Optimized intermolecular potential for nitriles based on Anisotropic United Atoms model

An extension of the Anisotropic United Atoms intermolecular potential model is proposed for nitriles. The electrostatic part of the intermolecular potential is calculated using atomic charges obtained by a simple Mulliken population analysis. The repulsion-dispersion interaction parameters for methyl and methylene groups are taken from transferable AUA4 literature parameters [Ungerer et al., J. Chem. Phys., 2000, 112, 5499]. Non-bonding Lennard-Jones intermolecular potential parameters are regressed for the carbon and nitrogen atoms of the nitrile group (–C≡N) from experimental vapor-liquid equilibrium data of acetonitrile. Gibbs Ensemble Monte Carlo simulations and experimental data agreement is very good for acetonitrile, and better than previous molecular potential proposed by Hloucha et al. [J. Chem. Phys., 2000, 113, 5401]. The transferability of the resulting potential is then successfully tested, without any further readjustment, to predict vapor-liquid phase equilibrium of propionitrile and n-butyronitrile

Fast, automated measurement of nematode swimming (thrashing) without morphometry

Background: The "thrashing assay", in which nematodes are placed in liquid and the frequency of lateral swimming ("thrashing") movements estimated, is a well-established method for measuring motility in the genetic model organism Caenorhabditis elegans as well as in parasitic nematodes. It is used as an index of the effects of drugs, chemicals or mutations on motility and has proved useful in identifying mutants affecting behaviour. However, the method is laborious, subject to experimenter error, and therefore does not permit high-throughput applications. Existing automation methods usually involve analysis of worm shape, but this is computationally demanding and error-prone. Here we present a novel, robust and rapid method of automatically counting the thrashing frequency of worms that avoids morphometry but nonetheless gives a direct measure of thrashing frequency. Our method uses principal components analysis to remove the background, followed by computation of a covariance matrix of the remaining image frames from which the interval between statistically-similar frames is estimated. Results: We tested the performance of our covariance method in measuring thrashing rates of worms using mutations that affect motility and found that it accurately substituted for laborious, manual measurements over a wide range of thrashing rates. The algorithm used also enabled us to determine a dose-dependent inhibition of thrashing frequency by the anthelmintic drug, levamisole, illustrating the suitability of the system for assaying the effects of drugs and chemicals on motility. Furthermore, the algorithm successfully measured the actions of levamisole on a parasitic nematode, Haemonchus contortus, which undergoes complex contorted shapes whilst swimming, without alterations in the code or of any parameters, indicating that it is applicable to different nematode species, including parasitic nematodes. Our method is capable of analyzing a 30 s movie in less than 30 s and can therefore be deployed in rapid screens. Conclusion: We demonstrate that a covariance-based method yields a fast, reliable, automated measurement of C. elegans motility which can replace the far more time-consuming, manual method. The absence of a morphometry step means that the method can be applied to any nematode that swims in liquid and, together with its speed, this simplicity lends itself to deployment in large-scale chemical and genetic screens. </p

AhR transcriptional activity in serum of Inuits across Greenlandic districts

<p>Abstract</p> <p>Background</p> <p>Human exposure to lipophilic persistent organic pollutants (POPs) including polychlorinated dibenzo-<it>p</it>-dioxins/furans (PCDDs/PCDFs), polychlorinated biphenyls (PCBs) and organochlorine pesticide is ubiquitous. The individual is exposed to a complex mixture of POPs being life-long beginning during critical developmental windows. Exposure to POPs elicits a number of species- and tissue-specific toxic responses, many of which involve the aryl hydrocarbon receptor (AhR). The aim of this study was to compare the actual level of integrated AhR transcriptional activity in the lipophilic serum fraction containing the actual POP mixture among Inuits from different districts in Greenland, and to evaluate whether the AhR transactivity is correlated to the bio-accumulated POPs and/or lifestyle factors.</p> <p>Methods</p> <p>The study included 357 serum samples from the Greenlandic districts: Nuuk and Sisimiut (South West Coast), Qaanaaq (North Coast) and Tasiilaq (East Coast). The bio-accumulated serum POPs were extracted by ethanol: hexane and clean-up on Florisil columns. Effects of the serum extract on the AhR transactivity was determined using the Hepa 1.12cR mouse hepatoma cell line carrying an AhR-luciferase reporter gene, and the data was evaluated for possible association to the serum levels of 14 PCB congeners, 10 organochlorine pesticide residues and/or lifestyle factors.</p> <p>Results</p> <p>In total 85% of the Inuit samples elicited agonistic AhR transactivity in a district dependent pattern. The median level of the AhR-TCDD equivalent (AhR-TEQ) of the separate genders was similar in the different districts. For the combined data the order of the median AhR-TEQ was Tasiilaq > Nuuk ≥ Sisimiut > Qaanaaq possibly being related to the different composition of POPs. In overall, the AhR transactivity was inversely correlated to the levels of sum POPs, age and/or intake of marine food.</p> <p>Conclusion</p> <p>i) We observed that the proportion of dioxin like (DL) compounds in the POP mixture was the dominating factor affecting the level of serum AhR transcriptional activity even at very high level of non DL-PCBs; ii) The inverse association between the integrated serum AhR transactivity and sum of POPs might be explained by the higher level of compounds antagonizing the AhR function probably due to selective POP bioaccumulation in the food chain.</p

Inhibition of E2-induced expression of BRCA1 by persistent organochlorines

BACKGROUND: Environmental persistent organochlorines (POCs) biomagnify in the food chain, and the chemicals are suspected of being involved in a broad range of human malignancies. It is speculated that some POCs that can interfere with estrogen receptor-mediated responses are involved in the initiation and progression of human breast cancer. The tumor suppressor gene BRCA1 plays a role in cell-cycle control, in DNA repair, and in genomic stability, and it is often downregulated in sporadic mammary cancers. The aim of the present study was to elucidate whether POCs have the potential to alter the expression of BRCA1. METHODS: Using human breast cancer cell lines MCF-7 and MDA-MB-231, the effect on BRCA1 expression of chemicals belonging to different classes of organochlorine chemicals (the pesticide toxaphene, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and three polychlorinated biphenyls [PCB#138, PCB#153 and PCB#180]) was measured by a reporter gene construct carrying 267 bp of the BRCA1 promoter. A twofold concentration range was analyzed in MCF-7, and the results were supported by northern blot analysis of BRCA1 mRNA using the highest concentrations of the chemicals. RESULTS: All three polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin reduced 17β-estradiol (E2)-induced expression as well as basal reporter gene expression in both cell lines, whereas northern blot analysis only revealed a downregulation of E2-induced BRCA1 mRNA expression in MCF-7 cells. Toxaphene, like E2, induced BRCA1 expression in MCF-7. CONCLUSION: The present study shows that some POCs have the capability to alter the expression of the tumor suppressor gene BRCA1 without affecting the cell-cycle control protein p21(Waf/Cip1). Some POCs therefore have the potential to affect breast cancer risk

Xenohormone transactivities are inversely associated to serum POPs in Inuit

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Determining Peptide Partitioning Properties via Computer Simulation

The transfer of polypeptide segments into lipid bilayers to form transmembrane helices represents the crucial first step in cellular membrane protein folding and assembly. This process is driven by complex and poorly understood atomic interactions of peptides with the lipid bilayer environment. The lack of suitable experimental techniques that can resolve these processes both at atomic resolution and nanosecond timescales has spurred the development of computational techniques. In this review, we summarize the significant progress achieved in the last few years in elucidating the partitioning of peptides into lipid bilayer membranes using atomic detail molecular dynamics simulations. Indeed, partitioning simulations can now provide a wealth of structural and dynamic information. Furthermore, we show that peptide-induced bilayer distortions, insertion pathways, transfer free energies, and kinetic insertion barriers are now accurate enough to complement experiments. Further advances in simulation methods and force field parameter accuracy promise to turn molecular dynamics simulations into a powerful tool for investigating a wide range of membrane active peptide phenomena

Can an hour or two of sun protection education keep the sunburn away? Evaluation of the Environmental Protection Agency's Sunwise School Program

BACKGROUND: Melanoma incidence is rising at a rate faster than any other preventable cancer in the United States. Childhood exposure to ultraviolet (UV) light increases risk for skin cancer as an adult, thus starting positive sun protection habits early may be key to reducing the incidence of this disease. METHODS: The Environmental Protection Agency's SunWise School Program, a national environmental and health education program for sun safety of children in primary and secondary schools (grades K-8), was evaluated with surveys administered to participating students and faculty. RESULTS: Pretests (n = 5,625) and posttests (n = 5,028) were completed by students in 102 schools in 42 states. Significant improvement was noted for the three knowledge variables. Intentions to play in the shade increased from 68% to 75%(p < 0.001) with more modest changes in intentions to use sunscreen. Attitudes regarding healthiness of a tan also decreased significantly. CONCLUSIONS: Brief, standardized sun protection education can be efficiently interwoven into existing school curricula, and result in improvements in knowledge and positive intentions for sun protection

The Rts1 Regulatory Subunit of Protein Phosphatase 2A Is Required for Control of G1 Cyclin Transcription and Nutrient Modulation of Cell Size

The key molecular event that marks entry into the cell cycle is transcription of G1 cyclins, which bind and activate cyclin-dependent kinases. In yeast cells, initiation of G1 cyclin transcription is linked to achievement of a critical cell size, which contributes to cell-size homeostasis. The critical cell size is modulated by nutrients, such that cells growing in poor nutrients are smaller than cells growing in rich nutrients. Nutrient modulation of cell size does not work through known critical regulators of G1 cyclin transcription and is therefore thought to work through a distinct pathway. Here, we report that Rts1, a highly conserved regulatory subunit of protein phosphatase 2A (PP2A), is required for normal control of G1 cyclin transcription. Loss of Rts1 caused delayed initiation of bud growth and delayed and reduced accumulation of G1 cyclins. Expression of the G1 cyclin CLN2 from an inducible promoter rescued the delayed bud growth in rts1Δ cells, indicating that Rts1 acts at the level of transcription. Moreover, loss of Rts1 caused altered regulation of Swi6, a key component of the SBF transcription factor that controls G1 cyclin transcription. Epistasis analysis revealed that Rts1 does not work solely through several known critical upstream regulators of G1 cyclin transcription. Cells lacking Rts1 failed to undergo nutrient modulation of cell size. Together, these observations demonstrate that Rts1 is a key player in pathways that link nutrient availability, cell size, and G1 cyclin transcription. Since Rts1 is highly conserved, it may function in similar pathways in vertebrates

An evaluation of potential reference genes for stability of expression in two salmonid cell lines after infection with either Piscirickettsia salmonis or IPNV

<p>Abstract</p> <p>Background</p> <p>Due to the limited number of species specific antibodies against fish proteins, differential gene expression analyses are vital for the study of host immune responses. Quantitative real-time reverse transcription PCR (qRT-PCR) is one of the most powerful tools for this purpose. Nevertheless, the accuracy of the method will depend on the careful selection of genes whose expression are stable and can be used as internal controls for a particular experimental setting.</p> <p>Findings</p> <p>The expression stability of five commonly used housekeeping genes [beta-actin (<it>ACTB</it>), elongation factor 1-alpha (<it>EF1A</it>), ubiquitin (<it>UBQ</it>), glyceraldehyd-3-phosphate dehydrogenase (<it>GAPDH</it>) and tubulin alpha (<it>TUBA</it>)] were monitored in salmonid cell lines CHSE-214 and RTS11 after infection with two of the most fastidious fish pathogens, the facultative bacterium <it>Piscirickettsia salmonis </it>and the aquabirnavirus IPNV (Infectious Pancreatic Necrosis Virus). After geNorm analysis, <it>UBQ </it>and <it>EF1A </it>appeared as the most stable, although <it>EF1A </it>was slightly upregulated at late stages of <it>P. salmonis </it>infection in RTS11. <it>ACTB </it>instead, showed a good performance in each case, being always considered within the three most stable genes of the panel. In contrast, infection-dependent differential regulation of <it>GAPDH </it>and <it>TUBA </it>was also demonstrated.</p> <p>Conclusion</p> <p>Based on the data presented here with the cell culture models CHSE-214 and RTS11, we suggest the initial choice of <it>UBQ</it>, <it>ACTB </it>and <it>EF1A </it>as reference genes in qRT-PCR assays for studying the effect of <it>P. salmonis </it>and IPNV on the host immune response.</p

Intratumoral heterogeneity of microRNA expression in rectal cancer

Introduction: An increasing number of studies have investigated microRNAs (miRNAs) as potential markers of diagnosis, treatment and prognosis. So far, agreement between studies has been minimal, which may in part be explained by intratumoral heterogeneity of miRNA expression. The aim of the present study was to assess the heterogeneity of a panel of selected miRNAs in rectal cancer, using two different technical approaches. Materials and Methods: The expression of the investigated miRNAs was analysed by real-time quantitative polymerase chain reaction (RT-qPCR) and in situ hybridization (ISH) in tumour specimens from 27 patients with T3-4 rectal cancer. From each tumour, tissue from three different luminal localisations was examined. Inter- and intra-patient variability was assessed by calculating intraclass correlation coefficients (ICCs). Correlations between RT-qPCR and ISH were evaluated using Spearman's correlation. Results: ICCsingle (one sample from each patient) was higher than 50% for miRNA-21 and miRNA-31. For miRNA-125b, miRNA-145, and miRNA-630, ICCsingle was lower than 50%. The ICCmean (mean of three samples from each patient) was higher than 50% for miRNA-21(RT-qPCR and ISH), miRNA-125b (RT-qPCR and ISH), miRNA-145 (ISH), miRNA-630 (RT-qPCR), and miRNA-31 (RT-qPCR). For miRNA-145 (RT-qPCR) and miRNA-630 (ISH), ICCmean was lower than 50%. Spearman correlation coefficients, comparing results obtained by RT-qPCR and ISH, respectively, ranged from 0.084 to 0.325 for the mean value from each patient, and from -0.085 to 0.515 in the section including the deepest part of the tumour. Conclusion: Intratumoral heterogeneity may influence the measurement of miRNA expression and consequently the number of samples needed for representative estimates. Our findings with two different methods suggest that one sample is sufficient for adequate assessment of miRNA-21 and miRNA-31, whereas more samples would improve the assessment of miRNA-125b, miRNA-145, and miRNA-630. Interestingly, we found a poor correlation between the expression estimates obtained by RT-qPCR and ISH, respectively