Uncovering human transcription factor interactions associated with genetic variants, novel DNA motifs, and repetitive elements using enhanced yeast one-hybrid assays

Identifying transcription factor (TF) binding to noncoding variants, uncharacterized DNA motifs, and repetitive genomic elements has been difficult due to technical and computational challenges. Indeed, current experimental methods such as chromatin immunoprecipitation are capable of only testing one TF at a time and motif prediction algorithms often lead to false positive and false negative predictions. Here, we address these limitations by developing two approaches based on enhanced yeast one-hybrid assays. The first approach allows to interrogate the binding of >1,000 human TFs to single nucleotide variant alleles, short insertions and deletions (indels), and novel DNA motifs; while the second approach allows for the identification of TFs that bind to repetitive DNA elements. Using the former approach, we identified gain of TF interactions to a GG→AA mutation in the TERT promoter and an 18 bp indel in the TAL1 super-enhancer, both of which are associated with cancer, and identified the TFs that bind to three uncharacterized DNA motifs identified by the ENCODE Project in footprinting assays. Using the latter approach, we detected the binding of 75 TFs to the highly repetitive Alu elements. We anticipate that these approaches will expand our capabilities to study genetic variation and under-characterized genomic regions.https://doi.org/10.1101/459305First author draf

The in vitro metabolism of 17-hydroxy-11-desoxycorticoserone (Reichstein's compound S).

Thesis (M.A.)--Boston UniversityAlthough it has been known that ∆4-pregnene-17a, 21-diol- 3, 20-dione (Reichstein's compoundS) is an intermediate in the biosynthesis of hydrocortisone (13), little was known of other aspects of its metabolism. The in vitro technique for study was chosen in the hope that the isolation of metabolites would permit an interpretation of the mechanism involved in the metabolism of compoundS. Since it has been demonstrated that liver contains a relatively high concentration of the necessary enzyme systems for the transformation of steroids, preparations of this tissue were selected for this in vitro metabolism study. Compound S was incubated with several types of enzyme preparations of rat liver, and products containing reduced structural configurations involving positions 3 and 5 were isolated. The utilization of silica gel column and paper chromatographic resolution has permitted the purification and resolution of two steroid metabolites in crystalline form. [TRUNCATED

The in vitro metabolism of a typical adrenocortical steroid (11-deoxycortisol)

Thesis (Ph.D.)--Boston UniversityThe liver has been well established as being one of the most important organs involved in the metabolism of adrenocortical steroids. Liver perfusion, incubations with liver slices, homogenates and purified liver preparations all effect extensive changes on the steroid nucleus. In order to develop a step-wise scheme for the in vitro metabolism of a typical adrenocortical steroid and to localize enzyme systems effecting single metabolic transformations, 11-deoxycortisol was incubated with various rat liver homogenate fractions and the conversion products isolated and identified. The liver fractions were obtained by differential centrifugation of liver homogenates. This procedure afforded a 6000 x g supernatant, a 6000 x g residue (corresponding to mitochondria in sedimentation properties), a 78,000 x g residue (corresponding to microsoaes in sedimentation properties) and a 78.000 x g particulate free supernatant. Each tissue preparation was incubated with 11-deoxycortisol at 37° c. for two hours with flasks open to the atmosphere. A tissue to steroid ratio of approximately 1000:1, based on the initial weight of the liver, was employed [TRUNCATED]

Enteral nutrition protects children undergoing allogeneic hematopoietic stem cell transplantation from blood stream infections

Enteral Nutrition (EN) is recommended as first line nutritional support for patients undergoing Allogeneic Hematopoietic Stem Cell Transplantation (allo-HSCT), but only few studies exist in the literature which compare EN to Parenteral Nutrition (PN) in the paediatric population. Forty-two consecutive paediatric patients undergoing allo-HSCT at our referral centre between January 2016 and July 2019 were evaluated. Post-transplant and nutritional outcomes of patients receiving EN for more than 7 days (EN group, n = 14) were compared with those of patients receiving EN for fewer than 7 days or receiving only PN (PN group, n = 28). In the EN group, a reduced incidence of Blood Stream Infections (BSI) was observed (p = 0.02) (n = 2 vs. n = 15; 14.3% vs. 53.6%). The type of nutritional support was also the only variable independently associated with BSI in the multivariate analysis (p = 0.03). Platelet engraftment was shorter in the PN group than in the EN group for a threshold of > 20*109/L (p = 0.04) (23.1 vs 35.7 days), but this correlation was not confirmed with a threshold of > 50*109/L. The Body Mass Index (BMI) and the BMI Z-score were no different in the two groups from admission to discharge. Our results highlight that EN is a feasible and nutritionally adequate method of nutritional support for children undergoing allo-HSCT in line with the present literature. Future functional studies are needed to better address the hypothesis that greater intestinal eubyosis maintained with EN may explain the observed reduction in BSI

An internet-based approach for lifestyle changes in patients with NAFLD: Two-year effects on weight loss and surrogate markers

Background & Aims Interventions aimed at lifestyle changes are pivotal for the treatment of non-alcoholic fatty liver disease (NAFLD), and web-based programs might help remove barriers in both patients and therapists. Methods In the period 2010–15, 716 consecutive NAFLD cases (mean age, 52; type 2 diabetes, 33%) were treated in our Department with structured programs. The usual protocol included motivational interviewing and a group-based intervention (GBI), chaired by physicians, dietitians and psychologists (five weekly meetings, n = 438). Individuals who could not attend GBI entered a web-based intervention (WBI, n = 278) derived from GBI, with interactive games, learning tests, motivational tests, and mail contacts with the center. The primary outcome was weight loss ≥10%; secondary outcomes were alanine aminotransferase within normal limits, changes in lifestyle, weight, alanine aminotransferase, and surrogate markers of steatosis and fibrosis. Results GBI and WBI cohorts had similar body mass index (mean, 33 kg/m2), with more males (67% vs. 45%), younger age, higher education, and more physical activity in the WBI group. The two-year attrition rate was higher in the WBI group. Healthy lifestyle changes were observed in both groups and body mass index decreased by almost two points; the 10% weight target was reached in 20% of WBI cases vs. 15% in GBI (not significant). In logistic regression analysis, after adjustment for confounders and attrition rates, WBI was not associated with a reduction of patients reaching short- and long-term 10% weight targets. Liver enzymes decreased in both groups, and normalized more frequently in WBI. Fatty liver index was reduced, whereas fibrosis remained stable (NAFLD fibrosis score) or similarly decreased (Fib-4). Conclusion WBI is not less effective than common lifestyle programs, as measured by significant clinical outcomes associated with improved histological outcomes in NAFLD. eHealth programs may effectively contribute to NAFLD control. Lay summary In patients with non-alcoholic fatty liver disease, participation in structured lifestyle programs may be jeopardized by job and time constraints. A web-based intervention may be better suited for young, busy patients, and for those living far from liver units. The study shows that, following a structured motivational approach, a web-based, interactive intervention coupled with six-month face-to-face meetings is not inferior to a standard group-based intervention with respect to weight loss, adherence to healthy diet and habitual physical activity, normalization of liver enzymes, and stable surrogate markers of fibrosis

Organophosphorous Pesticide Detection in Olive Oil by Using a Miniaturized, Easy-to-Use, and Cost-Effective Biosensor Combined with QuEChERS for Sample Clean-Up

En el laboratorio de curtición de pieles de la Facultad de Ciencias Pecuarias de la ESPOCH, se evaluó la obtención de napa de cordero para vestimenta con la aplicación de tres porcentajes de anilina, modelados bajo un Diseño Completamente al Azar en arreglo bifactorial empleando 45 unidades experimentales. En la evaluación de las resistencias físicas se reportaron diferencias altamente significativas (P < 0.05); registrándose los mejores resultados para adherencia (83,27%), desgarro (71,87 N/cm2) y porcentaje de elongación (86,73%), con la aplicación de 5% de anilina (T3). Respuestas similares se identificaron al evaluar las calificaciones sensoriales ya que se establecieron las mejores respuestas con la aplicación del 5% de anilina (T3), para blandura (4,53 puntos), redondez y efecto resorte (4,40 puntos). Finalmente en el análisis del beneficio costo se determinaron los mejores resultados con la adición de 5% de anilina por cuanto el beneficio costo fue de 1.23 que quiere decir que por cada dólar invertido se espera una rentabilidad del 23%. Por lo que se recomienda aplicar a la fórmula del teñido el 5% de anilina (T3), ya que se elevan significativamente las resistencias físicas y calificaciones sensoriales obteniéndose por lo tanto mayor rentabilidad de la napa de cordero

Gluten deprivation: What nutritional changes are found during the first year in newly diagnosed coeliac children?

Aim: A gluten-free diet (GFD) can expose children to excessive calories and fat intake. The study is intended to verify whether and how food intake, laboratory parameters, and growth are modified by a year of GFD. Methods: In 79 CD (coeliac disease) children (mean age 7.9 \ub1 3.8 years, 52 females, 27 males) diagnosed over 24 months, 24-h food diaries, food-frequency patterns, anthropometric and laboratory parameters (mainly blood sugar, insulin, lipid profile, and homocysteine) were prospectively collected before and during the first year of GFD. Nutrient intakes were compared over time and with recommendations. They were also used as regressors to explain the levels and changes of metabolic and growth variables. p-values &lt; 0.05 were considered statistically significant. Results: Average macronutrient intake did not change during the year. Caloric intake remained below 90% (p 64 0.0001) and protein intake above 200% (p 64 0.0001) of recommendations. Lipid intake was stable at 34% of overall energy intake. Unsaturated fats increased (less omega-6 and more omega-3 with a ratio improvement from 13.3 \ub1 5.5 to 8.8 \ub1 3.1) and so did fibers, while folate decreased. The children who experienced a containment in their caloric intake during the year, presented a slower catch-up growth. Some differences were found across gender and age groups. In particular, adolescents consumed less calories, and females more omega-3. Fiber and simple sugar intakes emerged as implicated in lipid profile shift: fibers negatively with triglycerides (TG) (p = 0.033), simple sugars negatively with high-density lipoprotein (HDL) (p = 0.056) and positively with TG (p = 0.004). Waist-to-height ratio was positively associated with homocysteine (p = 0.018) and Homeostasis Model Assessment (p = 0.001), negatively with fibers (p = 0.004). Conclusion: In the short run, GFD is nutritionally very similar to any diet with gluten, with some improvements in unsaturated fats and fiber intake. Along with simple sugars containment, this may offer CD patients the opportunity for a fresh start. Caloric intakes may shift and should be monitored, especially in adolescents