bluuebunny/crossref_metadata_2025 huggingface dataset:
Incorporation of realistic intracellular calcium (Ca2+) dynamics into a hypoglossal motoneuron computational model
Authors: Horn Kyle Graham , Solomon Irene C.
Published: June 2021

Hypoglossal motoneurons (HMs) play an important modulatory role in numerous behaviors, including breathing, swallowing, suckling, maintenance of upper airway patency, and speech. Failure to maintain upper airway patency has been implicated as a contributor to SIDS and OSA, the later of which is associated with a reduction in serotonergic drive to HMs during sleep. Serotonin predominantly acts through Ca2+‐dependent secondary messenger systems that leverage Ca2+ levels far from those seen at Ca2+ microdomains near the cell membrane. As a first step to better understand the role of Ca2+ in HMs, we have extended our HM mathematical model to incorporate both cystolic and microdomain Ca2+ compartments. Using this model, we demonstrate that the cystolic Ca2+ concentration during realistic HM firing replicates what is observed in biological data. Furthermore, since previous work from our laboratory has implicated Ca2+ as a major modulator of firing frequency, we also report the effects on firing frequency and excitability observed with this extended HM model. With such an extension, we will be able to further explore secondary messenger systems and their effects on other HM behaviors. Supported by HL63175.
The Effect of Milk Components on the Immune Response to a Pneumonia Vaccine: A Randomized Placebo‐controlled Clinical Trial
Authors: Fischer Laura K. , German J. Bruce , Gershwin M. Eric
Published: June 2021

Interventions aimed at improving immune function in the elderly could significantly reduce their risk of infectious disease and resulting mortality. The immune benefits of breast milk consumption in human infants are well established and suggest bovine milk components may also help enhance immune function in older populations. The objectives of this study were to evaluate the effects of a milk‐protein supplement on immune function in people at risk of immune senescence (IS). We enrolled 160 subjects over age 50 who were randomized to consume a milk‐based or control supplement for 8 weeks. Immune function was evaluated by measuring the antibody response to a pneumonia vaccine. IgG antibodies were measured before and after vaccine by multi‐analyte immunodetection system. Seroconversion was defined as a 4‐fold increase in titer. Risk of IS was evaluated by measuring a previously described marker of immune function, indolamine 2,3‐dioxygenase (IDO). IDO activity was defined as the ratio of the serum concentration of enzyme product (kynurenine) to substrate (tryptophan), measured by HPLC. Results from this study show a milk intervention significantly (p<0.05) increased the antibody response to 5 vaccine serotypes and the average seroconversion rate. These results highlight the relevance of diet for improving age‐related immune decline in individuals at risk of immune senescence.

Grant Funding Source: Dairy Management, Inc, National Dairy Council, the UC Discovery Grant
Blood pressure stabilization as a therapeutic approach to reverse high altitude induced loss of physical capacity
Authors: Schroeder Thies , Hanna Gabi , Irwin David , Hamilton Karyn , Shan Siqing , Zhao Yulin , Radiloff Daniel
Published: June 2021

Disruption of blood pressure homeostasis is a known complication of acute exposure to high altitude. We found that rats that are acutely exposed to hypobaric hypoxia and treated with a combination of the cardiac stimulant theophylline and the pulmonary vasodilator sitaxsentan run longer until exhaustion on a rodent exercise system. In addition to studying hemodynamic effects of this treatment, we have explored if other cardiac stimulants and pulmonary vasodilators have similar beneficial effects compared to theophylline and sitaxentan. Indeed, amphetamine derivatives also increase time run to exhaustion if combined with a pulmonary vasodilator. The vasodilators ambrisentan and bambuterol also prolong time run to exhaustion in rats if combined with a cardiac stimulant. The combination of theophylline and sitaxsentan reverses hypoxia‐induced blood pressure change in an anesthetized, hypoxic rat model. Theophylline does not increase vascular leak in exercising rats exposed to hypobaric hypoxia, indicating relative safety of the treatment. We demonstrate that the pairing of a cardiac stimulant with a pulmonary vasodilator provides synergism, compared to individual drugs. Other drug families therefore might be used interchangeably in this therapeutic approach. With preclinical data showing no toxicity, human studies are currently conducted to test the safety and efficacy of these drug combinations.
Receptors for Advanced Glycation End‐Products (RAGE) are Transcriptionally Inhibited by FoxA2 in Diverse Pulmonary Cell Types
Authors: Farmer Adam W. , Reynolds Paul R.
Published: June 2021

The receptor for advanced glycation end‐products (RAGE) has been developmentally identified on the membranes of ATI cells and has been implicated in the spreading and adherence that characterize the transitioning of ATII cells to ATI cells. We have previously demonstrated that TTF‐1 and Egr‐1, transcriptional regulators that function during lung development, regulate RAGE transcription. However, the precise pattern of transcription factors that limit RAGE expression to pulmonary‐specific cell types remained unclear. In the current study, transcriptional regulation of RAGE was assessed by transfection of ATII‐like alveolar epithelium (A‐549), bronchiolar epithelium (Beas2B), and macrophages (RAW 264.7) with a luciferase reporter containing 0.9‐kb of the mouse RAGE promoter. In addition to the reporter construct, each cell type was transfected with FoxA2, a winged double helix DNA binding protein that influences respiratory epithelial cell differentiation. In each case, FoxA2 repressed RAGE transcription in vitro. Mutagenesis of a proximal promoter region revealed that FoxA2 directly interacts with the coding sequence for RAGE. These data show that FoxA2 may negatively regulate RAGE expression by respiratory epithelium during pulmonary development. This work was supported by a grant from the Flight Attendant's Medical Research Institute (FAMRI, PRR) and a BYU Mentoring Environment Grant (PRR).
Acute Angiotensin II Effects On NCC are Dependent on WNK4
Authors: Ko Benjamin , Hansen Lauren , Mistry Abinash , Mallick Rickta , Hoover Robert
Published: June 2021

Acutely, angiotensin II (AII) is known to affect sodium chloride cotransporter (NCC) abundance and surface expression (SE). A possible role for WNK4 in mediating effects of AII on NCC has been described. To study this in a mammalian cell model, RNAi was used on mDCT15 cells to generate a WNK4 knock‐down cell line (WNK4KD). Thiazide‐sensitive 22Na+ uptake assays in mDCT15 cells showed a dose response to AII, peaking at 10−11M. mDCT15 cells showed a time dependent increase in activity, with peak response at 60 min (40±4%, p<0.01). An angiotensin receptor blocker eliminated this effect. WNK4 knock down in WNK4KD cells was measured via real time PCR to be 50±3%. WNK4KD cells did not show a statistically significant increase in activity with AII. NCC SE as measured by biotinylation and densitometry increased in mDCT15 cells with AII, peaking at 30 min (100±14%, p<0.01). There was a modest but statistically significant increase in SE at 30 min in WNK4KD cells (43±15%, p<0.05). These findings indicate a critical role for WNK4 in mediating acute effects of AII on NCC and provide a foundation to examine the mechanisms by which this occurs.

Support K08 DK081728(BK), R01 DK‐085097, VA Research Service (RH)
Community Dynamics in the Mouse Gut Microbiota: Role of NHE3
Authors: Engevik Melinda Anne , Shull Gary E , Worrell Roger T
Published: June 2021

Microbial dysbiosis plays a key role in complex diseases such as Inflammatory Bowel Diseasse (IBD), obesity and acute pancreatitis. A better understanding of how the gut microbiota are assembled and maintained is critical to the treatment of these complex chronic diseases. Na+/H+ exchanger isoform 3 (NHE3) plays an integral role in intestinal Na+ absorption and NHE3−/− mice exhibit chronic diarrhea. Quantitative PCR (qRT‐PCR) using 16S ribosomal DNA bacterial primers revealed that NHE3−/− mice exhibit microbial dysbiosis in both the lumen and mucosa‐associated bacteria showing decreased Firmicutes and increased Bacteriodetes from terminal ileum to distal colon. Luminal ammonia was increased in NHE3−/−proximal and distal colon indicating ammonia may be affecting bacterial proliferation in the colon. Bacteriodetes is able to increase mucosal glycan fucosylation and develop new niches. In NHE3−/−, fut2 mRNA, which encodes α‐1,2‐fucosyltransferase, was increased in the terminal ileum. fut2 mRNA correlated with increased glycan fucosylation in the terminal ileum of NHE3−/− mice. The increased colonization of the terminal ileum by Bacteriodetes may account for some of the observed downstream changes. These data show that differences in intestinal ion transport result in microbial dysbiosis which then can further alter the host epithelium. Supported by NIH DK050594 to GES and DK079979 to RTW.
An Endogenous Activator Protein of H+, K+‐ATPase Regulates Acid Secretion in Isolated Rabbit Gastric Glands
Authors: Nandi Jyotirmoy , Zinkievich J Michael , Ray Tushar
Published: June 2021

We previously showed that an endogenous activator protein (HAF) stimulated gastric H+, K+‐ATPase activity in vitro. The present study evaluates the direct role of HAF on intercellular modulation of gastric H+ transport using an HAF‐specific antibody (HAF‐Ab).

Methods

Cytosolic and membrane fractions of rabbit parietal cells (PC) were analyzed by Western blot using polyclonal HAF‐Ab. The HAF‐Ab was subsequently tested for its effects on H+, K+‐ ATPase activity in PC membrane fractions, and histamine‐stimulated H+ secretion in permeabilized gastric glands (PGG). H+ secretion was measured indirectly using [14C] aminopyrine (AP) uptake ratio.

Results

HAF‐Ab recognized 2 closely apposed bands near the 40 kDa region of the apical and tubulovesicle membranes, and cytosolic fractions. The catalytic α and β subunits of H+, K+‐ ATPase were not recognized by HAF‐Ab. HAF‐Ab dosedependently inhibited HAF‐stimulated membrane H+, K+‐ATPase activity, and decreased AP uptake in histamine‐stimulated PGG. In the latter study, immunofluorescence revealed migration of cytosolic HAF towards the cannelicular plasma membrane, where H+, K+‐ATPase is located.

Conclusions

HAF molecules migrate to the secretory plasma membrane following histamine stimulation, and close contact between HAF and H+, K+‐ATPase is essential in the process of gastric acid secretion. HAF‐Ab was found to be specific for HAF, but not for the α and β subunits of H+, K+‐ATPase. Since HAF‐Ab inhibited H+, K+‐ATPase and histamine‐stimulated acid secretion, this suggests, for the first time, evidence for a direct regulatory role of endogenous HAF on gastric H+ transport.
Genetic and Environmental Influences on Body Composition and Obesity in Chinese Children: a Twin Study in China
Authors: Li Ji , Liu Huijuan , Liu Xia , Wang Youfa
Published: June 2021

Introduction

Knowledge about the genetic influence on body composition (BC) is limited, especially among Asian children.

Objectives

to examine the relative contributions of genetic and environmental factors to BC traits among Chinese children, and assess if heritability estimates vary due to puberty.

Methods

620 twins (161 monozygotic (MZ) pairs, 149 dizygotic (DZ) pairs, 296 boys and 324 girls aged 7–15 y) were enrolled. Heritability was calculated by structural equation models.

Results

Intra‐class correlations for anthropometric and BC indicators were greater among MZ and same sex DZ twins than opposite sex DZ twins. The additive genetic factors were statistically significant in all genetic models, except for fat free mass index and waist‐to‐height ratio (WHtR), while the common environmental components were not significant for BMI. The age‐sex‐adjusted heritability estimates were low or moderate, ranging from 16.3% (WHtR) to 28.4% (BMI), 35.3% (body fat percentage), 68.0% (fat mass (FM)), 51.7% (overweight and obesity), and 22.7% (central obesity). Heritability among pubertal children was lower for waist circumference and WHtR, but greater for FM and FM index than among prepubertal ones.

Conclusions

BC including obesity status was significantly influenced by genetic factors in Chinese twins. The estimated proportion of genetic control for anthropometric and BC measures varied considerably.

Grant Funding Source: supported by the Johns Hopkins BSPH Faculty Innovation Award, the P&G Fellowship, the NIH/NIDDK (P60 DK0079637), and JMCHCH
Claudin 2 protein expression is increased in human necrotizing enterocolitis
Authors: De Plaen Isabelle G , Chou Pauline M , Weber Christopher R
Published: June 2021

We have previously shown that an increase in intestinal permeability precedes NEC in a neonatal mouse model and this permeability change is associated with increased expression of claudin 2 and internalization of claudins 2, 4 and 7 and occludin. Whether these proteins are altered in human NEC remains unknown. In this study, we examined the expression of claudin 2, 4, occludin, and ZO‐1 in control and NEC tissues by immunohistochemistry. Samples from 11 colon controls, 10 colon NEC, 9 small intestine controls and 10 small intestine NEC cases were used for the study. Staining intensity was scored semi‐quantitatively from 0 to 3 by two separate investigators and average scores determined. Claudin 2 expression was low in the colon of control tissues (crypt expression 1.0 ± 0.1, surface expression 1.0 ± 0.2). In small intestinal tissue, there was a decreasing gradient of expression from the crypts to the surface (crypt expression 1.5 ± 0.3 vs surface expression 0.7 ± 0.1, P <0.05). Compared to control, the expression of claudin 2 was dramatically increased in the crypts of NEC colon (2 ± 0.2 vs 1.0 ± 0.1, p<0.001) and NEC small intestine (2.3 ± 0.2 vs. 1.5 ± 0.3, P <0.05. In contrast, occludin and ZO‐1 expression and distribution were similar in control and NEC tissues. In conclusion, the expression of claudin 2 but not of 4, occludin, and ZO‐1 is increased in human NEC. Alteration in claudin 2 protein might play a role in NEC pathogenesis.
Increasing seafood in the USDA Food Patterns increases eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and other nutrients
Authors: OˈConnell Kellie M , Kuczynski Kevin , Guenther Patricia M , Rimm Eric B , Pérez-Escamilla Rafael , Britten Patricia
Published: June 2021

In response to scientific evidence that seafood has important health benefits, the 2010 Dietary Guidelines Advisory Committee investigated the impact of increasing seafood in the USDA Food Patterns on eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and overall nutrient adequacy. Three scenarios were modeled: 4 oz/wk of seafood higher in EPA+DHA (HI); 8 oz/wk of HI seafood and seafood lower in EPA+DHA (LO) in proportions currently consumed; and 12 oz/wk of LO seafood. The USDA Food Patterns and National Nutrient Database for Standard Reference (SR 22) were the basis for analysis. To meet desired EPA+DHA levels, amounts of seafood in the Protein Foods group were increased, replacing some meat and poultry, while eggs, processed soy products, nuts, and seeds were held constant. All scenarios met almost all goals for nutrient adequacy, paralleling the base USDA Food Patterns. With more seafood in the patterns, selenium and vitamins D and B‐12 were substantially increased. EPA+DHA levels in the 2000 calorie pattern were 292 mg/d (4 oz HI), 253 mg/d (8 oz LO+HI), and 201 mg/d (12 oz LO). Seafood can be increased to 8 oz/wk of LO and HI species as currently consumed to achieve 250 mg/d of EPA+DHA, while maintaining the nutrient adequacy of the USDA Food Patterns and improving vitamin D levels. The recommendation to increase seafood intake was included in the 2010 Dietary Guidelines for Americans. Supported by USDA/CNPP.
Inflammation in postpartum women is inversely related to transferrin saturation, but is not correlated with ferritin or hepcidin
Authors: Jorgensen Josh M , Yang Zhenyu , Lonnerdal Bo , Chantry Caroline J , Allen Lindsay H , Dewey Kathryn G
Published: June 2021

The relationship of inflammation to indicators of iron status in postpartum women is not well documented. We examined associations between markers of iron status (hemoglobin (Hb), ferritin, hepcidin, and transferrin saturation (TfSat)) and inflammation (C‐reactive protein (CRP) and α‐1‐acid glycoprotein (AGP)) in 114 lactating women enrolled in a randomized placebo controlled trial of iron‐containing prenatal vitamin‐mineral supplements. Blood samples were collected before and after 3 mo of supplementation. At baseline (x̄ = 13.8 d postpartum), 58% of women had CRP > 5 mg/L and 78% had AGP > 1 g/L. At the final study visit (n=95; x̄ = 17.3 wk postpartum), these values dropped to 26% and 29%. There were positive correlations between CRP and AGP at the baseline (r=0.66, P<0.0001) and final (r=0.57, P<0.0001) time points. Neither CRP nor AGP was significantly correlated with ferritin or hepcidin at either baseline or final. CRP and AGP were negatively correlated with TfSat at both baseline (r=−0.45, P<0.0001 and r=−0.32, P<0.001) and final (r=−0.43, P<0.0001 and r=−0.34, P<0.001). Hb was negatively correlated with CRP and AGP at baseline (r=−0.32, P<0.001 and r=−0.20, P=0.03), but not at the final time point. In postpartum women, ferritin and hepcidin were not influenced by inflammation, while TfSat decreased with increasing inflammation. Hb was inversely related to inflammation at 2 wk postpartum, but not at 17 wk.

Grant Funding Source : Center for Health and Nutrition Research at UC Davis
Weighting the Factors Associated with Obesity in Children: an International Perspective Toward a Unified Model
Authors: Gregori Dario , Baldi Ileana , Ghidina Marco
Published: June 2021

Beside genetic predisposition, several factors have been proposed to promote overweight and eventually obesity in children, from the socio‐ and built‐ environment down to behavioural attitudes. How this model is shared by different cultural settings is unclear and less investigated.

Using a unified protocol for data collection, a cross‐sectional study has been performed on 960 children in India, Italy, Germany, France, UK, Argentina, Mexico and Brazil. Children CDC z‐scores for Body Mass Index have been evaluated in association with several known factors influencing overweight and obesity (maternal and neonatal aspects, socio‐economic familiar status, BMI of parents, physical activity, nutrition habits, screening activities). Based on a random‐effect mixed effect model and the Kullback‐Leibler Entropy Measure, the capability to explain variability in BMI of such factors has been computed. Percentages of explained variation are given in the table.

Capability of proposed factors to capture variability in BMI is significantly higher in UK (p=0.03) than in other countries, being significantly lower in an emerging country like India (p=0.042): more intense research should be specifically targeted to capture risk factors specific for the cultural setting in addition to the general ones.

Bouchard, 2007.
PROLYL‐OLIGOPEPTIDASE INHIBITION AMELIORATES EXPERIMENTAL HYPERTENSION AND DECREASES INFLAMMATORY CYTOKINES IN MICE LACKING ANGIOTENSIN CONVERTING ENZYME N DOMAIN ACTIVITY
Authors: Ong Frank S , Shen Xiao Z , Fuchs Sebastien , Bernstein Kenneth E
Published: June 2021

Using gene targeting, ACE domain‐specific knockout mouse lines N‐KO & C‐KO were created. After 14 day Ang II infusion, BP was 27 mm Hg (p<0.0001) higher in N‐KO vs. WT/C‐KO. N‐KO kidney Ang II levels were 1.5‐fold those of WT/C‐KO (p<0.015). Peritoneal macrophage cytokine expression in response to Ang II, measured as percentage of F4/80+/TNFαhigh cells vs total F4/80+, was 40.3% ± 4.7% for N‐KO and 19.8% ± 2.8% for WT (p<0.005). In response to LPS, N‐KO macrophages made 4‐fold the TNFα as C‐KO/WT cells (p<0.001). AcSDKP, a substrate specifically cleaved by the N‐domain, is made by the enzyme prolyl‐oligopeptidase (POP). Daily injections of the POP inhibitor S‐17092 were given to N‐KO and WT mice. This reduced the LPS‐mediated N‐KO macrophage production of TNFα to that of cells from WT. The hypertensive response to Ang II was also tested in N‐KO and WT mice treated with daily S‐17092. POP inhibition had no effect on the BP of WT, but reduced the BP of N‐KO to that of WT (p<0.01). Furthermore, FACS analysis of peritoneal macrophages showed a decreased percentage of F4/80+/TNFαhigh cells in N‐KO mice treated with daily S‐17092. The lack of N domain activity in ACE exhibited a pro‐inflammatory role in experimental hypertension via cytokine regulation, accumulation of renal Ang II, and increased BP response. Inhibition of POP eliminated the differences in macrophage TNFα expression and blood pressure response between N‐KO and WT mice.
Structural and functional implications of plakoglobin compensation due to β‐catenin loss in the liver
Authors: Wickline Emily Diane , Monga Satdarshan P.
Published: June 2021

β‐Catenin is important in liver homeostasis as a part of Wnt signaling and cell‐cell adhesions. We previously reported that hepatocyte‐specific β‐catenin knockout (KO) mice show higher plakoglobin (PG; γ‐catenin) levels over wild‐type (WT) livers and also maintain intact adherens junctions (AJ) as a result of PG compensation. Since PG is a desmosomal protein we wanted to see if its increase in the KOs comes at the expense of desmosomes. Here we show no change in the desmosomes in the KO livers by protein analysis and TEM. Next we wanted to address functionality of PG's compensation for β‐catenin loss especially in Wnt signaling. We show that knocking down PG in Hep3B human hepatoma cells only slightly decreases TCF reporter (TOPflash) activity, indicating that PG is able bind TCF and participate in Wnt signaling. However, knock down of β‐catenin shows a dramatically greater drop in TOPflash activity. Next, we wanted to determine if changes in PG affect proliferation and migration. Changes in PG levels do not affect thymidine incorporation rates. However, scratch‐wound assays reveal that β‐catenin knock down (and subsequent PG increase) increases migration of Hep3Bs. Also, PG knock down decreases migration, indicating that PG may play a role in cell motility. Thus, β‐catenin loss is compensated by PG at the AJ without affecting desmosomes; however, the functions of β‐catenin as a part of the Wnt pathway remain unfulfilled by PG. Moreover, changes in PG levels also affect the migratory potential of human hepatoma cells. Research support: NIH grant 1R01DK62277.
Age‐dependent Impairments in Rat Bladder Regeneration
Authors: Burmeister David , Peyton Charles , AbouShwareb Tamer , Bergman Chris , Andersson Karl-Erik , Christ George
Published: June 2021

Models of de novo organ regeneration in mammals may provide guidance for harnessing natural regenerative capacities. We previously reported that removal of ~70% of the bladder (subtotal cystectomy; STC) in 12 week old female rats induces regeneration leading to functional recovery. In this study, 12 month (“older”) and 12 week (“younger”) old female rats underwent STC, and were followed up to 26 wks post‐STC with in vivo urodynamics, and pharmacological and histological analysis ex vivo. STC in older animals led to a higher mortality/morbidity rate (26%) than younger animals (11%), with histologic evidence of kidney damage. Bladder capacity 26 weeks post‐STC was 56% of age‐matched controls (AMC) in older rats, and 96% of AMCs in younger rats. There was no significant residual volume found in any group. Responses of bladder strips to several physiologically relevant stimuli were lower in younger STC animals than those from control bladders, but higher than older STC animals. Histology revealed age‐related differences post‐STC with respect to inflammation, angiogenesis, fibrosis, and proliferation. The impaired regeneration observed in older animals stands in stark contrast to the rather complete regeneration observed in younger animals. As such, this rodent model may aid in elucidating the basic biology of mammalian organ regeneration and identifying targetable pathways for optimizing regenerative capacity.
A microRNA pulldown approach uncovers regulation of p53 activity and growth factor signaling by miR‐34a
Authors: LAL ASHISH , THOMAS MARSHALL , Navarro Francisco , Li Xiao Ling , Lieberman Judy
Published: June 2021

A simple biochemical method to isolate mRNAs pulled down with a transfected, biotinylated microRNA was used to identify direct target genes of miR‐34a, a tumor suppressor gene. The method reidentified most of the known miR‐34a regulated genes expressed in K562 and HCT116 cells. The transcripts pulled down with miR‐34a were highly enriched for their roles in growth factor signaling and regulation of p53 activity. These genes regulate multiple signal transduction pathways that orchestrate the proliferative response to external growth stimuli. Multiple candidate miR‐34a‐regulated genes participate in RAS‐RAF‐MAPK signaling. Ectopic miR‐34a expression reduced basal ERK and AKT phosphorylation and enhanced sensitivity to serum growth factor withdrawal, while cells genetically deficient in miR‐34a were less sensitive. Several novel direct targets of miR‐34a were experimentally validated, including genes that participate in growth factor signaling as well as genes that regulate p53 activity. Thus miR‐34a tempers the proliferative and pro‐survival effect of growth factor stimulation by interfering with growth factor signal transduction and downstream pathways required for cell division.
Sister acts ‐ coordinating DNA replication & cohesion establishment
Authors: Jallepalli Prasad V
Published: June 2021

The ring‐shaped cohesin complex links sister chromatids and plays crucial roles in homologous recombination and mitotic chromosome segregation. In cycling cells, cohesin's ability to generate such linkages is restricted to S phase and depends on so‐called establishment factors that are intimately connected to DNA replication. Paramount among these are the Eco1‐related family of acetyltransferases. By applying high‐resolution microscopy, somatic cell gene‐targeting, and next‐generation sequencing approaches, we have begun to delineate how human Eco1 orthologs are targeted to chromosomes, how their acetyltransferase activity is coupled to cell cycle progression, and how their acetylation of cohesin's Smc3 subunit triggers cohesion establishment.. Our data suggest a model whereby Smc3 acetylation regulates a ‘gate’ within cohesin that otherwise interferes with DNA replication fork passage, and whose closure is crucial for cohesin's stable interaction with replicated DNA molecules.
Inhibition of selenocysteine tRNA[Ser]Sec aminoacylation provides evidence that aminoacylation is required for regulatory methylation of this tRNA
Authors: Carlson Bradley , Kim Jin Young , Xu Xue-Ming , Yoo Min Hyuk , Zeng Yu , Chen Shawn , Gladyshev Vadim , Lee Byeong Jae , Hatfield Dolph
Published: June 2021

Selenium is incorporated into protein as selenocysteine (Sec), the 21st amino acid in the genetic code. There are two isoforms of Sec tRNA[Ser]Sec that differ by a single methyl group, Um34, at the wobble position. The non‐Um34 isoform supports the synthesis of a subclass of selenoproteins, designated housekeeping, while the Um34 isoform supports the expression of another subclass, designated stress‐related selenoproteins. We investigated the relationship between tRNA[Ser]Sec aminoacylation and Um34 synthesis which is the last step in the maturation of this tRNA. Mutation of the discriminator base at position 73 in tRNA[Ser]Sec dramatically reduced aminoacylation with serine, as did an inhibitor of seryl‐tRNA synthetase, SB‐217452. Neither precluded the synthesis of any other base modifications on tRNA[Ser]Sec following microinjection and incubation of the mutant tRNA[Ser]Sec transcript, or the wild type transcript along with inhibitor, in Xenopus oocytes. The data demonstrate that Sec tRNA[Ser]Sec must be aminoacylated for Um34 addition. The fact that selenium is required for Um34 methylation suggests that Sec must be attached to its tRNA for Um34 methylation. This would explain why selenium is essential for the function of Um34 methylase and provides further insights into the hierarchy of selenoprotein expression. This research was supported by the Intramural Research Program of the NIH, NCI, CCR.
Effects of vitamin A status on the responses of the hepatic genes’ expression to insulin and retinoic acid in primary hepatocytes from Zucker lean and fatty rats
Authors: Chen Wei , Chen Guoxun
Published: June 2021

Insulin‐mediated regulation of hepatic gene expression plays a key role in glucose and lipid homeostasis. This study was aimed to examine the hepatic gene expression in response to insulin and retinoic acid (RA) treatments in primary hepatocytes from Zucker rats of different vitamin A (VA) status. Primary hepatocytes were isolated from Zucker lean (ZL) or fatty (ZF) rats fed either vitamin A sufficient (VAS) or deficient diet (VAD) for 8 weeks after weaning. The mRNA levels of Gck, Pck1, Srebp‐1c and Pklr after insulin and RA stimulation for 6 hours were determined using real‐time PCR. In hepatocytes of ZL rats on either diet, insulin synergized with RA to induce the expression of Gck and Srebp‐1c. This was impaired in hepatocytes of ZF rats on VAS, but partially recovered in those on VAD diet. In lean hepatocytes, the Pck1 expression was suppressed by insulin and induced by RA respectively. The insulin‐suppressed Pck1 expression was impaired in hepatocytes from ZF rats fed VAS diet and partially restored in that fed VAD diet. The RA‐induced Pck1 expression was normal in hepatocytes from ZF rats fed VAS, and was enhanced in that fed VAD. The Pklr expression was similar between ZL and ZF and not affected by VA status. We conclude that VA status modulate the insulin‐ and RA‐regulated hepatic gene expression in hepatocytes of ZF rats. It suggests that micronutrients contribute to the hepatic insulin response.

Grant Funding Source: AHA
Evaluation of MyPlate mini‐poster for older Latino adults: MiPlato para Adultos Mayores
Authors: Bobroff Linda B , Minton Emily , Diehl David C , Diaz Xiomara , Keith Mary , Medina-Solorzano Ada , Gylfadottir Unnur K
Published: June 2021

Culturally appropriate nutrition education materials are in high demand for the large and growing population of Latinos in the U.S. This study assessed older Latino adult responses to consumer messages, graphics, and foods included in the draft MyPlate for Older Adults mini‐poster for Latinos, MiPlato para Adultos Mayores (MPAM). Six focus groups (FGs) were conducted by trained moderators and a nutritionist observer with a convenience sample of 47 Latinos, 60 years or older, in 3 Florida counties. Two FGs were offered per county (one each in English and Spanish) to 35 women and 12 men, representing 9 Caribbean and Latin American countries. Discussions were tape‐recorded, transcribed, and analyzed for common themes. Participants found MPAM to be informative, appealing, and culturally appropriate. They made recommendations for additional foods including papaya and fresh fish, and reduction in size of the red meat graphic. Daily food group recommendations were found to be practical. Participants liked the variety of silhouettes depicting physically active older adults and consumer messages were found to be clear, useful, and well organized. MPAM is being incorporated into lessons and educational materials for older Latino adults. The mini‐poster can be used by nutrition educators to teach MyPlate concepts to older Latino adults in a variety of educational settings. Funding provided by IFAS Dean for Extension, UF.
Genistein‐mediated inhibition of mammary stromal adipocyte differentiation limits expansion of mammary stem/progenitor cells by paracrine signaling
Authors: Montales Maria Theresa E. , Scanlon Samantha , Nakatani Hajime , Matsuda Tsukasa , Simmen Rosalia C.M.
Published: June 2021

Mammary adiposity may contribute to breast cancer development and progression by releasing cytokines and other inflammatory mediators that promote mammary epithelial proliferation. We evaluated the effects of soy isoflavone genistein (GEN) on the adipogenic differentiation of a SV40‐immortalized mouse mammary stromal fibroblast like cells (MSF) treated with differentiating agents insulin, hydrocortisone and troglitazone (DM). MSF cultured in DM with and without extracellular matrix for 14 days showed reduced differentiation (2–3 fold) into mature adipocytes with 40nM but not 2uM GEN, as evaluated by Oil Red O staining. Addition of sera from GEN‐fed adult mice to MSF similarly reduced adipogenic differentiation (4–5 fold), relative to sera from casein‐fed mice. Lipid accumulation was decreased by GEN (40nM>2uM). Expression levels of lipogenic enzyme genes fatty acid synthase and PPAR‐γ were down‐regulated by 40nM GEN. Estrogen receptor (ER)‐β, an inhibitor of PPAR‐γ transcriptional activity, but not ER‐α transcript levels were increased by 40 nM GEN. Conditioned media from GEN‐treated MSF reduced anchorage‐independent mammosphere formation of human breast cancer cells MCF‐7. Thus, dietary factor control of mammary adiposity, possibly mediated by ER‐β, may limit mammary stem/progenitor cell expansion and protect against breast cancer.

Grant Funding Source: USDA‐CRIS 6251‐5100002‐06S; CUMG/ACHRI
Polyphenol‐rich Aronia melanocarpa (chokeberry) extract regulates expression of cholesterol and lipid metabolism genes in Caco‐2 cells
Authors: Kim Bohkyung , Park Youngki , Taheri Rod , Kimball Kerilyn , Roto Anna , Lee Jiyoung , Bolling Bradley
Published: June 2021

Chokeberry, Aoronia melanocarpa, is a rich source of polyphenols. We investigated the effect of an anthocyanin‐rich chokeberry extract (CBE) on the expression of genes involved in intestinal lipid metabolism in Caco‐2 cells, a human intestinal cell line. CBE had a Total Phenols value of 745 mg gallic acid equivalents/g and contained 12% anthocyanins. Cells were incubated with 50 or 100 μg/mL of CBE for 24 h and gene expression was measured by quantitative realtime PCR. ATP binding cassette transporter (ABC) A1, scavenger receptor class B, type 1, Niemann‐Pick C1 Like 1, and HMG‐CoA reductase mRNA abundance was significantly reduced by CBE. In contrast, CBE induced LDL receptor (LDLR) and ABCG8 to levels 2.5‐ and 1.5‐fold of controls. Western blot analysis confirmed increased LDLR protein level. Expression of genes involved in lipid metabolism and chylomicron assembly, including sterol regulatory element binding protein 1c, fatty acid synthase, and acyl‐CoA oxidase and microsomal triglyceride transfer protein, was significantly decreased by CBE in a dose‐dependent manner. Our data suggest that CBE may play an important role in the flux of cholesterol as well as lipid metabolism in the intestine. Supported by College of Agriculture and Natural Resources and Research Foundation at the University of Connecticut.

Grant Funding Source : College of Agriculture and Natural Resources and Research Foundation at the University of Connecticut.
Thioredoxin reductase 1: Role in oxidative stress within cancer cells
Authors: Yoo Min-Hyuk , Carlson Bradley A , Tsuji Petra A , Gladyshev Vadim N , Hatfield Dolph L
Published: June 2021

Many selenoproteins (SPs) have roles in intracellular redox regulation due to the presence of selenocysteine at its catalytic core; e.g., thioredoxin reductase 1 (TR1) is a major antioxidant enzyme in mammals that functions in regulating thioredoxin redox status. This oxidoreductase is enriched in many malignant cells and has been proposed as a target for cancer therapy. We previously assessed the role of TR1 in the malignant process by knocking down its expression in cancer cells resulting in reversing of many of the malignant phenotypes. Cancer cells are proposed to have a more efficient anti‐oxidative system than normal cells and thioredoxin and glutathione comprise two of the major enzymatic, anti‐oxidative systems in mammals. To further elucidate TR1's role in anti‐oxidative systems in cancer cells, we targeted the removal of TR1 and Trx1 expression in NIH3T3 and DT cells (DT cells were derived from NIH 3T3 cells by over‐expressing oncogenic k‐ras). Our data have provided new insights into the expanded role of TR1 in cancer cells and the interrelationships between anti‐oxidative systems which will be presented. This research was supported by the Intramural Research Program of the NIH, NCI, CCR.
Consumption of one egg per day reduces the expression of HMG‐CoA reductase in peripheral mononuclear cells (PBMC) to maintain LDL cholesterol concentrations
Authors: Fernandez Maria Luz , Ata Shymaa , Park Youngki , Lee Jiyoung
Published: June 2021

Thirty three subjects (men and women, 40–75 y) were recruited and randomly assigned to consume either 1 egg per day (n = 13) (203 additional mg cholesterol/d) (EGG) or the equivalent amount of egg substitute (n = 9) (0 additional mg cholesterol/d)(SUB) for 12 wk. We isolated plasma and peripheral blood mononuclear cells both at baseline and wk 12 to determine plasma lipids and the expression of genes involved in maintaining plasma LDL cholesterol (LDL‐C) and genes associated with reverse cholesterol transport. Plasma LDL‐C was reduced over time for all subjects in the EGG (134 ± 27 to 129 ± 30 mg/dL) and SUB (127 ± 14 to 113 ± 30 mg/dL) groups (p< 0.05). HDL cholesterol (HDL‐C) was increased by 17% in the EGG (P < 0.05) while no changes were observed in the SUB group. Dietary records indicated that subjects in the EGG were consuming 408 mg cholesterol/d compared to 165 mg in the SUB group. The expression of ATP binding cassette (ABC) AI and ABCG1 transporters or the LDL receptor were not affected over time or by diet. In contrast, the expression of HMG‐CoA reductase was decreased from 2.97 ± 2.7 to 1.37 ± 1.44 to arbitrary units (P<0.025) in the EGG and from 2.5 ± 2.1 to 1.04 ± 0.75 arbitrary units in the SUB group (p <0.025). These results suggest that the observed decreases in LDL‐C in both groups can be associated with a suppression of HMG‐CoA reductase gene expression [supported by the Egg Nutrition Center].
Role of General Control Nonderepressible 2 (GCN2) kinase in mediating responses to dietary methionine restriction
Authors: Plaisance Eric P , Van Nancy , Orgeron Manda , McDaniel Alexandra N , Behrens Phillip H , Gettys Thomas W , Anthony Tracy G
Published: June 2021

Restriction of dietary methionine from normal levels (0.86%) to 0.17% produces a coordinated series of biochemical and physiological responses that improve biomarkers of metabolic health, limit fat accretion, and enhance insulin sensitivity. GCN2 kinase serves as a sensor to detect reductions in essential amino acids and initiates integrated behavioral and transcriptional responses that mediate metabolic adaptation. The role of GCN2 in mediating the responses to dietary methionine restriction (MR) was assessed using GCN2−/− mice. Dietary MR for 8 wks produced a comparable reduction in hepatic triglyceride and increase in food consumption in wild type (WT) and GCN2−/− mice, while the normal diet‐induced increase in plasma adiponectin was actually enhanced in GCN2 nulls. The transcriptional responses produced by dietary MR in brown adipose tissue did not differ between WT and GCN2 null mice. In contrast, the diet‐induced increase in energy expenditure and reductions in fasting insulin and hepatic SCD‐1 expression observed in WT mice were partially compromised in GCN2 nulls, and the normal reductions in adiposity, plasma leptin and glucose were absent in GCN2−/− mice. The present findings suggest that GCN2 is an important component of the nutrient sensing mechanism(s) linking restriction of dietary methionine to the metabolic adaptations produced by the diet. Work supported by NIH: 2 P20 RR021945 (TWG), 2 U24 DK076169 (TWG), 1 F32 DK088513 (EPP), 1 RO1 HD070487 (TGA).

Grant Funding Source: NIH
Leucine Pulse Increases Skeletal Muscle Protein Synthesis during Continuous Feeding in Neonatal Pigs
Authors: Boutry Claire , Suryawan Agus , El-Kadi Samer W , Wheatley Scott M , Orellana Renan A , Nguyen Hanh V , Davis Teresa A
Published: June 2021

Continuous orogastric tube feeding is used for infants unable to maintain an oral diet. Orogastric bolus feeding has a greater stimulatory effect on muscle protein synthesis than continuous feeding in neonatal pigs. Leucine administration stimulates protein synthesis, but its effects during continuous feeding have not been investigated. Twenty piglets (8‐d‐old) received formula for 24h continuously (CON) or by bolus (BOL), i.e., every 4h for 15min. For the CON+LEU group, leucine was administered parenterally (800μmol•kg−1•h−1) every 4h for 1h. The leucine pulse increased circulating leucine (P<0.0005), but had no effect on isoleucine and valine concentrations. Circulating insulin, glucose, and most other essential amino acids increased 15min after the BOL meal (P<0.05) and returned to baseline by 2h, but were unchanged in CON and CON+LEU groups. Protein synthesis rates in longissimus dorsi, gastrocnemius and soleus muscles were greater in the CON+LEU and BOL than in the CON group (P<0.005). In the BOL and CON+LEU groups, there was an increase in the phosphorylation of ribosomal protein S6 kinase 1 and the formation of active eIF4E.eIF4G complex in skeletal muscle (P<0.05). In conclusion, administration of a leucine pulse during continuous orogastric feeding increases skeletal muscle protein synthesis in neonatal pigs by stimulating translation initiation (NIH AR444474 and USDA/ARS 6250‐51000‐055).

Grant Funding Source: NIH‐USDA/ARS
Pre‐ and Post‐weaning diets with low omega‐6 to omega‐3 fatty acid ratio reduced plasma lipid levels and increased cholesterol efflux capacity in the offspring of C57BL/6 mice
Authors: Balogun Kayode Adeniyi , Randunu Raniru Sergaevna , Cheema Sukhinder Kaur
Published: June 2021

We investigated whether maternal and post‐weaning dietary ratios of omega (n)‐6 to n‐3 PUFA alter lipid metabolism of the offspring and whether the effects are gender specific. Female C57BL/6 mice were fed semi‐purified diets (20% w/w fat) containing n‐6 to n‐3 PUFA ratios of 5:1, 15:1 or 30:1 before mating, during pregnancy, and until weaning. Offspring were continued on their mothers’ diets for 4 months. Plasma lipid profile of the offspring was measured at weaning and at 4 months. HDL cholesterol efflux capacity of plasma was assayed using J774 cells. Male offspring from 5:1 group had a relatively lower plasma cholesterol at weaning and at 4 months (p<0.05). At weaning, the male offspring showed no difference in plasma triglyceride (TAG) and nonesterified fatty acids (NEFA) however there was a relative reduction at 4 months in the 5:1 group (p<0.001). Females in the 5:1 group had a relatively lower TAG and NEFA at weaning and at 4 months (p<0.05). The 15:1 females had lower LDL‐cholesterol at weaning and at 4 months; 5:1 males had lower levels at weaning with no differences at 4 months. HDL‐cholesterol was the same in all the groups; however plasma from the 5:1 males showed higher cholesterol efflux compared to 30:1 (p < 0.05). Our data suggest that maternal diets low in n‐6 to n‐3 PUFA ratio reduce plasma lipid levels, the effect is enhanced if the post‐weaning diet is the same and there are gender differences. Supported by NSERC

Grant Funding Source: NSERC
Growth factor retention on decellularized rat liver matrices derived from normal and regenerating liver
Authors: Shupe Thomas , Zimmerman Cynthia , Petersen Bryon
Published: June 2021

We have reported a method for the decellularization of intact rat livers by perfusion with increasing concentrations of detergents. This procedure results in a decellularized organ that maintains an intact capsule and retains an appropriate distribution of extracellular matrix proteins and associated mucopolysaccharides. Immunoassay demonstrated that many growth factors were retained on the decellularized matrix. Several growth factors known to regulate hepatocyte phenotype, including PDGF, IGFBP‐3 and BMP‐7, were found to be increased in matrices from regenerating livers. Equilibration of the decellularized matrix in serum increased the levels of many of these factors. Decellularization by an alternative, enzyme based method resulted in the retention of a different repertoire of growth factors. The liver progenitor cell line, WB344, was able to colonize these matrices and differentiate into mature hepatocytes, but no differentiation was observed on plastic alone. These data indicate that matrices generated from livers in different growth states and by different methods may each be best suited for specific applications.
Effect of phosphatidyl choline liposomes on the relative bioavailability of fat soluble micronutrients in salad vegetables
Authors: Flendrig Leonard M , Zhou Yang , Quadt Frits , Mulder Theo , Schuurbiers Eric , White Wendy S
Published: June 2021

The purpose of this study was to determine the effects of phosphatidyl choline (PC) multilamellar vesicles (in absence of other lipids) in salad dressing on: 1) the absorption of α‐carotene, β‐carotene, lutein, lycopene, and tocopherols in salad vegetables; 2) the absorption of retinyl palmitate formed from the ingested α‐ and β‐carotene. Twelve healthy men each consumed 3 identical salads with 3 different salad dressings containing: 1) zero lipids (placebo), 2) 16g sunflower oil (positive control), 3) 1.25g tocopherol‐stripped soybean PC liposomes (treatment). The order in which the salads were consumed was randomly assigned; salads were separated by ≥ 2 week. Blood samples were collected at baseline and 2, 3, 4, 5, 6, and 9 h postprandially. Chylomicron fractions were analyzed by HPLC with coulometric array electrochemical detection. All six fat soluble micronutrients showed positive trends in relative bioavailability when comparing PC liposomes with zero lipid treatments. Statistical significance was reached for lutein (P < 0.05), beta‐carotene (P < 0.0003), and retinyl palmitate (P < 0.03). The full fat positive control produced substantial responses for all six micro‐nutrients, which validated the study protocol. In conclusion, the lecithin liposomes added to fat‐free salad dressing enhanced the intestinal absorption of fat soluble micronutrients.
Crosstalk mechanism between injured epithelia and bone marrow
Authors: Tamai Katsuto , Kikuchi Yasushi , Iinuma Shin , Fujita Ryo , Manjyo Nao , Kaneda Yasufumi
Published: June 2021

In the epidermis, resident stem/progenitor cells are involved in the complete renewal of the stratified squamous epithelium every 2–4 weeks and thus contribute to tissue homeostasis. In patients with the inherited chronic blistering skin disorder, recessive dystrophic epidermolysis bullosa (RDEB), however, the epidermal stem cell pool is continually depleted as a result of detachment of full‐thickness epidermis due to lack of the basement membrane protein, type VII collagen (Col 7). Nevertheless, the epidermis in RDEB continues to be renewed, suggesting there may be a mechanism to supplement epithelial stem/progenitor cells from extracutaneous sources. Here, we demonstrate that the detached epithelium from a Col 7‐null mouse model of RDEB will release abundant high mobility group box 1 (HMGB1), which mobilizes PDGFRα‐positive msenchymal stem cells from bone marrow into the circulation to repair the blistered skin. We believe that this study has broader implications for clinical medicine in identifying a means of mobilizing and recruiting particular bone marrow cells germane to tissue repair.