bluuebunny/crossref_metadata_2025 huggingface dataset:
Toddler Feeding Practices in Early Head Start and Migrant/Seasonal Early Head Start Mothers
Authors: Voorheis Grace , Papathakis Peggy , Nicholson Lisa
Published: June 2021

Objective

Measure baseline data on parental feeding practices of low‐income, Latina mothers of toddlers (1‐3 years) enrolled in Head Start in order to develop parent education.

Methods

Sixty‐six mothers in Early Head Start (EHS, n=25) and Migrant/Seasonal Early Head Start (MSHS, n=41) completed a 24 item Likert scale (1‐5 with 1=Never/Not confident) to assess self‐efficacy, behavior and intentions regarding 4 domains of toddler feeding practice. These were scheduling, self‐regulation, indulging, and parental role modeling. Mean Likert scores for each question were analyzed by subgroup.

Results

Mothers in both groups were similar in their parental feeding practices; they had low self‐efficacy for allowing their child to play with food and high self‐efficacy for discerning when their child was full. Both groups indicated they kept a regular meal schedule more frequently than a snack schedule and infrequently encouraged their child to eat more when he/she has indicated fullness. Self‐efficacy regarding avoiding indulging their child's favorite foods was higher in EHS mothers than MSHS mothers (p=0.041). For MSHS mothers, indulging their toddler with sweets (p=0.024) and role modeling eating fruits (p=0.014) were significantly more frequent than in EHS mothers. EHS mothers allowed their child to snack all day more frequently than MSHS mothers (p=0.013).

Conclusion

Indulging in favorite foods appears more common in families in MSHS than in EHS. Frequent consumption of fruit is higher in MSHS than EHS families, potentially because these mothers are employed in fruit‐picking industry. Future educational interventions should focus on encouraging authoritative feeding styles to avoid indulgence and maintain a schedule throughout the day.
Can Nutrition Education Support Academic Achievement? Evidence Supporting Integration of Food & Nutrition Education into Intermediate Curricula
Authors: Carraway‐Stage Virginia , Kolasa Kathryn , Diaz Sebastian , Duffrin Melani
Published: June 2021

Several nutrition education (NE) curricula use integrative approaches, however few studies have investigated its impact on academic achievement. Demonstration that NE instruction can improve achievement in science and mathematics is necessary for improving teacher engagement in obesity prevention activities. The purpose of this study was to explore associations between science (SK), mathematics (MK), and nutrition knowledge (NK) to provide evidence that integrating food/nutrition education in the curriculum supports academic achievement. A secondary analysis of a quasi‐experimental study was conducted using 33 fourth‐grade classrooms across North Carolina and Ohio. Researchers analyzed data using multiple linear regression. Post‐test‐NK score served as the dependent variable, baseline‐NK, post‐test SK and MK served as the independent, or predictor variables. Results revealed that the hypothesized model predicted post‐NK for full sample (F (437)=149.4, p<.001; Adjusted R2=.51), and intervention [F(268)=111.1; p<.001] and comparison groups [F(169)=39.1; p<.001]. For the three domains, SK and MK were predictive of NK with students exposed to the food‐based curriculum demonstrating stronger associations among the domains of knowledge. Using an integrative food‐based curriculum can significantly improve SK and MK among fourth‐graders and may help teachers meet multiple academic content standards, while efficiently using limited classroom time and increasing the amount of NE provided in the classroom.
Self‐Efficacy is Associated with Healthy Eating Behaviors for Fifth Grade Students
Authors: Hall Elisha , Chai Weiwen , Koszewski Wanda , Albrecht Julie
Published: June 2021

Objective

To assess the relationships between three integral constructs of the Social Cognitive Theory (SCT), knowledge, self‐efficacy, and behavior, and specific self‐efficacy and behavior variables for healthy eating among fifth grade students.

Methods: Ninety‐eight fifth grade students recruited from four local elementary schools completed the Healthy Habits Survey, a 37‐item nutrition and physical activity survey covering knowledge, self‐efficacy, behavior and demographics. Relationships between knowledge, self‐efficacy, and behavior were assessed using Pearson Correlation Coefficients. Differences in behavior and knowledge scores between participants with high and low self‐efficacy were examined using a t‐test.

Results: Total scores for self‐efficacy and behaviors were positively correlated (r=0.32, P=0.002); however, total knowledge scores were not associated with self‐efficacy (r=0.02, P=0.88) or behavior scores (r=0.19, P=0.10). In terms of specific eating behaviors, more than two thirds of fifth grade students reported that they did not eat any fruits or vegetables or ate them only once on a typical day, aligning with trends seen nationally on low fruit and vegetable consumption in children.

Conclusions

Results indicate that self‐efficacy is significantly associated with behavior, therefore should be integrated when creating interventions for behavior change. In addition, specific behaviors such as fruit and vegetable consumption continue to fall below dietary recommendations and need continued programming to target this issue.

*This research was supported by USDA NIFA Growing Healthy Kids through Health Communities grant; 2011‐67001‐30011.
Therapeutic Use of Ground Flaxseeds for Type 2 Diabetes
Authors: Ricklefs Kristin , Johnston Carol , Sweazea Karen
Published: June 2021

Type 2 diabetes (T2D) is an independent risk factor for cardiovascular disease. Hyperglycemia and chronic low‐grade inflammation, characteristics of T2D, additionally contribute to vascular dysfunction. Prior studies have shown that plant‐based fibers reduce inflammation and hemoglobin A1c (HbA1c) as well as attenuate T2D‐associated dyslipidemia. The objective of this 8‐week parallel‐arm controlled feeding trial is to determine if ground flaxseed (28 g/d) supplementation to a normal diet will decrease HbA1c concentrations as well as attenuate cardiovascular risk markers in subjects with T2D as compared to a control group who consumed ground psyllium (9 g/d). Fiber content of the supplements was matched as fiber is independently correlated with decreased postprandial blood glucose. To date n=4 subjects per group have completed the study. Results were analyzed using 2‐way RM ANOVA. At 8 weeks flaxseed significantly reduced waist circumference (‐1.25±0.29; p=0.034), systolic BP (‐7.0 +‐ 2.71; p=0.004), and diastolic BP (‐3.75±1.89; p=0.003). There were no significant differences within group following 8 weeks of ground psyllium. Since fiber content was matched, other components of flaxseeds such as folic acid or alpha‐linoleic acid may be responsible for the observed differences within groups. The preliminary findings therefore suggest ground flaxseed may have therapeutic potential for subjects with T2D to regulate factors which contribute to T2D complications such as BMI and blood pressure, although additional subjects will be required to confirm these findings. This study is funded by a grant from the AmeriFlax Council.
Serum fatty acids concentrations are associated with plasma adipokines during pregnancy
Authors: Lepsch Jaqueline , Pinto Thatiana , Farias Dayana , Franco‐Sena Ana , Rebelo Fernanda , Lima Natália , Vaz Juliana , Cunha Marcelo , Kac Gilberto
Published: June 2021

Objective

To investigate if serum concentrations of total saturated (SFA), polyunsaturated (PUFA) and its fractions are associated with plasma adiponectin and leptin concentrations during pregnancy.

Methods

Prospective cohort of 221 pregnant women followed at 5‐13th, 20‐26th and 30‐36th gestational weeks. Blood samples were collected after 12 hours of fasting. FA concentrations were determined using fast gas‐liquid chromatography. Plasma adiponectin (U/ml) and leptin (ng/dl) were estimated using ELISA. Statistical analyses included mean concentrations of the adipokines according to tertiles of FA distribution and longitudinal linear mixed‐effect models, adjusted for BMI, time and total energy intake.

Results

Mean adiponectin was lower in the 3th tertile of SFA when compared to the 2nd and 1st tertiles (2nd trimester: 1st tertile=11.4; 2nd=10.4; 3rd=9.26). Mean of leptin was higher in the 3th tertile of SFA compared to the 2nd and 1st tertiles (2nd trimester: 1st tertile=27.5; 2nd=33.3; 3rd=36.1). The serum concentrations of SFA (β=‐0299, p=0.022) and its fractions were negative and significantly associated with adiponectin [(14:0; β=‐0.021, p=0.012); (16:0; β=‐0.004, p=0.002); (18:0; β=‐0.009, p=0.051)]. Total PUFA n‐6 (β=0.003, p<0.001) and 18:2n6 (β=0.002, p=0.005) were positively associated with adiponectin. SFA (β=1.092, p=0.017), 14:0 (β=0.092, p=0.002) and 20:3n‐6 (β=0.055, p=0.006) was positively associated with leptin.

Conclusions

Total serum SFA and fractions were negatively associated with adiponectin and positively associated with leptin and total n‐6 PUFA was positively associated with adiponectin during pregnancy.
Cardiometabolic Profiles and Gene Expression Following Fish Oil Supplementation: Is There a Role for FADS Genotypes?
Authors: Roke Kaitlin , Zulyniak Michael , Gerling Christopher , Whitfield Jamie , Spriet Lawrence , Mutch David
Published: June 2021

Consumption of omega‐3 fatty acids (FAs) modifies cardiometabolic risk factors in hyperlipidemic and diabetic individuals; however, their effects in young healthy adults are unclear. We examined the impact of fish oil supplements on blood cardiometabolic markers, FA composition, eicosanoid profiles and gene expression. We then examined whether these endpoints varied with an individual's fatty acid desaturase (FADS) genotype. Young male adults (18 – 25 yrs) consumed daily fish oil supplements (up to 3g omega‐3 FAs) for 12 weeks. Significant reductions in blood triglycerides were seen by week 12. Paradoxically, two omega‐6 derived eicosanoids (PGF2α and TXB2) were increased with supplementation. Whole blood gene expression revealed no differences in key genes regulating eicosanoid production, i.e., ALOX5, ALOX12, ALOX15, COX‐2 and iPLA2. FA analysis showed increases in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) content in serum and red blood cells (RBCs), and decreases in arachidonic acid (AA). Compared to major allele carriers, individuals with FADS minor allele variants experienced greater increases in EPA levels in serum and RBCs during supplementation. There were no genotype differences for DHA or AA levels. Collectively, our results showed that fish oil supplements alter the eicosanoid profile independent of changes in gene expression. Additionally, we demonstrated that changes in FA levels after supplementation are influenced by an individual's FADS genotype. Further investigations regarding the relationship between fish oil supplements, eicosanoid profiles, and FADS genotype are warranted.

Grant Funding Source: Supported by NSERC
Role of Trifunctional Enzyme MTHFD1 in Maintenance of DNA Stability
Authors: Kamynina Elena , Field Martha , Stover Patrick
Published: June 2021

MTHFD1 is a trifunctional enzyme possessing methyleneTHF dehydrogenase (D), methenylTHF cyclohydrolase (C) and formylTHF synthetase (S) activities. We have recently demonstrated that MTHFD1 translocates to the nucleus in S phase where it plays a key enzymatic role providing one‐carbon units for dTMP synthesis. An inborn error of metabolism associated with mutations in the human methylenetetrahydrofolate dehydrogenase 1 (MTHFD1) gene has been recently identified. The proband presented with severe combined immune deficiency (SCID), megaloblastic anemia and neurologic abnormalities. In this study, we examined the impact of MTHFD1 loss‐of‐function on DNA stability. Patient fibroblasts exhibited increased DNA damage relative to control fibroblasts. γH2AX levels, a marker of double‐strand breaks in DNA, were markedly increased in MTHFD1‐deficient patient fibroblasts relative to both control lines as well as the adenosine deaminase deficient fibroblast lines. γH2AX staining increased in patient fibroblasts when cultured in folate‐deficient medium as compared to fibroblasts cultured in folate‐replete medium, indicating that the MTHFD1‐deficient fibroblasts are sensitized to folate deficiency‐induced DNA damage.
Niacin Intakes and Status of Healthy Adults Living in Republic of Korea
Authors: Kim Young‐Nam , Choi Ji Young , Cho Youn‐Ok
Published: June 2021

The purpose of this study was to determine niacin status of Korean adults. Three consecutive 24‐hour recalls and urine samples were collected from 149 healthy adults (70 males and 79 females), aged 20‐65y, living in Seoul metropolitan area, Republic of Korea. The mean dietary niacin intakes (mean/SD) were 17.65/4.70 mg/day and 9.24/2.13 mg/1000 kcal. Total niacin intake (dietary plus supplemental) was 21.51/11.38 mg/day. The dietary intake of men was significantly higher than that of women, but no significant difference in total intake was determined by gender. Only 5% of subjects consumed total niacin less than the Estimated Average Requirements for Koreans. Average urine niacin (N1‐methylnicotinamide) excretion was 1.45/1.16 μg/g creatinine. Urinary niacin of women was significantly higher than that of men (p<0.01). There were no significant correlations between intakes and urinary excretion of niacin. Only 13% of subjects were niacin deficient (urine niacin <0.5 μg/g creatinine), but urinary niacin excretions of almost half of subjects were 0.5‐<1.59 μg/g creatinine, which is representative of low niacin status. Therefore, although niacin intake is adequate compared to the Dietary Reference Intakes for Koreans, niacin status may not be inadequate in Korean adults. Supported by 2014 research fund of National Research Foundation of Korea (NRF‐2011‐0021273).
Effect of High‐Dose Vitamin D on Antimicrobial Peptides in Ventilator‐Dependent Critically Ill Patients
Authors: Jones J , Alvarez J , Han J , Hao L , Chiang E , Brown L , Tangpricha V , Martin G , Ziegler T
Published: June 2021

Objective

Vitamin D has been shown to influence indexes of immunity, including antimicrobial peptides (AMPs). Vitamin D deficiency is common in critical illness, but the impact of vitamin D status or the response to treatment with high‐dose vitamin D on AMPs in this population is unknown. Thus, we evaluated the impact of two doses of vitamin D3 (D3) versus placebo on AMPs in ventilated, critically ill subjects.

Methods

Thirty critically ill, ventilated subjects were enrolled in a randomized, double‐blind controlled trial of enteral high‐dose D3 (total dose = 250,000 IU or 500,000 IU) given in divided doses over 5 consecutive days. Plasma 25‐hydroxyvitamin D [25(OH)D] was measured by chemiluminescence. Plasma levels of AMPs, cathelicidin (LL‐37) and human beta‐defensin‐2 (hBD‐2), were measured by ELISA, and PBMC LL‐37 mRNA by quantitative PCR at baseline, days 7 and 14 after study entry. Descriptive statistics (mean ± SD), Spearman correlations, and repeated measures ANOVA were used for analysis.

Results

The study population was 60% male, mean age was 63.5 ± 17.3 yrs, and 43% were vitamin D deficient at baseline. D3 administration significantly increased plasma 25(OH)D levels from baseline to day 14 vs placebo (500,000 IU: 20.0 ± 7.3 to 62.4 ± 16.7 ng/mL; 250,000 IU: 23.2 ± 7.8 to 50.4 ± 29.4 ng/mL; placebo: 21.5 ± 12.2 to 21.5 ± 11.3 ng/mL, respectively; p < 0.01). Baseline 25(OH)D levels were unrelated to baseline AMP indexes. Plasma LL‐37, hBD‐2, and PBMC LL‐37 mRNA did not change over time in any group.

Conclusions

Baseline vitamin D status did not influence plasma LL‐37 and hBD‐2 levels or PBMC LL‐37 mRNA in critically ill, ventilated adults. High‐dose D3 administration markedly increased 25(OH)D levels, but did not alter production of AMPs.
The Impact of High‐Dose Vitamin D on Hemoglobin Concentrations in Critical Illness
Authors: Smith Ellen , Jones Jennifer , Han Jenny , Alvarez Jessica , Chiang Ellen , Martin Greg , Ziegler Thomas , Tangpricha Vin
Published: June 2021

Objective

Anemia and vitamin D deficiency are highly prevalent in critically ill patients, and vitamin D may be involved in the regulation of iron recycling. The purpose of this study was to examine the impact of vitamin D therapy on hemoglobin concentrations in critically ill patients.

Methods

We conducted a secondary analysis of a randomized, double‐blinded, placebo‐controlled trial of high‐dose vitamin D3 (D3) therapy in critically ill adults requiring mechanical ventilation (N=30). Participants were randomized to receive a total enteral dose of 500,000 IU D3, 250,000 IU D3, or placebo administered over 5 days. Blood was drawn weekly throughout hospitalization for up to 4 weeks. Linear mixed effects models were used to assess change in hemoglobin concentration by treatment group over time.

Results

The prevalence of anemia at baseline was 93.3%. Baseline mean hemoglobin and plasma 25‐hydroxyvitamin D concentrations were 9.7 g/dL (SD=1.9) and 21.4 ng/mL (SD=9.1), respectively, and did not differ significantly across treatment groups. In the 250,000 IU D3 group, change in hemoglobin over time did not differ from placebo (P=0.5). However, in the group who received 500,000 IU D3, hemoglobin concentrations increased by 0.8 g/dL (SE=0.3) per week (P=0.005).

Conclusion

In this population of critically ill patients, treatment with 500,000 IU D3 over 5 days increased hemoglobin concentrations during hospitalization, while no significant change in hemoglobin was observed in the 250,000 IU D3 or placebo groups. These findings support a role for vitamin D in improving hemoglobin concentrations in critical illness, which should be confirmed in larger studies.

Funding: T32 DK7734‐17, R21 HL110044
Effects of Ester‐C®and Ascorbic Acid on Gastrointestinal Outcomes: A Randomized, Double‐Blind Trial
Authors: Ye Qian , Combs Maile , Mitmesser Susan
Published: June 2021

Background

Optimum vitamin C intake is not always attainable solely via food consumption, therefore supplementation may be desirable. However, vitamin C supplementation is not without notable gastrointestinal discomfort resulting from its acidic nature, which may affect overall quality of life.

Objective

To compare the effects of Ester‐C® (EC) and Ascorbic Acid (AA) on gastrointestinal symptoms and quality of life in subjects with acid‐sensitivity.

Methods

In this double‐blind, cross‐over study, 50 healthy adults with acid‐sensitivity were randomized to receive vitamin C (1000 mg for Day 1‐5, 2000 mg for Day 6‐10) provided by either AA or EC. Gastrointestinal side effects as measured by Gastrointestinal Symptom Rating Scale (GSRS) and the associated changes in quality of life as measured by Medical Outcomes Short Form‐36 (SF‐36) were assessed.

Results

An intention‐to‐treat analysis was conducted on 50 subjects (11 males, 39 females). After ten days of treatment, AA resulted in significant worsening of Abdominal pain (p=0.02), Diarrhea (p=0.047), and total GSRS score (p=0.019), while there were no significant changes in EC group. The Role Physical and Bodily Pain scores were significantly better in EC group compared to AA group over the ten‐day period (p = 0.004 and 0.008, respectively).

Conclusion

Ester‐C® is better tolerated than Ascorbic Acid in individuals with acid‐sensitivity.
Neutrophil Elastase Responses in Smokers and Nonsmokers Consuming Dried Plums
Authors: Nelson Stephanie , Zawilski Alexandra , McGill Brittany , McIntosh Marian , Kern Mark , Hong Mee Young , Hooshmand Shirin
Published: June 2021

Neutrophil elastase, a marker of pulmonary inflammation, is secreted by macrophages within minutes of cigarette smoking to combat inflammation. Excessive levels of neutrophil elastase have been implicated in the pathogenesis of emphysema. Oxidative stress induced by smoking destroys the inhibitors of neutrophil elastase, further exacerbating the uncontrolled release and activity of the enzyme. To determine the effect of dried plums, a high antioxidant food, on neutrophil elastase, nonsmokers (n=14) and smokers (n=5) underwent two trials in which they consumed 100g dried plums at one visit and an isocaloric control food (muffins) during the other. Neutrophil elastase concentrations were evaluated in blood samples that were taken at baseline and at 60, 75, 90, and 120 minutes post‐feeding, and smokers smoked one cigarette at 45 minutes post‐feeding. Results demonstrated that there were no significant changes in neutrophil elastase from baseline in nonsmokers at any time point. However, smokers experienced a significant decrease in neutrophil elastase 90 minutes after consuming dried plums (p<0.05) that did not occur after consuming muffins. The results suggest that dried plums may control the secretion and activity of neutrophil elastase, which could potentially prevent the pathogenesis of emphysema associated with smoking cigarettes. This study was funded by the California Dried Plum Board.
Sulforaphane Reduces Inflammatory Gene Expression in Microglia from Aged Mice
Authors: Townsend Brigitte , Rytych Jennifer , Johnson Rodney
Published: June 2021

Increased oxidative stress resulting from heightened microglial activation is associated with age‐related cognitive impairment, neurodegenerative disease, and response to immune stimulus. The primary compensatory mechanism for neutralizing oxidative stress is through the nuclear factor E2‐related factor 2 (Nrf2) pathway which regulates transcription of antioxidant response element (ARE) genes. Here, BV2 cells and brain CD11b+ microglial cells isolated from young adult and aged mice were used to investigate the effects of sulforaphane (SFN), a potent Nrf2‐inducing bioactive, on ARE and inflammatory gene expression associated with increased age. We hypothesized that SFN would upregulate ARE genes in primary microglial cells and that this would attenuate elevated proinflammatory cytokine expression in microglia from aged mice. In support of our hypothesis, SFN upregulated antioxidant genes NAD(P)H dehydrogenase, quinone 1 (NQO1), heme oxygenase‐1 (HMOX1), and the modulatory subunit of glutamate‐cysteine ligase, modifier subunit (GCLM) in BV2 cells and primary microglia. Primary microglia from aged mice had higher expression of interleukin (IL‐)1β, IL‐6, and inducible nitric oxide synthase (iNOS) than adult controls, and SFN reduced IL‐1β and iNOS in microglia from aged mice. Taken together these data indicate that SFN is a potential therapeutic supplement that may be beneficial for reducing microglia‐mediated neuroinflammation and oxidative stress associated with aging.

Supported by NIH RO1AG16710
Obesity‐associated Inflammation Contributes to Adipose Stromal Cells Pro‐fibrotic Phenotype
Authors: Springer N , van den Berg M , Seo B R , Fischbach C
Published: June 2021

Introduction

Obesity is linked to breast cancer pathogenesis and physical changes to the adipose microenvironment may be important. We have observed increased myofibroblasts, cells derived from adipose stromal cells (ASC) that deposit extracellular matrix (ECM), and a correlation with macrophagic inflammation in obese fat. Our objective is to assess a functional link between inflammation, ECM stiffness and ASC differentiation.

Methods

Bone marrow macrophages (BMM) or RAW 264.7 cells were cultured on hydrogels mimicking the mechanical properties of lean (1 kPa) or obese (10 kPa) adipose ECM. BMM morphology and response to LPS were judged by immunofluorescence. RAW 264.7‐conditioned media (CM) tested the effect of macrophage‐secreted factors on ASC (3T3‐L1) differentiation into adipocytes and myofibroblasts. Non‐CM and TGFβ served as controls. Adipocyte and myofibroblast differentiation were quantified by lipid droplet diameter and αSMA immunofluorescence, respectively.

Results

BMM cultured on 10 kPa v. 1 kPa gels had increased cytoplasmic extensions and actin stress fibers. BMM from 10 kPa gels trended to reduced iNOS expression post‐LPS, which was negated by inhibiting mechanosignaling. CM increased αSMA and reduced lipid droplet size (p<0.05). Myofibroblast differentiation was similar for both stiffnesses, but CM from 10 kPa gels reduced adipogenesis more (p<0.05) and comparably to TGFβ positive control.

Conclusions

Stiffness alters macrophage function and macrophage‐secreted factors promote myofibroblast and hinder adipocyte differentiation of ASC. Experiments are underway to confirm macrophage mechanosignaling as a mechanism.

Funding

NIH T32ODO011000 & 1R01CA185293, CMM U54CA143876
LEAP: Launch into Education About Pharmacology. A Pharmacology‐based Enrichment Program for College Students at Duke
Authors: Schwartz‐Bloom Rochelle , Godin Elizabeth , Perez Anthony , Wormington Stephanie , Barger Michael , Snyder Kate , Linnenbrink‐Garcia Lisa
Published: June 2021

We developed an enrichment program to introduce Duke rising sophomores interested in science to the field of pharmacology. LEAP included a summer 2‐week mini‐course in pharmacology followed by a fall research experience. In the past 3 years we have had 227 students in the summer course; 87 of them took the Fall research course. The summer course met from 9‐4 daily, covering basic principles of pharmacology the first week and focusing on the pharmacotherapy of 4 diseases the second week. The teaching included a variety of methodologies (lecture, small group inquiry‐based learning, discussions, and creative group work). Students worked on a hypothetical research proposal of their own choice during the 2 weeks and presented their plan at a poster session. In the Fall, students proposed a research project on a pharmacologic topic of interest, and with the guidance of graduate student or postdoc mentors, pursued their own research plan, including planning the experiments, ordering reagents, carrying out the research, and analyzing the data. At the end of the semester, all students gave an oral presentation of their work. We hope that introducing pharmacology to students early in their college experience will foster an interest in this field for future graduate study. Funding: NIH GM090534.
Mobile Handheld Devices Applications to Teach Rational Prescribing and Safe Medication Practices
Authors: Rajasekaran Senthil Kumar , Smith Julia , Engwall Keith
Published: June 2021

With the growing emphasis on evidence‐based medicine, medical students face the challenge of assimilating a huge volume of new information during medical school. Among the most significant trends in medical education is the emergence of high quality medical applications for smart phones and tablets. As digital natives, millennial medical students respond well to this approach.

First year medical students were chosen for this activity. We chose three apps (Micromedex, Epocrates and DynaMed) and for each app, the students were assigned questions related to a clinical case and a corresponding app to answer them. The tasks included a drug interaction check, adverse effect check, cost effective alternative check and patient advice and reference the levels of evidence (I, II and III). Also, students were directed to use the apps to perform similar tasks with the prescription drugs that their family or friends were taking. The students were asked to fill out an optional online pre‐ and post‐session survey rating their level of awareness and expertise in using these apps. Out of 75 students, 42 answered both surveys.

Preliminary results indicate that self‐described ratings of application expertise rose substantially, as did their knowledge about critical clinical information relevant to prescribing practices. Students' knowledge of these applications also impacted their understanding of drug indications, efficacy, reactions, and interactions. Our results suggest that pedagogical interventions like these may hold potential to improve safe medication practices in clinical settings.
G protein‐coupled receptor kinase 4 gamma‐mediated activation of NF‐κB
Authors: Kanassatega Rhye‐Samuel , Liu Tammy , Alsamarah Abdelaziz , Luo Yun , Andresen Bradley
Published: June 2021

Polymorphisms within G protein‐coupled receptor kinase 4 gamma (GRK4γ) (R65L, A142V, A486V) are linked to hypertension in epidemiological studies and laboratory studies in mice. However, the mechanistic signal transduction pathway(s) of wild‐type (wt) and polymorphic GRK4γ remains unknown. Recent studies suggest a role of protein‐protein interactions regulating non‐classical roles of GRK4γ, such as interaction with and activation of the NF‐κB cascade. We hypothesized that polymorphic GRK4γ abnormally interacts with, and activates, the NF‐κB cascade. To address this hypothesis molecular modeling and wet lab experiments were conducted to generate models of GRK4γ and elucidate the molecular mechanism underlying GRK4γ activation of NF‐κB. All‐atom molecular dynamics (MD) computer simulation revealed the following shifts in the bundle RH domain from wt GRK4γ: R65L = 6.3 Å, A142V = 1.3 Å, and A486V = 5.1 Å. MD will allow detailed investigation of the conformational dynamics of protein‐protein binding. Luciferase assays containing each NF‐κB promoter indicates only polymorphic GRK4γ increases NF‐κB activity but only in the presence of serum. However, wt and polymorphic GRK4γ interact with NF‐κB essential modulator (NEMO) suggesting that this pathway either requires exogenous activation or is not the only pathway via which GRK4γ can activate NF‐κB. Current studies are designed to examine GRK4γ‐mediated nuclear translocation of each NF‐κB family member as well as the role of IκB and IκK family members. Identification of the structural‐mediated signaling of the polymorphisms may create new directions for personalized hypertension therapy.
Phenobarbital Inactivates Insulin Receptor to Elicit CAR‐dependent Decrease or –Independent Increase of Hepatic Gluconeogenesis
Authors: Yasujima Tomoya , Moore Rickie , Negishi Masahiko
Published: June 2021

Nuclear receptor CAR (constitutive active/androstane receptor, NR1I3) is a transcription factor activated by various drugs such as phenobarbital (PB). Upon activation, CAR suppresses hepatic gluconeogenesis as well as induces drug metabolism. Recently,epidermal growth factor receptor (EGFR) was characterized as an initial target of PB to elicit the cellular signal that activates CAR (Mutoh et al., Sci Signal. 6, ra31, 2013). Since PB is known to antagonize insulin receptor (IR) (Hwang et al., Biochem Biophys Res Commun. 135, 501‐6, 1986), here we have examined whether or not PB regulates CAR and/or gluconeogenesis through IR. PB treatment dephosphorylated IR in human hepatoma‐derived HepG2 and in mouse primary hepatocytes. This dephosphorylation appeared to be caused by PB binding to IR as indicated by competitive binding assays using 125I‐insulin. PB‐induced dephosphorylation resulted in inactivation of the down‐stream signal molecules RACK1, ERK1/2, AKT and FOXO1. Inactivation of RACK1 and ERK1/2 is consistent with CAR‐mediated repression of gluconeogenesis (Kodama et al., Mol Cell Biol. 24, 7931‐40, 2004) whereas that of AKT and FOXO1 can be a PB signaling that suppressed gluconeogenesis in CAR‐independent manner. Utilizing CAR KO mice, we have now been investigating to demonstrate that blood glucose levels are, in fact, regulated by these CAR‐dependent and –independent mechanisms in the livers in vivo.
Mechanism and Regulation of GPCR Endocytosis in vivo using Rhodopsin as a Model
Authors: Sun Wesley , Ferng Darwin , Shieh Bih‐Hwa
Published: June 2021

We explored the mechanisms and regulation of rhodopsin endocytosis in Drosophila by examining the trafficking itineraries of the major rhodopsin (Rh1) in vivo using transgenic flies expressing a modified Rh1 containing a C‐terminal mCherry tag (Rh1‐mCherry). By fluorescence microscopy, Rh1‐mCherry was detected in the visual organelles (rhabdomeres) of photoreceptors. Upon continued light stimulation, Rh1‐mCherry display light‐dependent endocytosis, and becomes enriched in the cytoplasm of photoreceptors. Internalization of rhodopsin may be mediated by the clathrin‐mediated endocytosis (CME). CME requires the presence of a minor phospholipid, phosphoinositol‐4, 5‐bisphosphate (PIP2) in the membrane. Interestingly, perturbation of the PIP2 biosynthesis fails to prevent the endocytosis of Rh1‐mCherry in rdgA mutants. The rdgA gene encodes diacylglycerol kinase critical for the recycling of PIP2.

We also explored the contribution of two visual arrestins, Arr1 and Arr2, as adaptor proteins in promoting the endocytosis of Rh1. We introduced Rh1‐mCherry into either arr1 or arr2 mutant background and investigated how trafficking is affected. We show that internalization of Rh1‐mCherry is drastically reduced in the arr2 mutant. Arr2 is expressed 7fold more abundant than Arr1. Based on the finding, we conclude that Arr2 is critical for orchestrating the light‐dependent endocytosis of Rh1‐mCherry. Importantly, this Arr2‐mediated event does not appear to belong to the classical CME as it is insensitive to a reduction of PIP2 in the membrane.
The specificity of Gβγ subunits regulating exocytosis through the adrenergic α2a receptor
Authors: Yim Yun Young , Betke Katherine , McDonald Hayes , Gilsbach Ralf , Chen Yunjia , Wang Qin , Hein Lutz , Schey Kevin , Hamm Heidi
Published: June 2021

Modulation of neurotransmitter exocytosis by Gi/o‐coupled GPCRs, such as the α2a adrenergic receptor (α2a‐AR), is a universal regulatory mechanism used to avoid overstimulation and to influence circuitry. In addition to interaction with voltage‐gated calcium channels, Gβγ release through α2a‐AR directly interact with SNARE protein to inhibit exocytosis. It has been shown that these regulatory mechanisms by both presynaptic α2a‐AR in adrenergic (auto α2a‐AR) and non‐adrenergic neurons (hetero α2a‐AR) can go awry in neurological disorders such as anxiety and working memory deficits. In this project, we investigate whether auto α2a‐AR in sympathetic neurons use the same Gβγ as hetero α2a‐AR in other neuronal types. Gβγ specificity was determined by co‐IP Gβγ with α2a‐AR receptors and SNARE from synaptosomes, followed by mass spectrometry. We hypothesize that specific Gβγ's interact with activated auto‐ and hetero‐α2a‐ARs and inhibit exocytosis by interacting with SNARE. We have identified Gβ1, Gβ2, Gβ4, Gβ5, Gγ2, Gγ3, Gγ4, Gγ7, Gγ12, and Gγ13 which preferentially interact with the activated auto α2a‐AR and SNARE. We also expect to identify Gβ or Gγ's specific for hetero α2a‐AR. These studies may identify the Gβγ‐SNARE interaction as a new therapeutic target to modulate exocytosis in combination with drugs targeted to Gi/o‐coupled GPCRs. This work was supported by the National Institutes of Health (EY10291, MH101679, and T32).
Preferential Binding of the Marine β‐carboline Manzamine A to the NTDK Domain of RSK1
Authors: Mayer Alejandro , Hall Mary , Lach Joseph , Clifford Jonathan , Chandrasena Kevin , Canton Caitlin , Kontoyianni Maria
Published: June 2021

We reported that manzamine A (MZA) inhibited LPS‐treated rat microglia TXB2 (IC50<0.016 µM) and O2‐ (IC50=0.1 µM) generation (BMC Pharmacology 5(1) 6, 2005), LPS‐treated human BMΦ TXB2 (apparent IC50<0.7 µM) (Inflam Res. 53(S3): S217, 2004) and rat p90 ribosomal S6 kinase 1 (RSK1) with a 10‐fold selectivity in potency versus RSK2 (IC50 of 15.01 μM and 108.4 μM, respectively) in biochemical studies (Program 545.5, Society of Neuroscience, 2009). We also hypothesized that MZA's differential binding and selectivity toward RSK1 and RSK2 could be explored by computational docking experiments employing both the N‐terminal (NTDK) and C‐terminal kinase domains (CTKD). Results: The RSK1‐MZA (NTDK and CTKD‐termini) complexes appeared to have stronger interactions and preferable energetics compared to respective RSK2‐MZA complexes. In addition, our computational strategy suggested that MZA bound preferentially to the NTDK domain of RSK1 rather than the CTKD domain. RSK is a vertebrate family of cytosolic serine‐threonine kinases that act downstream of the ras‐ERK1/2 (extracellular‐signal‐regulated kinase 1/2) pathway, which phosphorylates substrates shown to regulate several cellular processes including growth, survival and proliferation. Consequently, our findings lead us to hypothesize that there may be additional novel pharmacological properties of MZA and the 80 currently known manzamine‐type alkaloids isolated from several sponge genera. Support by the Office of Research, Midwestern University (AMSM) is gratefully acknowledged.
Pharmacological Activation of Fxr Attenuates Pregnancy‐Induced Repression of Hepatic and Ileal Bile Acid Transporters in Mice
Authors: Moscovitz Jamie , Kong Bo , Guo Grace , Aleksunes Lauren
Published: June 2021

The Farnesoid X Receptor (Fxr) controls bile acid homeostasis by coordinately regulating the expression of bile acid synthesis enzymes (Cyp7a1) and transporters in the intestine (Asbt, Ostβ) and liver (Ntcp, Oatp1a4, Bsep, Mrp3, Ostβ), as well as endocrine (Fgf15) and transcription (Shp) factors. We have demonstrated that expression of bile acid synthesis enzymes and transporters is dysregulated in pregnant mice. To determine the role of Fxr in the adaptive gene responses of pregnancy, mRNA and protein expression were quantified in livers and intestines from wild‐type and Fxr‐null mice, as well as wild‐type mice administered the Fxr agonist GW4064 (GW). In response to pregnancy, ileal Fgf15, Asbt and Ostβ mRNAs decreased by 25 to 57% in wild‐type mice. Expression of these genes was also constitutively reduced 40 to 99% in Fxr‐null mice, although unchanged by pregnancy. In the liver, pregnancy‐mediated repression of Bsep mRNA was observed in wild‐type, but not Fxr‐null mice. Similar to wild‐types, pregnancy further reduced hepatic Shp, Ntcp, Oatp1a4 and Mrp3 mRNA levels of Fxr‐null mice. Treatment of pregnant mice with GW induced hepatic Shp, Ntcp, Bsep, Mrp3 and Ostβ, and ileal Fgf15 and Ostβ mRNA levels by 20 to 300%. GW also enhanced protein levels of Ntcp and Bsep in pregnant mice. Likewise, the induction of Cyp7a1 in pregnant mice was reduced by GW to levels lower than virgin controls. Collectively, these data implicate both intestinal and hepatic Fxr signaling in pregnancy‐related procholestatic gene changes, and suggest that Fxr activation may restore bile acid homeostasis in maternal cholestasis. Supported by AFPE, R01ES020522, GM104037 and T32ES007148.
A Role for CHAC1 in Acetaminophen Hepatotoxicity
Authors: Crawford Rebecca , Rowe Josef , Prescott Eugenia , Mungrue Imran
Published: June 2021

Acetaminophen (APAP) is a commonly used pain reliever and a well‐established, dose‐related hepatotoxin. Hepatotoxicity occurs when metabolism of excessive APAP generates high concentrations of the reactive metabolite N‐acetyl‐p‐benzoquinone imine (NAPQI). NAPQI depletes cellular glutathione (GSH) and forms protein adducts leading to cellular stress, mitochondrial dysfunction, and, ultimately, oncotic necrosis of Zone 3 hepatocytes. The endoplasmic reticulum (ER) stress pathway is important to maintain cellular protein folding abilities in times of stress. However, extended or aberrant ER stress signaling can lead to apoptosis through the PERK‐ATF4‐CHOP pathway. Our research explores the regulation and contribution of CHAC1, an enzyme within this pathway, in response to APAP overdose.

As purified mouse and yeast CHAC1 was shown to degrade GSH, the consequences of CHAC1 overexpression on GSH levels in human liver was explored in a human hepatoma cell line (HepG2). Overexpression of CHAC1 by transient transfection leads to a ~50% reduction in total GSH levels. As depletion of GSH is an important step in the pathology of APAP hepatotoxicity, the expression of CHAC1 mRNA in HepG2 cells was examined following 24‐hour treatment with 30mM APAP. CHAC1 mRNA was induced five‐fold by APAP, along with other components of the PERK‐ATF4‐CHOP pathway. Interestingly, overexpression of CHAC1 in HepG2 cells treated with APAP led to increased toxicity versus APAP alone, implicating a role for CHAC1 in response to APAP. Overall, these data implicate a new role for CHAC1 in the pathology of drug‐induced liver injury.
12‐LOX Modulates FcγRIIa‐mediated Platelet Response
Authors: Yeung Jennifer , Tourdot Benjamin , Fernandez‐Perez Pilar , Maloney David , Holman Theodore , McKenzie Steven , Holinstat Michael
Published: June 2021

FcγRIIa is central to the pathophysiology of a number of immune‐mediated thrombocytopenia and thrombosis (ITT) syndromes. ITT is associated with significant morbidity and mortality due to unwanted platelet‐mediated vessel occlusion. Therapeutic approaches for prevention of ITT remain a challenge since the currently approved treatment reduces, but does not eliminate incidence of thrombosis and is complicated by prolonged bleeding risk. Hence novel therapeutic strategies for prevention of ITT are warranted. Our lab has demonstrated that platelet 12‐lipoxygenase (12‐LOX), an enzyme that oxidizes free fatty acid, is an important regulator of FcγRIIa–mediated platelet activation in humans and mice. The pharmacological inhibition of 12‐LOX in human and genetic ablation of 12‐LOX in transgenic mouse (FcR/ALOX12‐/‐) platelets resulted in significant decrease in activation of FcγRIIa‐mediated effectors (PLCγ2, calcium mobilization, Rap1, and PKC) as well as platelet aggregation. To further evaluate the role of 12‐LOX regulation in this pathway, a pull‐down of 12‐LOX was performed to assess protein interactions that are required for FcγRIIa‐mediated platelet activation. A number of novel proteins were found to interact with 12‐LOX in an activation‐dependent manner. Our data supports an essential role for 12‐LOX in regulating FcγRIIa‐mediated platelet function through its complex formation, and demonstrates 12‐LOX as a viable novel therapeutic target to prevent immune‐mediated thromboses such as heparin‐induced thrombocytopenia and thrombosis (HITT) and sepsis.
Ex Vivo Expansion and Characterization of Human Umbilical Cord Blood CD34+ Stem Cells
Authors: Zhang Yu , Shen Bin , Jiang Wenhong , Ren Zhihua , Dai Wei , Jiang Yongping
Published: June 2021

Limited CD34+ cell number in human umbilical cord blood (hUCB) samples stimulated the development of in vitro HSC expansion. hUCB CD34+ cells cultured with cytokine cocktails were increased about 7.3 folds after 4 days' culture in vitro. For in vivo study, CD34+ cells obtained directly from hUCB or from in vitro expansion were transplanted into NOD/SCID mice (n=4 for each group). Four groups of mice were injected with saline, 0.4 million unexpanded CD34+ cells, 0.4 million in vitro expanded CD34+ cells, and 2.9 million in vitro expanded CD34+ cells, respectively. Peripheral blood (PB) was collected and multi‐lineage differentiation was assessed at 1, 3, and 8 weeks post transplantation. CD34+ cells were decreased below 1% in group 2 and 3 but not in group 4 after 3 weeks. On the other hand, CD45+, CD41+, CD71+and CD15+ populations were increased in groups 2‐4 compared with those of 1 week. 8 weeks post transplantation, human CD34+ cells were hardly detected in PB from any groups. However, PB and bone marrow cells expressing human cell surface markers of various hematopoietic lineages were detectable in groups 2‐4 [Figure 1]. These results indicated that CD34+ cells from hUCB either before or after expansion in vitro are capable of differentiation into hematopoietic cells of various lineages in vivo. Our study also indicates that in vitro expanded CD34+ cells are successfully long term engrafted to bone marrow of mice [Figure 2]. An efficient experimental method has been developed for the expansion of hUCB CD34+ cells in vitro and these cells can be potentially explored for various applications in the clinic.
Optimized Apyrase Improves 1 Week Myocardial Function and Blood Flow and Decreases Hemorrhage When Given Before Clopidogrel in Dogs with Coronary Reperfusion
Authors: Abendschein Dana , Baum Pamela , Nguyen Annie , Jeong Soon , Chen Ridong , Zheng Jie
Published: June 2021

Early coronary reperfusion attenuates myocardial infarction, but flow‐limiting rethrombosis and increased bleeding with use of P2Y12 antagonists limits the benefits of therapy. We tested whether an optimized human recombinant apyrase (APT102), which degrades circulating ATP and ADP causing inhibition of platelet activation and generation of cardioprotective adenosine, improves reperfusion blood flow and functional recovery over 7 d without increasing bleeding when given in combination with clopidogrel. Conscious dogs sedated with morphine had coronary thrombotic occlusion induced by electrical current followed by fibrinolysis with IV rt‐PA (Activase, 1 mg/kg) an average of 91min after ischemia. Animals received either clopidogrel (4 mg/kg, PO) or IV APT102 (1.0 mg/kg) 10 min before rt‐PA plus heparin and aspirin (4 mg/kg), and then daily clopidogrel (1 mg/kg) for 7 d. Bleeding time was measured over 24 h and then hemorrhage around surgical incisions was assessed daily. Myocardial function and regional microvascular perfusion were quantified with magnetic resonance imaging at baseline, 24 h and 1 wk. Coronary flow (implanted probe) was at or above baseline with APT102 followed by clopidogrel (n=4), but remained below baseline in dogs given clopidogrel alone (n=5). Ejection fraction and microvascular perfusion both increased back to or above baseline by 1 wk with the combination, but not clopidogrel alone. Bleeding times returned to baseline before clopidogrel in the combination group. Interestingly, APT102 + clopidogrel had less hemorrhage from incisions requiring pressure bandages (1/4) than clopidogrel alone (3/5). We conclude that APT102 improves reperfusion blood flow and myocardial function compared with standard P2Y12 inhibition without increasing bleeding. Moreover, APT102 may actually decrease hemorrhage when used before P2Y12 inhibition. APT102 warrants further study as a conjunctive agent to optimize reperfusion therapy.
Genetic Depletion of T cell S1P Receptor 1 Exhibits Improved Cardiac Function and Less Fibrosis in Streptozotocin‐Induced Type 1 Diabetic Mice
Authors: Abdullah Chowdhury , Wang Xiuqing , Jin Zhuqiu
Published: June 2021

Introduction

T lymphocyte infiltration into myocardium has been observed in patients with diabetes. Sphingosine 1‐phosphate 1 (S1P1) regulates mature T cells egress from lymphoid organ to blood and peripheral organs. The role of T lymphocytes in diabetic hearts remained elusive. Methods and Results: Depletion of T cell S1P1 receptors was attained by breeding of S1P1loxP/loxP mice with Lck‐driven Cre transgenic mice. T cell‐specific S1P1 receptor knock‐out mice (TCS1P1KO) and littlemate wild‐type mice (WT) were divided into vehicle control and streptozotocin (STZ) groups. Type 1 diabetes was induced by five days injection (i.p.) of STZ at a dose of 50 mg/kg. Control mice were injected buffer only. Body weight and fasting blood glucose level was measured at the beginning, 4, and 11 weeks. Mature CD4 and CD8 T cells in blood as well as regulatory T cells (Tregs) and Th17 cell in both spleen and blood were counted by flow cytometry. Masson's Trichrome staining was conducted to assess fibrosis. Cardiac contractile force was measured by Langendorff perfusion at the end of 11 weeks. TCS1P1KO mice had reduced CD4 T cells (1.15+/‐0.30% vs 25.06+/‐0.64% in WT, P<0.01, n=4‐5 per group) and CD8 T cells (2.09+/‐042% vs 14.72+/‐038% in WT, P<0.01) in blood. Treg cells increased in blood in WT STZ mice (1.75+/‐0.20% vs 0.39+/‐0.18% in KO STZ mice). TCS1P1KO STZ mice exhibited less cardiac fibrosis and enhanced cardiac contractile force compared to WT STZ mice.

Conclusion

Selective depletion of T cell S1P1 reduces mature T lymphocytes in circulation and exerts improved cardiac function and less myocardial fibrosis in diabetic hearts.