Alternative Usage Research Articles

"Freedom herself is very agile, very co-dependent, and a lovely person": The School Identities of High School Aged Youth with Communication Differences

Keywordsadolescent identities, speech disabilities, high school structures, facilitated communication, alternative and augmentative communicationAbstractThree users of Facilitated Communication, between the ages were observed and interviewed (utilizing methods found in Biklen & Bogdan, 2007, along with their parents and speech pathologist, about their lives, in particular their school lives. The study was concerned with the intersection between disability identity, Facilitated Communication / alternative and augmentative communication usage, and adolescent identities with special focus on how this intersection impacts these students at school. Data was collected and coded utilizing theories both from Nancy Lesko's conceptualization of the construction of adolescence (2001) and Martha Sheridan's work on lifeworld identities (2008).The study reveals how inclusion and exclusion in and out of school impact student attitudes toward school in general and academic work specifically, as well as independence / interdependence / co-dependence issues.

Tumor-specific usage of alternative transcription start sites in colorectal cancer identified by genome-wide exon array analysis

BackgroundApproximately half of all human genes use alternative transcription start sites (TSSs) to control mRNA levels and broaden the transcriptional output in healthy tissues. Aberrant expression patterns promoting carcinogenesis, however, may arise from alternative promoter usage.ResultsBy profiling 108 colorectal samples using exon arrays, we identified nine genes (TCF12, OSBPL1A, TRAK1, ANK3, CHEK1, UGP2, LMO7, ACSL5, and SCIN) showing tumor-specific alternative TSS usage in both adenoma and cancer samples relative to normal mucosa. Analysis of independent exon array data sets corroborated these findings. Additionally, we confirmed the observed patterns for selected mRNAs using quantitative real-time reverse-transcription PCR. Interestingly, for some of the genes, the tumor-specific TSS usage was not restricted to colorectal cancer. A comprehensive survey of the nine genes in lung, bladder, liver, prostate, gastric, and brain cancer revealed significantly altered mRNA isoform ratios for CHEK1, OSBPL1A, and TCF12 in a subset of these cancer types.To identify the mechanism responsible for the shift in alternative TSS usage, we antagonized the Wnt-signaling pathway in DLD1 and Ls174T colorectal cancer cell lines, which remarkably led to a shift in the preferred TSS for both OSBPL1A and TRAK1. This indicated a regulatory role of the Wnt pathway in selecting TSS, possibly also involving TP53 and SOX9, as their transcription binding sites were enriched in the promoters of the tumor preferred isoforms together with their mRNA levels being increased in tumor samples.Finally, to evaluate the prognostic impact of the altered TSS usage, immunohistochemistry was used to show deregulation of the total protein levels of both TCF12 and OSBPL1A, corresponding to the mRNA levels observed. Furthermore, the level of nuclear TCF12 had a significant correlation to progression free survival in a cohort of 248 stage II colorectal cancer samples.ConclusionsAlternative TSS usage in colorectal adenoma and cancer samples has been shown for nine genes, and OSBPL1A and TRAK1 were found to be regulated in vitro by Wnt signaling. TCF12 protein expression was upregulated in cancer samples and correlated with progression free survival.

수송용 대체연료로서 바이오메탄의 잠재적 타당성 연구

Biogas production and utilization are an emerging alternative energy technology. Biogas is produced from the biological breakdown of organic matter through anaerobic digestion. Biogas can be utilized for various energy sectors such as space heating, electricity generation and vehicle fuel. Especially, to be utilized as vehicle fuel, raw biogas needs to be upgraded that is mainly the removal of carbon dioxide to increase the methane content up to more than 95 ~ 97 vol% in some cases, similar to the composition of fossil-based natural gas. Usage of Biogas as a fuel of vehicles have an effect of reducing <TEX>$CO_2$</TEX> emission compared to fossil fuels. Biomethane which is produced by upgrading of biogas is regarded as a good alternative energy and usage of clean energy is encouraged to deal with air pollution and waste management as well as production of clean energy. Recently, biogas projects for vehicle fuel are newly being launched and Korea government have also announced a plan for investment to develop biogas as a transport fuel. In this study, it is aimed to examine the potential feasibility of biomethane as a transport fuel. As a results, the status of biomethane, quality standard, quality characteristics, and upgrading technology of biogas were investigated to evaluate of biogas as a vehicle fuel of transportation.

RNA-binding protein HuR autoregulates its expression by promoting alternative polyadenylation site usage

RNA-binding protein HuR modulates the stability and translational efficiency of messenger RNAs (mRNAs) encoding essential components of the cellular proliferation, growth and survival pathways. Consistent with these functions, HuR levels are often elevated in cancer cells and reduced in senescent and quiescent cells. However, the molecular mechanisms that control HuR expression are poorly understood. Here we show that HuR protein autoregulates its abundance through a negative feedback loop that involves interaction of the nuclear HuR protein with a GU-rich element (GRE) overlapping with the HuR major polyadenylation signal (PAS2). An increase in the cellular HuR protein levels stimulates the expression of long HuR mRNA species containing an AU-rich element (ARE) that destabilizes the mRNAs and thus reduces the protein production output. The PAS2 read-through occurs due to a reduced recruitment of the CstF-64 subunit of the pre-mRNA cleavage stimulation factor in the presence of the GRE-bound HuR. We propose that this mechanism maintains HuR homeostasis in proliferating cells. Since only the nuclear HuR is expected to contribute to the auto-regulation, our model may explain the longstanding observation that the increase in the total HuR expression in cancer cells often correlates with the accumulation of its substantial fraction in the cytoplasm.

Surveys of complementary and alternative medicine usage: A scoping study of the paediatric literature

To conduct a scoping review of paediatric CAM utilisation literature, in order to establish the depth and breadth of the field and identify gaps in knowledge regarding the epidemiology of CAM use. A CAM and paediatric search strategy was developed and run in three databases (Medline, Embase, Amed) in June 2006. It was revised and rerun in February 2010. Utilisation studies about paediatric CAM were selected if they had an English or French abstract. Data were extracted into an excel table by one individual. 152 paediatric CAM utilisation studies were identified as of February 2010, 136 were fulltext English. There was variation in how CAM was defined: vitamins were included in 35 studies, excluded in 12 studies and not mentioned in 40; while prayer was included in 64, excluded in 9, and not mentioned in 39. Over half of the studies identified were from North America. The majority of studies were conducted in conventional health settings. The four most commonly studied subpopulations were cancer, asthma, autistic spectrum disorder, and attention deficit hyperactivity disorder. Eighteen percent of studies did not report on the period of assessment used. Twenty-seven percent of studies inquired about adverse events and 32% inquired about costs and insurance coverage. There are substantial paediatric utilisation data available but some subpopulations remain poorly researched. The need for transparent utilisation data remains important in order to help prioritize safety and efficacy research.

Truncated IRAG variants modulate cGMP-mediated inhibition of human colonic smooth muscle cell contraction

Nitric oxide (NO) induces relaxation of colonic smooth muscle cells predominantly by cGMP/cGMP-dependent protein kinase I (cGKI)-induced phosphorylation of the inositol 1,4,5-trisphosphate receptor (IP(3)R)-associated cGMP kinase substrate (IRAG), to block store-dependent calcium signaling. In the present study we analyzed the structure and function of the human IRAG/MRVI1 gene. We describe four unique first exon variants transcribed from individual promoters in diverse human tissues. Tissue-specific alternative splicing with exon skipping and alternative splice donor and acceptor site usage further increases diversity of IRAG mRNA variants that encode for NH(2)- and COOH-terminally truncated proteins. At the functional level, COOH-terminally truncated IRAG variants lacking both the cGKI phosphorylation and the IP(3)RI interaction site counteract cGMP-mediated inhibition of calcium transients and relaxation of human colonic smooth muscle cells. Since COOH-terminally truncated IRAG mRNA isoforms are widely expressed in human tissues, our results point to an important role of IRAG variants as negative modulators of nitric oxide/cGKI-dependent signaling. The complexity of alternative splicing of the IRAG gene impressively demonstrates how posttranscriptional processing generates functionally distinct proteins from a single gene.

RNA Splicing Manipulation: Strategies to Modify Gene Expression for a Variety of Therapeutic Outcomes

Antisense oligomers initially showed promise as compounds to modify gene expression, primarily through RNaseH induced degradation of the target transcript. Expansion of the field has led to new chemistries capable of invoking different mechanisms, including suppression of protein synthesis by translational blockade and gene silencing using short interfering RNAs. It is now apparent that the majority of the eukaryotic genome is transcribed and non-protein coding RNAs have been implicated in the regulation of gene expression at many levels. This review considers potential therapeutic applications of antisense oligomers to modify gene expression, primarily by interfering with the process of exon recognition and intron removal during gene transcript splicing. While suppression of gene expression will be necessary to address some conditions, it is likely that antisense oligomer splice modification will have extensive clinical application. Pre-mRNA splicing is a tightly co-ordinated, multifactorial process that can be disrupted by antisense oligomers in a highly specific manner to suppress aberrant splicing, remove exons to by-pass nonsense or frame-shifting mutations or influence exon selection to alter spliceoform ratios. Manipulation of splicing patterns has been applied to a diverse range of conditions, including b-thalassemia, Duchenne muscular dystrophy, spinal muscular atrophy and certain cancers. Alternative exon usage has been identified as a major mechanism for generating diversity from a limited repertoire of genes in higher eukaryotes. Considering that the majority of all human primary gene transcripts are reportedly alternatively spliced, intervention at the level of pre-mRNA processing is likely to become increasingly significant in the fight against genetic and acquired disorders.

Comparative Analysis of mRNA Isoform Expression in Cardiac Hypertrophy and Development Reveals Multiple Post-Transcriptional Regulatory Modules

Cardiac hypertrophy is enlargement of the heart in response to physiological or pathological stimuli, chiefly involving growth of myocytes in size rather than in number. Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the “fetal gene program”. Here, using a genome-wide approach we systematically defined genes and pathways regulated in short- and long-term cardiac hypertrophy conditions using mice with transverse aortic constriction (TAC), and compared them with those regulated at different stages of embryonic and postnatal development. In addition, exon-level analysis revealed widespread mRNA isoform changes during cardiac hypertrophy resulting from alternative usage of terminal or internal exons, some of which are also developmentally regulated and may be attributable to decreased expression of Fox-1 protein in cardiac hypertrophy. Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing. Moreover, we found 3′UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy. Taken together, our results comprehensively delineated gene and mRNA isoform regulation events in cardiac hypertrophy and revealed their relations to those in development, and suggested that modulation of mRNA isoform expression plays an importance role in heart remodeling under pressure overload.

An exploratory study of treated-bed nets in Timor-Leste: patterns of intended and alternative usage

BackgroundThe Timor-Leste Ministry of Health has recently finalized the National Malaria Control Strategy for 2010-2020. A key component of this roadmap is to provide universal national coverage with long-lasting insecticide-treated nets (LLINs) in support of achieving the primary goal of reducing both morbidity and mortality from malaria by 30% in the first three years, followed by a further reduction of 20% by end of the programme cycle in 2020 [1]. The strategic plan calls for this target to be supported by a comprehensive information, education and communication (IEC) programme; however, there is limited prior research into household and personal usage patterns to assist in the creation of targeted, effective, and socio-culturally specific behaviour change materials.MethodsNine separate focus group discussions (FGDs) were carried out in Dili, Manatuto, and Covalima districts, Democratic Republic of Timor-Leste, in July 2010.These focus groups primarily explored themes of perceived malaria risk, causes of malaria, net usage patterns within families, barriers to correct and consistent usage, and the daily experience of users (both male and female) in households with at least one net. Comprehensive qualitative analysis utilized open source analysis software.ResultsThe primary determinants of net usage were a widespread perception that nets could or should only be used by pregnant women and young children, and the availability of sufficient sleeping space under a limited number of nets within households. Both nuisance biting and disease prevention were commonly cited as primary motivations for usage, while seasonality was not a significant factor. Long-term net durability and ease of hanging were seen as key attributes in net design preference. Very frequent washing cycles were common, potentially degrading net effectiveness. Finally, extensive re-purposing of nets (fishing, protecting crops) was both reported and observed, and may significantly decrease availability of nighttime sleeping space for all family members if distributed nets do not remain within the household.ConclusionsEmphasizing that net usage is acceptable and important for all family members regardless of age or gender, and addressing the complex behavioural economics of alternative net usages could have significant impacts on malaria control efforts in Timor-Leste, as the country's programmes make progress towards universal net coverage.

Identification of a novel Sp1 splice variant as a strong transcriptional activator

The transcription factor Sp1 regulates expression of numerous genes involved in many cellular processes. Different post-transcriptional modifications can influence the transcriptional control activity and stability of Sp1. In addition to these modifications, alternative splicing isoforms may also be the basis of its distinct functional activities. In this study, we identified a novel alternative splice isoform of Sp1 named Sp1c. This variant is generated by exclusion of a short domain, which we designate α, through alternative splice acceptor site usage in the exon 3. The existence of this new isoform was confirmed in vivo by Western blotting analysis. Although at very low levels, Sp1c is ubiquitously expressed, as seen in its full-length Sp1. A preliminary characterization of Sp1c shows that: (a) Sp1c works as stronger activator of transcription than full-length Sp1; (b) percentage of HEK293 Sp1c-overexpressing cells is higher in G1 phase and lower in S phase than percentage of HEK293 Sp1-overexpressing cells.

Estrogen receptor-alpha in the Alzheimer's disease brain

The decline in estrogen levels around the menopause was suggested to be an important factor in the pathogenesis of Alzheimer's disease (AD). Therefore, we investigated whether estrogen receptors (ER) may change in the brain of AD patients. ER alpha and its splice variants were studied at the mRNA level using polymerase chain reaction and DNA sequencing in the frozen human brain samples and at the protein level in the paraffin embedded sections using immunocytochemistry. Nuclear ER alpha expression was increased in the cholinergic basal forebrain (the nucleus basalis of Meynert (NBM) and the vertical limb of the diagonal band of Broca (VDB)) and in the hypothalamic nuclei but was diminished in the hippocampus of AD cases. These alterations in nuclear ER alpha staining could not be related to the occurrence of neurofibrillary tangles or to the changes in the local estrogen biosynthesis. We further identified 62 ER alpha mRNA splice variants in the brain of AD patients andelderly control donors and classified them according to the type of deletion and the presence ofinserts. Forty-eight of these ER alpha splice forms were found for the first time. The average number of the ER alpha mRNA splice variants per brain area was significantly lower in the AD cases compared with control donors. The incidence of large deletions with alternative usage of 5' and 3' splice sites inside exons was more prevalent in AD female cases. Immunoreactivity of the hippocampal ER alpha splice variant TADDI was decreased in the hippocampus, the NBM and the tuberomamillary nucleus of the hypothalamus of AD women. Our data show that alternative splicing of the ER alpha mRNA is reduced in the AD brain and more significantly in AD female cases. High incidence of the dominant negative del.7 isoform in the brain of the elderly suggests diminished estrogen signaling through both canonical ER alpha and beta. It is plausible that the brain area-specific alterations in nuclear ER alpha may be related to the differential degree of the tau pathology in the hypothalamus, basal forebrain and hippocampus that may affect the intracellular trafficking of the receptor.

Extensive relationship between antisense transcription and alternative splicing in the human genome

To analyze the relationship between antisense transcription and alternative splicing, we developed a computational approach for the detection of antisense-correlated exon splicing events using Affymetrix exon array data. Our analysis of expression data from 176 lymphoblastoid cell lines revealed that the majority of expressed sense-antisense genes exhibited alternative splicing events that were correlated to the expression of the antisense gene. Most of these events occurred in areas of sense-antisense (SAS) gene overlap, which were significantly enriched in both exons and nucleosome occupancy levels relative to nonoverlapping regions of the same genes. Nucleosome occupancy was highly correlated with Pol II abundance across overlapping regions and with concomitant increases in local alternative exon usage. These results are consistent with an antisense transcription-mediated mechanism of splicing regulation in normal human cells. A comparison of the prevalence of antisense-correlated splicing events between individuals of Mormon versus African descent revealed population-specific events that may indicate the continued evolution of new SAS loci. Furthermore, the presence of antisense transcription was correlated to alternative splicing across multiple metazoan species, suggesting that it may be a conserved mechanism contributing to splicing regulation.

A Novel Cold-Active Lipase from Candida albicans: Cloning, Expression and Characterization of the Recombinant Enzyme

A novel lipase gene lip5 from the yeast Candida albicans was cloned and sequenced. Alignment of amino acid sequences revealed that 86–34% identity exists with lipases from other Candida species. The lipase and its mutants were expressed in the yeast Pichia pastoris, where alternative codon usage caused the mistranslation of 154-Ser and 293-Ser as leucine. 154-Ser to leucine resulted in loss of expression of Lip5, and 293-Ser to leucine caused a marked reduction in the lipase activity. Lip5-DM, which has double mutations that revert 154 and 293 to serine residues, showed good lipase activity, and was overexpressed and purified by (NH4)2SO4 precipitation and ion-exchange chromatography. The pure Lip5-DM was stable at low temperatures ranging from 15–35 °C and pH 5–9, with the optimal conditions being 15–25 °C and pH 5–6. The activation energy of recombinant lipase was 8.5 Kcal/mol between 5 and 25 °C, suggesting that Lip5-DM was a cold–active lipase. Its activity was found to increase in the presence of Zn2+, but it was strongly inhibited by Fe2+, Fe3+, Hg2+ and some surfactants. In addition, the Lip5-DM could not tolerate water-miscible organic solvents. Lip5-DM exhibited a preference for the short-and medium-chain length p-nitrophenyl (C4 and C8 acyl group) esters rather than the long chain length p-nitrophenyl esters (C12, C16 and C18 acyl group) with highest activity observed with the C8 derivatives. The recombinant enzyme displayed activity toward triacylglycerols, such as olive oil and safflower oil.

Chromosome-wide mapping of DNA methylation patterns in normal and malignant prostate cells reveals pervasive methylation of gene-associated and conserved intergenic sequences

BackgroundDNA methylation has been linked to genome regulation and dysregulation in health and disease respectively, and methods for characterizing genomic DNA methylation patterns are rapidly emerging. We have developed/refined methods for enrichment of methylated genomic fragments using the methyl-binding domain of the human MBD2 protein (MBD2-MBD) followed by analysis with high-density tiling microarrays. This MBD-chip approach was used to characterize DNA methylation patterns across all non-repetitive sequences of human chromosomes 21 and 22 at high-resolution in normal and malignant prostate cells.ResultsExamining this data using computational methods that were designed specifically for DNA methylation tiling array data revealed widespread methylation of both gene promoter and non-promoter regions in cancer and normal cells. In addition to identifying several novel cancer hypermethylated 5' gene upstream regions that mediated epigenetic gene silencing, we also found several hypermethylated 3' gene downstream, intragenic and intergenic regions. The hypermethylated intragenic regions were highly enriched for overlap with intron-exon boundaries, suggesting a possible role in regulation of alternative transcriptional start sites, exon usage and/or splicing. The hypermethylated intergenic regions showed significant enrichment for conservation across vertebrate species. A sampling of these newly identified promoter (ADAMTS1 and SCARF2 genes) and non-promoter (downstream or within DSCR9, C21orf57 and HLCS genes) hypermethylated regions were effective in distinguishing malignant from normal prostate tissues and/or cell lines.ConclusionsComparison of chromosome-wide DNA methylation patterns in normal and malignant prostate cells revealed significant methylation of gene-proximal and conserved intergenic sequences. Such analyses can be easily extended for genome-wide methylation analysis in health and disease.

Cover Picture: Protein folding in vivo

AbstractCover illustration: Protein folding in vivo. This special issue, edited by Marc Blondel (Brest, France) and Mónica Marín (Montevideo, Uruguay) includes review and original articles on co‐translational folding, alternative codon usage and the impacts of protein folding on protein production. The drawing on the cover illustrates the ribosome as a platform for co‐translational folding of newly synthesized proteins. © HRATCH ARBACH 2011

Abstract 3428: Coordinated regulation of mesenchymal epithelial transition in the PMC42-LA breast cancer cell line variant

Abstract PMC42 cells were derived from a breast cancer pleural effusion and exhibit stem cell-like features when first characterized (Whitehead, Bertoncello et al. 1983; Whitehead, Monaghan et al. 1983). The PMC42-LA subline (Ackland, Michalczyk et al. 2001; Ackland, Newgreen et al. 2003) is distinctly less mesenchymal than parental PMC42-ET cells, but both exhibit EGF- and TGFβ-inducible mesenchymal-like change. Together they provide an ideal model in which to study the regulation of epithelial-mesenchymal plasticity in a breast cancer context. We have assessed in vitro parameters including migration, 3D colony formation and EMT marker expression, and undertook both miRNA profiling (mirVana probe set V1, Ambion) and alternative splice usage (Affymetrix human 1.0 ST whole genome exon arrays) analysis. Several miRNAs were expressed differently in the two sublines and/or up- or down-regulated in response to EMT-inducing treatments. Relative to the mesenchymal ET subline, the, miR-200 family members were elevated in the epithelial LA subline, where a corresponding loss of Zeb1 expression was seen. The LA subline also showed reduced E-Cadherin promoter methylation, and increased methylation of the Zeb1 promoter. Manipulation of the miR 200 family affected the EMT marker expression in these cells, as did Zeb1 suppression with shRNA. Matrigel invasion was also inhibited with Zeb1 knockdown. Additional gene expression changes and alternative splice usage, promoter methylation changes, and miR expression changes are under study. These studies may identify molecules and pathways that are important in cell specification across the epithelial mesenchymal axis, and represent diagnostic and therapeutic targets in breast cancer. Ackland, M. L., A. Michalczyk, et al. (2001). “PMC42, A novel model for the differentiated human breast.” Exp Cell Res 263(1): 14-22. Ackland, M. L., D. F. Newgreen, et al. (2003). “Epidermal growth factor-induced epithelio-mesenchymal transition in human breast carcinoma cells.” Lab Invest 83(3): 435-48. Whitehead, R. H., I. Bertoncello, et al. (1983). “A new human breast carcinoma cell line (PMC42) with stem cell characteristics. I. Morphologic characterization.” J Natl Cancer Inst 70(4): 649-61. Whitehead, R. H., P. Monaghan, et al. (1983). “A new human breast carcinoma cell line (PMC42) with stem cell characteristics. II. Characterization of cells growing as organoids.” J Natl Cancer Inst 71(6): 1193-203. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3428. doi:10.1158/1538-7445.AM2011-3428

Sustainability issues related to the engineering geology of long linear developments

The development of long linear structures such as roads, rail roads, tunnels, canals and pipelines often has unique engineering geology challenges. These include geological modeling, the identification of material strength and support factors, stability and risk issues, material excavation characteristics and the proposal of techniques for overcoming geotechnical problems, which are normally assessed as part of the conventional engineering geological investigation. An additional factor that is becoming increasingly important but is seldom included in investigations is the sustainability of the geotechnical inputs, in contrast to the sustainability of the project which is generally included. Sustainability issues revolve around the non-renewable nature of most construction resources and there is no doubt that the injudicious use of these construction materials and construction water is not sustainable in the long term: it is thus essential that the engineering geo-logical investigation should take cognizance of such issues and be adapted to provide the design engineer with the information that will maximize the sustainability options. This will also require a closer on-going relation-ship between the engineering geologist and the design engineer. This paper highlights significant sustainability issues (note that these differ from conventional environmental issues) and suggests some mitigating solutions. The sustainability issues discussed include primarily material and water usage, with some reference to energy conservation (mostly through alternative material usage and processing techniques and transportation).

The use of complementary and alternative medicine among chronic renal failure patients

The aim of this study is to evaluate the factors affecting the use and frequency of use of complementary and alternative medicine among chronic renal failure patients. The use of complementary and alternative medicine in the general population and patients with chronic renal failure has increased significantly. Despite this, there is limited information concerning the use of complementary and alternative medicine among chronic renal failure patients. Cross-sectional survey. The research was carried out at the nephrology and internal medicine outpatient clinics. Two hundred and six chronic renal failure patients admitted to the outpatient clinics were included in the study. Mean outcomes measures were the frequency and type of complementary and alternative medicine use, demographic and disease-related characteristics affecting complementary and alternative medicine use and the reasons for using complementary and alternative medicine. The data were evaluated by Pearson's chi-square test and Fisher's exact test. While 2·9% of the patients had been using complementary and alternative medicine before the renal disease occurred, 25·2% of the patients reported that they had at least once used complementary and alternative medicine methods after the renal disease occurred. A significant difference was found between complementary and alternative medicine usage and age, gender, place of living, occupational status and educational background (p < 0·05). While most of the patients using complementary and alternative medicine (78·3%) stated that they used such methods as a cure for their disease, 46·1% used body-mind techniques. The results of our study showed that one-fourth of the chronic renal failure patients were using complementary and alternative medicine, mainly body-mind techniques. In addition, the study proved that most of the patients do not discuss their complementary and alternative medicine usage with their doctors and nurses. It is essential that nephrology doctors and nurses should ask specific questions about complementary and alternative medicine usage while taking anamnesis on the patients' disease and nutritional status and that the nephrology team should expand their knowledge on complementary and alternative medicine methods to ensure patient and treatment safety.

Identification of C-terminally and N-terminally truncated estrogen receptor α variants in the mouse

We re-examined mouse ERα mRNA variants using rapid amplification of cDNA ends (RACE) and RT-PCR. Our analysis showed the presence of several mRNA variants containing unique 5′- or 3′-nucleotide sequences. We mapped the cDNA sequences on the mouse genome, and identified four novel 3′-terminal and 5′-leader exons in the intronic region between exons 4 and 5. RT-PCR analysis revealed that the expression patterns of the C-terminally truncated ERα products (CTERPs) were similar to that of Wild-type ERα and that the N-terminally truncated ERα products (NTERPs) appeared to have different expression profiles. Moreover, we constructed expression vectors and analyzed the subcellular localization and the transcriptional activation abilities of the variant proteins in transfected HEK293 cells using immunocytochemistry and luciferase reporter assay. The CTERP variants localized in the nuclei and constitutively activated estrogen response element (ERE)-driven promoters, while the NTERP variant was located in the extra-nuclear regions and had no ability to activate the ERE promoters in the presence or absence of 10 nM estradiol. Our results indicate that the mouse ERα gene is more complex than previously thought in terms of genomic organization and that alternative splicing and alternative usage of intronic promoters contribute to the remarkable diversity of ERα mRNAs and proteins.

Computational detection of alternative exon usage.

Background: With the advent of the GeneChip Exon Arrays, it is now possible to extract “exon-level” expression estimates, allowing for detection of alternative splicing events, one of the primary mechanisms of transcript diversity. In the context of (1) a complex trait use case and (2) a human cerebellum vs. heart comparison on previously validated data, we present a transcript-based statistical model and validation framework to allow detection of alternative exon usage (AEU) between different groups. To illustrate the approach, we detect and confirm differences in exon usage in the two of the most widely studied mouse genetic models (the C57BL/6J and DBA/2J inbred strains) and in a human dataset. Results: We developed a computational framework that consists of probe level annotation mapping and statistical modeling to detect putative AEU events, as well as visualization and alignment with known splice events. We show a dramatic improvement (∼25 fold) in the ability to detect these events using the appropriate annotation and statistical model which is actually specified at the transcript level, as compared with the transcript cluster/gene-level annotation used on the array. An additional component of this workflow is a probe index that allows ranking AEU candidates for validation and can aid in identification of false positives due to single nucleotide polymorphisms. Discussion: Our work highlights the importance of concordance between the functional unit interrogated (e.g., gene, transcripts) and the entity (e.g., exon, probeset) within the statistical model. The framework we present is broadly applicable to other platforms (including RNAseq).