In applications of item response theory, assessment of model fit is a critical issue. Recently, limited-information goodness-of-fit testing has received increased attention in the psychometrics literature. In contrast to full-information test statistics such as Pearson’s X2 or the likelihood ratio G2, these limited-information tests utilise lower order marginal tables rather than the full contingency table. A notable example is Maydeu-Olivares and colleagues’ M2 family of statistics based on univariate and bivariate margins. When the contingency table is sparse, tests based on M2 retain better Type I error rate control than the full-information tests and can be more powerful. While in principle the M2 statistic can be extended to test hierarchical multidimensional item factor models (e.g., bifactor and testlet models), the computation is non-trivial. To obtain M2, a researcher often has to obtain (many thousands of) marginal probabilities, derivatives, and weights. Each of these must be approximated with high-dimensional numerical integration. We propose a dimension reduction method that can take advantage of the hierarchical factor structure so that the integrals can be approximated far more efficiently. We also propose a new test statistic that can be substantially better calibrated and more powerful than the original M2 statistic when the test is long and the items are polytomous. We use simulations to demonstrate the performance of our new methods and illustrate their effectiveness with applications to real data.
model fit; multidimensional item response theory; hierarchical item factor analysis; bifactor model; testlet response model; two-tier model; dimension reduction
Genotyping arrays are a cost effective approach when typing previously-identified genetic polymorphisms in large numbers of samples. One limitation of genotyping arrays with rare variants (e.g., minor allele frequency [MAF] <0.01) is the difficulty that automated clustering algorithms have to accurately detect and assign genotype calls. Combining intensity data from large numbers of samples may increase the ability to accurately call the genotypes of rare variants. Approximately 62,000 ethnically diverse samples from eleven Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) Consortium cohorts were genotyped with the Illumina HumanExome BeadChip across seven genotyping centers. The raw data files for the samples were assembled into a single project for joint calling. To assess the quality of the joint calling, concordance of genotypes in a subset of individuals having both exome chip and exome sequence data was analyzed. After exclusion of low performing SNPs on the exome chip and non-overlap of SNPs derived from sequence data, genotypes of 185,119 variants (11,356 were monomorphic) were compared in 530 individuals that had whole exome sequence data. A total of 98,113,070 pairs of genotypes were tested and 99.77% were concordant, 0.14% had missing data, and 0.09% were discordant. We report that joint calling allows the ability to accurately genotype rare variation using array technology when large sample sizes are available and best practices are followed. The cluster file from this experiment is available at www.chargeconsortium.com/main/exomechip.
AIM: To investigate functional duodenal abnormalities in functional dyspepsia (FD) and the role of serotonin (5-hydroxytryptamine, 5-HT) in mucosal ion transport and signalling.
METHODS: Duodenal mucosal biopsies were obtained from 15 patients with FD and 18 healthy controls. Immunohistochemistry was used to study the number of 5-HT-containing cells and real-time polymerase chain reaction for expression of 5-HT receptors 1A, 1B, 2A, 2B, 3A, 3B, 3C, 3D, 3E, 4 and 7, as well as expression of the serotonin re-uptake transporter (SERT) gene SLC6A4 and tryptophan hydroxylase 1 (TPH1). Biopsies were mounted in Ussing chambers for evaluation of basal and 5-HT-stimulated short-circuit current (SCC).
RESULTS: Conductance was lower in FD [42.4 ± 4.7 mS/cm2 (n = 15) vs 62.5 ± 4.5 mS/cm2 (n = 18), P = 0.005]. 5-HT induced a dose dependent rise in SCC in both FD (n = 8) and controls (n = 9), the rise was lower in FD (P < 0.001). Mean number of 5-HT stained cells per high power field was the same [34.4 ± 8.4 in FD (n = 15) and 30.4 ± 3.7 in controls (n = 18), P = 0.647]. The following genes were highly expressed: 5-HT receptor HTR3E, HTR4, HTR7, SERT gene (SLC6A4) and TPH1. Differences in expression levels were observed for HTR3E (higher expression in FD, P = 0.008), HTR7 (lower expression in FD, P = 0.027), SLC6A4 (higher expression in FD, P = 0.033) and TPH1 (lower expression in FD, P = 0.031).
CONCLUSION: Duodenal ion transport in response to exogenous 5-HT is abnormal in FD patients and associated with high expression of the HTR3E receptor and the serotonin transporter.
Dyspepsia; Ion transport; Gene expression; Duodenum; Epithelium; Serotonin
For the primary prevention of sudden cardiac death, guidelines provide left ventricular ejection fraction (EF) criteria for implantable cardioverter defibrillator (ICD) placement without specifying the technique by which it should be measured. We sought to investigate the potential impact of performing cardiovascular magnetic resonance (CMR) for EF on ICD eligibility.
The study population consisted of patients being considered for ICD implantation who were referred for EF assessment by CMR. Patients who underwent CMR within 30 days of echocardiography were included. Echocardiographic EF was determined by Simpson’s biplane method and CMR EF was measured by Simpson’s summation of discs method.
Fifty-two patients (age 62±15 years, 81% male) had a mean EF of 38 ± 14% by echocardiography and 35 ± 14% by CMR. CMR had greater reproducibility than echocardiography for both intra-observer (ICC, 0.98 vs 0.94) and inter-observer comparisons (ICC 0.99 vs 0.93). The limits of agreement comparing CMR and echocardiographic EF were – 16 to +10 percentage points. CMR resulted in 11 of 52 (21%) and 5 of 52 (10%) of patients being reclassified regarding ICD eligibility at the EF thresholds of 35 and 30% respectively. Among patients with an echocardiographic EF of between 25 and 40%, 9 of 22 (41%) were reclassified by CMR at either the 35 or 30% threshold. Echocardiography identified only 1 of the 6 patients with left ventricular thrombus noted incidentally on CMR.
CMR resulted in 21% of patients being reclassified regarding ICD eligibility when strict EF criteria were used. In addition, CMR detected unexpected left ventricular thrombus in almost 10% of patients. Our findings suggest that the use of CMR for EF assessment may have a substantial impact on management in patients being considered for ICD implantation.
Implantable cardioverter defibrillators; Ejection Fraction; Echocardiography; Cardiovascular Magnetic Resonance
Traditional estimators of item response theory (IRT) scale scores ignore uncertainty carried over from the item calibration process, which can lead to incorrect estimates of standard errors of measurement (SEM). Here, we review a variety of approaches that have been applied to this problem and compare them on the basis of their statistical methods and goals. We then elaborate on the particular flexibility and usefulness of a Multiple Imputation (MI) based approach, which can be easily applied to tests with mixed item types and multiple underlying dimensions. This proposed method obtains corrected estimates of individual scale scores, as well as their SEM. Furthermore, this approach enables a more complete characterization of the impact of parameter uncertainty by generating confidence envelopes (intervals) for item tracelines, test information functions, conditional SEM curves, and the marginal reliability coefficient. The MI based approach is illustrated through the analysis of an artificial data set, then applied to data from a large educational assessment. A simulation study was also conducted to examine the relative contribution of item parameter uncertainty to the variability in score estimates under various conditions. We found that the impact of item parameter uncertainty is generally quite small, though there are some conditions under which the uncertainty carried over from item calibration contributes substantially to variability in the scores. This may be the case when the calibration sample is small relative to the number of item parameters to be estimated, or when the IRT model fit to the data is multidimensional.
IRT; IRT scoring; calibration; standard error of measurement; multiple imputation; confidence envelope
Full-information item bifactor analysis is an important statistical method in psychological and educational measurement. Current methods are limited to single group analysis and inflexible in the types of item response models supported. We propose a flexible multiple-group item bifactor analysis framework that supports a variety of multidimensional item response theory models for an arbitrary mixing of dichotomous, ordinal, and nominal items. The extended item bifactor model also enables the estimation of latent variable means and variances when data from more than one group are present. Generalized user-defined parameter restrictions are permitted within or across groups. We derive an efficient full-information maximum marginal likelihood estimator. Our estimation method achieves substantial computational savings by extending Gibbons and Hedeker’s (1992) bifactor dimension reduction method so that the optimization of the marginal log-likelihood only requires two-dimensional integration regardless of the dimensionality of the latent variables. We use simulation studies to demonstrate the flexibility and accuracy of the proposed methods. We apply the model to study cross-country differences, including differential item functioning, using data from a large international education survey on mathematics literacy.
hierarchical factor model; item response theory; multidimensional IRT; item factor analysis; differential item functioning
The pathogenesis of colorectal neoplasia is still unresolved but has been associated with alterations in epithelial clearance of xenobiotics and metabolic waste products. The aim of this study was to functionally characterize the transport of cyclic nucleotides in colonic biopsies from patients with and without colorectal neoplasia.
Cyclic nucleotides were used as model substrates shared by some OATP- and ABC-transporters, which in part are responsible for clearance of metabolites and xenobiotics from the colonic epithelium. On colonic biopsies from patients with and without colorectal neoplasia, molecular transport was electrophysiologically registered in Ussing-chamber set-ups, mRNA level of selected transporters was quantified by rt-PCR, and subcellular location of transporters was determined by immunohistochemistry.
Of four cyclic nucleotides, dibuturyl-cAMP induced the largest short circuit current in both patient groups. The induced short circuit current was significantly lower in neoplasia-patients (p = 0.024). The observed altered transport of dibuturyl-cAMP in neoplasia-patients could not be directly translated to an observed increased mRNA expression of OATP4A1 and OATP2B1 in neoplasia patients. All other examined transporters were expressed to similar extents in both patient groups.
OATP1C1, OATP4A1, OATP4C1 seem to be involved in the excretory system of human colon. ABCC4 is likely to be involved from an endoplasmic-Golgi complex and basolateral location in goblet cells. ABCC5 might be directly involved in the turnover of intracellular cAMP at the basolateral membrane of columnar epithelial cells, while OATP2B1 is indirectly related to the excretory system. Colorectal neoplasia is associated with lower transport or sensitivity to cyclic nucleotides and increased expression of OATP2B1 and OATP4A1 transporters, known to transport PGE2.
Cyclic-AMP compartmentalization; Human colonic biopsy; OATP-; ABC-; PGE2-transporters
SNP genotyping arrays have been useful for many applications that require a large number of molecular markers such as high-density genetic mapping, genome-wide association studies (GWAS), and genomic selection. We report the establishment of a large maize SNP array and its use for diversity analysis and high density linkage mapping. The markers, taken from more than 800,000 SNPs, were selected to be preferentially located in genes and evenly distributed across the genome. The array was tested with a set of maize germplasm including North American and European inbred lines, parent/F1 combinations, and distantly related teosinte material. A total of 49,585 markers, including 33,417 within 17,520 different genes and 16,168 outside genes, were of good quality for genotyping, with an average failure rate of 4% and rates up to 8% in specific germplasm. To demonstrate this array's use in genetic mapping and for the independent validation of the B73 sequence assembly, two intermated maize recombinant inbred line populations – IBM (B73×Mo17) and LHRF (F2×F252) – were genotyped to establish two high density linkage maps with 20,913 and 14,524 markers respectively. 172 mapped markers were absent in the current B73 assembly and their placement can be used for future improvements of the B73 reference sequence. Colinearity of the genetic and physical maps was mostly conserved with some exceptions that suggest errors in the B73 assembly. Five major regions containing non-colinearities were identified on chromosomes 2, 3, 6, 7 and 9, and are supported by both independent genetic maps. Four additional non-colinear regions were found on the LHRF map only; they may be due to a lower density of IBM markers in those regions or to true structural rearrangements between lines. Given the array's high quality, it will be a valuable resource for maize genetics and many aspects of maize breeding.
Cardiac resynchronization therapy (CRT) can be a valuable treatment for heart failure. However, there are high nonresponse rates using current CRT inclusion criteria.
To assess the value of three-dimensional echocardiography (3DE) in predicting response to CRT.
Functional assessments and 3DE were performed in heart failure patients pre-CRT, 24 h post-CRT and six to 12 months after CRT. The dyssynchrony index (DI) was calculated as the SD of the time to minimum volume in 16 left ventricle segments corrected by heart rate. Response to CRT was defined as functional improvement (alive at late follow-up with improvement by one New York Heart Association class) and a decrease in left ventricular end-systolic volume by 15% or greater at six to 12 months follow-up.
A total of 53 patients were enrolled. Average 3DE acquisition time was less than 5 min. Seventy-two per cent of patients showed functional improvement, while 43% showed functional and echocardiographic evidence of response. Baseline DI and the decrease in DI at 24 h were both correlated with reverse remodelling. Responders had higher baseline DI values compared with nonresponders (mean 16.8 versus 7.1, P<0.001), and showed a greater decrease in DI values at 24 h (mean decrease 7.9 versus 0.7, P<0.001). All responders had baseline DI values of greater than 10 (negative predictive value of 100%). A decrease in the DI value by more than 5 at 24 h in patients with a baseline DI of greater than 10 identified responders with a positive predictive value of 83%.
3DE may be valuable in predicting response to CRT. A baseline DI cut-off of greater than 10 in our patients excluded reverse remodelling to CRT. In addition, the decrease in DI at 24 h had a high positive predictive value for long-term response to CRT.
Biventricular pacing; Cardiac resynchronization therapy; Dyssynchrony; Heart failure; Three-dimensional echocardiography; Ventricular remodelling
The extraordinary phenotypic diversity of dog breeds has been sculpted by a unique population history accompanied by selection for novel and desirable traits. Here we perform a comprehensive analysis using multiple test statistics to identify regions under selection in 509 dogs from 46 diverse breeds using a newly developed high-density genotyping array consisting of >170,000 evenly spaced SNPs. We first identify 44 genomic regions exhibiting extreme differentiation across multiple breeds. Genetic variation in these regions correlates with variation in several phenotypic traits that vary between breeds, and we identify novel associations with both morphological and behavioral traits. We next scan the genome for signatures of selective sweeps in single breeds, characterized by long regions of reduced heterozygosity and fixation of extended haplotypes. These scans identify hundreds of regions, including 22 blocks of homozygosity longer than one megabase in certain breeds. Candidate selection loci are strongly enriched for developmental genes. We chose one highly differentiated region, associated with body size and ear morphology, and characterized it using high-throughput sequencing to provide a list of variants that may directly affect these traits. This study provides a catalogue of genomic regions showing extreme reduction in genetic variation or population differentiation in dogs, including many linked to phenotypic variation. The many blocks of reduced haplotype diversity observed across the genome in dog breeds are the result of both selection and genetic drift, but extended blocks of homozygosity on a megabase scale appear to be best explained by selection. Further elucidation of the variants under selection will help to uncover the genetic basis of complex traits and disease.
There are hundreds of dog breeds that exhibit massive differences in appearance and behavior sculpted by tightly controlled selective breeding. This large-scale natural experiment has provided an ideal resource that geneticists can use to search for genetic variants that control these differences. With this goal, we developed a high-density array that surveys variable sites at more than 170,000 positions in the dog genome and used it to analyze genetic variation in 46 breeds. We identify 44 chromosomal regions that are extremely variable between breeds and are likely to control many of the traits that vary between them, including curly tails and sociality. Many other regions also bear the signature of strong artificial selection. We characterize one such region, known to associate with body size and ear type, in detail using “next-generation” sequencing technology to identify candidate mutations that may control these traits. Our results suggest that artificial selection has targeted genes involved in development and metabolism and that it may have increased the incidence of disease in dog breeds. Knowledge of these regions will be of great importance for uncovering the genetic basis of variation between dog breeds and for finding mutations that cause disease.
Forward and reverse genetics now allow researchers to understand embryonic and postnatal gene function in a broad range of species. Although some genetic mutations cause obvious morphological change, other mutations can be more subtle and, without adequate observation and quantification, might be overlooked. For the increasing number of genetic model organisms examined by the growing field of phenomics, standardized but sensitive methods for quantitative analysis need to be incorporated into routine practice to effectively acquire and analyze ever-increasing quantities of phenotypic data. In this study, we present platform-independent parameters for the use of microscopic x-ray computed tomography (microCT) for phenotyping species-specific skeletal morphology of a variety of different genetic model organisms. We show that microCT is suitable for phenotypic characterization for prenatal and postnatal specimens across multiple species.
volumetric x-ray computed tomography; microCT; phenomics; fetus; phenotyping; embryogenesis; imaging; mouse
The availability of array-based genotyping platforms for single nucleotide polymorphisms (SNPs) for the canine genome has expanded the opportunities to undertake genome-wide association (GWA) studies to identify the genetic basis for Mendelian and complex traits. Whole blood as the source of high quality DNA is undisputed but often proves impractical for collection of the large numbers of samples necessary to discover the loci underlying complex traits. Further, many countries prohibit the collection of blood from dogs unless medically necessary thereby restricting access to critical control samples from healthy dogs. Alternate sources of DNA, typically from buccal cytobrush extractions, while convenient, have been suggested to have low yield and perform poorly in GWA. Yet buccal cytobrushes provide a cost-effective means of collecting DNA, are readily accepted by dog owners, and represent a large resource base in many canine genetics laboratories. To increase the DNA quantities, whole genome amplification (WGA) can be performed. Thus, the present study assessed the utility of buccal-derived DNA as well as whole genome amplification in comparison to blood samples for use on the most recent iteration of the canine HD SNP array (Illumina).
In both buccal and blood samples, whether whole genome amplified or not, 97% of the samples had SNP call rates in excess of 80% indicating that the vast majority of the SNPs would be suitable to perform association studies regardless of the DNA source. Similarly, there were no significant differences in marker intensity measurements between buccal and blood samples for copy number variations (CNV) analysis.
All DNA samples assayed, buccal or blood, native or whole genome amplified, are appropriate for use in array-based genome-wide association studies. The concordance between subsets of dogs for which both buccal and blood samples, or those samples whole genome amplified, was shown to average >99%. Thus, the two DNA sources were comparable in the generation of SNP genotypes and intensity values to estimate structural variation indicating the utility for the use of buccal cytobrush samples and the reliability of whole genome amplification for genome-wide association and CNV studies.
Tumor initiation has been attributed to haploinsufficiency at a single locus for a large number of cancers. Patched1 (Ptc1) was one of the first such loci, and Ptc1 haploinsufficiency has been asserted to lead to medulloblastoma and rhabdomyosarcoma in mice.
To study the role of Ptc1 in cerebellar tumor development and to create a preclinical therapeutic platform, we have generated a conditional Ptc1 haploinsufficiency model of medulloblastoma by inactivating Ptc1 in Pax7-expressing cells of the cerebellum.
These mice developed exclusively medulloblastoma. We show that despite the presence of transcription of Ptc1, Ptc1 protein is nearly undetectable or absent in tumors. Our results suggest that Ptc1 loss of function is complete, but achieved at the protein level rather than by the classic genetic two-hit mechanism or a strict half-dosage genetic haploinsufficiency mechanism. Furthermore, we found that bortezomib, a 26S proteasome inhibitor, had a significant anti-tumor activity in vitro and in vivo, which was accompanied by restoration of Ptc1 protein and downregulation of the hedgehog signaling pathway. The same effect was seen for both human and mouse medulloblastoma tumor cell growth.
These results suggest that proteasome inhibition is a potential new therapeutic approach in medulloblastoma.
Medulloblastoma; Haploinsufficiency; Hedgehog; Patched1; Proteasome; Bortezomib
The pathogenesis for colorectal cancer remains unresolved. A growing body of evidence suggests a direct correlation between cyclooxygenase enzyme expression, prostaglandin E2 metabolism and neoplastic development. Thus further understanding of the regulation of epithelial functions by prostaglandin E2 is needed. We hypothesized that patients with colonic neoplasia have altered colonic epithelial ion transport and express functionally different prostanoid receptor levels in this respect.
Patients referred for colonoscopy were included and grouped into patients with and without colorectal neoplasia. Patients without endoscopic findings of neoplasia served as controls. Biopsy specimens were obtained from normally appearing mucosa in the sigmoid part of colon. Biopsies were mounted in miniaturized modified Ussing air-suction chambers. Indomethacin (10 μM), various stimulators and inhibitors of prostanoid receptors and ion transport were subsequently added to the chamber solutions. Electrogenic ion transport parameters (short circuit current and slope conductance) were recorded. Tissue pathology and tissue damage before and after experiments was assessed by histology.
Baseline short circuit current and slope conductance did not differ between the two groups. Patients with neoplasia were significantly more sensitive to indomethacin with a decrease in short circuit current of 15.1 ± 2.6 μA·cm-2 compared to controls, who showed a decrease of 10.5 ± 2.1 μA·cm-2 (p = 0.027). Stimulation or inhibition with theophylline, ouabain, bumetanide, forskolin or the EP receptor agonists prostaglandin E2, butaprost, sulprostone and prostaglandin E1 (OH) did not differ significantly between the two groups. Histology was with normal findings in both groups.
Epithelial electrogenic transport is more sensitive to indomethacin in normal colonic mucosa from patients with previous or present colorectal neoplasia compared to colonic mucosa from control patients. Stimulated epithelial electrogenic transport through individual prostanoid subtype receptors EP1, EP2, EP3, and EP4 is not significantly different between neoplasia diseased patients and controls. This indicates that increased indomethacin-sensitive mechanisms in colonic mucosa from neoplasia diseased patients are not related to differences in functional expression of EP receptor subtypes.
The dissection of complex traits of economic importance to the pig industry requires the availability of a significant number of genetic markers, such as single nucleotide polymorphisms (SNPs). This study was conducted to discover several hundreds of thousands of porcine SNPs using next generation sequencing technologies and use these SNPs, as well as others from different public sources, to design a high-density SNP genotyping assay.
A total of 19 reduced representation libraries derived from four swine breeds (Duroc, Landrace, Large White, Pietrain) and a Wild Boar population and three restriction enzymes (AluI, HaeIII and MspI) were sequenced using Illumina's Genome Analyzer (GA). The SNP discovery effort resulted in the de novo identification of over 372K SNPs. More than 549K SNPs were used to design the Illumina Porcine 60K+SNP iSelect Beadchip, now commercially available as the PorcineSNP60. A total of 64,232 SNPs were included on the Beadchip. Results from genotyping the 158 individuals used for sequencing showed a high overall SNP call rate (97.5%). Of the 62,621 loci that could be reliably scored, 58,994 were polymorphic yielding a SNP conversion success rate of 94%. The average minor allele frequency (MAF) for all scorable SNPs was 0.274.
Overall, the results of this study indicate the utility of using next generation sequencing technologies to identify large numbers of reliable SNPs. In addition, the validation of the PorcineSNP60 Beadchip demonstrated that the assay is an excellent tool that will likely be used in a variety of future studies in pigs.
A wealth of genetic associations for cardiovascular and metabolic phenotypes in humans has been accumulating over the last decade, in particular a large number of loci derived from recent genome wide association studies (GWAS). True complex disease-associated loci often exert modest effects, so their delineation currently requires integration of diverse phenotypic data from large studies to ensure robust meta-analyses. We have designed a gene-centric 50 K single nucleotide polymorphism (SNP) array to assess potentially relevant loci across a range of cardiovascular, metabolic and inflammatory syndromes. The array utilizes a “cosmopolitan” tagging approach to capture the genetic diversity across ∼2,000 loci in populations represented in the HapMap and SeattleSNPs projects. The array content is informed by GWAS of vascular and inflammatory disease, expression quantitative trait loci implicated in atherosclerosis, pathway based approaches and comprehensive literature searching. The custom flexibility of the array platform facilitated interrogation of loci at differing stringencies, according to a gene prioritization strategy that allows saturation of high priority loci with a greater density of markers than the existing GWAS tools, particularly in African HapMap samples. We also demonstrate that the IBC array can be used to complement GWAS, increasing coverage in high priority CVD-related loci across all major HapMap populations. DNA from over 200,000 extensively phenotyped individuals will be genotyped with this array with a significant portion of the generated data being released into the academic domain facilitating in silico replication attempts, analyses of rare variants and cross-cohort meta-analyses in diverse populations. These datasets will also facilitate more robust secondary analyses, such as explorations with alternative genetic models, epistasis and gene-environment interactions.
Colonic diverticular disease is a bothersome condition with an unresolved pathogenesis. It is unknown whether a neuroepithelial dysfunction is present. The aim of the study was two-fold; (1) to investigate colonic epithelial ion transport in patients with diverticulosis and (2) to adapt a miniaturized Modified Ussing Air-Suction (MUAS) chamber for colonic endoscopic biopsies.
Biopsies were obtained from the sigmoid part of the colon. 86 patients were included. All patients were referred for colonoscopy on suspicion of neoplasia and they were without pathological findings at colonoscopy (controls) except for diverticulosis in 22 (D-patients). Biopsies were mounted in MUAS chambers with an exposed area of 5 mm2. Electrical responses to various stimulators and inhibitors of ion transport were investigated together with histological examination. The MUAS chamber was easy to use and reproducible data were obtained.
Median basal short circuit current (SCC) was 43.8 μA·cm-2 (0.8 – 199) for controls and 59.3 μA·cm-2 (3.0 – 177.2) for D-patients. Slope conductance was 77.0 mS·cm-2 (18.6 – 204.0) equal to 13 Ω·cm2 for controls and 96.6 mS·cm-2 (8.4 – 191.4) equal to 10.3 Ω·cm2 for D-patients. Stimulation with serotonin, theophylline, forskolin and carbachol induced increases in SCC in a range of 4.9 – 18.6 μA·cm-2, while inhibition with indomethacin, bumetanide, ouabain and amiloride decreased SCC in a range of 6.5 – 27.4 μA·cm-2, and all with no significant differences between controls and D-patients. Histological examinations showed intact epithelium and lamina propria before and after mounting for both types of patients.
We conclude that epithelial ion transport is not significantly altered in patients with diverticulosis and that the MUAS chamber can be adapted for studies of human colonic endoscopic biopsies.
A bold new effort to disrupt every gene in the mouse genome necessitates systematic, interdisciplinary approaches to analyzing patterning defects in the mouse embryo. We present a novel, rapid, and inexpensive method for obtaining high-resolution virtual histology for phenotypic assessment of mouse embryos. Using osmium tetroxide to differentially stain tissues followed by volumetric X-ray computed tomography to image whole embryos, isometric resolutions of 27 μm or 8 μm were achieved with scan times of 2 h or 12 h, respectively, using mid-gestation E9.5–E12.5 embryos. The datasets generated by this method are immediately amenable to state-of-the-art computational methods of organ patterning analysis. This technique to assess embryo anatomy represents a significant improvement in resolution, time, and expense for the quantitative, three-dimensional analysis of developmental patterning defects attributed to genetically engineered mutations and chemically induced embryotoxicity.
Developmental biology is entering the digital age, thanks to advancements in imaging technologies, instrumentation, and software. These advancements are converging with discoveries in developmental biology to deliver unprecedented insight into how human development is impacted by the products we use, the environment we live in, and our genetic composition. Industrial societies are becoming increasingly concerned with the exposure of women and their unborn fetuses to pharmaceuticals and commonplace household chemicals. In addition, understanding genetic causes of birth defects is now possible through the isolation of specific genes, which can be efficiently disrupted in embryos, and subsequently observed for birth defects. Such studies of embryotoxicity typically involve the use of mouse embryos. However, evaluation of mouse embryos in the past has involved expensive and cumbersome external inspection and thin sectioning for view under the microscope. As such, developmental biologists have eagerly anticipated the advent of tools that would allow them to routinely assess the complex and dynamic organization of embryos using techniques that are fast and inexpensive. In this article, the authors introduce a rapid, high-quality, and inexpensive technique for the three-dimensional visualization of mouse embryos using X-ray computed tomography that is ideally suited for researchers in pharmaceutical, industrial, and academic laboratories.
The data provided to the Genetic Analysis Workshop 14 (GAW 14) was the result of a collaboration among several different groups, catalyzed by Elizabeth Pugh from The Center for Inherited Disease Research (CIDR) and the organizers of GAW 14, Jean MacCluer and Laura Almasy. The DNA, phenotypic characterization, and microsatellite genomic survey were provided by the Collaborative Study on the Genetics of Alcoholism (COGA), a nine-site national collaboration funded by the National Institute of Alcohol and Alcoholism (NIAAA) and the National Institute of Drug Abuse (NIDA) with the overarching goal of identifying and characterizing genes that affect the susceptibility to develop alcohol dependence and related phenotypes. CIDR, Affymetrix, and Illumina provided single-nucleotide polymorphism genotyping of a large subset of the COGA subjects. This article briefly describes the dataset that was provided.
To replicate, a retrovirus must synthesize a cDNA copy of the viral RNA genome and integrate that cDNA into a chromosome of the host. We have investigated the role of a host cell cofactor, HMG I(Y) protein, in integration of human immunodeficiency virus type 1 (HIV-1) and Moloney murine leukemia virus (MoMLV) cDNA. Previously we reported that HMG I(Y) cofractionates with HIV-1 preintegration complexes (PICs) isolated from freshly infected cells. PICs depleted of required components by treatment with high concentrations of salt could be reconstituted by addition of purified HMG I(Y) in vitro. Here we report studies using immunoprecipitation that indicate that HMG I(Y) is associated with MoMLV preintegration complexes. In mechanistic studies, we show for both HIV-1 and MoMLV that each HMG I(Y) monomer must contain multiple DNA binding domains to stimulate integration by HMG I(Y)-depleted PICs. We also find that HMG I(Y) can condense model HIV-1 or MoMLV cDNA in vitro as measured by stimulation of intermolecular ligation. This reaction, like reconstitution of integration, depends on the presence of multiple DNA binding domains in each HMG I(Y) monomer. These data suggest that binding of multivalent HMG I(Y) monomers to multiple cDNA sites compacts retroviral cDNA, thereby promoting formation of active integrase-cDNA complexes.
Picture archiving and communications systems (PACS) have inescapably altered the face of radiology. Images are available to radiologists and clinicians alike, nearly instantaneously. For patient care management, service has improved, but without inclusion of input from radiologists. Effecting timely report availability requires reorganization of radiology. In a hospital-wide PACS environment, we undertook to render a preliminary report on all nonprocedural computed radiography examinations within 30 minutes in a teaching environment. Two periods of time in the same month were analyzed, one before reorganization and one after. Of 686 reports, 117 were examined with a statistical significance of α=.05 (95% confidence) and a power of 90%. Average times for examination acquisition to preliminary report availability on the PACS decreased from 5 hours to 31 minutes. Standard deviation in report generation times decreased from 8 hours to 30 minutes. This preliminary study suggests that business process reengineering can effect improvement in information flow within a teaching facility resulting in radiologists rejoining the patient care management team. Successes, pitfalls, and future requirements are discussed.
picture archiving and communications system (PACS); radiology reports; diagnostic radiology; department management