bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of Mono‐Acyl‐Glycerol Lipase by JZL‐184 Results in Glucolipotoxicity in Pancreatic β‐Cells
Authors: Berdan Charles , Corkey Barbara E , Deeney Jude T
Published: June 2021

Objective

Chronic exposure of pancreatic β‐cells to elevated glucose and fatty acid (FA) levels (24–48 hrs) results in glucolipoxity (GLT), a condition mimicking type 2 diabetes with elevated basal secretion and impaired glucose stimulated insulin secretion (GSIS). Acute exposure to mono‐oleoyl‐glycerol (MOG) results in rapid onset (<1hr) of GLT in both clonal β‐cells and dissociated rat islets. Our goal was to determine the effect of altered lipid cycling on β‐cell function using JZL, a mono‐acyl‐glycerol (MAG) lipase inhibitor.

Methods

Acute and chronic incubations of clonal β‐cells (INS‐1 832/13) and dissociated rat islets with the MAG lipase inhibitor JZL‐184 were performed. Insulin secretion (HTRF), glycerol release (NADH luciferase) and intracellular lipid accumulation (Nile red) were measured.

Results

JZL (1–10uM) acutely inhibited GSIS and KCl/diazoxide‐stimulated insulin secretion in both dissociated rat islets and INS‐1 cells. Chronic JZL (10uM) exposure in the presence of FA (0.1 mM) increased intracellular lipid droplets and accelerated the development of GLT in INS‐1 cells. Under these same conditions JZL reduced glycerol release indicating MAG lipase was inhibited.

Conclusion

Inhibition of MAG lipase resulted in altered insulin secretion mimicking chronic effects of elevated glucose and FA.

Funded by NIH.
The miR‐696 contributes to insulin resistance by reducing mitochondrial function and antioxidant capacity in skeletal muscle cells
Authors: Queiroz Andre Lima , Sousa Thais Amaral , Teodoro Bruno Gonçalves , Bomfim Lucas M , Sampaio Igor H , Silveira Leonardo R
Published: June 2021

Insulin resistance is associated to type II diabetes but its mechanism is not fully understood. Micro‐RNAs have been described as non‐coding molecules associated with post‐transcriptional regulation of several physiological processes including diabetes. We have observed that miR‐696 is regulated in an opposite manner compared to PGC‐1α leading to a proposition that the miR‐696 may be involved to insulin resistance. Mice mioblast cells (C2C12) were treated with palmitic acid (500μM) for 24 h. The insulin resistance was confirmed by evaluation of Akt phosphorylation after insulin incubation by western blot. Expression of mature miR‐696 was evaluated by real‐time RT‐PCR using TaqMan® MicroRNA Assay. The expression of microRNAs was normalized with the SnoR‐202. Gene expression of PGC1α was evaluated by real time RT‐PCR using Sybr Green. Analysis of oxygen consumption was performed by oxygraph (Hansatech) using CCCP and oligomycin as controls. We observed that PGC1α transcription was markedly reduced in insulin resistant cells, an effect that was reflected by a low mitochondrial respiration, reduced antioxidant capacity and elevated miR‐696 level. In contrast, the PGC1α transcription was markedly elevated after AICAR treatment as reflected by elevated mitochondrial respiration and antioxidant capacity and low miR‐696 level. Interestingly, the inhibition of miR‐696 transcription in insulin resistant cells re‐established the insulin response, PGC1α transcription and mitochondrial function indicating an association of miR‐696 and mitochondrial biogenesis. Therefore, we are suggesting that miR‐696 contributes to insulin resistance reducing mitochondrial biogenesis and mitochondrial function in skeletal muscle cells.
Inhibition of secretory Phospholipase A2 Group IIa by thyroid hormone
Authors: Sharma Pragya , Thakran Shalini , Deng Xiong , Elam Marshall B , Park Edwards A
Published: June 2021

Secretory phospholipase A2 group IIa (PLA2g2a) is involved in multiple biological processes including inflammation and atherosclerosis. In this study, we investigated the regulation of PLA2g2a by thyroid hormone (T3). T3 stimulates expression of genes through the binding of the nuclear T3 receptor (TR) to T3 response elements (TRE). The liganded TRβ recruits transcriptional coactivators to stimulate gene expression. However, as many genes are down regulated by T3 as are stimulated. We found that T3 suppressed the expression of the PLA2g2a gene in hepatocytes, human HepG2 cells and in vivo in rats. To gain insight into the mechanism of transcriptional repression, we cloned the PLA2g2a promoter. We identified a negative TRE (nTRE) in the promoter that bound the T3 receptor as a heterodimer with the retinoid X receptor (RXR). Mutation of the nTRE eliminated the ability of T3 to inhibit PLA2g2a. Knockdown of the nuclear corepressors (NCoR) blocked the T3 inhibition of PLA2g2a. In conclusion our experiments demonstrate a novel inverse recruitment mechanism where liganded TRβ recruits corepressors to inhibit PLA2g2a expression. These results suggest that in addition to the well known role of T3 in regulating genes involved in hepatic metabolism T3 may also modulate the expression of inflammatory genes.

This work was supported by National Institute of Health.
Effects of Aloe vera components on the Renin‐Angiotensin System in human mesangial cells
Authors: Arita Lilian S , Santos Lilian , Yokota Rodrigo , Tanae Mirtes Midori , Aragão Danielle S , Nogueira Marie Doki , Banchi Rafael Silva , Matsumoto Larissa Emi , Arita Danielle Yuri , Cunha Tatiana Sousa , de Riggio Lima‐Landman Maria Teresa , Casarini Dulce Elena
Published: June 2021

The local Renin‐Angiotensin System (RAS) is related to the development of diabetic nephropathy. Studies show that Aloe vera has hipoglicemic and antioxidant effects. Aloe emodin (AE), a component of A. vera, increases glycogen synthesis and also glucose transport via the PKI3 intracellular pathway. We evaluate if the Aloe vera components have effects on angiotensin I‐converting enzyme (ACE) activity in human mesangial cells exposed to normal (NG) and high glucose (HG) medium. Two components, P9 and P9a, were semi‐purified from A. vera by High Performance Liquid Chromatography (HPLC) and were eluated at the same retention time of the standard AE. Cells were treated with insulin, P9, P9a and AE at 1, 100 and 1000 pg/mL and ACE activity was evaluated using Z‐Phe‐His‐Leu‐OH as substrate. All treatments were compared to control. Cells under NG treated with P9, P9a and AE showed decreased ACE activity. In HG, high doses of P9 increased ACE activity and AE tend to present the same profile. These data suggest that the semi‐purified fractions contain the Aloe emodin component. Further studies are needed to elucidate the role of A. vera over RAS under physiological and pathological conditions, which may occur by sinergistic or isolated compound action. It could be important in A. vera terapeutic effects research, for instance, in diabetic nephropathy.
A Modular Approach to the Histone H2A Family of Antimicrobial Peptides
Authors: Fischer Alexandra L , Bustillo Maria , Elmore Donald E
Published: June 2021

Hipposin and buforin II (BF2) are naturally occurring antimicrobial peptides (AMPs). Although BF2 is essentially identical to the middle section of hipposin, BF2 is a membrane translocating peptide and hipposin is a membrane permeabilizing peptide. To investigate this difference, we compared the structure, antimicrobial activity, translocating ability, and membrane permeabilizing ability of hipposin, BF2, parasin (analogous to the N‐terminal section of hipposin), and buforin I (essentially parasin plus BF2). Hipposin and BF2 had the highest activities against bacteria, followed by buforin I then parasin. Cellular permeabilization assays demonstrated that hipposin, buforin I, and parasin all display membrane permeabilizing ability, while translocation assays showed that only BF2 was membrane translocating. These observations imply that the addition of the parasin fragment to BF2 alters its antibacterial mechanism. Ongoing studies are examining the activity and mechanisms of other portions of the hipposin peptide.
Engineering a specific phosphatidylcholine binding site motif into a phosphatidylinositol‐specific phospholipase C
Authors: Goldstein Rebecca , Cheng Jiongjia , Stec Boguslaw , Gershenson Anne , Roberts Mary Fedarko
Published: June 2021

Peripheral membrane proteins can be targeted to specific organelles or the plasma membrane by differential recognition of phospholipid headgroups. While molecular determinants of specificity for phosphatidylserine and phosphoinositide headgroups are well‐defined, specific recognition of the headgroups of zwitterionic lipids, including phosphatidylcholine (PC), is less well characterized. For PC, the tetramethylammonium provides a handle that can be recognized by 2 to 4 aromatic residues located within 4–5 Å of the quaternary amine. Engineering this π‐box into secreted phosphatidylinositol‐specific phospholipase C from Staphylococcus aureus, which lacks affinity for PC, results in dramatically enhanced binding to PC‐rich vesicles, with a much lower affinity for phosphatidylethanolamine‐rich vesicles. Crystal structures of the S. aureus PI‐PLC N254Y/H258Y detect choline as well as dibutyroylphosphatidylcholine molecules binding to the mutated protein via π‐cation interactions. This simple PC recognition motif may be engineered into a variety of secondary structures providing a general method for anchoring proteins to membranes with zwitterionic phospholipids. This work was supported by N.I.H grant R01 GM60418.
Regulation of the ATP Hydrolysis and Transport Cycles of the P‐Glycoprotein Multidrug Transporter by Sterols and Phospholipids
Authors: Sharom Frances J , Clay Adam T , Lu Peihua
Published: June 2021

Multidrug resistance in many human cancers is linked to expression of the P‐glycoprotein multidrug transporter (Pgp, ABCB1), an ABC protein that exports many structurally diverse compounds from cells. Since Pgp pumps lipophilic substrates out of the membrane, the lipid environment plays an important role in regulating its activity. Using purified Pgp functionally reconstituted into proteoliposomes, we investigated the interaction of Pgp with sterols, and the effect of various defined phospholipids and sterols on basal ATP hydrolysis, Hoechst 33342‐stimulated ATP hydrolysis, and Hoechst 33342 transport. Fluorescence quenching showed that Pgp interacted with the fluorescent sterols, dehydroergosterol (DHE) and NBD‐cholesterol, with high affinity. Cholesterol hemisuccinate (CHS) greatly stabilized the basal ATPase activity of Pgp in detergent and proteoliposomes of some phospholipids, but DHE did not. CHS also had a pronounced effect on thermal unfolding of Pgp, as determined by CD spectroscopy. The stabilizing effects of CHS on drug‐stimulated ATP hydrolysis and drug transport were strongly dependent on bilayer phospholipid composition, and were distinct from those noted for basal ATP hydrolysis. This suggests that the basal ATPase and drug transport cycles of Pgp are distinct, and regulated differently by the lipid environment.

Funded by the Canadian Cancer Society (grant #700248).
The role of Cyclin Dependent Kinase 5 in insulin exocytosis
Authors: Trye Alice , Aguanno Ann
Published: June 2021

Cyclin dependent kinase 5 (CDK5) is an atypical member of the cyclin dependent kinase family, in that it is not a regulator of the cell cycle. CDK5 has been shown to play an active role in the terminal differentiation of neurons and dysregulation of CDK5 has been implicated in a number of neurodegenerative diseases including Alzheimer's Disease (AD). An emerging role for CDK5 has recently been identified in some non‐neuronal tissues. For example, in the pancreas CDK5 activity is involved in insulin exocytosis; whether it is a positive or negative regulator of the process, however, is still unclear. Our lab studies the role of CDK5 in mammalian development and has previously shown a negative effect on the developing neuronal phenotype when CDK5 activity is inhibited. We are now exploring the consequences of inhibiting CDK5 activity in the developing pancreas. Specifically, we employ the non‐insulin producing acinar cell line, AR42J, which can be transdifferentiated into an insulin‐positive cell in response to growth factors, and analyze insulin exocytosis by immunochemical methods. Here we report the results of chemical inhibition of CDK5 activity during this transdifferentiation process. Our preliminary findings shed light on the role of CDK5 in this process, allowing us to explore the function of this neuronal‐specific protein in both pancreatic development and pathologies.
Tumor suppressor p53 status as a determinant of estrogen receptor beta signaling in breast cancer
Authors: Das Gokul M , Mukhopadhyay Utpal Kumar , Bansal Sanjay , Medisetty Rajesh , Swetzig Wendy M , Wickramasekera Nadi
Published: June 2021

Tumor suppressor p53 and estrogen receptor have opposite roles in the onset and progression of breast cancer. The biological actions of estrogens are mediated by genetically distinct estrogen receptors (ER) alpha and beta. In addition to its expression in several ERalpha‐positive breast cancers and normal mammary cells, ERbeta is usually expressed in ERalpha‐negative cancers including triple‐negative breast cancer, but its role in oncogenesis remains unclear. Using glutathione‐S‐transferase (GST)‐pull down and coimunoprecipitation assays, we have demonstrated direct binding of ERbeta to p53 and have delineated the domains required for the interaction. Using the highly sensitive proximity ligation assay (PLA), we show ERbeta‐p53 interaction in situ. Surprisingly, we found that ERbeta has opposite functions depending on the mutational status of p53. A combination of proliferation and apoptosis assays, RNAi technology, quantitative chromatin immunoprecipitation (qChIP), and quantitative real‐time PCR showed that in the cellular context of wild type p53, ER beta is pro‐proliferative, whereas in the context of mutant p53, ER beta is anti‐proliferative. Our finding that ERbeta can elicit opposite functions dependent on the p53 context provides significant insight into the controversy whether ERbeta is pro‐or anti‐tumorigenic with important implications in therapeutic targeting of ERbeta.
Novel Signaling Pathways Triggered by a Human Alpha1a‐Adrenoceptor Genetic Variant
Authors: Oganesian Anush , Kleine‐Brueggeney Maren , Gradinaru Irina , Babaeva Ekaterina , Schwinn Debra A
Published: June 2021

Alpha1‐adrenergic receptors (α1ARs) play an important role in cardiovascular disease. We reported that naturally occurring genetic variant of human α1aAR, with G247R SNP (247R) identified in hypertensive patient, leads to constitutive hyperproliferation of fibroblasts via βarrestin/MMP‐dependent EGFR‐transactivation.

Here we examine if the 247R‐triggered hyperproliferation is generalizable in other cell types. Surprisingly, expression of 247R in cardiomyoblasts results in morphological and phenotypical changes to fibroblast‐like cells. RNA and protein expression levels of FSP1, DDR2, MMP2, vimentin, Troponin I, βMHC confirm fibroblast‐like phenotype and cells exhibit agonist‐induced hypertrophy and constitutive, serum‐independent 2–3‐fold increased proliferation. Interestingly, hypertrophy but not proliferation is inhibited by prazosin, an α1aAR‐inverse agonist, suggesting two independent pathways triggered by 247R: proliferation is caused by Gq‐independent, but βarrestin/Src‐dependent EGFR transactivation while hypertrophy is Gq‐dependent.

In summary, our data suggest that the naturally occurring human α1aAR genetic variant triggers βarrestin/MMP/ADAM dependent EGFR transactivation pathway leading to hyperproliferation or hypertrophy in different cardiovascular cell types and is a novel hypertension‐associated signaling mechanism.
Apigenin and Kaempferol inhibit LPS‐induced inflammatory responses by regulating intracellular translocation of RNA‐binding protein HuR in macrophages
Authors: Zhang Xiaoxuan , Wang Guangji , Liu Runping , Zhang Luyong , Hylemon Phillip B , Pandak William M , Zhou Huiping
Published: June 2021

Inflammation is a key player in numerous human diseases including metabolic diseases and cardiovascular diseases. Natural flavonoids are the major components of traditional Chinese medicine and have been used to treat various inflammatory diseases. However, the underlying cellular/molecular mechanisms remain unknown. Apigenin and kaempferol are major, naturally‐occurring plant flavonoids and abundantly present in common fruits and vegetables. The aim of this study was to examine the effect of apigenin and kaempferol on LPS‐induced inflammatory response and further identify the underlying cellular/molecular mechanisms in macrophages.

Methods

Cultured mouse J774A.1 macrophages and human THP‐1‐derived macrophages were used in this study. The mRNA and protein levels of IL‐1β, IL‐6, TNF‐α, MCP‐1, and COX‐2 were detected by real time RT‐PCR, Western blot and ELISA, respectively. The intracellular distribution of RNA binding protein HuR was determined by Western blot and immunofluorescence staining.

Results

Both Apigenin and Kaempferol significantly inhibited LPS‐induced expression of proinflammatory cytokines (IL‐1β, IL‐6, TNF‐α, MCP‐1) and COX‐2. LPS‐mediated cytoplasmic translocation of HuR was inhibited by apigenin and Kaempferol.

Conclusion

Apigenin and Kaempferol inhibit LPS‐induced inflammatory responses through regulating intracellular translocation of HuR in macrophages.
FERTILITY ACTIVITY OF AQUEOUS LEAF EXTRACT OF TAPINANTHUS BANGWENSIS ON OVULATION IN FEMALE‐SPRAGUE DAWLEY RATS
Authors: Onadeko Damilola , Nwoke Stella C , Magbagdeola Abiola O , Ebuehi Albert O. , Imaga Ngozi O , Ajayi Godwin O. , Gbotolorun Stella C
Published: June 2021

The fertility activity of Tapinanthus bangwensis in female rats were investigated in the present study. Twenty four cyclic female rats were administered orally at 1g/kg body weight between 140 and 200g. This was confirmed by checking their vaginal smear for four consecutive cycles.

The animals were divided into eight groups of three rats each, group 1 animals served as the control and received equivalent volume of distilled water. Group 2 animals received clomiphene citrate( a fertility enhancing drug), group 3 received duofem(a contraceptive drug), Group 4,5 and 6 received the aqueous leaf extract at 100mg/kg, 200mg/kg and 300mg/kg respectively. group 7 received Vit E and group 8 received Folic acid. They were all administered orally at 9:00am on proestrus to determine it's effect on ovulation. Results revealed a significant difference at P<0.05 between the groups that received the extract at 100mg/kg, 200mg/kg and 300mg/kg. when compared to the control groups. The extract group gave higher mean values of 13.1 as compared to the control of mean value 7.3. Our result gives added scientific support to the use of Tapinanthus bangwensis in the treatment of infertility related problems in women.
Expression of a metacaspase from the fungus Schizophyllum commune
Authors: Loew Ethan Bailey , Fox Kristin M
Published: June 2021

Metacaspases are a family of arginine/lysine specific peptidases known to induce programmed cell death in plants, protists, and fungi. Current research suggest that metacaspases contain a cysteine/histidine catalytic dyad in their active site, are calcium activated, and do not necessarily require proteolytic processing for catalytic activity. No specific substrate of metacaspases has been identified. To learn more about the structure and function of metacaspases, we are interested in expressing scp3, one of five putative metacaspases from the fungus Schizophyllum commune. Previous work in our lab has shown that the metacaspases are difficult to express in Escherichia coli. Because of similar difficulties with expression of active caspases, most are expressed as their individual p10 and p20 subunits, and then unfolded, combined, and refolded to produce active protein. To test this method with metacaspases, the p10 and p20 subunits of scp3 were amplified individually and cloned into pGEM® T‐Easy (Promega). The p10 and p20 subunits were then subcloned into the expression vector pQE‐80L (Qiagen) and successfully expressed in E. coli M15 cells. Work continues to fold the subunits and quantify their enzymatic activity.
Cypher/ZASP, a PKA Scaffolding Protein to Regulate the Phosphorylation of L‐type Calcium Channel
Authors: Cheng Hongqiang , Lin Changsong , Yin Xiang , Guo Xiaogang , Chen Ju , Ke Yuehai
Published: June 2021

PKA signaling is important in phosphorylation regulation of some key calcium handling proteins and its activity is compartmentalized through A‐kinase anchoring proteins. It has been demonstrated that Cypher/ZASP, a PDZ‐LIM domain protein, plays pivotal structural roles in the integrity of sarcomeres and several of its mutations are identified to associate with myopathies including dilated cardiomyopathy. Here we showed that Cypher/ZASP, interacting specifically with type II regulatory subunit RII of PKA through its ZM motif, was a typical A‐kinase anchoring protein. And Cypher/ZASP itself is phosphorylated at Ser265 and Ser296 by PKA. Furthermore, the PDZ domain of Cypher/ZASP interacts with L‐type calcium channel through its C‐terminal PDZ binding motifs. Cypher/ZASP expression facilitated PKA to phosphorylate L‐type calcium channel in vitro. Furthermore the phosphorylation of L‐type calcium channel at Ser1928 induced by isoproterenol was impaired at neonatal Cypher/ZASP null cardiomyocytes. Moreover, Cypher/ZASP interacted with the Ser/Thr phosphatase calcineurin which has been reported to be a phosphatase for L‐type calcium channel. Our data strongly suggests that Cypher/ZASP plays not only structural roles at sarcomeric integrity, also is an important signaling spot for activity regulation for the key calcium handling protein.
Beta synemin binds PKA type I upon beta‐adrenergic stimulation in HL‐1 cells
Authors: Prudner Bethany Cheree , Damron Derek S , Russell Mary A
Published: June 2021

While our lab previously established that the intermediate filament protein beta synemin is an A‐kinase anchoring protein (AKAP) for PKA type II, which plays a role in maintaining the specificity of the protein kinase A (PKA) signaling pathway; we found that beta synemin also binds PKA type I. Classically PKA type I was thought to be soluble due to low affinity for AKAPs. However, recent work has shown that several AKAPs are capable of binding PKA type I. Here we report yeast two hybrid and FACS‐FRET analyses and co‐immunoprecipitation data that demonstrate the binding of PKA type I to beta synemin. Additionally, this binding is dependent upon stimulation of the beta‐adrenergic (beta‐AR) pathway via isoproterenol. We hypothesize that beta synemin functions as a dual AKAP in which PKA type II is bound during resting conditions then released following beta‐AR stimulation. According to our model, the regulatory subunit of PKA type II is released from beta synemin, leaving the PKA binding site on beta synemin unoccupied and available for PKA type I binding. Though the physiological relevance of the intersection of the two types of PKA at beta synemin remains unclear, due to differences in biochemical characteristics including downstream substrates, the release of PKA type II and binding of type I to beta synemin would allow for a highly coordinated spatial and temporal response to beta‐AR stimulation.

NIH R01 HL65701 (DD)
Regulation of epithelial‐mesenchymal transition and apoptosis by dual specificity phosphatase‐2
Authors: Lin Shih‐Chieh , Hsiao Kuei‐Yang , Tsai Shaw‐Jenq
Published: June 2021

Our previous findings provide evidence to demonstrate that dual specificity phosphatase‐2 (DUSP2) is markedly reduced or even lost in most solid cancers. To further unravel the effect of DUSP2 downregulation in cancer progression, we overexpressed DUSP2 in cancer cells and performed microarray assay to document gene expression profile. Analysis of microarray data derived from DUSP2 overexpressed cancer cells revealed that many genes involved in angiogenesis, epithelial‐mesenchymal transition (EMT), cell migration, and apoptosis are regulated by DUSP2. Since EMT is a critical process for an increase in metastatic capacity and drug resistance, we tested whether loss‐of‐function of DUSP2 would induce EMT. Knock down of DUSP2 induced the expression of mesenchymal markers (N‐cadherin) and reduced the level of epithelial marker (E‐cadherin). Consistent with this notion, treatment with conditioned media collected from DUSP2 knockdown cells enhanced cancer cells migration while treatment with conditioned medium collected from DUSP2 overexpressed cells inhibited cell migration. Furthermore, we established a stable clone of HCT116 cells carrying doxycycline‐inducible DUSP2 minigene. Treatment of cells with doxycycline to induce DUSP2 expression resulted in upregulation of pro‐apoptotic genes such as Bcl2‐modifying factor and endonuclease/exonuclease/phosphatase family domain‐containing protein 1. As expect, induction of DUSP2 expression increased cell death. These data suggest that restoration of DUSP2 expression in cancer cells represents a potential therapeutic approach. In‐depth investigation of mechanisms responsible for DUSP2‐induced cell death will provide valuable information for designing novel treatment regimen for cancer therapy.
LT1002 Metalloantibody Uses Ca2+ Cofactor
Authors: Ward Aaron D , Yacoub Dina , Erasmus Frank , Huxford Tom
Published: June 2021

The biologically‐active lipid, Sphingosine‐1‐phosphate (S1P), is a signaling molecule present in cell membranes that promotes heart disease, and tumor growth. Murine, Immunoglobulin G (IgG) LT1002 and human IgG LT1009 specifically bind S1P and inhibit its effects. X‐ray crystallography revealed that two Ca2+ bridge LT1009 and its S1P antigen. This was the first direct structural evidence of metal ion bridging in an antibody: antigen complex. While the role of Ca2+ in the complex is clear, it is unknown to what extent Ca2+ binds in the absence of antigen and whether other biologically significant metals interact with the antibody. To this end, we have conducted quantitative analysis comparing the binding enthalpy, dissociation constant, and stoichiometry of LT1002 and nonmetal‐binding IgG binding to biologically relevant, divalent cations. Isothermal titration calorimetry (ITC) revealed 300uM [LT1002] bound 20mM [Ca2+] with a KD= 238uM with a LT1002:Ca2+ stoichiometry of 1:3. Ba2+, another metal ion that was observed to bind free LT1002 in previous studies, showed no binding to LT1002 by ITC and Ca2+ failed to bind to control murine IgG1. ITC data, in conjunction with the crystal structure of an antigen‐free antibody: Ca2+ complex, indicate that Ca2+ specifically binds to the epitope region of LT1002.

Supported by NIH MBRS‐IMSD Grant 5R25GM058906
Regulation of photoreceptor phosphodiesterase (PDE6) by the rod photoreceptor‐specific glutamic acid‐rich protein 2 (GARP2)
Authors: Yao Wei , Cote Rick H
Published: June 2021

As the central effector of the visual signaling pathway, photoreceptor phosphodiesterase (PDE6) is precisely regulated to control the sensitivity, speed and amplitude of the light response. This study seeks to define the regulatory role of the glutamic acid‐rich protein 2 (GARP2) that binds to PDE6 in rod photoreceptors. GARP2 is a splice variant of the rod photoreceptor cGMP‐gated channel β‐subunit. Using interaction assays, we identified the inhibitory γ‐subunit (Pγ) of the PDE6 holoenzyme (αβγγ) as the site of interaction with GARP2. Using Pγ truncation mutants, we found that GARP2 binds primarily to the N‐terminal region (a.a 1–30) of Pγ. The stoichiometry of GARP2 binding to PDE6 holoenzyme was determined to be 1.0 ± 0.2 mol GARP2 per mol PDE6 holoenzyme. In addition to suppressing PDE6 basal catalytic activity, GARP2 also accelerates cGMP dissociation from noncatalytic cGMP binding sites in the regulatory domain of PDE6. Together, these results indicate that GARP2 binding to the N‐terminal region of the inhibitory Pγ subunit can lower PDE6 catalytic activity in the dark‐adapted state, effectively reducing photoreceptor dark noise. In addition, GARP2 modulation of cGMP binding affinity to the PDE6 regulatory domains is consistent with a negative feedback role for GARP2 in PDE6 regulation following light activation of the phototransduction cascade. Supported by NIH grant EY‐05798 (to RHC).
Compound Seropositivity of neutralizing antibodies against Adipogenic Adenoviruses (ADV36+ and ADV 37+): no additive effect on overweight‐obesity and on Insulin Resistance (IRHOMA). The microbiome across the obesity and into the liver
Authors: Trovato Guglielmo M. , Catalano Daniela , Martines G. Fabio , Trovato Francesca M. , Garozzo Adriana , Pirri Clara , Tonzuso Antonia
Published: June 2021

A greater hazard of obesity and Insulin Resistance is associated with ADV36 adenovirus seropositivity (ADV36+). Human ADV37 has steatogenic liver effects in humans, but adipogenic effect only in animals: ADV37+patients do not show greater prevalence of obesity vs. ADV37/ADV36 seronegative subjects.

Aim

to investigate if compound ADV36/ADV37 seropositivity exerts a different effect on overweight‐obesity and IR vs. seronegative subjects and vs. ADV36‐ADV37 single seropositivity. 430 patients were studied, m 157, f 273, age 47.99 ± 15.50 years, BMI 28.14 ± 6.29. Dietary profile was assessed by Adherence to Mediterranean Diet Score (AMDS). Greater BMI (29.74 ± 7.21 vs. 26.47 ± 5.32;p: 0.004) and no difference for HOMA (2.42 ± 1.38 VS 2.28 ± 1.38; p: 0.591) are observed in ADV36+ subjects vs. ADV37+. Compound ADV36+/ADV37+ vs. ADV37+ lone seropositivity has greater BMI: 29.42 ± 6.13 vs. 26.47 ± 5.32; p: 0.001 and greater HOMA: 2.94 ± 2.31 vs. 2.28 ± 1.38; p: 0.029, exactly as the compound seropositivity vs. totally seronegative subjects: BMI: 29.42 ± 6.13 vs. 26.41 ± 5.99; p: 0.0001; HOMA: 2.94 ± 2.31 vs. 2.07 ± 1.39; p: 0.0001; no difference for compound seropositivity ADV36+/ADV37+ vs. ADV36+ lone group. No difference of AMDS was observed between any group.

Conclusion

ADV36+ and ADV37+ compound seropositivity, independently by difference of nutritional profile, has no additive effect for obesity and insulin resistance.
The Healthy Eating Index and the Intestinal Microbiome in HIV Infection
Authors: Volpe Gretchen , Sheehan Heidi , Tang Alice , Duffalo Chad , Dinh Duy , Landy David , Ward Honorine , Kane Anne , Wanke Christine
Published: June 2021

The objective of this analysis was to characterize the intestinal microbiota of HIV‐infected persons according to the Healthy Eating Index (HEI). We calculated HEI‐2005 scores from 0 (worst quality diet) to 100 (best quality diet) from food records of 50 HIV‐infected participants, via the Nutrition Data System for Research software (University of Minnesota). We used pyrosequencing of 16S rDNA amplicons from stool and the open source platform Qiime (http://qiime.sourceforge.net) to obtain percent relative abundance of phyla. First, we compared groups based on HEI below or at/above the sample median of 58. Those with HEI <58 were more likely to be female, to have lower CD4+ counts, to have higher relative abundance of Bacteroidetes, and to have been heavier smokers compared to those with HEI ≥58. Next, we used HEI as a continuous variable in multivariate analysis to determine associations of relative abundance of the two major phyla, Bacteroidetes and Firmicutes. Percent relative abundance of Bacteroidetes was negatively associated with HEI and positively associated with energy intake for body weight (kilocalories/kg); the inverse of these associations was found for % relative abundance Firmicutes. In conclusion, in our sample of HIV‐infected persons, higher HEI was associated with higher % relative abundance of Firmicutes and lower % relative abundance of Bacteroidetes. Supported by P30DA13868, P30AI42853, 1UL1RR025752–03, 5R01HL065947–10, 5T32AI007438–19, and 2T32AI07389–20.
Prebiotic fiber diet does not improve offspring ‘leaky gut’ from maternal low protein diet
Authors: Hallam Megan Carroll , Reimer Raylene A
Published: June 2021

Metabolic endotoxemia can initiate obesity and insulin resistance. Prebiotic fiber can reduce metabolic endotoxemia and intestinal permeability as measured by plasma lipopolysaccharide (LPS). Offspring of rats fed a low protein diet during pregnancy are at high risk of obesity which may be related to increased intestinal permeability. Our objective was to determine if prebiotic fiber minimized this effect. Virgin Wistar dams received AIN‐93G (normal protein, NP) or a low protein (LP) diet (7% wt/wt) during pregnancy. Female offspring were weaned onto control or a high prebiotic fiber diet (21% wt/wt 1:1 oligofructose and inulin) (HF). Body composition was analyzed along with plasma LPS concentrations at 24 wk of age. Gene expression in the colon was analyzed for tight junction protein‐1 (TJP‐1). There was no difference in overall body weight but lean mass was greater in offspring consuming HF (P=0.013). HF was associated with higher plasma LPS in LP‐exposed versus NP‐exposed offspring (P=0.009). TJP1 mRNA levels were higher in NP‐exposed offspring whether consuming C or HF (P=0.05). A maternal diet low in protein is associated with a ‘leaky gut’ in female offspring which has the potential for negative metabolic effects. Contrary to adult obesity, HF was not able to correct metabolic endotoxemia in female offspring of LP dams.
Selecting Items of a Food Behavior Checklist for the Development of Nutrition Quotient (NQ) for Children in Korea
Authors: Kang Myung‐Hee , Lee Jung‐Sug , Kim Hye‐Young P. , Kwon Sehyug , Choi Young‐Sun , Chung Hae Rang , Kwak Tong‐Kyung , Lee Hye‐Jin , Cho Yang‐Hee
Published: June 2021

The objective of this study is to select a simple and easy measurable food behavior checklist for the development of Nutrition Quotient (NQ) for children, which reflects children's diet quality, as well as to evaluate the validity of the items in the food behavior checklist (FBC). The first 36 items in the checklist were selected by a literature review and the Korean National Health and Nutrition Examination Survey data. 341 children of 5th and 6th grades at an elementary school participated in a one‐day dietary record survey, and later responded to 36 food behavior questions of the checklist. Pearson's correlation coefficients between the responses to the FBC items along with the mean nutrient intakes of the children were calculated. From the result, a second checklist with 22 items was selected. A survey was conducted by using the second checklist. 1,393 children in the 5th and 6th grades in six large cities in Korea, participated in the survey. Further, an exploratory factor analysis was performed. After the analysis, 19 items were finalized as the FBC items for the NQ. The final 19 FBC items were composed of 5 factors: ‘Balance’, ‘Diversity’, ‘Moderation’, ‘Regularity’, and ‘Practice’. This study is a significant first trial to establish a comprehensive system for evaluating children's food habit and diet quality in Korea. (This research was supported by a grant from Amway Korea Ltd. in 2011.)
White Potato consumption is positively associated with potassium intake
Authors: Storey Maureen L , Anderson Partricia
Published: June 2021

Intake of potassium (K), dietary fiber (DF), calcium (Ca), and vitamin D (vit D) are low enough to be of public health concern. White potatoes (WP) contribute DF and are an excellent source of potassium in the U.S. food supply, providing more potassium than most other commonly consumed vegetables. We examined nutrient intakes of children and adults using data from the National Health and Nutrition Examination Survey (NHANES) 2009–2010 and nutrients contributed by WP. Appropriate survey weights were used to calculate average daily intakes of total energy, protein, total fat, saturated fat, monounsaturated fatty acids, polyunsaturated fatty acids, cholesterol, carbohydrates, dietary fiber, vitamin A, vitamin C, thiamin, riboflavin, niacin, vitamin B6, folates, vitamin B12, vitamin K, calcium, phosphorus, magnesium, iron, zinc, copper, sodium, and potassium by males and females 2– 4, 5–8, 9–13, 14–18, 19–30, 31–50, 51–70, and 71+y. Intakes of K, DF, Ca and vit D were all below the RDA or Adequate Intake (AI) levels recommended by the Institute of Medicine. Multivariate regression analyses showed that, controlling for energy intake and demographic variables, WP consumption was positively associated with K intake. Controlling for the other variables in the model, each additional kilocalorie of WP consumed was associated with a 1.6 mg increase in K intake among adults 19+y and a 1.7 mg increase among children and teens 2–18y. These results suggest that children, adolescents and adults do not meet dietary recommendations for key nutrients and consumption of WP increases intake of K.

This study was supported by the non‐profit organization Alliance for Potato Research and Education.
Assessment of variation in methods to determine reported iron content of nutrition labels of commercial dark chocolate bars
Authors: Giusti Riana , Kazaks Alexandra
Published: June 2021

Objective

To assess variation in methods used by industry for determining iron content in dark chocolate bars to provide information for nutrition labeling.

Methods

Ten brands of dark chocolate bars (70–85% cocoa solids) were purchased from vendors on the US Pacific Coast (San Francisco, California to Victoria, BC). Reported values for iron content (%DV), controlled for serving size, were assessed. Producers were contacted to confirm specific analytical methods or other data utilized for product nutrition labels.

Results

For the chocolate bars assessed, reported iron content ranged from 4–39 %DV (Mode = 10, SD = 11.7). Label values for iron content were obtained by the following methods: 2 producers used USDA Nutrient Laboratories databases, 4 used analytical testing, 1 was exempt from labeling, 3 did not provide test methods.

Conclusion

Iron, a class II nutrient, is required by FDA to be listed as a percentage of the 18mg daily value (%DV). There is a high level of variability in how dark chocolate producers obtain data for product nutrition labels. This highlights a need for standardized methods to determine iron content for labeling dark chocolate bars. As a result of this analysis, we plan to perform and compare several accepted methods for determining the iron content in dark chocolate bars. Data will be compared to nutrition labels to assess any true variation of iron content in dark chocolate bars.

Grant Funding Source: Center for Student Research Bastyr University
Environmental barriers to children's summer outdoor play
Authors: Worobey John , Lelah Lydia , Gaugler Randy
Published: June 2021

Although the health benefits of physical activity are well documented, it appears that even children may exhibit levels of physical activity that are insufficient to prevent or reduce excess weight gain. In light of the current child obesity epidemic, our objective was to survey children to identify the main reasons they might give for not engaging in outdoor play in the summer, ostensibly a time when school is out and weather is most favorable. For this study, 300 children (age 6–14) were surveyed at four county/4‐H fairs and asked to indicate how important various factors (e.g., heat, traffic, mosquitoes) were in preventing them from playing outside. The reasons ranked highest were bad weather, bullies/gangs, fear of strangers, no street lights, and it being too hot. These results are somewhat consistent with studies that indicate neighborhood safety as a concern (Carver et al., 2008), but add weather conditions as a factor to be considered when seeking to promote outdoor activity. Allowing access to unused school gyms might be a useful strategy for facilitating large motor activity in summer months.

This project was supported by a Robert Wood Johnson Foundation Active Living Grant to JW.
Pooled Analysis of Impact of Lycium barbarum Fruit (Goji) Intake on Central Adiposity, Sleep Quality and General Well‐being: Randomized, Double‐blind, Placebo‐controlled Human Clinical Studies
Authors: Amagase Harunobu , Hsu Chiu‐Hsieh , Nance Dwight M
Published: June 2021

Obesity has negative impacts on sleep and other conditions. Lycium barbarum intake has been shown to improve general well‐being, sleep quality (SQ) and postprandial energy expenditure, and to reduce central adiposity in multiple randomized, double‐blind, placebo‐controlled trials. To assess possible interactions among these parameters, we analyzed pooled data from these trials (n=102, mean age=33 y) using 2‐sample t‐test and scatter plot with linear regression. SQ and general well‐being were evaluated by Pittsburgh Sleep Quality Index (PSQI) and subjective questionnaire, respectively, and waist circumference (WC) was assessed pre‐ and post‐dietary intervention with Lycium barbarum for 2–4 weeks. Results indicated that PSQI scores were correlated with SQ, increased general complaints about well‐being and increased WC. Lycium barbarum intake significantly improved SQ (p=0.021) and general well‐being (p=0.001), and reduced WC (p=0.002) compared to placebo. Correlations among these parameters were: sleep vs general well‐being, 0.31 (p=0.002); sleep vs WC, −0.04 (p=0.747); and general well‐being vs WC, − 0.25 (p=0.039). These correlations suggest that diet‐induced improvement in general well‐being is associated with improved SQ and is negatively correlated with changes in WC. These suggest that Lycium barbarum intake may stimulate metabolism and could contribute to an obesity management program.
Effects of the sea tangle‐added patty on postprandial blood glucose and lipid profiles in borderline‐hyperlipidemic adults
Authors: Lim Hyeon‐Sook , Kim Hwa‐Hyun
Published: June 2021

Sea tangle (Laminaria japonica) has been well known to have a hypolipidemic effect in human as well as in animals. We found that the sea tangle‐added patty decreased postprandial plasma glucose and lipids levels in adults with normal blood lipid profiles in the previous study. In this study, we intended to evaluate the effects of the sea tangle‐added patty on postprandial blood glucose and lipid profiles in borderline‐hyperlipidemic (cholesterol≥200 mg/dL and/or triglyceride ≥150 mg/dL) adults. Fourteen subjects voluntarily participated and each subject provided written consent. Experimental patty (E) was made by adding 2.25 g of sea tangle powder instead of eliminating 1.125 g each of pork and chicken. Two hundred gram of Control patty (C) was provided with water to each subject who was in empty stomach for more than 12 hours and 4 mL of blood was collected intermittently at 0, 30, 60, 120, 180, and 240 minute after consuming the patty. After one week, the same amount of E patty was supplied and blood sampling was done as the first time. Before taking each patty, all subjects consumed the same meal for three days. E patty made blood glucose level increased less than C patty at 30 min, but there was no difference in serum C–peptide concentration. E patty also made serum total cholesterol concentration reduced more at 120 min and LDL‐C concentration lessened more at 120 min than C patty. However, HDL‐C level was maintained higher at 30 min, but kept lower at 180 min when consuming E patty than C patty. There was no significant difference in triglyceride concentration. The results indicate that the sea tangle‐added patty may improve postprandial plasma glucose and lipids profiles in borderline‐hyperlipidemic adults. (Supported by a grant from Chonnam National University, Korea)
Kaempferia galanga L. suppresses melanin synthesis through inhibition of the CREB/MITF in B16 cells
Authors: Jeon Byoung‐Kook , Mun Yeun‐Ja , Woo Won‐Hong , Lee Young‐Eun
Published: June 2021

Melanin is the main component determining the color of skin and also plays an important role in protecting skin from UV light. Kaempferia galanga Linn. (Zingiberaceae) presents many chemical constituents of the volatile oil extracted from the rhizome. In this study, we examined the effects of the methanolic extract of Kaempferia galanga L. (MKG) on melanogenesis and signaling pathway in B16F10 cells. MKG significantly inhibited ¥á‐melanocyte stimulating hormone (¥á‐MSH)‐induced melanin synthesis and tyrosinase activity, and also decreased ¥á‐MSH‐induced expression of microphthalmia‐associated transcription factor (MITF) and tyrosinase‐related proteins (TRPs). Moreover, MKG reduced the level of phosphorylated cAMP‐response element binding protein (CREB), activity‐regulated transcription factor that functions in melanogenesis. These findings suggest that inhibition of phospho‐CREB and MITF expression may lead to the suppression of melanogenesis in MKG‐treated B16 cells.

Grant Funding Source: None
Supercritical fluid extraction (SFE) of anti‐inflammatory polyacetylenes from celeriac (Apium graveolens L.)
Authors: Metzger Brandon T. , Waksmonski James C. , Thompson Amber L. , Barnes David M.
Published: June 2021

Celeriac is an agricultural root crop and source of secondary plant defense compounds called polyacetylenes (PAC's). PAC's inhibit lipid transport enzymes, up regulate phase II liver detoxification enzymes, are cytotoxic to cancer cells, and potent antiinflammatory modulators. SFE of PAC's has been reported, but extraction parameters have not been optimized. A full factorial experiment was designed to test extraction parameters of temperature, pressure, and ethanol co‐solvent addition at a solvent‐to‐ feed ratio of 100. The dried feed stock contained 4.5x's the amount of PAC's compared to purple carrots. Optimal extraction conditions were low temperature (40°C), high pressure (500 bar), and high ethanol co‐solvent (20% w/w). Pressure had a larger effect (F <0.0001) on extraction of falcarinol and falcarindiol than temperature (F = 0.13 and 0.02 respectively), which was also dependent on co‐solvent amount (F = 0.0009). Under optimal extraction conditions, 1.7% of the starting dry weight, or 47% of the total lipids were extracted and up to 43% of falcarindiol was obtained compared to exhaustive ethyl acetate extraction. Ethyl acetate extracts of celeriac inhibited (IC50) pro‐inflammatory cyclooxygenases (Cox‐1 and Cox‐2) at concentrations of 0.26 and 0.06 mg/mL respectively. SFE extraction of dried celeriac root offers potential as an industrial scale enrichment process of antiinflammatory PAC's.
Characterization and quantitation of bioactive compounds in date fruit seed using UPLC‐DAD‐ESI‐MS
Authors: Habib Hosam , Platat Carine , Ibrahim Wissam
Published: June 2021

Date fruit seeds have been shown to possess antioxidant activity and to constitute a significant source of antioxidants. However, the exact natures of these bioactive compounds and their quantities have not been thoroughly investigated. These compounds are being reported here for the first time in date seed. Two analyses were performed: 1) an analysis of simple phenolic compounds (phenolic acids, hydroxycinnamic acids, flavonols, flavones, flavan‐3‐ols monomers and dimers and trimers), 2) an analysis of all flavan‐3‐ ols (monomers, oligomers, and polymers proanthocyanidins) after depolymerization. The amount of total polyphenols before depolymerization was found to be 2.193 mg/g date seed. Protocatechuic acid was 0.079 mg/g, flavan‐3‐ols (dimer B1, trimer, catechin, trimer, dimer B2, and epicatechin) were 1.65 mg/g, flavones or flavonols (apigenin di‐C‐hexoside, quercetin glucoside derivative, flavonol or flavone derivative, Kampferol derivative, luteolin derivative) were 0.07 mg/g, while hydroxycinnamic acids (caffeoyl shikimic acid and caffeoyl shikimic acid) were 0.39 mg/g. The analysis of constituent units of proanthocyanidins after depolymerization revealed 50.18 mg/g flavan‐3‐ols with 93.28% epicatechin and 6.72% catechin. These results indicate that date seeds represent a very rich source of bioactive compounds that can be used as functional food additives and nutraceuticals.