Validation of Results from Knowledge Discovery: Mass Density as a Predictor of Breast Cancer

The purpose of our study is to identify and quantify the association between high breast mass density and breast malignancy using inductive logic programming (ILP) and conditional probabilities, and validate this association in an independent dataset. We ran our ILP algorithm on 62,219 mammographic abnormalities. We set the Aleph ILP system to generate 10,000 rules per malignant finding with a recall >5% and precision >25%. Aleph reported the best rule for each malignant finding. A total of 80 unique rules were learned. A radiologist reviewed all rules and identified potentially interesting rules. High breast mass density appeared in 24% of the learned rules. We confirmed each interesting rule by calculating the probability of malignancy given each mammographic descriptor. High mass density was the fifth highest ranked predictor. To validate the association between mass density and malignancy in an independent dataset, we collected data from 180 consecutive breast biopsies performed between 2005 and 2007. We created a logistic model with benign or malignant outcome as the dependent variable while controlling for potentially confounding factors. We calculated odds ratios based on dichomotized variables. In our logistic regression model, the independent predictors high breast mass density (OR 6.6, CI 2.5–17.6), irregular mass shape (OR 10.0, CI 3.4–29.5), spiculated mass margin (OR 20.4, CI 1.9–222.8), and subject age (β = 0.09, p < 0.0001) significantly predicted malignancy. Both ILP and conditional probabilities show that high breast mass density is an important adjunct predictor of malignancy, and this association is confirmed in an independent data set of prospectively collected mammographic findings

Control of Parasitophorous Vacuole Expansion by LYST/Beige Restricts the Intracellular Growth of Leishmania amazonensis

The intracellular protozoan Leishmania replicates in parasitophorous vacuoles (PV) that share many features with late endosomes/lysosomes. L. amazonensis PVs expand markedly during infections, but the impact of PV size on parasite intracellular survival is still unknown. Here we show that host cells infected with L. amazonensis upregulate transcription of LYST/Beige, which was previously shown to regulate lysosome size. Mutations in LYST/Beige caused further PV expansion and enhanced L. amazonensis replication. In contrast, LYST/Beige overexpression led to small PVs that did not sustain parasite growth. Treatment of LYST/Beige over-expressing cells with vacuolin-1 reversed this phenotype, expanding PVs and promoting parasite growth. The opposite was seen with E-64d, which reduced PV size in LYST-Beige mutant cells and inhibited L. amazonensis replication. Enlarged PVs appear to protect parasites from oxidative damage, since inhibition of nitric oxide synthase had no effect on L. amazonensis viability within large PVs, but enhanced their growth within LYST/Beige-induced small PVs. Thus, the upregulation of LYST/Beige in infected cells functions as a host innate response to limit parasite growth, by reducing PV volume and inhibiting intracellular survival

Mutations in MITF and PAX3 Cause “Splashed White” and Other White Spotting Phenotypes in Horses

During fetal development neural-crest-derived melanoblasts migrate across the entire body surface and differentiate into melanocytes, the pigment-producing cells. Alterations in this precisely regulated process can lead to white spotting patterns. White spotting patterns in horses are a complex trait with a large phenotypic variance ranging from minimal white markings up to completely white horses. The “splashed white” pattern is primarily characterized by an extremely large blaze, often accompanied by extended white markings at the distal limbs and blue eyes. Some, but not all, splashed white horses are deaf. We analyzed a Quarter Horse family segregating for the splashed white coat color. Genome-wide linkage analysis in 31 horses gave a positive LOD score of 1.6 in a region on chromosome 6 containing the PAX3 gene. However, the linkage data were not in agreement with a monogenic inheritance of a single fully penetrant mutation. We sequenced the PAX3 gene and identified a missense mutation in some, but not all, splashed white Quarter Horses. Genome-wide association analysis indicated a potential second signal near MITF. We therefore sequenced the MITF gene and found a 10 bp insertion in the melanocyte-specific promoter. The MITF promoter variant was present in some splashed white Quarter Horses from the studied family, but also in splashed white horses from other horse breeds. Finally, we identified two additional non-synonymous mutations in the MITF gene in unrelated horses with white spotting phenotypes. Thus, several independent mutations in MITF and PAX3 together with known variants in the EDNRB and KIT genes explain a large proportion of horses with the more extreme white spotting phenotypes

The Cytosolic Protein G0S2 Maintains Quiescence in Hematopoietic Stem Cells

Bone marrow hematopoietic stem cells (HSCs) balance proliferation and differentiation by integrating complex transcriptional and post-translational mechanisms regulated by cell intrinsic and extrinsic factors. We found that transcripts of G0/G1 switch gene 2 (G0S2) are enriched in lineage− Sca-1+ c-kit+ (LSK) CD150+ CD48− CD41− cells, a population highly enriched for quiescent HSCs, whereas G0S2 expression is suppressed in dividing LSK CD150+ CD48− cells. Gain-of-function analyses using retroviral expression vectors in bone marrow cells showed that G0S2 localizes to the mitochondria, endoplasmic reticulum, and early endosomes in hematopoietic cells. Co-transplantation of bone marrow cells transduced with the control or G0S2 retrovirus led to increased chimerism of G0S2-overexpressing cells in femurs, although their contribution to the blood was reduced. This finding was correlated with increased quiescence in G0S2-overexpressing HSCs (LSK CD150+ CD48−) and progenitor cells (LS−K). Conversely, silencing of endogenous G0S2 expression in bone marrow cells increased blood chimerism upon transplantation and promoted HSC cell division, supporting an inhibitory role for G0S2 in HSC proliferation. A proteomic study revealed that the hydrophobic domain of G0S2 interacts with a domain of nucleolin that is rich in arginine-glycine-glycine repeats, which results in the retention of nucleolin in the cytosol. We showed that this cytosolic retention of nucleolin occurs in resting, but not proliferating, wild-type LSK CD150+ CD48− cells. Collectively, we propose a novel model of HSC quiescence in which elevated G0S2 expression can sequester nucleolin in the cytosol, precluding its pro-proliferation functions in the nucleolus

AAV Exploits Subcellular Stress Associated with Inflammation, Endoplasmic Reticulum Expansion, and Misfolded Proteins in Models of Cystic Fibrosis

Barriers to infection act at multiple levels to prevent viruses, bacteria, and parasites from commandeering host cells for their own purposes. An intriguing hypothesis is that if a cell experiences stress, such as that elicited by inflammation, endoplasmic reticulum (ER) expansion, or misfolded proteins, then subcellular barriers will be less effective at preventing viral infection. Here we have used models of cystic fibrosis (CF) to test whether subcellular stress increases susceptibility to adeno-associated virus (AAV) infection. In human airway epithelium cultured at an air/liquid interface, physiological conditions of subcellular stress and ER expansion were mimicked using supernatant from mucopurulent material derived from CF lungs. Using this inflammatory stimulus to recapitulate stress found in diseased airways, we demonstrated that AAV infection was significantly enhanced. Since over 90% of CF cases are associated with a misfolded variant of Cystic Fibrosis Transmembrane Conductance Regulator (ΔF508-CFTR), we then explored whether the presence of misfolded proteins could independently increase susceptibility to AAV infection. In these models, AAV was an order of magnitude more efficient at transducing cells expressing ΔF508-CFTR than in cells expressing wild-type CFTR. Rescue of misfolded ΔF508-CFTR under low temperature conditions restored viral transduction efficiency to that demonstrated in controls, suggesting effects related to protein misfolding were responsible for increasing susceptibility to infection. By testing other CFTR mutants, G551D, D572N, and 1410X, we have shown this phenomenon is common to other misfolded proteins and not related to loss of CFTR activity. The presence of misfolded proteins did not affect cell surface attachment of virus or influence expression levels from promoter transgene cassettes in plasmid transfection studies, indicating exploitation occurs at the level of virion trafficking or processing. Thus, we surmised that factors enlisted to process misfolded proteins such as ΔF508-CFTR in the secretory pathway also act to restrict viral infection. In line with this hypothesis, we found that AAV trafficked to the microtubule organizing center and localized near Golgi/ER transport proteins. Moreover, AAV infection efficiency could be modulated with siRNA-mediated knockdown of proteins involved in processing ΔF508-CFTR or sorting retrograde cargo from the Golgi and ER (calnexin, KDEL-R, β-COP, and PSMB3). In summary, our data support a model where AAV exploits a compromised secretory system and, importantly, underscore the gravity with which a stressed subcellular environment, under internal or external insults, can impact infection efficiency

Relationship of depression, disability, and family caregiver attitudes to the quality of life of Kuwaiti persons with multiple sclerosis: a controlled study

<p>Abstract</p> <p>Background</p> <p>Assessment of subjective quality of life (QOL) of persons with multiple sclerosis (MS) could facilitate the detection of psychosocial aspects of disease that may otherwise go unrecognized. The objectives of the study were to (i) compare the QOL ratings of relapsing remitting (RRMS) and progressive (PMS) types of MS with those of a general population group and the impression of their family caregivers; and (ii) assess the association of demographic, clinical, treatment, depression, and caregiver variables with patients' QOL.</p> <p>Methods</p> <p>Consecutive clinic attendees at the national neurology hospital were assessed with the 26 -item WHOQOL Instrument, Beck's Depression Inventory and Expanded Disability Scale. Caregivers rated their impression of patients' QOL and attitudes to patients' illness.</p> <p>Results</p> <p>The 170 patients (60 m, 109 f) consisted of 145(85.3%) with RRMS and 25 with PMS, aged 32.4(SD 8.8), age at onset 27.1(7.7), EDSS score 2.9 (1.8), and 76% were employed. The patients were predominantly dissatisfied with their life circumstances. The RRMS group had higher QOL domain scores (P < 0.001), and lower depression(P > 0.05) and disability (P < 0.0001) scores than the PMS group. Patients had significantly lower QOL scores than the control group (P < 0.001). Caregiver impression was significantly correlated with patients' ratings. Depression was the commonest significant covariate of QOL domains. When we controlled for depression and disability scores, differences between the two MS groups became significant for only one (out of 6) QOL domains. Patients who were younger, better educated, employed, felt less sick and with lesser side effects, had higher QOL. The predictors of patients' overall QOL were disability score, caregiver impression of patients' QOL, and caregiver fear of having MS.</p> <p>Conclusion</p> <p>Our data indicate that MS patients in stable condition and with social support can hope to have better QOL, if clinicians pay attention to depression, disability, the impact of side effects of treatment and family caregiver anxieties about the illness. The findings call for a regular program of psychosocial intervention in the clinical setting, to address these issues and provide caregiver education and supports, in order to enhance the quality of care.</p

Local Increase of Arginase Activity in Lesions of Patients with Cutaneous Leishmaniasis in Ethiopia

The leishmaniases are a complex of diseases caused by Leishmania parasites. Currently, the diseases affect an estimated 12 million people in 88 countries, and approximately 350 million more people are at risk. The leishmaniases belong to the most neglected tropical diseases, affecting the poorest populations, for whom access to diagnosis and effective treatment are often not available. Leishmania parasites infect cells of the immune system called macrophages, which have the capacity to eliminate the intracellular parasites when they receive the appropriate signals from other cells of the immune system. In nonhealing persistent leishmaniasis, lymphocytes are unable to transmit the signals to macrophages required to kill the intracellular parasites. The local upregulation of the enzyme arginase has been shown to impair lymphocyte effector functions at the site of pathology. In this study, we tested the activity of this enzyme in skin lesions of patients presenting with localized cutaneous leishmaniasis. Our results show that arginase is highly upregulated in these lesions. This increase in arginase activity coincides with lower expression of a signalling molecule in lymphocytes, which is essential for efficient activation of these cells. These results suggest that increased arginase expression in the localized cutaneous lesions might contribute to persistent disease in patients presenting with cutaneous leishmaniasis

Antineoplastic effects of rosiglitazone and PPARγ transactivation in neuroblastoma cells

Neuroblastoma (NB) is the most common extracranial solid tumour in infants. Unfortunately, most children present with advanced disease and have a poor prognosis. In the present study, we evaluated the role of the peroxisome proliferator-activated receptor γ (PPARγ) agonist rosiglitazone (RGZ) in two NB cell lines (SK-N-AS and SH-SY5Y), which express PPARγ. Rosiglitazone decreased cell proliferation and viability to a greater extent in SK-N-AS than in SH-SY5Y. Furthermore, 20 μM RGZ significantly inhibited cell adhesion, invasiveness and apoptosis in SK-N-AS, but not in SH-SY5Y. Because of the different response of SK-N-AS and SH-SY5Y cells to RGZ, the function of PPARγ as a transcriptional activator was assessed. Noticeably, transient transcription experiments with a PPARγ responsive element showed that RGZ induced a three-fold increase of the reporter activity in SK-N-AS, whereas no effect was observed in SH-SY5Y. The different PPARγ activity may be likely due to the markedly lower amount of phopshorylated (i.e. inactive) protein observed in SK-N-AS. To our knowledge, this is the first demonstration that the differential response of NB cells to RGZ may be related to differences in PPARγ transactivation. This finding indicates that PPARγ activity may be useful to select those patients, for whom PPARγ agonists may have a beneficial therapeutic effect

Comparative Genomic Analysis of Pathogenic and Probiotic Enterococcus faecalis Isolates, and Their Transcriptional Responses to Growth in Human Urine

Urinary tract infection (UTI) is the most common infection caused by enterococci, and Enterococcus faecalis accounts for the majority of enterococcal infections. Although a number of virulence related traits have been established, no comprehensive genomic or transcriptomic studies have been conducted to investigate how to distinguish pathogenic from non-pathogenic E. faecalis in their ability to cause UTI. In order to identify potential genetic traits or gene regulatory features that distinguish pathogenic from non-pathogenic E. faecalis with respect to UTI, we have performed comparative genomic analysis, and investigated growth capacity and transcriptome profiling in human urine in vitro. Six strains of different origins were cultivated and all grew readily in human urine. The three strains chosen for transcriptional analysis showed an overall similar response with respect to energy and nitrogen metabolism, stress mechanism, cell envelope modifications, and trace metal acquisition. Our results suggest that citrate and aspartate are significant for growth of E. faecalis in human urine, and manganese appear to be a limiting factor. The majority of virulence factors were either not differentially regulated or down-regulated. Notably, a significant up-regulation of genes involved in biofilm formation was observed. Strains from different origins have similar capacity to grow in human urine. The overall similar transcriptional responses between the two pathogenic and the probiotic strain suggest that the pathogenic potential of a certain E. faecalis strain may to a great extent be determined by presence of fitness and virulence factors, rather than the level of expression of such traits