Effect of cholesterol depletion on the pore dilation of TRPV1

The TRPV1 ion channel is expressed in nociceptors, where pharmacological modulation of its function may offer a means of alleviating pain and neurogenic inflammation processes in the human body. The aim of this study was to investigate the effects of cholesterol depletion of the cell on ion-permeability of the TRPV1 ion channel. The ion-permeability properties of TRPV1 were assessed using whole-cell patch-clamp and YO-PRO uptake rate studies on a Chinese hamster ovary (CHO) cell line expressing this ion channel. Prolonged capsaicin-induced activation of TRPV1 with N-methyl-D-glucamine (NMDG) as the sole extracellular cation, generated a biphasic current which included an initial outward current followed by an inward current. Similarly, prolonged proton-activation (pH 5.5) of TRPV1 under hypocalcemic conditions also generated a biphasic current including a fast initial current peak followed by a larger second one. Patch-clamp recordings of reversal potentials of TRPV1 revealed an increase of the ion-permeability for NMDG during prolonged activation of this ion channel under hypocalcemic conditions. Our findings show that cholesterol depletion inhibited both the second current, and the increase in ion-permeability of the TRPV1 channel, resulting from sustained agonist-activation with capsaicin and protons (pH 5.5). These results were confirmed with YO-PRO uptake rate studies using laser scanning confocal microscopy, where cholesterol depletion was found to decrease TRPV1 mediated uptake rates of YO-PRO. Hence, these results propose a novel mechanism by which cellular cholesterol depletion modulates the function of TRPV1, which may constitute a novel approach for treatment of neurogenic pain.     

A Case Study of In Silico Modelling of Ciprofloxacin Hydrochloride/Metallic Compound Interactions

With the development of physiologically based absorption models, there is an increased scientific and regulatory interest in in silico modelling and simulation of drug–drug and drug–food interactions. Clinically significant interactions between ciprofloxacin and metallic compounds are widely documented. In the current study, a previously developed ciprofloxacin-specific in silico absorption model was employed in order to simulate ciprofloxacin/metallic compound interaction observed in vivo. Commercially available software GastroPlus™ (Simulations Plus Inc., USA) based on the ACAT model was used for gastrointestinal (GI) simulations. The required input parameters, relating to ciprofloxacin hydrochloride physicochemical and pharmacokinetic characteristics, were experimentally determined, taken from the literature or estimated by GastroPlus™. Parameter sensitivity analysis (PSA) was used to assess the importance of selected input parameters (solubility, permeability, stomach and small intestine transit time) in predicting percent drug absorbed. PSA identified solubility and permeability as critical parameters affecting the rate and extent of ciprofloxacin absorption. Using the selected input parameters, it was possible to generate a ciprofloxacin absorption model, without/with metal cation containing preparations co-administration, which matched well the in vivo data available. It was found that reduced ciprofloxacin absorption in the presence of aluminium hydroxide, calcium carbonate or multivitamins/zinc was accounted for by reduced drug solubility. The impact of solubility–permeability interplay on ciprofloxacin absorption can be observed in the ciprofloxacin–aluminium interaction, while in ciprofloxacin–calcium and ciprofloxacin–zinc interactions, effect of solubility was more pronounced. The results obtained indicate that in silico model developed can be successfully used to complement relevant in vitro studies in the simulation of physicochemical ciprofloxacin/metallic compound interactions.KEY WORDS: absorption profile, drug interaction, GastroPlus, permeability, solubility     

NLRP7, a nucleotide oligomerization domain-like receptor protein, is required for normal cytokine secretion and co-localizes with Golgi and the microtubule-organizing center

A hydatidiform mole (HM) is a human pregnancy with hyperproliferative placenta and abnormal embryonic development. Mutations in NLRP7, a member of the nucleotide oligomerization domain-like receptor family of proteins with roles in inflammation and apoptosis, are responsible for recurrent HMs. However, little is known about the functional role of NLRP7. Here, we demonstrate that peripheral blood mononuclear cells from patients with NLRP7 mutations and rare variants secrete low levels of IL-1β and TNF in response to LPS. We show that the cells from patients, carrying mutations or rare variants, have variable levels of increased intracellular pro-IL-1β indicating that normal NLRP7 down-regulates pro-IL-1β synthesis in response to LPS. Using transient transfections, we confirm the role of normal NLRP7 in inhibiting pro-IL-1β and demonstrate that this inhibitory function is abolished by protein-truncating mutations after the Pyrin domain. Within peripheral blood mononuclear cells, NLRP7 co-localizes with the Golgi and the microtubule-organizing center and is associated with microtubules. This suggests that NLRP7 mutations may affect cytokine secretion by interfering, directly or indirectly, with their trafficking. We propose that the impaired cytokine trafficking and secretion caused by NLRP7 defects makes the patients tolerant to the growth of these earlier arrested conceptions with no fetal vessels and that the retention of these conceptions until the end of the first trimester contribute to the molar phenotype. Our data will impact our understanding of postmolar choriocarcinomas, the only allograft non-self tumors that are able to invade maternal tissues.     

CoMEt: a statistical approach to identify combinations of mutually exclusive alterations in cancer

Cancer is a heterogeneous disease with different combinations of genetic alterations driving its development in different individuals. We introduce CoMEt, an algorithm to identify combinations of alterations that exhibit a pattern of mutual exclusivity across individuals, often observed for alterations in the same pathway. CoMEt includes an exact statistical test for mutual exclusivity and techniques to perform simultaneous analysis of multiple sets of mutually exclusive and subtype-specific alterations. We demonstrate that CoMEt outperforms existing approaches on simulated and real data. We apply CoMEt to five different cancer types, identifying both known cancer genes and pathways, and novel putative cancer genes.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0700-7) contains supplementary material, which is available to authorized users.     

Monitoring ungulates in steep non-forest habitat: a comparison of faecal pellet and helicopter counts

Faecal pellet counts have been widely used to monitor the abundances of introduced ungulates in New Zealand, but ground-based sampling cannot be conducted safely in the steep non-forest habitats that are common in New Zealand's Southern Alps. Helicopter counts may be an effective technique for monitoring ungulates in steep non-forest habitat. We evaluated the relationship between faecal pellet and helicopter counts of ungulates (primarily feral goat Capra hircus) at 12 non-forest sites in the Southern Alps. Within each site we counted the numbers of ungulates from a helicopter on three occasions and the number of intact faecal pellets along 30 transects. Mean observed densities of feral goats derived from helicopter counts ranged from 0.0 to 20.2 km^?2. There was a positive curvilinear (concave down) relationship between faecal pellet and helicopter counts. Compared with faecal pellet counts, helicopter counts were cheaper, could identify ungulate species and provided estimates of absolute density. Helicopter counts are a cost-effective method for monitoring ungulates in the steep non-forest habitats of New Zealand's Southern Alps.     

The effects of nimodipine and L-NAME on coronary flow and oxidative stress parameters in isolated rat heart

The aim of this study was to assess the effects of Ca2+ channel antagonist nimodipine (in concentration which competitive inhibited phosphodiesterase 1--PDE1) on oxidative stress alone or under inhibition of nitric oxide synthase by L-NAME in isolated rat heart. The hearts from male Wistar albino rats (n=18, BM about 200 g, age 8 weeks) were retrograde perfused according to the Langendorff technique at gradually increased constant perfusion pressure conditions (CPP, 40-120 cm H2O). The experiments were performed under control conditions, in the presence of Nimodipine (2 microM) or Nimodipine (2 microM) plus L-NAME (30 microM). Coronary flow (CF) varied in the autoregulatory range from 3.7 +/- 0.4 ml/min/g wt at 50 cm H2O to 4.35 +/- 0.79 at 90 cm H2O. Basal nitrite outflow, index of lipid peroxidation (measured as TBARS release) and superoxide anion release (O2-) (at 60 cm H2O) were 0.64 +/- 0.18 nmol/min/g wt, 0.55 +/- 0.13 micromol/min/g wt and 19.72 +/- 3.70 nmol/min/g wt, respectively. Nimodipine induced significant vasodilation at all values of CPP (from 26% at 40 cm H2O to 36% at 120 cm H2O) accompanied with significant decrease of nitrite outflow (from 59% at 40 cm H2O to 40% at 120 cm H2O), significant increase of TBARS above autoregulatory range (about 40%) and significant increase of O2- release (from 186% at 40 cm H2O to 117% at 120 cm H2O). However, perfusion with L-NAME completely reversed the effects of Nimodipine. Nimodipine-induced flow changes were decreased under L-NAME (from 3% at 40 cm H2O to 11% at 120 cm H2O) without changes in the autoregulatory range, accompanied with significantly increased nitrite outflow (from 69% at 40 cm H2O to 36% at 120 cm H2O) and TBARS release (almost 50%), as well as significantly decreased O2- release (from 50% at 40 cm H2O to 43% at 120 cm H20). Our findings show that effect of nimodipine on coronary flow should be significantly influenced by NO, TBARS and O2- release in isolated rat heart.     

LSID Tester,a tool for testing Life Science Identifier resolution services

Background  

Life Science Identifiers (LSIDs) are persistent, globally unique identifiers for biological objects. The decentralised nature of LSIDs makes them attractive for identifying distributed resources. Data of interest to biodiversity researchers (including specimen records, images, taxonomic names, and DNA sequences) are distributed over many different providers, and this community has adopted LSIDs as the identifier of choice.     

Characterization of 13 newly isolated strains of anaerobic, cellulolytic, thermophilic bacteria

Characteristics of 13 newly isolated thermophilic, anaerobic, and cellulolytic strains were compared with previously described strains of Clostridium thermocellum: ATCC 27405 and JW20 (ATCC 31549). Colony morphology, antibiotic sensitivity, fermentation end-products, and cellulose degradation were documented. All 13 strains were sensitive to erythromycin (5 μg/ml) and chloramphenicol (25 μg/ml), and all strains but one were sensitive to kanamycin (20 μg/ml). Polymerase chain reaction (PCR) amplification using primers based on gene sequences from C. thermocellum ATCC 27405 was successful for all 13 strains in the case of the hydrogenase gene and 11 strains in the case of phosphotransacetylase/acetate kinase genes. Ten strains amplified a product of the expected size with primers developed to be specific for C. thermocellum 16SrRNA primers. Two of the 13 strains did not amplify any product with the PCR primers designed for the phosphotransacetylase/acetate kinase and 16SrRNA primers. A MboI-like GATC- recognizing restriction activity was present in all of the five strains examined. The results of this study have several positive implications with respect to future development of a transformation system for cellulolytic thermophiles. Journal of Industrial Microbiology & Biotechnology (2001) 27, 275–280. Received 12 September 2000/ Accepted in revised form 20 November 2000     

Relationship of Psychiatric Diagnosis and Weight Loss Maintenance in Obese Breast Cancer Survivors

Objective: Obese breast cancer survivors are a unique population for weight loss counseling because both obesity and a diagnosis of breast cancer can increase the risk of depression. In this pilot study, weight loss maintenance was examined in obese breast cancer survivors with relationship to psychiatric diagnosis. Research Methods and Procedures: Forty‐eight subjects were enrolled. The intervention, which used individualized counseling for diet and exercise, lasted 24 months. After a 6‐month period of no contact with study subjects, a follow‐up body weight was obtained at 30 months. Results: The nine subjects who dropped out of the study before 12 months all failed to complete a structured psychiatric interview. Of the remaining 39 subjects, 9 had major depressive disorder, and 10 had a definable psychiatric disorder of lesser severity such as adjustment disorder. Subjects with any type of psychiatric diagnosis displayed significantly less weight loss at the 12‐month time‐point than those with no diagnosis (6.3% vs. 12.6% loss of baseline weight, respectively). At the 30‐month follow‐up visit, subjects with any psychiatric disorder had a mean weight loss of 1.2% of baseline weight compared with 7.8% weight loss in subjects with no diagnosis. Discussion: These results suggest that the presence of psychiatric disorders can interfere with weight loss. Therefore, recognition and treatment of psychiatric disorders may be important in attempts at weight reduction, and this will be especially important in populations such as cancer survivors, who seem to have higher rates of depression and other disorders than the general population.     

Combining Weight‐Loss Counseling with the Weight Watchers Plan for Obese Breast Cancer Survivors

Objective: The objective was to develop effective weight‐loss methods for women who have had breast cancer, because obesity may result in an adverse prognosis. Research Methods and Procedures: This randomized pilot study tested an individualized approach toward weight loss in obese women who have had a diagnosis of breast cancer. An individualized approach was applied either alone or combined with the commercial Weight Watchers program. Forty‐eight women (body mass index of 30 to 44 kg/m²) were enrolled. Results: Weight change after 12 months of intervention was as follows (mean ± SD): 0.85 ± 6.0 kg in the control group, ?2.6 ± 5.9 kg in the Weight Watchers group, ?8.0 ± 5.5 kg in the individualized group, and ?9.4 ± 8.6 kg in the comprehensive group that used both individualized counseling and Weight Watchers. Weight loss relative to control was statistically significant in the comprehensive group 3, 6, and 12 months after randomization, whereas weight loss in the individualized group was significant only at 12 months. Weight loss of 10% or more of initial body weight was observed in 6 of 10 women in the comprehensive group at 12 months. In the comprehensive and Weight Watchers–only groups, weight loss was significantly related to frequency of attendance at Weight Watchers meetings, and attendance was more frequent in the comprehensive group. Discussion: These data indicate that the most weight loss was achieved when the counseling approach combined both Weight Watchers and individualized contacts. This was effective even though most of the individualized contacts were by telephone.