bluuebunny/crossref_metadata_2025 huggingface dataset:
Caregiver's Knowledge of Sugar Sweetened Beverages is Associated with Availability of this Type of Beverages in the Home
Authors: Quesada Catalina , Schilling Elizabeth , Ferris Ann , Lora Karina
Published: June 2021

Educating caregivers about sugar‐sweetened beverages (SSBs) is potentially a cost effective intervention strategy for reducing child SSB consumption. Baseline data from a longitudinal study explored the association between caregiver's information about SSBs and availability of total SSBs in the home. Low income caregivers of preschoolers (n=472) provided demographic information and completed a home beverage inventory and an Information‐Motivation‐Behavioral Skills measure of child consumption of SSBs (IMB‐mCSSB). Regressions on the information subscale of the IMB‐mCSSB, revealed that minority caregivers demonstrated significantly less knowledge about SSBs than Whites (p<.05) and male caregivers were less informed about SSBs than females (p<.05). Although SSB availability did not differ significantly by race/ethnicity or gender, information about SSBs was associated with their availability in the home (p<.05). In this study, as information about SSBs increased, availability of SSBs in the home decreased. These results suggest that educating minority and male caregivers about SSBs may affect SSB purchasing behavior.
Lean body mass and loci containing genes involved in insulin and glucose regulation are associated with calorie intake in dogs
Authors: Jewell Dennis Edward , Brockman Jeffrey A. , Huentelman Matthew J. , Kingsley Christopher B. , Siniard Ashley L. , Richholt Ryan , Hall Jean A.
Published: June 2021

Body composition and food intake required to maintain body weight were measured in 311 dogs from 1 to 17 years of age. All dogs were provided with regular opportunities for socialization and environmental enrichment. Dogs experienced behavioral enrichment through daily interaction and play time with caretakers, and by daily opportunities to run and exercise with access to toys. Total fat and percent body fat were positively associated with age (P<0.01) whereas lean body mass was not (P>;0.7). Lean body mass was positively associated with food intake (r2=0.59, P<0.01) whereas body fat and age were not associated with food intake. Males had overall greater body mass, with more lean body mass and higher fat body mass, yet were lower in percent body fat than females. Whole genome association studies identified a locus containing the alpha adrenergic receptor which was associated with percent body fat. Also, a number of loci containing genes involved in insulin and glucose regulation were associated with caloric intake. These data support the hypothesis that the number of calories needed to maintain weight is a response to the amount of body lean and related to specific genes which are involved in insulin and glucose regulation.
Biochemical and metabolomic effects of perinatal choline deficiency in the piglet
Authors: Getty Caitlyn M , Dilger Ryan N
Published: June 2021

Choline plays an essential role in tissue development during gestation and early postnatal life, and the piglet (Sus scrofa) may serve as a translational animal model to study perinatal effects of choline deficiency. Our objective was to determine developmental changes in piglets exposed to differing levels of choline both prenatally and postnatally. A factorial arrangement of choline sufficient (CS; 50% above requirement) and choline deficient (CD; 50% below requirement) diets were provided during the prenatal (last 65 d of gestation) and postnatal (48 h after birth through 28 d of age) periods. An interactive effect of choline level and perinatal period was observed for body weight gain (P < 0.001), and prenatal CD piglets had smaller (P = 0.014) brains than prenatal CS piglets. Blood chemistry panels revealed generalized impairments in liver function in postnatal CD piglets. Overall, metabolomic analysis identified 276 biochemicals and revealed that choline deficiency in the pre‐ and postnatal periods altered plasma levels of metabolites related to choline biosynthesis, glycolysis, and lipid metabolism. Additionally, interactive effects were noted for glycolytic intermediates, carnitine‐related metabolites, and many choline‐containing lysolipids. We conclude that the piglet is a sensitive animal model for studying developmental aspects of perinatal choline deficiency.

U.S. Department of Agriculture Hatch Project ILLU‐538–319
Influence of a polyphenol‐enriched protein powder on exerciseinduced inflammation and oxidative stress in athletes: a metabolomics approach
Authors: Nieman David Christopher , Knab Amy , Shanely R. Andrew , Cialdella‐Kam Lynn , Gillitt Nicholas , Pappan Kirk , Lila Mary Ann
Published: June 2021

Polyphenol intake was tested as a countermeasure to inflammation and oxidative stress induced by 3‐d intensified training. A soy protein isolate matrix was sorbed with polyphenols (SPMP) from blueberry and green tea extracts (2,136 mg/d gallic acid equivalents). Runners (N=31) were randomized to 40 g/d SPMP or placebo for 17 d using double‐blinded methods, with 3 d running inserted at day 14 (2.5 h/d, 70% VO2max). Blood samples were collected pre‐ and post‐14 d supplementation, and immediately and 14 h after the third day of running, and analyzed at Metabolon (Durham, NC) using GC‐MS and LC‐MS. Metabolites characteristic of gut bacteria metabolism of phenolics were increased with SPMP (e.g., hippurate, catechol sulfate, 4‐methylcatechol sulfate,1.8‐, 1.7‐, 2.5‐fold, respectively, P<0.05), an effect which persisted for 14‐h recovery. Fatty acid oxidation and ketogenesis were induced by exercise in both groups, with more ketones at 14‐h recovery in SPMP (3‐hydroxybutyrate, 1.8‐fold, P<0.05). Traditional biomarkers for inflammation (CRP, cytokines) and oxidative stress (protein carbonyls) did not differ between groups. In conclusion, 17‐d SPMP enhanced ketogenesis and fat oxidation in runners during recovery from 3‐d heavy exertion, and was linked to a gut‐derived phenolic signature that may exert anti‐oxidative and anti‐inflammatory effects not captured by typical biomarkers.

Funded by Dole Foods
Monitoring sodium in commercial packaged and restaurant foods
Authors: Ahuja Jaspreet K , Pehrsson Pamela , Haytowitz David , Holden Joanne , Nickle Melissa , Showell Bethany , Thomas Robin , Hoy Kathy , Martin Carrie , Sebastian Rhonda , Moshfegh Alanna , Gillespie Cathleen , Cogswell Mary
Published: June 2021

As part of a collaborative initiative between USDA, CDC and FDA to monitor sodium in the US food supply, a systemic program to monitor commercial packaged and restaurant foods with added sodium is underway at Agricultural Research Service (ARS). It includes nationwide sampling and analyses of about 125 Sentinel Foods, identified by ARS scientists as primary indicators to assess changes in the food supply. An additional ~1200 foods that support the processing of dietary data in the national survey, What We Eat In America (WWEIA), NHANES are being monitored through review of the Nutrition Facts Panel. As of FY 2012, about 75% of the Sentinel Foods have been sampled and analyzed. Preliminary review of the data shows mixed trends in mean sodium content. For example, there were no significant changes for taco shells and fast food cheese and pepperoni pizzas, whereas thin crust cheese pizza increased by 10.7% (p=0.014). Reductions of 20–25% were seen in ketchup (p<0.005) and canned pasta with sauce (p=0.038). These updated data are publicly available for use by researchers and epidemiologists and released annually in the USDA National Nutrient Database for Standard Reference (SR) and biennially in the Food and Nutrient Database for Dietary Studies (FNDDS) for use in the WWEIA, NHANES.
The association between seafood consumption and blood mercury levels in adults age 20 years and up, 2007–2010
Authors: Nielsen Samara Joy , Kit Brian K. , Ogden Cynthia L.
Published: June 2021

Background

Seafood is a dietary source of mercury, a neurotoxin with adverse health consequences. No previous studies have examined associations between seafood consumption and blood mercury in the entire US adult population.

Objective

To determine the association between seafood consumption and blood mercury levels in US adults, using the most recent national data.

Design

Adults, ages 20 years and older, who participated in the 2007–2010 National Health and Nutrition Examination Survey. Frequency of fish and/or shellfish (seafood) consumption was determined using a limited food frequency questionnaire over the past thirty days. Geometric mean blood mercury levels for quartiles of seafood consumption were calculated overall and by sex. Trends in mean mercury levels by quartile of seafood consumption were tested using linear trend tests.

Results

There was a positive linear trend in blood mercury as seafood consumption increased (p<.001 for all trends). Geometric mean blood mercury was 0.5 ug/L among the lowest consumers of seafood compared to 2.1 ug/L among the highest consumers. Trends for men and women separately were also significant (p<.001).

Conclusion

In the US adult population blood mercury levels are highest among those who consume the largest amounts of seafood.
Cysteinyl‐glycine reduces mucosal pro‐inflammatory cytokine response to fMLP in a piglet model of intestinal atrophy
Authors: Nosworthy Matthew , Brunton Janet A.
Published: June 2021

PepT1 is a small intestinal (SI) di/tripeptide transporter also capable of transporting bacterial peptides. We measured the mucosal cytokine response to a pro‐inflammatory peptide (fMLP) when delivered with cysteinyl‐glycine (CG); CG competes for PepT1‐mediated uptake and has anti‐inflammatory potential. Pigs (n=6, 10 d) received parenteral nutrition (PN) for 4 d to induce SI atrophy; littermates (n=6) remained with the sow (S). Subsequently, five 10 cm loops of the distal SI were isolated and perfused for 3 h with one of: 1) 5 mM cysteine + 5 mM glycine (5 mM cys + gly) 2) 5 mM CG 3) 10 μM fMLP 4) 5 mM cys + gly + 10 μM fMLP 5) 5 mM CG + 10 μM fMLP. In both S and PN, SI exposed to fMLP had higher mucosal TNF‐α and IFN‐γ compared to unexposed loops (p<0.001), while IFN‐γ was higher in PN compared to S pigs (p < 0.01). CG + fMLP treatment lowered the IFN‐γ response in both S and PN (p <0.05), but neither group responded significantly to free cys + gly. Interestingly, free cys + gly reduced the TNF‐α response in S pigs (p<0.001), but not in the PN group. CG + fMLP lowered TNF‐α compared to fMLP alone in both groups (p<0.001) and in S pigs the response was significantly more abated than with cys + gly (p < 0.001). Thus, CG was more effective than free cys + gly at attenuating a bacterial peptide‐induced cytokine response particularly in the injured SI; this may be due to impaired free amino acid uptake or effective competitive inhibition of fMLP uptake. (NSERC)
Effects of heat stress on the response to metabolic challenges in growing pigs
Authors: Fernandez Maria Victoria Sanz , Johnson Jay S , Abuajamieh Mohannad , Stoakes Sara K , Lei Soi M , Gabler Anna L , Toledo Mateus Zucato , Rhoads Rob P , Baumgard Lance H
Published: June 2021

Heat stress (HS) decreases feed intake but unexplainably increases plasma insulin parameters in a variety of animal models. Whether HS alters post‐absorptive carbohydrate and lipid metabolism in growing pigs is unknown. Seven crossbred gilts (53±6 kg BW) were subjected to 2 periods: thermoneutral (TN; 20°C, 36% RH) for 4d, and HS (32°C, 23% RH) for 8d. All pigs received an intravenous glucose tolerance test (GTT) on d2 of TN and d1 and 6 of HS; and an epinephrine challenge (EC) on d3 of TN and d2 and 7 of HS. Heat stress increased rectal temperatures (P<0.01; 39.4 vs 40.4°C) and respiratory rates (P<0.01; 30 vs 94 bpm), and reduced feed intake by 37% (P<0.01). Heat stress decreased (P<0.05) basal glucose and insulin concentrations by 14% and 50%, respectively. Basal nonesterified fatty acids (NEFA) were initially elevated during HS (92%), but progressively declined to TN levels by d3 (P<0.01). On d1 HS, glucose disposal and insulin response to a GTT decreased (49% and 24%, respectively) compared to TN. By d6 HS, the insulin response remained unchanged, but glucose disposal increased compared to d1 (P<0.01). On d2, NEFA response to an EC did not differ from TN, but was reduced (53%; P<0.01) by d7. Glucose response to the EC did not change over time (P>;0.05). These data suggest that, despite a low plane of nutrition, decreased adipose tissue utilization and increased glucose oxidation might be key acclimation mechanisms to heat stress.
Association of serum 25OHD with race/ethnicity and quantitative measures of skin color in urban schoolchildren
Authors: Au Lauren Elizabeth , Harris Susan S , Dwyer Johanna T , Jacques Paul F , Sacheck Jennifer M
Published: June 2021

To determine the extent to which differences in skin color explain racial/ethnic differences in serum 25OHD levels in urban schoolchildren. Multivariate regression was used to determine associations of serum 25OHD, race/ethnicity and skin color, measured by reflectance colorimeter [Individual Typology Angle (ITA°, higher value corresponds to lighter skin)], in 307 schoolchildren aged 9–15 from the Boston area during winter. Nearly 60% of all children had inadequate 25OHD (<20 ng/mL) and only 5% achieved optimal 25OHD status (≥30 ng/ml). Prevalence of inadequate 25OHD differed by race/ethnicity (P<0.001): white (47%), black (74%), Hispanic (65%), Asian (89%), multi‐racial/other (53%). Mean (SD) ITA° skin color also varied by race/ethnicity (P<0.001): white 50.1° (10.1), black −10.7° (25.2), Hispanic 34.1° (10.7), Asian 28.7° (14.6), multi‐racial/other 31.3° (14.9). Serum 25OHD increased 0.6 ng/mL per 10° increase in ITA° value (P<0.001). However, addition of skin color to models that included race/ethnicity did not increase explanatory value of the model (R2 for models with and without skin color were 0.19). Most urban schoolchildren had inadequate 25OHD, but prevalence of inadequate status was higher in racial/ethnic minorities. Objective measures of skin color did not explain additional variability in 25OHD beyond that explained by parent‐reported race/ethnicity. Supported by NIH/ODS #3R01HL106160.

Grant Funding Source: NIH/ODS #3R01HL106160
Effect of Whole Wheat and its Fractions on Adiposity, Insulin Resistance, and Fatty Liver
Authors: Dos Santos Ana C.G. , Jonnalagadda Satya , Gallaher Daniel
Published: June 2021

Whole grains intake has been shown to decrease insulin resistance. Whether whole wheat decreases insulin resistance and, if so, which fraction of wheat is responsible, is unknown. Therefore we examined the effects of whole wheat and wheat fractions on adiposity, insulin resistance, and fatty liver, using Zucker Diabetic Fatty rats as model of obesity with diabetes. Rats were fed diets containing either 65% whole wheat flour, 54% refined wheat flour, 9.4% bran, 1.6% germ or 15% germ for 5 weeks. No differences in weight gain were observed between the obese control and the obese wheat‐fed groups. Adiposity, determined by DEXA scanning and fat pad weights, did not differ statistically between the obese control and any obese wheat groups. Insulin resistance, measured by glucose tolerance and insulin tolerance tests, showed that whole wheat and all wheat fractions, except bran (trend only), improved insulin resistance. Liver lipids were determined gravimetrically as a measure of fatty liver. Whole wheat and all wheat fractions decreased total liver lipids, whereas bran and germ also reduced liver cholesterol. The reduction in insulin resistance and fatty liver suggests that consumption of whole wheat or its fractions may reduce the risk of developing type 2 diabetes. Supported by General Mills, Inc.
Bone protective effects of dried plum is through increasing osteoprotegerin and suppressing sclerostin levels
Authors: Hooshmand Shirin , Brisco Jayme , Elam Marcus , Chai Sheau C , Arjmandi Bahram H.
Published: June 2021

Although several studies have confirmed the bone protective properties of dried plum, its exact mechanisms of action remain unclear. Recent research has shown that osteocytes may control bone formation via production of sclerostin and bone resorption via the receptor activator of nuclear factor kappa‐B ligand (RANKL) and its inhibitor osteoprotegerin (OPG). To investigate the mechanism of action of dried plum in reversing bone loss, we measured serum sclerostin and OPG levels in osteopenic postmenopausal women (n=160). Participants were randomly assigned to either the treatment group of 100g/day dried plum or a comparative control for one year. All participants received 500 mg Ca/400 IU vitamin D daily. Bone mineral density of lumbar spine, forearm, hip, and whole body were assessed at baseline and at the end of the study using DXA. Blood samples were collected at baseline and 12 months to assess bone biomarkers. Dried plum significantly increased BMD of ulna and spine in comparison to the control group. In comparison with corresponding baseline values, dried plum decreased serum sclerostin levels by −1.2% vs. +3.8% in the control group and increased OPG levels by +4.62% vs. −2.15% in controls. Hence, these preliminary data indicate that the positive effects of dried plum on bone are in part due to inhibition of sclerostin and promotion of OPG.
Bone loss during hematopoietic cell transplantation (HCT) in children
Authors: Bechard Lori J. , Feldman Henry A. , Gura Kathleen , Venick Robert , Guinan Eva C. , Gordon Catherine , Duggan Christopher
Published: June 2021

HCT may be deleterious to bone health and vitamin D status in children, but longitudinal studies are few. We conducted a multicenter, prospective cohort study to define time related changes in bone health markers during the first 100 days following allogeneic HCT in 26 children. Bone mineral density (BMD), bone mineral content (BMC), bone‐specific alkaline phosphatase (BSAP) and serum 25‐hydroxyvitamin D (25‐OHD) concentrations were measured at baseline, 30 days, and 100 days after HCT. Mean BMD and BMC Z‐scores (mean ± SD, −0.48 ± 1.09 and −0.59 ± 1.36, respectively) were normal at baseline. Repeated‐measures analysis revealed a 0.5 decline in BMD Z‐score (p<.0001) over the 100 day study period, when adjusted for age, sex, Tanner stage, lean mass, fat mass, resting energy expenditure, total energy intake, insulin sensitivity, and serum phosphorus. BMC and BSAP concentrations also declined significantly. Adjusted mean (SE) serum 25‐OHD concentrations declined from 30.1 ± 2.9 ng/ml at baseline, to 18.9 ± 2.0 ng/ml at 100 days after HCT. At 100 days after HSCT, 50% of patients were vitamin D deficient (serum 25‐ OHD <20 ng/ml). Significant bone loss and vitamin D deficiency occur in children in the first 100 days following allogeneic HCT. Strategies to attenuate acute bone loss in this setting are needed.

Grant Funding Source: This project was supported by the Massachusetts Vitamin Litigation Grant and NIH grants M01‐ RR02172, UL1 RR025758–01, M01‐RR00865, M01‐RR00188, and K24 HD 058795.
Rsu‐1 (Ras suppressor protein 1) ‐ A potential tumor suppressor in Hepatocellular Carcinoma
Authors: Donthamsetty shashi , Mars wendy , Michalopoulos George
Published: June 2021

One of the signaling pathways that has been implicated in hepatocellular carcinoma (HCC) development is the Ras pathway. However, Ras mutations are extremely rare in HCC. It is likely that proteins that control this pathway are deregulated/mutated leading to increased activation of Ras signaling. Our lab has shown that 10% of human HCC cases had deletions in Rsu‐1. This protein is known to regulate Ras signaling atleast in mammary tumors and glioblastoma. The present work evaluated the potential role of Rsu‐1 as a tumor suppressor in HCC. Our preliminary data have shown that 1. Rsu‐1 is either absent or downregulated in most human HCC samples. 2. Most of the human HCC cell lines (Hep3B, HepG2, and SNU 449) examined also show down regulation of Rsu‐1 as compared to normal human hepatocytes which is associated with increased Ras activation. The only cell line that had Rsu‐1 levels comparable to human hepatocytes was HUH7 3. Overexpression of Rsu‐1 in HCC cells inhibits cell growth and migration in vitro and 4. Surprisingly, Rsu‐1 does not seem to regulate the activation of Ras; instead, it inhibits the downstream target of Ras called RhoGTPase. Based on these preliminary data Rsu‐1 seems to be a novel regulator of Ras signaling in the hepatocytes and may act as a tumor suppressor by inhibiting the downstream target of Ras signaling pathways called RhoGTPase.
Rotation and reorganization of dissection groups promotes professionalism and enhances cadaver based learning
Authors: Mork Amy Lovejoy , Klement Brenda , Paulsen Douglas F. , Wineski Lawrence E.
Published: June 2021

In order to promote both professionalism and exposure to human anatomical variation, the integrated first year medical curriculum at Morehouse School of Medicine has developed a program combining periodic dissection group rotation and reorganization with regular group quizzes. Reorganization involves changing the membership of each dissection team; rotation involves moving teams to new cadavers. Group quizzes are shared scoring demonstrations of each team's dissections. Group restructuring ensures each dissecting team is composed of academically and demographically diverse individuals, repeatedly introducing students to new learning cohorts. Unlike traditional static group assignments, dissection team reorganization advances professionalism by tightly linking individual and group responsibility and accountability, emphasizing cooperative work habits within shifting environments, and underscoring the need for ongoing development of interpersonal communication and tolerance. Dissection of multiple cadavers throughout the course highlights human anatomical variation, one of the major educational benefits of cadaver based gross anatomy. Group quizzes serve to further promote cooperative engagement in active learning and support among group members. Student buy‐in to this methodology has been very positive, and we regard this dissection system as highly successful.

Grant Funding Source: Supported in part by NIH PO3 1B04017 and G12‐RR 03034
Genetic Approaches to Cancer
Authors: Elledge Steve , Pavlova Natasha , Davoli Teresa , Solimini Nicole
Published: June 2021

Breast cancer is a collection of diseases with distinct clinical behaviors and underlying genetic causes. We have searched genetically for genes that have cancer relevant phenotypes including genes that alter cellular proliferation, cellular transformation, cell survival, cellular senescence, and genes that are essential for the proliferation of cancer cells. We have approached this by the generation of libraries of shRNAs for loss of function experiments and libraries of ORFs for gain of function experiments. We will discuss these technologies and how they can be applied to the functional dissection of genes important for breast cancer. Using these new technologies we have identified new oncogenes and tumor suppressors. We find that tumor cells selectively delete negative growth regulators and suggest that the deletion of clusters of these genes may drive tumorigenesis by haploinsufficiency. These recurrent deletions also avoid deletion of one copy of many essential genes through haploinsufficiency. We put forward the Cancer Gene Island model that postulates that tumors select for hemizygous loss of islands of gene enriched in negative regulators of proliferation and depleted in essential genes to promote tumor cell proliferation through cumulative haploinsufficiency.
Evolution of RNA Binding Proteins and Their Regulatory Networks
Authors: Herschlag Dan
Published: June 2021

In recent years the ubiquity of RNA as an important player in regulation has become apparent. Keene's seminal ‘RNA Operon’ hypothesis posited that RNAs encoding proteins that function or localize together would be bound by common proteins, and this hypothesis was strongly supported by our work on RNA targets of several proteins, which showed that individual RNA binding proteins (RBPs) can bind hundreds of related RNA targets. Looking deeper at the patterns of RNA binding sites, we can now identify families of RNAs that are combinatorially bound by RBPs and likely function together. Conversely, we have also identified RBPs that appear to compete for common RNA binding sites. Interestingly, these RNA targets fall into two groups –functional sites of high specificity that are positioned within a particular region of the mRNA and are evolutionarily conserved– and a group that appear to be less strongly bound, are not constrained to a particular mRNA region, are subject to greater competition, and are not conserved. We are also able to trace the evolutionary history of sets of RNA binding sites and the RBPs that occupy these sites. This information improves the ability to identify RBP targets and, more profoundly, reveals fascinating aspects of how changes in RBPs and their targets are part of the ‘rewiring’ process that underlies adaptation and specialization to form distinct species through evolution.
Nutrition in Medicine: Effectiveness of a web‐based curriculum for obstetrics and gynecology residents
Authors: Kaplan Barrie Suskin , Karkowsky Chavi Eve , Kohlmeier Martin , Dayal Ashlesha , Chazotte Cynthia , Landsberger Ellen
Published: June 2021

OBJECTIVES

To assess the effectiveness and feasibility of a web‐based nutrition education curriculum on improvement of clinical practice skills of Obstetrics and Gynecology (OBGYN) residents.

STUDY DESIGN

Pre & post‐intervention surveys compared practices and confidence level of OBGYN residents for nutritional issues before and after an interactive web‐based nutrition curriculum*. The intervention consisted of 3 modules from the Nutrition in Medicine/Nutrition Education for Practicing Physicians program. Data was analyzed using paired T tests & chi‐square tests for non‐paired categorical variables. P value <0.05 was considered significant.

RESULTS

36/44 of the residents participated. After completing the modules, residents reported they were more likely to complete many of the discussed skills. Provider confidence in counseling regarding nutritional issues improved in all areas. More residents were willing to refer patients to bariatric surgery. 35/36 found the modules informative and most (86.1%) believed their clinical skills improved.

CONCLUSION

The web‐based nutritional curriculum improved the clinical skills of OBGYN residents. This is a feasible and acceptable method of learning for practicing physicians.
The Metabolic Fingerprints of Prostate Cancer
Authors: Priolo Carmen , Pyne Saumyadipta , McDunn Jonathan , Rose Joshua , Ravasz Erzsebet , Zadra Giorgia , Photopoulos Cornelia , Scaglia Natalia , DeMarzo Angelo , Loda Massimo
Published: June 2021

Cancer cells undergo profound metabolic changes as a result of either abnormal signaling pathways or mutations occurring in genes encoding metabolic enzymes. A fundamental unanswered question is whether all oncogenic drivers harness a similar metabolic response or whether each oncogenic event results in its own specific metabolic program.

Mass spectrometry‐based metabolomics was performed on immortalized human prostate epithelial cells transformed by the MYC and Akt oncogenes, transgenic mice driven by the same oncogenes under the control of a prostate‐specific promoter and human prostate tumors that were characterized for the expression and activation of these proteins, respectively. Genomic and gene expression analyses of the prostate tumors were also carried out. Through metabolite profiling and metabolite set enrichment analysis ran across the three datasets (human prostate cells, transgenic mice and human prostate tumors), we found that Akt1 drives primarily aerobic glycolysis while MYC does not elicit a Warburg‐like effect and significantly enhances glycerophospholipid synthesis instead. This regulation is Gleason grade‐ and pathological stage‐independent.

Our data provides evidence that prostate tumors exhibit metabolic fingerprints of their molecular phenotypes, which may have significant impact on both diagnostics and targeted therapeutics.

Funding: NIH/NCI R01 CA131945
Evidence of autophagy activity in stage V seminiferous epithelium
Authors: Young J'Nelle (Ellie) , Saber Nelly , Vogl A. Wayne
Published: June 2021

Autophagy involves the breakdown of cytosolic components in conjunction with lysosomes. In this study, we test the prediction that tubulobulbar complexes (TBCs) formed by Sertoli cells in rat testes are associated with autophagy. TBCs are cytoskeleton‐related structures that are formed in association with intercellular junctions in the seminiferous epithelium (SE). TBCs are most defined at apical junctions between Sertoli cells and spermatids, although they also are found at basal junctions between adjacent Sertoli cells. TBCs contain a tubular region surrounded by a cuff of actin and a distal region that is associated with cisternae of endoplasmic reticulum. Mounting evidence suggests that Sertoli cells internalize junction proteins via the budding off of double‐membrane vesicles from TBCs followed by fusion with lysosomes resulting in degradation or recycling of junction proteins to newly forming junctions. We used immunological probes to explore the potential of autophagy activity in the SE. Immunostaining of the epithelium suggests that autophagy marker LC3A is not present at apical TBCs in rat testes as predicted but rather, is present elsewhere in the cells, particularly during Stage V of spermatogenesis. Our results indicate that autophagy is not associated apical junction disassembly in Sertoli cells.

Grant Funding Source: NSERC
Role of the heart beat during early s‐looping of the early embryonic vertebrate heart
Authors: Molinaro Kateri , Chu‐LaGraff Quynh , Ramasubramanian Ashok
Published: June 2021

The embryonic vertebrate heart goes through a process known as c‐looping during its development where it forms a distinctive c shape between 36 (stage 10) and 48 (stage 12) hours of incubation. Early s‐looping begins at 48 hours and continues until 56 hours of incubation (stage 16). At this stage, the future ventricle moves to a position below what will become the atrium. Previous studies on c‐looping indicate that the heartbeat is not necessary for c‐looping. However, the role of the heartbeat during early s‐looping is unknown. Here we investigate the role of the heartbeat in the formation of the heart at this stage. Using verapamil, a potent calcium‐channel blocker, chick embryos were isolated and incubated in chick media containing 30μM verapamil for 5 hours. Embryos were imaged before the addition of and immediately after the addition of verapamil, and after the 5 hour incubation. Preliminary studies indicate that verapamil causes the heart to swell and become stiff, resembling a water balloon. These preliminary results suggest that verapamil causes swelling and stiffness similar to heart edema, a known side effect of verapamil and other calcium channel blockers. We will present findings detailing whether or not s‐looping requires verapamil‐mediated heart contractions.
Control of adipogenic transformation in vascular smooth muscle cells by the transglutaminase 2/b‐catenin signaling axis
Authors: Beazley Kelly E , Williams Katelyn , Nurminskaya Maria V
Published: June 2021

In atherosclerosis, smooth muscle cells (SMCs) undergo a unique process known as “phenotype modulation,” transitioning from a quiescent, contractile phenotype to an adipocyte‐like state. Increased b‐catenin signaling has been observed in atherosclerotic plaques, and deficiency of transglutaminase 2 (TG2) – an endogenous activator of b‐catenin – reduces lipid accumulation in the mouse model of atherosclerosis. Here we examined whether inhibition of the TG2/b‐catenin signaling axis in SMCs can prevent their phenotypic transition. Adipogenesis was induced in aortic SMCs using a high‐lipid culture medium, and detected histologically by Oil Red‐O and on the molecular level by expression of adipogenic markers. Pronounced adipogenic differentiation was detected in 6 day‐cultures. Activation of both TG2 enzyme and b‐catenin signaling occurs 4 days earlier, suggesting that this signaling axis may mediate the transition. Inhibition of TG2 by genetic ablation or pharmacologically suppressed both lipid accumulation and b‐catenin activation in aortic SMCs. In addition, direct inhibition of b‐catenin signaling with siRNA for b‐catenin efficiently prevented SMC adipogenesis. Together our results support a key role for the TG2/b‐catenin signaling axis in the transition of SMCs to an adipogenic phenotype. This research was supported by NIH grants R01HL093305 and T32AR007592.
The role of APOE genotype on neuropathological changes in human's olfactory bulb
Authors: Aboud Orwa , Griffin Sue T , Biedermann JoAnn , Mrak Robert E
Published: June 2021

Changes in olfaction often precede clinical signs and symptoms of Alzheimer's disease (AD) and other neurodegenerative disorders. We have shown self‐propagating cycles in which proinflamatory cytokines IL‐1 and S100B induce each other in glia and induce excess expression of the precursors of Aβ plaques and neurofibrillary tangles.

Anatomical and pathological features were determined in frozen and paraffin‐embedded tissue sections of olfactory bulbs resected from autopsy tissue of 15 patients (5 APOE ε3,3 neurologically normal, 5 APOE ε3,3 and 5 APOE ε4,4 AD patients) with an age range from 68 to 85 yrs.

Microscopical assessment of the olfactory bulb showed that there were fewer glomeruli and the glomerular layer was thinner in AD patients than in controls. Olfactory bulbs of all APOE ε4,4 AD patients had many Aβ plaques, and most of them were “senile” type, mature dense core neuritic plaques compared to the diffuse non‐neuritic plaques present in some APOE ε3,3 AD patients. Neurologically normal controls had fewer plaques than AD, and those plaques were of the diffuse, non‐neuritic type, and there were fewer activated GFAP immunoreactive glia overexpressing S100B than were present in either of the AD patient groups.

Our findings suggest that olfaction should be tested and noted for assessing the disease progression, especially in those with APOE ε4,4 genotype.

Grant Funding Source: NIA‐AG12411 and the Windgate Foundation
A potential role for LIM protein Ajuba in repression of DNA damage response in human cells
Authors: Kalan Sampada , Matveyenko Anastasiya , Loayza Diego
Published: June 2021

Mammalian telomeres are specialized DNA‐protein structures located at each end of chromosomes. Telomeric DNA is bound by a six‐protein complex called shelterin, which prevents DNA damage response activation at the ends.

In addition to shelterin, several accessory proteins associate with telomeres and assist in proper functioning of the complex. Our laboratory has discovered interaction between shelterin and LIM domain proteins. Of those, TRIP6 and LPP participate in the repression of the DNA damage response at telomeres.

Here, using immunoprecipitation, we show that another LIM protein, Ajuba, associates with POT1 in tumor cells. Ajuba was detected at telomeres by chromatin immunoprecipitation. We also discovered that depletion of Ajuba by siRNA caused decrease in cell proliferation and increase in DNA damage response. An overall increase in p53‐BP1 damage foci was observed in nuclei of Ajuba‐depleted cells. By using FACS analysis, we observed an accumulation of cells in S phase. An activation of p53 with concomitant hyperphosphorylation of Rb was observed. Our results point to a vital role of Ajuba in the repression of endogenous DNA damage response in tumor cells, and could provide insights in understanding of senescence or immortalization of human cells.

This research was funded by National Center for Research Resources (NCRR), a component of the National Institutes of Health (NIH).
The X‐ray Crystal Structure of Escherichia Coli RNA Polymerase σ70 Holoenzyme
Authors: Murakami Katsuhiko
Published: June 2021

Escherichia coli RNA polymerase (RNAP) is the best‐studied bacterial RNAP and has been used as the model RNAP for potential RNAP‐targeting antibiotic screening and evaluation. However, the X‐ray crystal structure of E. coli RNAP has been limited to individual domains. In this study, E. coli RNAP core enzyme (E) and holoenzyme (Eσ70) were prepared from overexpressed proteins. The holoenzyme was crystallized and solved its X‐ray structure at 3.8 Å resolution. The E. coli holoenzyme structure shows the σ region 1.1 (σ1.1) and α subunit C‐terminal domain (αCTD) structures for first time in any bacterial RNAP. The structural comparison between the E. coli and Thermus RNAPs shows that the ω subunit structure is different in E. coli from that in Thermus that may be related to the fact that the only E. coli RNAP is able to respond ppGpp for transcription regulation. The σ1.1 is positioned at the RNAP DNA binding channel and completely blocks DNA entry to the RNAP active site. The structure reveals that the σ1.1 contains a basic patch on its surface, which may play an important role in the DNA interaction to facilitate the transition from the closed to open promoter complexes. The αCTD is positioned next to the σ domain 4 (σ4) with a fully stretched linker between the N‐ and C‐terminal domains. E. coli RNAP crystals can be conveniently prepared from overexpressed proteins, allowing further structural studies of bacterial RNAP mutants, including antibiotic resistant RNAPs.
Expression of the mouse Lin28a gene is epigenetically regulated by histone modification
Authors: Pang Alan L.Y. , Title Alexandra C. , Rennert Owen M.
Published: June 2021

The RNA‐binding protein Lin28a is considered to be a “stemness” marker. Lin28a is expressed predominantly in pluripotent cells and cancer cells. However, the regulation of its expression has not been thoroughly studied. We found that the Lin28a promoter DNA was transcriptionally active in non Lin28a‐expressing cells, which made us to hypothesize that the expression of Lin28a is regulated epigenetically. By bisulfite‐sequencing analysis, we did not observe a correlation between the absence of gene expression and Lin28a promoter DNA methylation. Consistently, Lin28a expression was not activated in non‐Lin28a expressing cells by 5‐aza‐2′‐deoxycytidine treatment. Instead, treatment of NIH/3T3 cells with histone deacetylase inhibitor trichostatin A (TSA) induced Lin28a expression and made the Lin28a promoter DNA sensitive to nuclease digestion, suggesting that Lin28a expression was mediated by histone acetylation and a relaxation of chromatin. A significant enrichment of H3K9Ac and reduced occupancy of H3K9Me2 was detected on Lin28a promoter in Lin28a‐expressing cells. The opposite pattern of histone modifications was observed when Lin28a expression was silenced. The histone modification pattern also influenced the abundance of RNA polymerase II on Lin28a promoter. Taken together, our results indicate that Lin28a expression in mouse cells is epigenetically regulated by histone modification.
Serum and glucocorticoid inducible kinase regulates hERG channels through inhibition of Nedd4–2 and activation of Rab11 recycling endosomes
Authors: Lamothe Shawn Michael , Zhang Shetuan
Published: June 2021

The human ether‐a‐go‐go‐related gene (hERG) encodes the rapidly activating delayed rectifier potassium channel (IKr). Loss of function of hERG channels due to mutations or drug‐induced blockage can cause long QT syndrome, leading to ventricular arrhythmias or sudden death. Physical or emotional stress is known to affect ion channel activity. Stress hormones such as cortisol regulate the expression of various genes including the serum‐ and glucocorticoid‐inducible kinase (SGK). Using patch clamp, Western blot, co‐immunoprecipitation, and immunocytochemistry methods, we demonstrate that SGK isoforms SGK1 and SGK3 increased hERG current (IHERG) and the expression level of mature hERG proteins. We recently showed that mature hERG channels are degraded by ubiquitin ligase Nedd4–2 via enhanced ubiquitination. We observed that SGK1 and SGK3 overexpression enhanced Nedd4–2 phosphorylation which is known to inhibit Nedd4–2 activity. However, removal of the Nedd4–2 effect on hERG channels by disrupting Nedd4–2 target motif in hERG reduced but did not eliminate the SGK‐induced increase in hERG expression. Additional disruption of Rab11 proteins led to a complete elimination of SGK‐mediated hERG increase. We conclude that SGK isoforms 1 and 3 enhance hERG stability by inhibiting Nedd4–2 ubiquitin ligase and activating Rab11 recycling endosomes.

Supported by: Canadian Institutes of Health Research (CIHR)
Transthyretin regulates lysosomal degradation of hippocampal 14–3‐3ζ
Authors: Vieira Marta , Saraiva Maria João
Published: June 2021

Transthyretin (TTR) is a 55kDa homotetrameric protein mainly synthetized in liver and choroid plexus. It is the carrier protein of thyroxine and retinol in plasma and cerebrospinal fluid (CSF), and has also been shown to have neuroprotective properties both in the central and peripheral nervous system. Studies in TTR null mice (TTR−/−) revealed that absence of TTR reduces signs of depressive‐like behavior, increases neuropeptide Y levels, delay in nerve regeneration in nerve injury conditions; these mice present impaired memory when compared with wild type animals.14–3‐3 proteins very important in many cellular processes such as signal transduction, cell‐cycle regulation, cell survival, cellular trafficking, cytoskeletal organization, protein synthesis, redox‐regulation and protein folding. Their absence is also related with deficits in memory and learning. The analysis of the relationship between these two proteins is the main objective of this work. We found that hippocampus of young TTR null mice presented lower levels of 14–3‐3ζ protein, but no changes in gene expression when compared to TTR wild type littermates were noted. Cellular studies ascribed this finding to increased degradation of 14–3‐3ζ in lysosomes in the absence of TTR. Taken together, these results suggest that the lower levels of 14–3‐3ζ found in vitro and in vivo in the absence of TTR, are attributable to increased degradation in the lysosomal compartment. We hypothesize that the memory deficits found in TTR null mice might be relate to hippocampal 14–3‐3ζ deficiency.
Characterization of the Non‐catalytic Inhibitory Effect of Ubp6 on Proteasome Function
Authors: Roychowdhury Anasuya , Robert Jessica , Lee Byung‐Hoon , Finley Daniel J.
Published: June 2021

Selective protein degradation in eukaryotes is mediated primarily by the ubiquitin‐proteasome system (UPS) in which ubiquitin is attached to a target protein to signal its degradation by the proteasome. Therefore, the proteasome, a multisubunit protease, is a central enzyme in the system and controls the levels of hundreds of proteins. However, little attention has been given to how the proteasome is regulated. One important mechanism of proteasome regulation involves reversibly‐associated deubiquitinating enzymes (DUBs). One of these DUBs is the yeast protein Ubp6 (whose mammalian ortholog is Usp14). Interestingly, in addition to its catalytic activity of deubiquitination, Ubp6, has a second, highly unexpected function: a capacity to delay the degradation of ubiquitinated proteins (termed the non‐catalytic inhibitory effect). The degradation delay caused by Ubp6 might serve to ensure that the deubiquitinating activity of Ubp6 has adequate time to regulate substrate commitment to degradation. However, the mechanism of this inhibitory effect is unknown. In search of this mechanism we have characterized noncatalytic proteasome inhibition by Ubp6 with the benefit of site‐directed mutagenesis, quantitative biochemical assays, and yeast genetics.
Purification of the des‐(31,32) and des‐(64,65) intermediates of proinsulin conversion
Authors: Mackin Robert B
Published: June 2021

The biosynthesis of insulin requires proteolytic processing of a larger precursor, proinsulin (PI), by three different proteases: proprotein convertase 1 (PC1), PC2 and carboxypeptidase E. The conversion process must proceed through one of two intermediates, des‐(31,32)‐PI or des‐(64,65)‐PI, to reach the final product, insulin. I am interested in examining the structure of the intermediates to help explain the specificity of the enzyme‐substrate interactions involved in PI conversion. I have previously reported the use of partial trypsin/carboxypeptidase B digestion to generate the intermediates, and the use of anion‐exchange chromatography to separate the intermediates from the substrate and products using anion‐exchange chromatography. The intermediates have now been separated from one another and purified from miscleaved PI using reversed‐phase high performance liquid chromatography. Starting from 5 mg of PI, 0.4 mg of des‐(31,32)‐PI at 99% purity and 0.2 mg of des‐(64,65)‐PI at 90% purity can be obtained. It should therefore be possible to scale up purification of the intermediates for structural characterization and examination of PI cleavage specificity.
Comparison of TaqMan and SYBR Green qPCR Methods for Quantitative Gene Expression in Plants
Authors: Cao Heping , Shockey Jay M.
Published: June 2021

Quantitative real‐time‐PCR (qPCR) is widely used for gene expression analysis due to its large dynamic range, tremendous sensitivity, high sequence‐specificity, little to no post‐amplification processing, and sample throughput. TaqMan and SYBR Green qPCR assays are two frequently used methods. However, direct comparison of both methods using the same primers and biological samples is still limited. We compared both assays using seven RNAs from the seeds, leaves, and flowers of tung tree (Vernicia fordii) which produces high‐value industrial oil. High quality RNA was isolated from tung tissues, as indicated by high rRNA ratio and RNA integrity number. qPCR primers and TaqMan probes were optimized. Under optimized conditions, both qPCR assays gave high correlation co‐efficiency and similar amplification efficiency, but SYBR Green qPCR over‐estimated the expression levels regardless of the genes and tissues tested. This is validated using well‐known Dgat2 and Fadx gene expression in tung tissues. The results demonstrate that both assays are reliable for determining gene expression in tung tissues and that TaqMan assay is more sensitive but generates lower calculated expression levels than SYBR Green assay. This study suggests that any discussion of gene expression levels needs to be linked to which qPCR method is used in the analysis.
Modulation of cellular and mitochondrial reactive oxygen species production by external magnetic fields
Authors: Martino Carlos Federico , Castello Pablo
Published: June 2021

This work studied the modulation of production of extracellular, intracellular, and mitochondrial superoxide and hydrogen peroxide (H2O2) in cultured yeast and isolated mitochondria exposed to external static and weak radio frequency magnetic fields. Cells and isolated mitochondria were exposed to a combination of magnetic fields: 150 μT static fields (Case I); 45 μT static fields in perpendicular to 10 MHz RF signal (Case II); 45 μT static fields in parallel to 10 MHz RF signal (Case III). Controls were exposed to 45 μT static field. H2O2 production was measured by fluorometric technique. The production of H2O2 was evaluated in the presence of inhibitors of the mitochondria respiratory chain. Superoxide production was measured by a set of hydroxylamine spin probes with EPR technique. CAT1‐H detected only extramitochondrial and extracellular superoxide. MitoTEMPO‐H was used to target intramitochondrial superoxide in intact cells. CMH was used to detect superoxide in the cytoplasm. The results presented here suggest that, a) weak static and RF magnetic fields modulate the production of mitochondrial reactive oxygen species, b) complex I of the electron transport chain is involved in the H2O2 production. These findings shed light on the mechanism through which external weak magnetic fields modulate ROS production by modifying mitochondrial functioning.