1 cells. Bisulfite sequencing showed higher methylation levels of CpG sites upstream of the miR-376c gene in HuCCT1 relative to HIBEpiC cells. Combined treatment with the DNA-demethylating agent 5-aza-2′-deoxycytidine and the histone deacetylase inhibitor trichostatin A significantly upregulated the expression of miR-376c in HuCCT1 cells. We revealed that epigenetic repression of miR-376c accelerated EGF-dependent cell migration through its target GRB2 in HuCCT1 cells. These findings suggest that miR-376c functions as a tumor suppressor. Since metastasis is the major cause of death in ICC, microRNA manipulation could lead to the development of novel anti-cancer therapy strategies for ICC.     

Travellers as sentinels: Assaying the worldwide distribution of polymorphisms associated with artemisinin combination therapy resistance in Plasmodium falciparum using malaria cases imported into Scotland

There is growing evidence that Plasmodium falciparum parasites in southeastern Asia have developed resistance to artemisinin combination therapy. The resistance phenotype has recently been shown to be associated with four single nucleotide polymorphisms in the parasite’s genome. We assessed the prevalence of two of these single nucleotide polymorphisms in P. falciparum parasites imported into Scotland between 2009 and 2012, and in additional field samples from six countries in southeastern Asia. We analysed 28 samples from 11 African countries, and 25 samples from nine countries in Asia/southeastern Asia/Oceania. Single nucleotide polymorphisms associated with artemisinin combination therapy resistance were not observed outside Thailand and Cambodia.     

On computational efficiency of the hybrid Monte Carlo method applied to the multicanonical ensemble

Abstract

In the present paper, computational efficiency of the hybrid Monte Carlo (HMC) method applied to the multicanonical ensemble is studied; the HMC is an equation of motion guided Monte Carlo method. As in the standard HMC for the canonical ensemble, the multicanonical HMC calculations with high acceptance ratio show better efficiency; about 60% acceptance yields the best performance for the system examined.     

Optimization of 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidines to generate a highly selective PI3Kδ inhibitor

Chemical optimization of the 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidine (THPP) scaffold was conducted with a focus on cellular potency while maintaining high selectivity against PI3K isoforms. Compound 11f was identified as a potent, highly selective and orally available PI3Kδ inhibitor. In addition, 11f exhibited efficacy in an in vivo antibody production model. The desirable drug-like properties and in vivo efficacy of 11f suggest its potential as a drug candidate for the treatment of autoimmune diseases and leukocyte malignancies.     

A Food-Derived Flavonoid Luteolin Protects against Angiotensin II-Induced Cardiac Remodeling

Oxidative stress has been implicated in cardiac remodeling (cardiac fibrosis and hypertrophy), which impairs cardiac function and metabolism; therefore, it is anticipated antioxidative compounds will have protective properties against cardiac remodeling. Luteolin (3’,4’,5,7-tetrahydroxyflavone), a widely distributed flavonoid found in many herbal extracts including celery, green pepper, perilla leaves and seeds, and chamomile, is a known to be a potent antioxidant and was previously demonstrated to exert an antifibrotic effect in the lungs and the liver. In this study, we clearly demonstrate that oral pretreatment with the higher-luteolin diet (0.035% (wt/wt)) protected against cardiac fibrosis and hypertrophy as well as a hyperoxidative state in Ang II-infused rats. In cardiac tissue, increased gene expression levels of TGFβ1, CTGF, Nox2, Nox4, ANP, and BNP induced by Ang II were restored by oral pretreatment of this high-luteolin diet. In cultured rat cardiac fibroblasts, H₂O₂-induced TGFβ1 expression and the phosphorylation of JNK were suppressed by luteolin pretreatment. In conclusion, food-derived luteolin has protective actions against Ang II-induced cardiac remodeling, which could be mediated through attenuation of oxidative stress.