bluuebunny/crossref_metadata_2025 huggingface dataset:
Estimates of Body Composition in Drug Naïve Asymptomatic HIV‐infected Rural Kenyan Women
Authors: Ernst Judith Ann , Ettyang Grace , Katschke Adrian , Lewis Brian , Neumann Charlotte
Published: June 2021

Objective

Bioelectrical impedance analysis (BIA), as a method to estimate fat free mass (FFM) and % body fat (%BF), is compared to the anthropometric (Anthro) method using skinfold (SF) and body circumference (BC) measures in HIV‐infected women in rural Kenya.

Methods

In 138 women, (WHO Stage 1 or 2; CD4 > 250 cells/uL), height, weight, SF, BC and BIA were obtained with standardized procedures. Fat area of the arm (MAFA), muscle area of the arm (MAMA), thigh (MTMA), calf (MCMA) and sum of SF were calculated from standard equations using BC and SF. Estimates of FFM, %BF and total body water (TBW) from BIA measures of resistance were calculated with standard equations.

Results

The mean (SD) for age, BMI and CD4 was 36.4(7.5) yrs, 22.3(3.6); 11% < 18.5, and 492(191) cells/uL, respectively.

Anthro compared to BIA for estimates of lean and fat mass

Anthro
MAMA
MTMA
MCMA
MAMA
MAFA
Sum
SF

BIA
FFM
FFM
FFM
TBW
%BF
%BF
__

r
0.61
0.70
0.71
0.65
0.84
0.87
__

All correlations p<0.001

Conclusions

BIA, when compared to the method using anthropometry measures, has a strong relationship to fat and lean mass, at baseline, in drug naïve asymptomatic HIV‐infected rural Kenyan women enrolled in an 18 month randomized controlled field trial that measures the impact of added protein and energy on body composition. Supported in part by USAID Grant No. PCE‐G‐00‐98‐00036‐00 and 1R01HD57646‐01A1 (CFDA #93.865)
Inter‐ and intra‐strain variability for survival time after hemorrhage in inbred rats
Authors: Rose Rajiv , Oh Taesung , Calderon Mariam , Klemcke Harold
Published: June 2021

In an effort to identify genes influencing survival time after hemorrhage (STaH) we earlier reported 8‐fold differences in STaH among 15 inbred rat strains, indicating a genetic basis for STaH (Klemcke et al., Shock 29:748, 2008). Subsequently, we identified that the inbred rat strains differed in blood volume (BV) which could account for differences in STaH. Thus, to re‐address the question of genetic influences on STaH, 5 inbred rat strains [Brown Norway (BN/Mcwi, n=9), Dark Agouti (DA, n=8), Fawn Hooded Hypertensive (FHH, n=10), Lewis (Lew, n=9) and Salt Sensitive (SS, n=9)] were bled using strain specific BVs. For each rat, 47% of its BV was removed and STaH was measured. Threefold differences for STaH among the strains (p<0.0001) were observed (Table). Rats of an inbred strain are genetically very similar with a small degree of genetic variability. Thus, it was of interest to observe considerable intra‐strain variation for STaH (Table).

Strain
Mean STaH ± SEM (min)
Range (min)
Coefficient of Variation (%)

BN/Mcwi
52 ± 5
35 – 79
28

DA
190 ± 29
42 – 240
43

FHH
113 ± 24
31 – 240
66

LEW
95 ± 19
62 – 240
60

SS
114 ± 17
70 – 235
44

Environmental conditions were highly controlled in this study, thus: 1) observed inter‐strain variability confirms the presence of a genetic component to STaH; and 2) intra‐strain variability may be explained by alternatives such as epigenetic mechanisms. (Funding provided by USAMRMC).
A Study of GABA in Bivalve Molluscs
Authors: Cochran Tierra , Brown Cherryle , Mathew Keron , mathieu Sadchla , Carroll Margaret A , Catapane Edward J
Published: June 2021

The nervous systems of most studied bivalves contain serotonin and dopamine in their ganglia that serve as neurotransmitters regulating various physiological functions such as heart rate, foot movement, reproduction and gill lateral cilia beating. GABA is a major inhibitory neurotransmitter found in the CNS of vertebrates and ganglia of many invertebrates. Its presence and functions have not been well studied in bivalves. Here we used a HPLC method with pre‐column derivatization and fluorescence detection to look for GABA in ganglia of Crassostrea virginica, Mercenaria mercenaria and Mytilus edulis. We also examined effects of GABA on beating of gill lateral cilia in C. virginica and M. edulis. GABA was detected in low ng amounts in each ganglia of each of the bivalves. In C. virginica and M. edulis GABA had no direct effect on lateral cilia activity when applied directly to the gill. However, when serotonin was applied to the cerebral ganglia, GABA, whether applied before or after serotonin, blocked the normal excitatory response of serotonin on the beating of the cilia. Furthermore, the GABA antagonist bicuculline methchloride blocked the effects of GABA in the cerebral ganglia of C. virginica. This study is showing that GABA is present and has a neurophysiological role in bivalve ganglia. This work was supported by grants 2R25GM0600309 of NIGMS, 0516041071 of NYSDOE and 0622197 of NSF.
Purple‐fleshed potatoes suppress colon‐systemic oxidative stress/inflammatory markers via alternations in the gut bacterial signature
Authors: Reddivari Lavanya , Kim Sung Woo , Radhakrishnan Sridhar , Yang Peiying , Knight Robert , Vanamala Jairam
Published: June 2021

Purple‐fleshed potatoes (PP) are rich in putative anti‐oxidant/antiinflammatory anthocyanins. We hypothesized that PP, even after processing, suppress colon‐systemic oxidative stress/inflammatory markers by altering gut bacterial signature and metabolite profile. To test this hypothesis, 40 pigs on high‐fat diet (HFD) for 12 wk were provided with one of the five diets: HFD, white‐fleshed potato (WP) or PP chips at 10/20% for 5 wk. Serum, distal colon mucosa/digesta and fecal samples were utilized to analyze oxidative stress (MDA; colorimetric assay) and inflammatory (TNFα; ELISA and prostaglandins; LC‐MS/MS) markers, gut bacteria (pyrosequencing) and metabolites (LC‐MS). All diets, except WP (20%), suppressed (p = 0.04) systemic levels of TNFα compared to HFD. However, only PP suppressed MDA and colonic PGE2 levels (p ≤ 0.05). Potato diets increased members of Clostridia and Lachnospiraceae. Only PP increased distal colon bacteroidetes to firmicutes ratio (BFR) compared to HFD. This is particularly important as BFR is reduced during obesity. Indeed, MDA and TNFα elevated during obesity positively correlated with Firmicutes (0.518, p = 0.002; 0.380, p = 0.024, respectively) and MDA negatively correlated with BFR (−0.413, p = 0.036). Preliminary metabolomics data revealed 4350 metabolites in both distal colon and feces and the correlations with gut bacteria are being carried out.

Grant Funding Source: National Research Initiative Grant 2009–55200‐05197 from the USDA National Institute for Food and Agriculture (2009–2012)
Relationship between Parental Feeding and Child Eating Styles in Low‐income Families with Preschool Age Children
Authors: Vollmer Rachel L , Mobley Amy R
Published: June 2021

While parental feeding styles have been associated with children's dietary intake and weight status, the objective of this study was to determine how a child's eating style is related to their parent's feeding style. Parents of preschool children (n=43) completed the Caregiver Feeding Style Questionnaire to determine parental feeding style and the Child Eating Behaviour Questionnaire (CEBQ) to determine their child's eating style. Authoritarian (35%) and indulgent (30%) feeding styles were most common, and the authoritative (14%) feeding style was least common among parents. Children of indulgent parents had higher CEBQ scores for enjoyment of food (r=.453, p= 0.01) and lower scores for slowness in eating subscales (r=‐.430, p=0.01), which have both been linked to higher child BMI. Children of authoritarian parents had higher CEBQ scores for satiety responsiveness (r=.673, p=0.01) and slowness in eating (r=.600, p=0.01), and lower scores for enjoyment of food (r=‐.477, p=0.01), which have been associated with desirable child weight status. Interestingly, children of authoritarian parents scored higher on emotional over eating (r=.522, p=0.01), emotional under eating (r=.343, p=0.05), and food fussiness (r=.481, p=0.01), which may be why these parents use a controlling feeding style. Further research should determine the direction of the relationship between parent feeding and child eating styles.

Grant Funding Source: Subcontract to USDA/AFRI Grant 2010–85215‐20662
Molecular mechanisms of hypolipidemic effects of curcumin
Authors: Zingg Jean‐Marc , Hasan Syeda T. , Azzi Angelo , Meydani Mohsen
Published: June 2021

Some phytochemicals and micronutrients may directly scavenge free radicals or they may modulate expression of antioxidant genes. Alternatively, they may modulate lipid metabolic genes in tissues and reduce plasma lipid levels and thus indirectly prevent lipid‐mediated oxidative and endoplasmatic stress. Here we analyzed the molecular mechanisms by which the polyphenol curcumin may influence the elevated oxidative‐ and lipid‐mediated stress present during inflammation, obesity and atherosclerosis. In LDLr−/−mice fed a high fat diet (HFD), we found that curcumin (500, 1000, 1500 mg/kg diet, for 4 months) reduced plasma lipid levels what may indirectly contribute to inhibition of fatty acid transport proteins (CD36/FAT, FABP4/aP2) in peritoneal macrophages. Curcumin increased the HFD‐induced suppressed cAMP levels in tissues (liver, adipose). In THP‐1 monocytes, curcumin (5 and 10 μM) increased the level of cAMP and activated the transcription factor CREB. Since the cAMP/protein kinase A (PKA)/CREB pathway plays an important role in lipid homeostasis, energy expenditure and thermogenesis by increasing lipolysis and fatty acid β‐oxidation, an increase in cAMP levels induced by curcumin could be at the basis of its hypolipidemic and anti‐atherosclerotic effects, and similar to forskolin or caffeine also explain its effect on suppressing weight gain. Supported by USDA Contract #58–1950‐0–014.
Acute phytoecdysteroid treatment increases PI3k‐Akt signaling in aged mouse skeletal muscle
Authors: Zwetsloot Kevin A , Lawrence Marcus M , Arthur Susan T , Nieman David C , Lila Mary Ann , Grace Mary , Shanely R. Andrew
Published: June 2021

Activation of the PI3k‐Akt pathway leading to protein synthesis is diminished in aged skeletal muscle. Phytoecdysteroids, in particular 20‐hydroxyecdysone (20E), from the plant Ajuga turkestanica increases protein synthesis in C2C12 muscle cells. We previously reported that 21 mo old mice fed phytoecdysteroids for 28 d display larger fiber cross‐sectional area in the triceps brachii and plantaris muscles. The objective of this study was to determine the extent to which an acute dose of phytoecdysteroids activates PI3k‐Akt signaling in skeletal muscle of aged mice. Aged male C57BL/6 mice (20 mo) received a single gavage dose (50 mg/kg bw) of A. turkestanica extract (ATE), 20E, or vehicle (CT) and skeletal muscles were harvested 90 min later. Western blotting performed on gastrocnemius muscles revealed a significant 52% increase in Akt phosphorylation in 20E‐treated skeletal muscle of aged mice, compared to CT (p=0.006). The phosphorylation of p70, 4EBP1, and S6 ribosomal protein (downstream targets in the PI3k‐Akt pathway) were unaltered at this time point. In conclusion, these data suggest that a single dose of 20E activates Akt, a central component of the PI3k‐Akt pathway, in sedentary aged skeletal muscle.

Supported in part by a USDA Kannapolis Scholars Fellowship.
Physical activity behaviors and cognitions of parents of preschoolers (1021.6)
Authors: Spaccarotella Kim , Martin‐Biggers Jennifer , Hongu Nobuku , Worobey John , Byrd‐Bredbenner Carol
Published: June 2021

This study examined PA behaviors, cognitions, barriers, and strategies for coping with barriers by 137 parents of preschoolers via survey (n=137) and focus groups (n=23). Parents spent 4.2±2.2 SD days/week engaging in vigorous play with children, with 2/3 spending <3 hours/week in vigorous play. Time spent in active play rose significantly (p<0.01) as parent education increased but did not differ based on primary language (n=84 English, n=50 Spanish), location (n=65 AZ, n=72 NJ), or parent age. Preschoolers spent 2.4±1.7SD hours/day watching TV. Focus group parents reported using TV to occupy children while performing chores, during inclement weather, and before bed. Most believed TV time should be limited but were unsure of alternatives. All felt active play with their children was important for modeling healthy behaviors. Many considered educational TV exempt from TV time limits. Commonly‐cited barriers to increasing time spent in active play with their children included lack of indoor space, lack of safe outdoor play areas, limited time, and health concerns with being outdoors, such as asthma and insect bites. Coping strategies included active video games, school recreation programs, dancing to music, and visiting indoor playgrounds. Results suggest that parents with less education may especially benefit from strategies that help overcome barriers to modeling positive PA behaviors to their young children.

Grant Funding Source: Supported by USDA, National Institute of Food and Agriculture, Grant Number 2011‐68001‐30170
Osmotic regulation and tissue localization of the myo‐inositol biosynthesis pathway in tilapia (Oreochromis mossambicus) larvae (1182.6)
Authors: Sacchi Romina , Gardell Alison , Fernando Villarreal , Li Johnathon , Kültz Dietmar
Published: June 2021

The myo‐inositol biosynthesis (MIB) pathway converts glucose‐6‐phosphate to the compatible osmolyte myo‐inositol (MI), which protects cells from osmotic stress. We exposed tilapia larvae just after yolk sac resorption to various hypersaline environments and recorded robust induction of the enzymes that constitute the MIB pathway, myo‐inositol‐phosphate synthase (MIPS) and inositol monophosphatase 1 (IMPA1). Strong up‐regulation of these enzymes is evident at both mRNA (quantitative real‐time PCR) and protein (densitometric analysis of Western Blots) levels. The highest level of induction of MIB enzymes occurs at the highest salinity that larvae were exposed to (90ppt). Less severe osmotic stress causes proportionally lesser induction of the MIB pathway. Two distinct MIPS mRNA variants are present in tilapia larvae and both are induced at comparable levels for all the salinity challenges (34, 70 and 90ppt). Immunohistochemical localization of IMPA1 protein in sagittal sections of salinity stressed and control larvae identified tissues that are particularly potent in inducing the MIB pathway. These tissues include the skin, gills, eye (retina) and heart. We conclude that the MIB pathway plays an important role in protecting larval stages of tilapia from hyperosmotic stress. Therefore, tilapia larvae represent excellent in vivo models for studying physiological mechanisms by which the MIB pathway is regulated. Supported by NSF grant IOS‐1049780.

Grant Funding Source: NSF IOS‐1049780
Optimizing two‐component light‐switchable systems for controlling gene expression in Escherichia coli (991.2)
Authors: Stork Devon , Schmidl Sebastian , Tabor Jeff
Published: June 2021

Light‐induced gene expression is a powerful tool for manipulating cells because it allows for precise spatial and temporal control of cellular behavior. A light‐sensitive two‐component system based on the Synechocystis PCC6803 phytochrome CcaS and the cognate response regulator CcaR has been previously constructed in Escherichia coli.1 The chromophore‐binding domain of CcaS performs a reversible photoconversion between a green (~535 nm) and a red (~672 nm) light absorbing form. The green form phosphorylates CcaR, which then induces expression of the cpcG2 promoter by binding to a conserved direct repeat designated the G‐box. Green light‐induced transcription from PcpcG2 was about ~4.3‐fold for this original construct.

In order to engineer a constitutively expressed green/red light‐switchable two‐component system with improved fold‐inducibility, we built different ccaS expression cassettes using the constitutive promoter J23106 and RBSs with various translation initiation rates. This library was first screened against an aTc‐inducible expression system for CcaR resulting in a ~40 fold‐inducibility. After screening different ccaR expression cassettes using the constitutive promoter J23100 and RBSs with various translation initiation rates, a fold induction as high as ~80‐fold for a specific CcaS/CcaR‐expression pair could be observed.

There are over 10,000 two‐component systems in nature, and discovering the design and expression rules for these systems will allow engineering of a wide variety of genetic circuits for practical applications. The increase in fold‐induction in this system makes light‐induction a more powerful method of spatiotemporal control of living cells and aids precise cellular studies in synthetic and systems biology.

Grant Funding Source: Supported by an NSF award from the Division of Biological Infrastructure No. 1004853.
Glial Cell Line Derived Neurotrophic Factor Protein Secretion by Skeletal Muscles is Altered by Activity of L‐Type Calcium Channels
Authors: Boynton Alicia M. , Spitsbergen John M.
Published: June 2021

Glial cell line‐derived neurotrophic factor (GDNF) is an important signaling molecule throughout the peripheral nervous system. GDNF protein is produced and secreted by skeletal muscle cells and helps maintain motor neuron innervation at the neuromuscular junction. Treatment with exogenous GDNF prevents denervation which occurs with aging and neurodegenerative disease. The therapeutic potential of GDNF cannot be fully explored without understanding the mechanisms by which GDNF protein production is regulated. Although the specific cellular events necessary for regulation of GDNF secretion remains to be elucidated, previous studies from our laboratory have shown that increased skeletal muscle activity significantly alters GDNF protein levels in vivo and in vitro. The primary objective of this study was to determine the role of calcium in regulating GDNF protein secretion by skeletal muscle cells. Skeletal muscle cells (C2C12) were grown in culture and allowed to differentiate into myotubes which display a contractile phenotype. The following treatments were given, Nifedipine to block the voltage‐gated L‐type calcium channels, and Bay K8644 to enhance calcium influx through L‐type calcium channels. Both Western Blot and ELISA were employed for protein identification and GDNF protein quantification respectively. Preliminary results indicate that treatment of myotubes with Bay K8644 (100μM) caused an increase in GDNF secretion to 170% of that in untreated controls. Treatment of cells with Nifedipine blocked the stimulatory effect of Bay K8644, and treatment with Nifedipine alone (10μM and 100μM) inhibited GDNF secretion to 77% of that in control cells. The results suggest that secretion of GDNF protein by C2C12 myotubes is regulated by intracellular calcium levels. Acquiring a greater understanding of the role that calcium plays in regulating GDNF production may help to identify potential sites for therapeutic intervention to increase or decrease GDNF production.

Support or Funding Information

NSF Grant DBI‐1062883 and NIH Grant 1R15AG022908‐01A2
Crystal structures of acetylated HRas K104 mimic K104Q and mutant K104A suggest unique role of K104 in interlobe communication across HRas
Authors: Marcus Kendra , Johnson Christian , Sanchez Jaquelyn , Mattos Carla
Published: June 2021

Ras GTPases belong to a subfamily of signal transduction switches found in the Ras superfamily. Three isoforms of Ras (H‐, K‐, and N‐Ras) are expressed to regulate cellular proliferation, growth, and apoptosis. Unregulated activity of these proteins is closely related to disease, particularly the growth of tumors.1 The GTP/GDP cycle is a common regulatory feature of signaling output across GTPases. Exchange of GDP for GTP within the Ras active site results in an active signaling cascade. GAPs accelerate the hydrolysis of GTP to GDP, turning off the signal. In addition to exchange factors and activating proteins, the Ras GTPase cycle is also controlled by post‐translational modifications. An acetylation mimic (K104Q) in KRas G12V has been shown to negate oncogenic growth of the double mutant cell lines. However, KRas K104A/G12V does not reduce the oncogenicity of the cell line.2 The purpose of this study is to investigate the structural‐functional implications of K104Q and K104A mutations in HRas through X‐ray crystallography and to obtain their intrinsic hydrolysis rates through kinetic experiments with γ‐32P‐GTP. These data show a novel hydrogen‐bonding network present in K104Q, but not WT or K104A HRas. Intrinsic hydrolysis of GTP to GDP is significantly impaired in HRas K104Q, but is not affected by a K104A mutation. The unique network may control helix 5 motility in HRas, resulting in a lower intrinsic hydrolysis rate. Connectivity from K104Q towards helix 5 passes through a region of amino acids that are different between the H‐, K‐, and N‐ Ras isoforms. This suggests that Ras acetylation of K104 may affect the function of the isoforms in subtly different ways.

Support or Funding Information

This work is funded by a grant from the NSF MCB‐1244203
Defining Thermophilic Glucan Dikinases in Cyanidioschyzon merolae
Authors: Brizzee Corey O. , Husodo Satrio , Gentry Matthew S.
Published: June 2021

Objective

The goal of this project is to elucidate the mechanism of action of starch dikinases in carbohydrate metabolism using the red alga Cyanidioschyzon merolae (Cm) as a model system.

Significance

Highly regulated reversible starch phosphorylation is key for starch metabolism in photosynthetic organisms such as plants and algae. Defining the enzymes that are involved in reversible starch phosphorylation in C. merolae will allow us to engineer strategies to harness starch production in a more efficient manner for many industrial applications.

Methods

Bioinformatics to identify conserved functional domains with orthologs from different kingdoms. Radiometric kinase assays to measure specificity and kinetic parameters of starch dikinases. Differential Scanning Fluorimetry (DSF) to characterize glucan binding and protein thermal stability. Cell biology techniques such as gene suppression, synchronized cultures, mRNA extraction, and immunofluorescence. Structural analysis through Small Angle X‐Ray Scattering (SAXS), protein purification, and western analysis.

Results

We show here that C. merolae GWD shares conserved functional domains with orthologs from many different kingdoms. Validating Cm‐GWD is a functional glucan dikinase, results show robust Cm‐GWD specificity towards phosphorylating the C6 position of glucose residues. Radiometric‐ATP incorporation assays reveals that recombinant Cm‐GWD is as efficient as Arabidopsis thaliana (At)‐GWD at phosphorylating starch. Lastly, we utilize SAXS to structurally characterize each domain of the multi‐domain Cm‐GWD.

Conclusion

We have defined C. merolae growth and optimal starch accumulation parameters as well as cell localization of enzymes involved in reversible starch phosphorylation. Furthermore, we have characterized gene expression of Cm‐GWD via mRNA levels during synchronous cultures. Additionally, this study exposes biochemical targets to enhance starch metabolism in planta. The biochemical properties of Cm‐GWD enable increased starch metabolism not only in C. merolae but in exogenous plants.

Support or Funding Information

KSEF grants KSEF‐2268RDE‐014 and KSEF‐2971‐RDE‐017; NSF Grants IIA‐1355438 and MCB‐1252345
Perceived Onset of Obesity in Sleeve Gastrectomy Candidates
Authors: Robinson Katie , Rowitz Blair , Donovan Sharon M. , Teran‐Garcia Margarita
Published: June 2021

Sleeve gastrectomy (SG) procedures are becoming increasingly common in response to the rising prevalence of morbid obesity. Post‐operatively, SG patients experience positive outcomes beyond weight loss, including metabolic disease resolution and improved quality of life, which may be impacted by previous life experiences, such as onset and duration of obesity. The degree of influence of these factors is unknown. Therefore, the aim of this research is to describe the characteristics of a cohort of SG candidates and to highlight differences between individuals who reported earlier onset of weight gain and obesity with those who experienced weight gain later in life. Candidates for SG (n=28, 29% male, 46.8 ± 9.0 yrs) completed surveys in which they identified their perceived age of obesity onset and their child body size using a set of child images that represented varying body mass index (BMI) percentiles. Three‐day food logs, physical activity records and body composition via bioelectrical impedance were collected prior to the preoperative diet, on the morning of surgery and one month post‐operatively.

At baseline, mean BMI was 48.4 ± 7.4 kg/m2 and body fat percentage was 50.9 ± 5.0%. The most common reported cause of weight gain was poor eating habits (82%), followed by physical inactivity (75%), genetics (46%), stress (36%) and depression (29%). The most common comorbidities were hypertension (54%), hypercholesterolemia (43%), anxiety (36%), type 2 diabetes (32%) and depression (32%). Nine women and two men identified with the child's image that was greater than the 90th percentile (OW). These participants were compared to those who perceived themselves to be normal weight children. Women who reported that they were OW as children had higher BMIs at initial visit (p=0.002). Participants who reported OW as children reported less minutes walking (p=0.05) and no differences in diagnosed comorbidities.

Men lost 5.8 ± 1.7 kg and 1.9 ± 0.6 BMI units and women lost 5.5 ± 1.8 kg and 2.0 ± 0.8 BMI units following the two week pre‐operative diet. Men lost significantly more body fat (1.75 ± 2.0 %) than women (0.05 ± 1.3%). Weight loss during pre‐operative diet was not significantly different based on perceived onset of obesity or child body size perception. Although data collection in this cohort is ongoing, our findings suggest that perceived child size and onset of weight gain is associated with adult BMI, however, it does not predict the ability of an individual to lose weight pre‐operatively.

Support or Funding Information

This research was supported by the Agriculture and Food Research Initiative of the USDA National Institute of Food and Agriculture as part of the AFRI Childhood Obesity Prevention Challenge (2011‐67001‐30101) to the Division of Nutritional Sciences at the University of Illinois.
The Influence of Commensal Microflora on Small Intestinal Inflammation and Wnt Pathway in High Fat‐ and Genetically‐induced Obese Apc1638N Mice
Authors: Liu Wei , Crott Jimmy W. , Li Jinchao , Pfalzer Anna C. , Mason Joel B. , Liu Zhenhua
Published: June 2021

Obesity, which has reached epidemic proportions worldwide, is an established risk factor for some cancers, but the mechanisms responsible for this relationship remain elusive. Our previous study demonstrated that obesity‐induced inflammation elevates Wnt‐signaling, a pro‐tumorigenic pathway that is altered in ~90% of colon cancers. The objective of this study was to examine the role of the intestinal microflora in the increases in inflammation and Wnt‐signaling observed in both high fat‐ and genetically‐induced obese state, and thereby the intestinal tumorigenesis in the Apc1638N mouse model. Three experimental groups with 2 different genotypes and 2 dietary treatments were used: Apc+/1638NLepr+/+ fed a low fat diet (10% fat, Apc‐LFD), Apc+/1638NLepr+/+ fed a high fat diet (60% fat, Apc‐HFD), and Apc+/1638N Leprdb/db fed a low fat diet (Apc‐DbDb). All animals (9~12 mice/group) received experimental diets for 16 weeks from 8 to 24 weeks of age. Because tumors occur exclusively in the small intestine in the Apc1638N model, we examined the microbial composition, inflammatory status and the expressions of Wnt pathway‐specific genes in this organ, as opposed to the large intestine. The relative abundances of 19 bowel cancer associated bacterial taxa were examined by quantitative real‐time PCR. The prevalence of Turicibacter and Desulfovibrio decreased, but Faecalibacterium prausnitizii (F.prausnitizii) increased significantly in high fat diet‐induced obese mice (p < 0.05); whereas in genetically‐induced obese mice, the prevalence of Bifidobacterium, Akkermansia muciniphila (A.muciniphila) and Eubacterium rectale (E.rectale) decreased, but Peptostrptococcus, and Escherichia coli(E.coli) increased (p < 0.05). Both high fat‐ and genetically‐induced obesity increased small intestinal inflammation as indicated by elevated protein levels of IFNγ, IL‐6 or TNF‐α. The changes of inflammatory cytokine transcripts, including TNF‐α, were associated with abundance of Turicibacter, Bifidobacterium, A.muciniphila, E.rectale, and E.coli, as well as the expression of several genes (Wnt5a, Akt, β‐catenin, JNK1, etc.) within the Wnt pathway cascade (p < 0.05). These findings demonstrate that the composition of the small intestinal microflora is affected differentially in high fat diet‐ and genetically‐induced obesity, but both are associated with an elevation of intestinal inflammation and alterations in the expression of Wnt‐signaling related genes.

Support or Funding Information

This project is supported by USDA grants (2014‐67017‐21762, MAS00454), SPHHS Deans Incentive Grant; and the Agricultural Research Service of the United States Department of Agriculture, Cooperative Agreements #58‐1950‐0‐014 and #58‐1950‐4‐003
Long Term Hypoxia Reduces Ca2+ Wave Function In Basilar Arterial Myocytes of Fetal and Adult Sheep
Authors: Reid Casey , Romero Monica , Francis Michael , Taylor Mark S , Longo Lawrence D , Hessinger David A , Wilson Sean M
Published: June 2021

Ca2+ entry through L‐Type channels mediate contraction of basilar arterial myocytes and whole‐cell Ca2+ waveforms are thought to be integral to this process. Based on previous evidence, we hypothesize that modifications, due to both long term hypoxia (LTH) and ontogeny, will affect the influence of depolarization on intracellular Ca2+ waves. Specifically, fetal arteries will have reduced Ca2+ signals while LTH will further reduce depolarization induced Ca2+ responses. In order to test this hypothesis, we examined Ca2+ wave activity using confocal fluorescence imaging techniques on basilar arterial myocytes in an en face preparation of low and high altitude fetal (FN, FH) and adult (AN, AH) sheep, where sheep were placed at 3,801 m for >100 days. Ca2+ waves were evaluated with and without 30mM K (30K), which was used to depolarize the myocytes. LTH decreased the area under the curve (AUC) independent of animal age. These changes in AUC were due to a faster decay in the Ca2+ signal. Depolarization with 30K increased Ca2+ wave amplitude and duration in all groups except for AH. These observations illustrate that both altitude and ontogeny play important roles in affecting various aspects of the Ca2+ signals. This likely contributes to the changes in arterial contractility that were previously observed with ontogeny and LTH and potentially other mechanisms pertaining to cerebral blood flow.

Support or Funding Information

Support from NIH and NSF
Anti‐MBG antibodies attenuate MBG‐induced anti‐proliferative and anti‐angiogenic milieu in cytotrophoblast cell model
Authors: Afroze Syeda H , Larrick James W , Wright Susan C , Drever Nathan N , Allen Steven R , Sharma Vikram , Mendelsohn Andrew R , Wages John M , Zawieja David C , Kuehl Thomas J , Uddin Mohammad N
Published: June 2021

Preeclampsia (PreE) is a hypertensive pregnancy disorder, which occurs in approximately 10% of all gestations. Recently, a digitalis‐like factor, marinobufagenin (MBG) has been implicated as a causative factor in preE. We demonstrated that MBG inhibits the proliferation, migration, and invasion of cytotrophoblast (CTB) cells. We have identified a novel human monoclonal antibody with higher specificity than Digibind for MBG. We assessed the attenuation of MBG‐induced CTBs dysfunction by three anti‐MBG antibodies: 206–208, H1L2, and 3e9.

Methods

A panel of anti‐MBG antibodies with potential as human therapeutic agents was developed by Panorama Research, Inc.; Sunnyvale, CA (206–208, H1L2). H1L2 was a humanized version of previously described anti‐MBG murine antibody 3e9. 206–208 was identified in a human phage antibody library. Human CTBs were treated with DMSO (vehicle) or 0.1, 1, 10 or 100 nM of MBG for 48 h. Some cells were pretreated with 206–208, H1L2, or 3e9 for 2h, while others were co‐treated with these antibodies prior to MBG treatment. Culture media were collected for analysis of pro‐angiogenic and anti‐angiogenic factors. Cell viability was measured using a CellTiter Assay kit. Cell lysates were utilized to evaluate the expression of proliferating cell nuclear antigen (PCNA) and p38 MAPK phosphorylation by western blot. Levels of vascular endothelial growth factor (VEGF), placental growth factor (PlGF), soluble fms‐like tyrosine kinase‐1 (sFlt‐1), and soluble endoglin (sEng) were measured in culture media using ELISA kits. Statistical comparisons were performed using analysis of variance with Duncan's post hoc test.

Results

MBG down‐regulated PCNA and up‐regulated p38 phosphorylation in CTBs treated with ≥1 nM MBG compared to basal (DMSO treatment) (*p < 0.05 for each). Secretion of sFlt‐1 and sEng were increased, while VEGF and PIGF were decreased in CTBs treated with ≥1.0 nM MBG (*p < 0.05 for each). Both anti‐MBG antibodies pretreatment and co‐treatment significantly up‐regulated PCNA and down‐regulated p38 phosphorylation, and corrected the angiogenic profile of CTBs (p < 0.05 for each). The anti‐proliferative and anti‐angiogenic effects of MBG were not due to a cytotoxic effect, as evaluated by a cell viability assay. MBG at 0.1 nM had no effect.

Conclusions

We found that all 3 anti‐MBG antibodies attenuate MBG‐induced anti‐proliferative and anti‐angiogenic milieu in cultured CTBs. Here we describe for the first time two fully human anti‐MBG antibodies, which can be targeted as therapeutic agents for the development of innovative treatment strategies for preE and potentially other disorders involving aberrant MBG signaling.

Support or Funding Information

NIH Small Business Innovation Research (SBIR) : 1R43HD079238‐01
Angiotensin Receptor Blocker ameliorates hyperlipidemia and increases lipid utilization in a rat model of metabolic syndrome
Authors: Garcia Jose Antonio , Lee Andrew , Ortiz Rudy
Published: June 2021

CD36 and lipoprotein lipase (LPL) are essential proteins involved in lipid mobilization and triglyceride storage in various cell types, including adipocytes. Accumulation of excess lipids in non‐adipose tissues leads to cell dysfunction, potentially cell death, and ultimately insulin resistance. Furthermore, activation of the renin‐angiotensin system (RAS) is implicated in the manifestation of metabolic syndrome, for which dyslipidemia is an associated factor. To address our hypothesis that blockade of the angiotensin II receptor (AT1) can improve lipid utilization, we measured body mass, retroperitoneal mass, adipose lipase activity, CD36 and LPL expression in five groups of rats; 1) Long‐Evans Tokushima Otsuka (LETO) normal diet (ND) (n=6), 2) Otsuka Long‐Evans Tokushima Fatty (OLETF) ND (n=8), 3) OLETF high‐fat diet (HFD, 62% fat in food) (n=8), 4) OLETF + angiotensin receptor blocker (ARB) (10 mg olmesartan/kg/d 6 wk) (n=8), and 5) OLETF HFD + ARB (n=7). Body mass increased 38% in OLETF relative to LETO; ARB decreased it by 20% relative to OLETF, and HFD+ARB decreased it by 37% relative to HFD. ARB decreased retroperitoneal fat 16% in relation to OLETF, and ARB+HFD decreased it 72% compared to HFD. CD36 decreased in response to a HFD and lipase activity increased with ARB treatment. Results suggest that AT1 activation increases lipid utilization, therefore, ameliorating the development of hyperlipidemia from a high fat diet.

Support or Funding Information

UC Leadership and Excellence through Advanced Degrees (UC LEADS) United States Department of Agriculture (USDA)
Bone as a buffer: Consequences for vertebrate design and anaerobic physiological performance
Authors: Warren Dan
Published: June 2021

Conventional wisdom usually states that mineralized tissues are primarily used by vertebrates as sites for respiratory and locomotory muscle attachment or for physical protection from predators. Often ignored is the role these tissues play in maintaining ionic and acid‐base homeostasis and how this might have shaped vertebrate evolution. We have argued that this latter function may have been important in shaping vertebrate design in early tetrapods, particularly the maintenance of dermal armor in early tetrapods, and is still important for anaerobic physiological performance in many extant vertebrates, especially in reptiles and amphibians. We use physiological, biomechanical, and mineralogical approaches to understand how vertebrate bone, especially that of turtles, interacts with and is affected by extreme deviations in acid‐base homeostasis. Our work, and that of others, has shown that the mineral composition of turtle bone can predict tolerance to extreme acidosis, and that the most derived of these vertebrates, the anoxia‐tolerant painted turtle, is able to utilize calcium and magnesium carbonates within its bone to prevent lethal decreases in body fluid pH in the face of a 180 mM lactic acid load, with minimal effect on whole‐bone material properties.

Support or Funding Information

This work is funded by a CAREER Award from the National Science Foundation.
Self‐Organizing Actomyosin Patterns on the Cell Cortex at Epithelial Cell‐Cell Junctions
Authors: Neufeld Zoltan
Published: June 2021

The behavior of actomyosin critically determines morphologically distinct patterns of contractility found at the interface between adherent cells. One such pattern is found at the apical region (zonula adherens) of cell‐cell junctions in epithelia, where clusters of the adhesion molecule E‐cadherin concentrate in a static pattern. Meanwhile, E‐cadherin clusters throughout lateral cell‐cell contacts display dynamic movements in the plane of the junctions. To gain insight into the principles that determine the nature and organization of these dynamic structures, we analyze this behavior by modeling the 2D actomyosin cell cortex as an active fluid medium. The numerical simulations show that the stability of the actin filaments influences the spatial structure and dynamics of the system. We find that in addition to static Turing‐type patterns, persistent dynamic behavior occurs in a wide range of parameters. In the 2D model, mechanical stress‐dependent actin breakdown is shown to produce a continuously changing network of actin bridges, whereas with a constant breakdown rate, more isolated clusters of actomyosin tend to form. The model qualitatively reproduces the dynamic and stable patterns experimentally observed at the junctions between epithelial cells.

Support or Funding Information

This work was supported by the National Health and Medical Research Council of Australia (1037320, 1044041, and 1067405), the Australian Research Council (DP120104667 and FT130100659), and the Kids Cancer Project of the Oncology Research Foundation.
The Role of Cell Contractility in Epithelial Morphogenesis
Authors: Nelson Celeste M. , Varner Victor D.
Published: June 2021

Cell‐generated mechanical forces drive many of the tissue movements and rearrangements that are required to transform simple populations of cells into the complex three‐dimensional geometries of mature organs. However, mechanical forces do not need to arise from active cellular movements. Here, I will describe recent studies that have illuminated the roles of passive mechanical forces resulting from mechanical instabilities between epithelial tissues and their surroundings. These mechanical instabilities cause essentially one‐dimensional epithelial tubes and two‐dimensional epithelial sheets to buckle or wrinkle into complex topologies containing loops, folds, and undulations in organs as diverse as the brain, the intestine, and the lung. Our work suggests a new class of tissue development ‐ buckling or wrinkling morphogenesis ‐ and that this morphogenetic mechanism may be broadly responsible for sculpting organ form. Harnassing these mechanical instabilities represents an intriguing strategy for engineering organs ex vivo.

Support or Funding Information

This work was supported by grants from the NIH (HL118532 and HL120142) and the NSF (CMMI‐1435853).
Green tea extract lowers NFκB‐mediated inflammation during nonalcoholic steatohepatitis in mice fed a high‐fat diet consistent with reduced Toll‐like receptor‐4 signaling
Authors: Li Jinhui , Chitchumroonchokchai Chureeporn , Kim Joshua B. , Moller Meredith V. , Bruno Richard S.
Published: June 2021

Green tea extract (GTE) reduces NFκB‐mediated inflammation and oxidative stress during nonalcoholic steatohepatitis (NASH). We hypothesized that its antiinflammatory activities that limit oxidative stress and liver steatosis would be mediated in a Toll‐like receptor‐4 (TLR4)‐ dependent manner. Wild‐type (WT) and loss‐of‐function TLR4‐mutant (TLR4m) mice were fed a high‐fat diet containing 0 or 2% (w/w) powdered GTE for 8 wk before assessing hepatic steatosis, oxidative stress, and NFκB‐mediated inflammation. TLR4m mice had lower (P<0.05) body mass compared to WT mice, but similarly high adipose mass, whereas body mass and adiposity were lowered by GTE regardless of genotype. GTE in WT mice reduced hepatic TLR4 and MCP‐1 mRNA, and protein levels of the phosphorylated‐p65 NFκB subunit, which were also lowered in TLR4m mice regardless of GTE. Oxidized:reduced glutathione and malondialdehyde were lowered by GTE in WT mice and similarly lowered in TLR4m mice regardless of GTE. Relative to WT mice, histological evidence of liver steatosis, hepatic triglyceride, and serum alanine aminotransferase were also lowered in WT mice fed GTE and these were similarly lowered in TLR4m mice regardless of GTE. However, GTE lowered lipogenic gene expression regardless of genotype, but their expression in TLR4 controls was unaffected compared to WT controls. TLR4m mice had reduced insulin resistance compared to WT mice, but GTE in WT mice lowered it to a greater extent by more substantially reducing fasting glucose. These findings suggest that GTE limits NFκB‐mediated inflammatory responses leading to liver steatosis and oxidative stress during NASH, consistent with a mechanism of reduced pro‐inflammatory signaling through TLR4/NFκB.

Support or Funding Information

Supported by USDA‐NIFA and Ohio Agricultural Research and Development Center
Controlling the Rate of Energy Transduction in a Protein System Through Coupling Pathway Redesign
Authors: Wilson Corey J.
Published: June 2021

Protein mediated energy transduction reactions are fundamental life‐processes that can be leveraged to produce novel bio‐electronic devices and advance our understanding of complex phenomena such as photosynthesis and cellular respiration. Energy transduction typically involves two fundamental reactions, electron transfer (EleT) and energy transfer (EngT). Thus, our ability to control (re‐design) the properties that influence EleT and EngT are of critical importance. In this study we used computational protein design to alter the coupling matrix, followed by complementary and extensive experimental characterization to study the role of the intervening medium between the acceptor and donor species during EleT and EngT reactions using the model system Pseudomonas aeruginosa azurin.

Support or Funding Information

This work was supported by NSF Award 1507385, to CJW
Comparison of the Effects of a Sweetened Beverage Intervention on Self‐Selected Food Intake
Authors: Widaman Adrianne M. , Stanhope Kimber L. , Chen Guoxia X. , Medici Valentina M. , Bremer Andrew A. , Lee Vivian , Nunez Marinelle V. , Havel Peter J. , Keim Nancy L.
Published: June 2021

Evidence suggests that the intake of added sugar increases the risk of chronic disease and should be targeted for reduction. It is unclear if all types of added sugar have equivalent effects on food intake. This prospective, blinded intervention study compared parallel groups consuming one of five types of beverage sweeteners: glucose (GLU), fructose (FRU), sucrose (SUC), high fructose corn syrup (HFCS) or aspartame (ASP). The beverages were provided at ~25% of estimated energy needs for weight maintenance for 2 weeks and consumed along with the subjects’ usual diets. Before (week 0) and at the end (week 2) of the intervention, subjects were admitted to a metabolic ward and as inpatients participated in a day‐long protocol designed to measure ad libitum food and beverage intake from buffets offered at breakfast, lunch, and dinner. One hundred and thirty men and women with an average age of 26.3 ± 6.3 y and a BMI of 25.3 ± 3.6 kg/m2 participated. The changes in food and beverage energy intake (week 2 – week 0) were analyzed using ANCOVA, adjusting for BMI, sex, and energy intake at week 0. Energy intake was significantly affected by sugar (p=0.0026, ANCOVA), with subjects consuming SUC (386 ± 96 kcal, p=0.0011, Tukeys) or HFCS (242 ± 89 kcal, p=0.022, Tukeys) significantly increasing their intake at 2 weeks compared with subjects consuming ASP (−169 ± 100 kcal) and compared with their week 0 intake (p=0.0001 and p=0.0072, respectively). Buffet energy intake did not change significantly for the groups consuming GLU (p=0.198), FRU (p=0.064), or ASP (p=0.096) between weeks 0 and 2. Further analysis of the influence of sex on buffet intake showed that the increases in energy intake at week 2 were mainly attributable to men. Men consuming SUC (580 ± 137 kcal, p=0.0072, Tukeys) or HFCS (426 ± 123 kcal, p=0.047, Tukeys) significantly increased their energy intake at 2 weeks compared with men consuming ASP (−176 ± 143 kcal). In contrast, for women, there were no differences among groups, or between week 0 and week 2 buffet energy intake, regardless of the sweetened beverage assigned. In conclusion, we found that compared with aspartame, the caloric sweeteners, HFCS and sucrose, commonly used to sweeten soft drinks and fruit drinks, increased total energy intake during a day‐long buffet protocol offered in a controlled research setting. The effect of these sweeteners was more pronounced in men.

Support or Funding Information

Supported by NIH 1R01 HL09133, 1R01 HL107256, RR024146, and K12 HD051958, and USDA Project 2032‐51530‐022
SIRT5 in cancer metabolic reprogramming
Authors: Lombard David B , Park Jeongsoon , Kumar Surinder
Published: June 2021

The sirtuins are a family of NAD+‐dependent protein deacylases that regulate metabolism and other diverse aspects of cell biology. Three sirtuins (SIRT3, SIRT4, and SIRT5) are present in the mitochondrial matrix. SIRT5 is an inefficient deacetylase, instead removing succinyl, malonyl, and glutaryl groups from lysines of its target proteins. To provide insight into SIRT5 functions, together with Dr. Yingming Zhao's group, we previously carried out proteomics profiling of the cellular succinylome (Park et al., Mol. Cell 2013). This revealed potential impacts of succinylation on enzymes involved in diverse aspects of mitochondrial metabolism. We tested the impact of SIRT5 on two substrates identified in our studies, Pyruvate Dehydrogenase Complex (PDC) and Succinate Dehydrogenase (SDH). SIRT5 inhibited biochemical activities of both complexes, and suppressed overall mitochondrial respiration in the presence of glucose, pyruvate, and succinate.

Metabolism in tumor cells is “reprogrammed” to meet the anabolic demands of uncontrolled cellular proliferation. Reprogramming of mitochondrial metabolism represents a key element of this metabolic rewiring. PDC and SDH are both implicated in neoplasia and cancer cell metabolic reprogramming. In particular, activation of PDC activity in cancer cells reverts tumor cell metabolism, inhibits malignant behavior, and induces apoptosis in diverse cancer types. Given the role of SIRT5 in regulating PDC and SDH, we hypothesized that SIRT5 might play a role in metabolic reprogramming in cancer. Our progress in testing this idea will be discussed.

Support or Funding Information

NIH R01GM101171 and UL1TR000433, DoD OC140123, the Glenn Foundation for Medical Research, the John S. and Suzanne C. Munn Cancer Fund of the University of Michigan Comprehensive Cancer Center, and the University of Michigan Department of Pathology.