bluuebunny/crossref_metadata_2025 huggingface dataset:
Q‐16, a Partial PPARg Agonist, Decreases Adiposity and Induces the Thermogenesis‐Related Genes in Obese Mice
Authors: Coelho Michella , Lima Caroline , Royer Carine , Pitta Ivan , Amato Angelica , Neves Francisco
Published: June 2021

Adipose tissue is classified as white (WAT) and brown (BAT). A third type of adipocyte, so‐called beige, was described as an inducible cell type in WAT, in a process named browning of WAT. Beige adipocytes are fully capable of expressing Ucp‐1 and executing thermogenesis. Full PPARγ activation by TZDs drives the appearance of Ucp‐1‐positive cells in WAT depots, but these drugs are associated with a number of adverse effects, including weight gain.We have described a partial PPARg agonist, GQ‐16, that does not induce weight gain, but have not been shown to promote browning of WAT. Objective: To evaluate the effects of GQ‐16 on browning of WAT in mice.

Methods

Mice were fed a control (C) or high‐fat (HFD) diet since weaning to promote obesity. At the age of 16 wk, they received GQ‐16 (40 mg/kg/d), rosiglitazone (RSG, 4 mg/kg/d) or vehicle. Weight gain, energy intake, and blood glucose levels were measured. Visceral WAT (vWAT) and subcutaneous WAT (sWAT) pads were excised for determination of adiposity, gene expression and histological analysis.

Results (p<0.05, 4 mice/group)

GQ‐16 treatment reduced blood glucose, weight gain and vWAT fat mass (C: 34±6, HFD: 50±4, RSG: 67±5, GQ‐16: 30±5 mg/g) to values similar than those of mice fed a C diet. Mice treated with GQ‐16 increased energy intake and decreased adipocyte size in both vWAT and sWAT. Surprisingly, in vWAT, GQ‐16 increased the expression of thermogenesis‐related genes (Ucp‐1, Cidea and Prmd16) and there was a trend towards increased expression of beige adipocyte‐selective genes.

Conclusion

Our findings possibly reflect increased thermogenesis induced by GQ‐16, and reinforce the potential partial PPARγ agonists as a new strategy to treat obese patients with T2DM.
Calcium Glycerophosphate (CGP) Preserves Transepithelial Integrity
Authors: DATTA PALIKA , Weis Margaret
Published: June 2021

We hypothesized that CGP would mitigate the effect of hypoxia, cytokines (CK) and/or α‐gliadin peptide 31‐55 (α‐GP, the gluten peptide fragment of celiac disease) to increase transepithelial permeability. All experiments used Caco‐2 monolayers with transepithelial electrical resistance (TEER) of 600‐700 Ω/cm2. All agents were added to the luminal side of the cell monolayer. In normoxic cells, TEER increased at 0.69 ± 0.11% / hour and mannitol flux was 28 ± 4 dpm / hour. In hypoxic cells, TEER decreased 19.4 ± 1.31 % / hour and mannitol flux increased to 162 ± 27 dpm / hour. 10 µM CGP, reduced hypoxia induced TEER loss to 10.6±1.86 % / hour and mannitol flux to 35±dpm / hour (p<0.001 compared to normoxia or hypoxia alone). CK stimulated TEER loss was 2.1 ± 0.31% / hour and mannitol flux was 54 ± 1 dpm / hour. 10 µM CGP reduced TEER loss to 1.4 ± 0.28% / hour, and mannitol flux to 13 ±1 dpm / hour (p<0.001 compared to control or to hypoxia alone). In separate experiments, α‐GP + CK increased mannitol flux from 21±1 to 39 ± 3 dpm / hour; α‐GP + CK + CGP (1 mM) group, mannitol flux was 32 ±1 dpm / hour (p<0.05 vs α‐GP + CK). In these same experiments, TEER loss was 2.72 ± 0.497% / hour in the peptide + CK group vs. 1.20 ± 0.327% /hour in the α‐GP + CK + CGP group (p<0.05 vs α‐GP + CK). E‐cadherin protein was 75.8±8.6% of control in the α‐GP + CK group, while in the α‐GP + CK + CGP treated cells, E‐cadherin was 101 ± 2.9% of control (p<0.05 vs α‐GP + CK. In summary, CGP showed a significant time and concentration dependent effect to attenuate increased gut permeability caused by hypoxia, CK, or α‐GP an effect that was observable even at 1µM. These factors may make it worthwhile to pursue calcium glycerophosphate as an adjunct to other therapies to prevent loss of gut epithelial integrity in hypoxia or celiac disease.
Identifying New Tumor Suppressor in Liver Cancer Using Unbiased High‐Throughput Approaches
Authors: Zhang Xi , Zhang Yuxia , Wang Li
Published: June 2021

Purpose

Tumor suppressor genes are often epigenetically silenced. This study identified a new tumor suppressor in hepatocellular cancer (HCC) using an unbiased high throughput approach.

Methods/Results

Studies from our laboratory has established nuclear receptor small heterodimer partner SHP as a tumor suppressor. We also established an inhibitory feedback regulatory loop between SHP and DNA methyltransferases (Dnmts). Methylated DNA‐IP‐on‐ChIP (MeDIP) revealed alterations in DNA methylation and RNA‐sequencing (RNA‐seq) revealed changes in gene expression. By integrating both studies using Bioinformatics tools and appropriate validation, we identified the mitochondrial protein pyruvate dehydrogenase kinase 4 (PDK4) as a putative tumor suppressor. PDK4 promoter was hypermethyalted and its expression downregualted in SHP‐/‐ mice. PDK4 was also silenced in human HCC. Treatment of HCC cell lines with the demethylating agent 5'‐aza ‐2'‐deoxycytidine (Aza) or histone deacetylase inhibitor trichostatin (TSA) induced PDK4 expression. The number of foci was decreased and the tumor sizes were reduced in HCC cells that stably overexpressing PDK4, whereas the opposite was found with PDK4 knockdown. PDK4 overexpression sensitized HCC cells to cell death induced by glucose deprivation by reducing the intracellular ATP contents.

Conclusions

Epigenetic silencing of PDK4 is associated with the development of HCC.

[Grant support] NIH DK080440, VA Merit Award 1I01BX002634.
Chronic Exposure To Microbial Stimuli Affects The Development Of The Intestinal Epithelium In Human Colonic Enteroids
Authors: Rodrigues Ely , Slobbe Lynn , Schultz Michael , Butt Grant
Published: June 2021

Adult stem cells and transient amplifying cells that support epithelial regeneration are key sites where bacteria could modulate epithelial regeneration and homeostasis. The aim of this research was to determine if modulation of adult intestinal stem cells by the bacterial components affected the development of the intestinal epithelium. Enteroids were grown from crypts isolated from the transverse colon of healthy individuals and transferred to Matrigel and growth media for 15 days in the presence and absence of lipopolysaccharide (LPS, 20 ng ml‐1). Organoid structure was assessed by light microscopy and gene expression by microarray, qPCR, Western Blotting and immunohistochemistry (IHC). LPS increased the total number of goblet cells to 20%+3% (n=5), compared with none in the untreated controls. Microarray analysis showed an increase in the goblet cell associated proteins MUC2, Clca1, Zg16, Mpgc60, microtubule‐associated protein and resistin‐like beta protein and also an increase in genes associated with cell cycle, growth and differentiation in the LPS treated enteroids. Western blot confirmed an increase in the goblet cell marker MUC2. IHC confirmed an increase in the number of MUC2 and TFF3 positive cells confirming a lineage change to give more goblet cells. Collectively, these data indicate that LPS modulates stem cell activity and effects intestinal epithelial development.
The Expression and Role of Melanopsin in Paralichthys dentatus
Authors: Akinnola Ifeolu , Kingston Alexandra , Robinson Phyllis , Cronin Thomas
Published: June 2021

Less than two decades ago a research on the light sensitive skin cells of Xenopus laevis led to the discovery of the visual pigment known as melanopsin. Now, it's known that melanopsin is expressed in the vertebrate retina, as well as in the brain and skin of certain species. The presence of melanopsin has been studied in several fish species. In fact, zebrafish have five melanopsin genes. Paralichthys dentatus (Summer flounder) is commonly found in the ocean water off the east coast of North America. A flat fish where the adult form is bilaterally asymmetrical, this flounder uses dynamic patterning for camouflage and signaling. It has been shown that there are visual opsin genes present within the dermal tissue of the flounder, suggesting the tissue is also light sensitive.

Previous work has shown that there is an expression of rhodopsin and cone opsin in the dermal tissue of the flounder. The activation of visual opsins within retinal cells causes hyperpolarization. In contrast, the activation melanopsin is known to cause cells to depolarize. We hypothesize that melanopsin is used in a “push‐pull” system, in addition to visual opsins, to regulate the flounder's dynamic patterning. In addition melanopsin also activates at a slower rate than visual opsins, making it a good candidate for our hypothesized “push‐pull” system. Our aim is to identify the melanopsin gene using a flounder transcriptome and confirm the expression of melanopsin within dermal and retinal tissue. We also hope to confirm the presence of the components needed for proper phototransduction within the cells melanopsin is expressed in.

Funding Source: NIH grant number 5T34GM008663‐18 to the UMBC MARC‐U STAR Program; The Office of Naval Research Basic Research Challenge grant number N00014‐10‐0989 to T.W.C.
Comparative Analysis of the Amino Terminus Region of Twist1 and Twist2 Proteins
Authors: Rodriguez Yacidzohara , González‐Mendez Ricardo , Cadilla Carmen
Published: June 2021

Twist proteins belong to the basic helix‐loop‐helix (bHLH) family of multifunctional transcriptional factors. ADD1, RunX2 and MEF2 have been shown to interact with TWIST2 in studies that suggest there are other elements or domains, besides the common bHLH, involved in these protein‐protein interactions. There have been attempts to characterize the roles of their N‐terminus, bHLH, and C‐terminus in protein‐protein interactions, more focus is needed at the N‐terminus. We analyzed the conservation of the N‐terminal region of different vertebrate Twist proteins by sequence comparison and motif analysis. We found two putative de novo motifs (SSSPVSP and SEEE), specifically in mammalian sequences. Mammalian Twist1 sequences contain two, already described, glycine rich motifs not present in Twist2 mammalian sequences or other vertebrate species. Disordered domain analysis and secondary structure prediction demonstrated that both the N‐ and C‐terminus are found in a disordered/flexible region. We investigated the origin and evolution of Twist proteins by phylogenetic analysis. First, a gene duplication event occurred to produce the two Twist paralogs, Twist1 being evolved first. Second, Twist2 vertebrate proteins seem to have a higher rate of sequence divergence when compared to Twist1 mammals since they do not contain the two additional glycine‐rich regions. Our study determined the relationship of Twist1 and Twist2 paralogs among mammals in vertebrates. Sequence conservation and diversity in the N‐terminus of mammalian Twist proteins provides a rationale for studies of the role of the N‐terminus in protein‐protein interactions. This research was supported through NIH grants T36GM008789 and P41 RR06009.
A YAP/TAZ‐Regulated Transcriptional Signature Associated with Oral Squamous Cell Carcinoma
Authors: Hiemer Samantha , Zhang Liye , Kartha Vinay , Packer Trevor , Almershed Munirah , Noonan Viki , Kukuruzinska Maria , Bais Manish , Monti Stefano , Varelas Xaralabos
Published: June 2021

Oral squamous cell carcinoma (OSCC) is a prevalent form of cancer that develops from the epithelium of the oral cavity. OSCC is on the rise worldwide, and the death rates associated with the disease are particularly high. Despite progress in the understanding of the mutational and expression landscape associated with OSCC, advances in deciphering these alterations for the development of therapeutic strategies have been limited. Further insight into the molecular cues that contribute to OSCC is therefore required.

Here we show that the transcriptional regulators YAP and TAZ, which are key effectors of the Hippo pathway, drive pro‐tumorigenic signals in OSCC. Regions of pre‐malignant oral tissues exhibit aberrant nuclear YAP accumulation, suggesting that dysregulated YAP activity contributes to the onset of OSCC. Supporting this premise, we found that nuclear YAP and TAZ activity drives OSCC cell proliferation, survival, and migration in vitro, and is required for OSCC tumor growth and metastasis in vivo. Analyses of the global gene expression changes associated with YAP and TAZ knockdown revealed roles in the control of genes important for pro‐tumorigenic signaling, including those encoding factors required for cell cycle progression and survival. Notably, the transcriptional signature regulated by YAP and TAZ significantly correlates with gene expression changes occurring in human OSCCs identified by “The Cancer Genome Atlas” (TCGA), emphasizing a central role for YAP and TAZ in OSCC biology. This study defines a YAP/TAZ‐regulated transcriptional program in OSCC, and reveals novel roles for nuclear YAP/TAZ activity in the onset and progression of this cancer.
Fatty Acids from the leaves of Clerodendrum volubile Suppresses Tumor Cell Migration and Invasion, Attenuates Oxidative Stress, and Regulates Cell Cycle Progression in Human Neuronal Glioblastoma Cells
Authors: Erukainure Ochuko , Ebuehi Osaretin , Choudhary Iqbal , Asraf Nadia , Naqvi Asma , Muhammad Aliyu , Zaruwa Moses , Elemo Gloria
Published: June 2021

Malignant gliomas have been shown to represent some of the greatest challenges in the management of cancer patients. In this study, the antiproliferative effect of the fatty acid components of Clerodendrum volubile leaves as well as its antioxidant effect on U‐87 MG (Human Neuronal Glioblastoma) cell lines was investigated. Fatty acids extracted from C. volubile leaf oil were subjected to GCMS analysis. U‐87 MG cells were cultured and treated with the fatty acids for 48 hrs, after which the antiproliferative effect was ascertained via MTT assay and cell viability analysis using BD FACS Calibur. Cell cycle was analyzed by flow cytometry on FACS Calibur. Western blotting was used in determining expression of proteins in the cell lines. Cell migration and invasion were analyzed using the migration and invasion transwell assay protocol. The treated cell lines were assessed for reduced glutathione (GSH) level, catalase, superoxide dismutase (SOD) and lipid peroxidation. The fatty acids significantly inhibited cell proliferation, arrested S phase, modulated expression of MMP – 9 and VEGF, attenuated oxidative stress, and suppressed migration and invasion in U‐87 MG cell lines. These results indicate the therapeutic potential of the fatty acids components of the leaves of Clerodendrum volubile on human neuronal glioblastoma cells.
Replication‐dependent repression of SV40 early transcription occurs by placement of a nucleosome over early regulatory elements
Authors: Milavetz Barry , Christensen Kendra , Balakrishnan Lata
Published: June 2021

We have previously shown that during the Simian Virus 40 (SV40) life‐cycle, DNA replication serves as an epigenetic switch by introducing H3K9me1 into newly replicated SV40 minichromosomes in order to repress early transcription in these newly‐replicated minichromosomes. While these studies demonstrated that epigenetics played a critical role in regulation of SV40 transcription, they did not directly address the exact mechanism of how the introduction of H3K9me1 repressed early transcription. We have now analyzed the location of nucleosomes in SV40 wild‐type 776 and the repression‐defective mutant cs1085 using a ChIP‐Seq like approach and confirmed that there is a significant increase in minichromosomes lacking a nucleosome over the early regulatory region in the cs1085 repression‐defective mutant minchromosomes compared to the wild‐type minichromosomes. In addition, we have shown that nucleosomes containing H3K9me1 are preferentially found within the early regulatory region in the wild‐type minichromosomes. These results suggest replication‐dependent repression of early transcription occurs by a mechanism in which nucleosomes specifically containing H3K9me1 are targeted to locations within the early regulatory region including the binding site of TBP and RNA Polymerase II. These specifically located nucleosomes would serve as a physical barrier to prevent the binding of the factors necessary for initiation of early transcription.
Leukocyte Integrins αLβ2, αMβ2 and αXβ2 as Collagen Receptors ‐ Receptor Activation and Recognition of GFOGER Motif
Authors: Lahti Matti , Heino Jyrki , Käpylä Jarmo
Published: June 2021

Integrins αLβ2, αMβ2 and αXβ2 are expressed on leukocytes. Their primary ligands are counter transmembrane receptors or plasma proteins, such as intercellular cell adhesion molecule‐1 (ICAM‐1) or components of complement system (iC3b, iC4b), respectively. Function blocking antibodies for these integrins may also reduce cell adhesion to collagens. To make the first systematical comparison of human αLβ2, αMβ2 and αXβ2 as collagen receptors, we produced the corresponding integrin αI domains both in wild‐type and activated form and measured their binding to collagens I‐VI. In the “closed” (wild‐type) conformation, the αLI and αMI domains bound with low avidity to their primary ligands, and the interaction with collagens was also very weak. Gain‐of‐function mutations αL I306G, αL K287C/K294C and αM I316G are considered to mimic “open”, activated αI domains. The binding of these activated αI domains to the primary ligands was clearly stronger and they also recognized collagens with moderate avidity (Kd<400nM). After activation, the αLI domain favored collagen I (Kd∼80nM) when compared to collagen IV. The integrin αXI domain acted in a very different manner since already in native, wild‐type form it bound to collagen IV and iC3b (Kd∼200‐400nM). Antibodies against αXβ2 and αMβ2 blocked promyelocytic leukemia cell adhesion to the collagenous GFOGER motif, a binding site for the β1 integrin containing collagen receptors. In brief, leukocyte β2 integrins may act as collagen receptors in a heterodimer specific manner.
Unraveling binding interactions of serotonin 5‐HT1B/1D receptor agonists on P‐glycoprotein: the gatekeeper of the brain
Authors: Wilt Laura , Roberts Arthur
Published: June 2021

Lack of drug penetration through the blood‐brain barrier (BBB) represents a formidable hurdle to treatment of central nervous system (CNS) diseases and the development of CNS drugs. The transporter P‐glycoprotein (Pgp) is the gatekeeper at the BBB and effluxes a wide range of chemically and structurally diverse neurotherapeutics. The triptans represent a chemical class of CNS agonists that show large differences in their Pgp efflux rates and transport properties, despite their similarities in molecular structure. We hypothesize that the variations of chemical structure and P‐glycoprotein transport rate in the triptan class are controlled by differences in the binding location on the transporter. Identifying the molecular features that are critical for transport will improve predictions of drug disposition and will lead to novel neurotherapeutics that have minimal transport. To gain insight into the molecular basis of triptan transport, binding interactions and efflux ratios were characterized within our biomimetic model consisting of reconstituted Pgp in artificial membranes. Specific ATPase activity, transport and intrinsic tryptophan fluorescence were used to investigate eletriptan (Relpax®) and zolmitriptan (Zomig®) binding interactions in our biomimetic model. We have successfully reconstituted Pgp and characterized the ATPase activity and transport with the probe drug verapamil. Eletriptan and zolmitriptan were identified to have different ATPase activity, transport rates and intrinsic tryptophan fluorescence, suggesting these neurotherapeutics interact with Pgp in distinct regions on the transporter.
EU‐OPENSCREEN: Novel chemical tool compounds for molecular biologists
Authors: Stechmann Bahne , Frank Ronald
Published: June 2021

Small molecules that can be applied as chemical 'tool' compounds (or 'probes') have become indispensable in basic research for the elucidation of fundamental biological mechanisms. They act directly with the protein‐of‐interest and often allow for the interrogation of biological processes that cannot be properly studied with traditional genetic or RNA interference approaches.

EU‐OPENSCREEN (www.eu‐openscreen.eu) is the largest emerging academic chemical biology research infrastructure initiative in Europe with the aim to collaboratively develop novel research tool compounds with independent cell biologists. As a joint effort of national networks in 16 European countries, EU‐OPENSCREEN offers access to high‐throughput screening platforms, chemistry services and a large compound collection. It welcomes molecular biologists who have a robust and suitable biological assay and are interested in collaboratively developing chemical tool compounds to validate their targets‐of‐interest. Selected assays are screened against a collection of more than 100,000 compounds, incl. confirmatory and counter screening, IC/EC50 determination, SAR (structure‐activity relationships) and QC of confirmed hit compounds.

EU‐OPENSCREEN will start operations in late 2015, but it can already look back on a growing number of transnational activities: joint screening projects, exchange of local compound libraries, development of new design principles for its compound collection; exchange of experimental data through its pilot database etc.
Coordinate regulation of ASK1/TGF‐β signaling contributes to improved glucose and lipid metabolism in mice
Authors: Ha Hyunjung , Seong Hyun‐A , Manoharan Ravi
Published: June 2021

We explore the molecular connections between ASK1 and TGF‐β signaling pathways and examine the physiological processes in which coordinate regulation of these two signaling pathways plays a critical role. ASK1 and TGF‐β signaling pathways are interconnected by complex containing Smad proteins (Smad2, 3, 4, and 7) and ZPR9, a zinc finger protein, under the control of apoptosis signal‐regulating kinase 1 (ASK1) and its upstream kinase, murine protein serine‐threonine kinase 38 (MPK38), and a signal for either ASK1 or TGF‐β activity is sufficient to activate both ASK1 and TGF‐β signaling pathways via phosphorylation of Smad(s) and ZPR9 by ASK1 and MPK38. The Smad(s)‐ZPR9 complex is required for the coordinate regulation of ASK1 and TGF‐β signaling in response to ASK1 or TGF‐β signals. We also defined Smad(s) as differential regulators for ASK1 and its downstream targets (MKK3, p38, and JNK), as well as ZPR9 as a regulator of Smad(s) stability. In the physiological setting, restoration of down‐regulated ASK1 and TGF‐β signaling in a genetic mouse model of obesity by adenoviral delivery of Smad3 or ZPR9 resulted in the amelioration of hyperglycemia and hyperlipidemia.
The Functions of miRNAs in Self‐renewal and Differentiation of Human Mesenchymal Stem Cells
Authors: Suh Nayoung , Park Seul‐Ki , Hwang Supyong , Jung Yideun
Published: June 2021

Human mesenchymal stem cells (hMSCs) have emerged as therapeutic promises for a wide range of diseases, but the regulatory mechanisms that control MSC fate determination and therapeutic efficacy remain poorly understood. MicroRNAs (miRNAs) are small non‐coding RNAs that regulate diverse biological processes at the post‐transcriptional level. We aimed to identify miRNAs enriched in hMSCs which modulate differentiation commitments. miRNA microarray analysis revealed that miR‐140‐3p and miR‐140‐5p were significantly upregulated in undifferentiated hMSCs from various tissue sources such as adipose, bone marrow, and umbilical cord, compared to human fibroblasts. Target prediction analysis and luciferase reporter assay revealed that miR‐140‐5p directly repress bone morphogenic protein 2 (BMP2), a potent inducer for osteogenetic differentiation. Furthermore, blocking miR‐140‐5p in human adipose‐derived mesenchymal stem cells (hADSCs) enhanced the expression of BMP2 and a subset of bone morphogenic protein (BMP) signaling components which is essential for osteogenetic lineage commitment. We propose that miR‐140‐5p can affect osteogenic lineage commitment in undifferentiated hADSCs at least in part by negatively regulate BMP2. §This work is supported by grants from KMEST (2012R1A1A3013555 and 2014R1A1A2059398).
Stonefish toxin defines a new branch of the perforin‐like superfamily
Authors: Ellisdon Andrew
Published: June 2021

The lethal factor in stonefish venom is Stonustoxin (SNTX), a cytolytic protein that induces hypotension and cardiovascular collapse in mammals. Here, we show that SNTX is a pore forming member of an undiscovered branch of the Cholesterol Dependent Cytolysin / Membrane Attack Complex/Perforin (CDC/MACPF) superfamily. Structural analyses suggest that the initial SNTX interaction with a target membrane is mediated by two C‐terminal PRYSPRY domains, which are closely related to the lipid and protein binding domains of the tripartite motif immune protein family. Two N‐terminal CDC/MACPF domains are arranged in a pre‐pore like assembly; these data provide high‐resolution insight into how SNTX and other CDC/MACPF proteins assemble into pores. Finally, our analyses reveal that SNTX‐like CDCs are distributed throughout eukaryotic life, and likely play a broader, and possibly immune‐related function, outside venom.
The yeast atlastin Sey1p may not be sufficient to drive homotypic ER fusion at its physiological concentration
Authors: Moon Yeojin , Lee Miriam , Ko Young‐Joon , Han Minsoo , Park Sangmin , Jun Youngsoo
Published: June 2021

The endoplasmic reticulum (ER) is one of the most important organelle in the cell. The ER has many functions including cytoplasmic calcium level control, protein quality control, and lipid synthesis. These functions are thought to be closely related with the unique structure of the ER, which forms a network of sheets and tubules that are maintained by homotypic ER membrane fusion. Although the exact mechanism and players of homotypic ER fusion is yet to be unveiled, the dynamin‐like GTPase atlastin is believed to mediate homotypic ER fusion by forming a homo‐dimer in trans between fusing membranes. In this study, we measured the physiological concentration of the yeast atlastin homolog Sey1p from isolated yeast microsomes and performed lipid mixing assays with Sey1p‐proteoliposomes bearing various concentrations of Sey1p. Our study revealed that the physiological ratio of lipids to Sey1p molecules was approximately 1.0 x 105 and that lipid mixing was hardly detectable for proteoliposomes bearing Sey1p at a 1:4000 molar ratio to lipids, which is still 25 fold abundant as compared to isolated microsomes. Thus, this study suggests that the yeast atlastin homolog Sey1p at its physiological concentration is not sufficient to drive homotypic ER fusion by itself and that Sey1p needs additional factors to support ER fusion in vivo.
Effect of ASIC‐BK Interaction on Migration of PC12 Cells
Authors: Springer Carola , Young Michael , Vega Quinn , Petroff Elena
Published: June 2021

Acid sensing ion channels (ASICs) and large conductance calcium and voltage‐activated potassium (BK) channels are expressed in normal glia as well as glioma tumors. At reduced pH, ASIC channels are activated and the inhibition of BK is relieved (Petroff et al., 2008). Lactic acidosis has been associated with cerebral trauma and tumors (Andersen et al. 1988 and Wike‐Hooley 1984). Our previous studies suggest that pH‐dependent relief of BK inhibition by ASICs increases cell proliferation in glial cells. We hypothesized that the activation of BK channels by a decrease in pH and may also promote cell migration. PC12, a rat pheochromocytoma cell line that expresses both ASIC and BK channels, was used as a model system. The cells were cultured at physiological pH 7.4 and reduced pH (7.0), in the presence and in the absence of 200nM of charybdotoxin, a BK channel blocker, and assessed for cell migration. Our data show that activation of BK channels by reduced pH or inhibition of BK channels by charybdotoxin have no effect on PC12 cell migration. These results indicate that in PC12 cells, the molecular mechanism involved in cellular proliferation by activation of BK channels might be independent from the mechanisms involved in cell migration. This work is supported by the R15 NIH grant to E.P.
Sorghum as a Biological Model for Studying the Effect of Microbial Interactions on Growth and Metabolic Activity in Mining‐Impacted Soil
Authors: Dhawi Faten , Datta Rupali , Wusirika Ramakrishna
Published: June 2021

The main challenge for plants growing in nutrient poor, metal‐contaminated soil is reduction in productivity due to nutrient deficiency as well as heavy metal toxicity. Our aim was to overcome these challenges using various microbial combinations, and study the potential beneficial impact of plant‐microbe interactions on plant physiological systems. In the current study, sorghum BTx623 seedlings planted in metal contaminated mining‐impacted soil were subjected to different microbial combinations. Following treatments were used: Plant growth promoting Bacteria (PGPB) isolated from metal contaminated sediments, PGPB + mycorrhizal fungi, mycorrhizal fungi alone and control group with no microbial treatment. The results indicate highest growth enhancement in plants treated with the mycorrhiza alone, followed by PGPB bacteria alone and PGPB and mycorrhiza mix in comparison with control. The increase in root biomass was accompanied with increase in uptake of primary nutrients, secondary nutrients and micronutrients in both shoots and roots. A total of 29 compounds were identified, out of which 23 showed significant changes in comparison to the control. The metabolites that changed significantly were those involved in starch and sucrose; and biosynthesis of amino acids such as alanine, aspartate and glutamate, galactose and valine, leucine and isoleucine.
Synthesis of Fluorescently‐Labeled Tobacco Etch Virus (TEV) RNA and Its Interactions with Pokeweed Antiviral Protein (PAP)
Authors: Rodriguez David , Domashevskiy Artem
Published: June 2021

Fluorescence spectroscopy has been extensively used to investigate protein‐protein and protein‐nucleic acid interactions. Changes in fluorescence intensity provide vital information about these macromolecular interactions. Availability of extrinsic fluorophores became invaluable for researchers monitoring these interactions. Here, we successfully synthesize a fluorescent anthranioyl‐m7GpppG‐capped tobacco etch virus (TEV) RNA, and examine its properties pertaining to interaction with pokeweed antiviral protein (PAP). PAP is a ribosome inactivating protein (RIP) and is an RNA N‐glycosidase that removes purines from the sarcin/ricin loop of large rRNA. This leads to the translational termination. PAP possesses antiviral properties, and it is thought to bind to the 5'‐cap of viral RNA and depurinate it down the stream of the cap. RIPs are thought to play an important role in plant defense mechanisms. We employ fluorescence spectroscopy and HPLC techniques to investigate and characterize Ant‐m7GpppG‐capped TEV RNA with PAP.
RNA Distribution During Flock House Virus Infection
Authors: Gopal Radhika , Short James , Schneemann Anette
Published: June 2021

The mechanisms involved in the specific packaging of multipartite genomes by (+) RNA viruses remain poorly understood. In order to gain insight into the location of genomic RNA encapsidation, we employed fluorescence in situ hybridization (FISH) to visualize the sub‐cellular distribution of (+) vRNA in cells infected with the model nodavirus, Flock House virus (FHV). During FHV assembly, genomic RNAs 1 and 2 are encapsidated into a T = 3 icosahedral capsid. Previously it has been shown that while vRNA replication and CP synthesis are initially targeted to different cellular locations (mitochondria and ER, respectively), these regions converge into perinuclear clusters late in infection. Our analysis of the cellular distribution of FHV RNA during infection, however, suggests that the vast majority of (+) RNA is trafficked towards the periphery of the cell relatively early during the infectious cycle (within 3 hours post infection) and does not relocate to the perinuclear region during mitochondrial condensation (15 hours post infection). In addition, the (+) RNA fits well within the region occupied by CP at the time points we tested. Our findings show that RNA transport precedes genome packaging and virus assembly. This suggests that genomic RNA is likely released from vRNA replication sites fairly soon after synthesis and trafficked to peripheral sites prior to encapsidation. Our analysis in mammalian cells also revealed that in the absence of B2, a small virus‐encoded protein known to bind dsRNAs, RNA2 is stalled in cytoplasmic aggregates while RNA1 trafficking is largely unaffected.
Exploring the Requirement for the CCR4‐NOT Deadenylase Complex Subunit Regena/NOT2 in MicroRNA‐mediated Gene Silencing
Authors: Reinke Catherine
Published: June 2021

A forward genetic screen was carried out using Drosophila melanogaster to determine the cellular components and molecular mechanisms that regulate microRNA (miRNA)‐mediated gene silencing. MiRNAs are non‐coding RNA molecules that interact with target‐gene messenger RNA transcripts via complementary base pairing, and silence target gene expression via interaction with the miRNA‐induced silencing complex (miRISC). Our screen utilized a Green Fluorescent Protein (GFP)‐based reporter to visualize miRNA‐mediated gene silencing. The GFP transgene contains miRNA binding sites, and is regulated by endogenous miRNAs. MiRNA‐mediated silencing is accomplished through translation block, mRNA degradation, or a combination of these mechanisms by miRISC. A requirement for the CCR4‐NOT complex subunit Regena/NOT2 in miRNA‐mediated gene silencing was discovered through genetic analysis of an isolated mutant. We are exploring the specific role of Regena/NOT2 in silencing by determining whether this CCR4‐NOT complex component is required for translational block, transcript degradation, or both. We are further characterizing a putative role for Regena/NOT2 in miRNA biogenesis and maturation, as has been demonstrated in Arabidopsis. Ongoing studies of additional mutants generated in the screen aim to identify the components of miRISC in different cellular contexts and with respect to different miRNA targets, to elucidate the complex molecular mechanisms of miRNA‐mediated silencing.
Influence of Obesity‐Related Genotype on Weight Loss Success and Body Composition Changes While Participating in an a 3‐Month Exercise and Weight Loss Program: Preliminary Findings
Authors: Coletta A , Sanchez B , O'Connor A , Dalton R , Springer S , Koozehchian M , Jung Y , Simbo S , Cho M , Goodenough C , Reyes A , Galvan E , Levers K , Wilkins K , Rasmussen C , Kreider R
Published: June 2021

This study examined whether genotype of some obesity‐related genes (FABP2, PPARG, ADRB2‐79, ADRB2‐46, ADRB3) influences success to different diets while participating in an exercise program. 40 sedentary women (41±13 yrs, 35.8±7.9 kg/m2) were matched according to genotypes favoring carbohydrate restricted and less restricted diets. Diets were 1,500 kcal/d with 20:35:45 and 30:25:45 percentages of C:F:P. Participants performed a circuit style resistance‐exercise program (4 d/wk) and a walking program (10,000 steps/d, 3 d/wk). Body weight and DEXA body composition measures were obtained at 0, 4, 8, & 12 wks, analyzed by MANOVA, and are presented as changes from baseline after 12 wks. An overall Wilks' Lambda time x genotype trend was observed (p=0.08) with no time x diet effects (p=0.57). Univariate analysis revealed significant differences between true (T) and false (F) matched genotype groups in fat mass (T: ‐3.81±2.5, F: ‐1.92±3.2 kg; p=0.013), FFM fat free mass (T:‐0.76±1.8, F: ‐2.16±2.3; p = 0.042) and body fat (T: ‐2.15±2.3, F: 0.01±2.1 %; p=0.002) with no differences in changes in body weight (T: ‐4.33±3.6, F: ‐3.91±3.9 kg, p=0.36). These preliminary findings suggest that women participating in a diet and exercise program may experience greater improvements in body composition when diet type is aligned with their genetic profile.
Effect of the Mediterranean Diet on DNA Methylation of Selected Genes in the PREDIMED‐Valencia Intervention Trial
Authors: Corella Dolores , Ordovas Jose , Sorli Jose , Asensio Eva , Ortega Carolina , Carrasco Paula , Portoles Olga , Coltell Oscar
Published: June 2021

We previously reported diet‐gene interactions of the Mediterranean diet (MedDiet) in modulating the effects of polymorphisms in the TCF7L2‐rs7903146 and FTO‐rs9939609 genes on stroke and diabetes. Our aims were: 1) to analyze the influence of adherence to the MedDiet at baseline on the DNA‐methylation of the TCF7L2 and FTO genes and 2) to study the impact of a one‐year intervention with MedDiet supplemented with extra virgin olive oil (EVOO) on the methylation of those same loci in comparison to that of a control diet. We conducted a pilot study (n=195) on the PREDIMED‐Valencia participants (aged 67+/‐7 y), a randomized controlled intervention trial. Quantitative DNA methylation analysis was carried out by MALDI‐TOF mass spectrometry at baseline and after 1‐year dietary intervention with MedDiet supplemented with EVOO and in the control group. We analyzed 31 CpG sites surrounding the SNPs. The polymorphisms were strongly associated with methylation levels (P<0.01 for the CpG site 7 in amplicon B of the TCF7L2‐rs7903146 and CpG site 8A of the FTO). At baseline, we obtained an inverse association between adherence to MedDiet and methylation levels in the TCF7L2. Of all the nutrients, folate intake presents the most significant associations with methylation levels of the several CpG islands of both genes (P<0.005). Intervention with MedDiet was significantly associated with hypomethylation of some CpG. But, they are also some loci associated with hypermethylation. That specificity requires further investigations.
Correlation Between Diet Macronutrients and Metabolic plus Cardiovascular Abnormalities in Spontaneously Hypertensive Rats
Authors: Oliveira Junior Silvio , Padovani Carlos , Rodrigues Sergio , Silva Nilza , Martinez Paula , Campos Dijon , Okoshi Marina , Okoshi Katashi , Cicogna Antonio
Published: June 2021

This study analyzed the association between dietary macronutrients and metabolic, hormonal, as well as cardiovascular responses in spontaneously hypertensive rats (SHR). SHR (n=20) were submitted to control (3.2 Kcal/g) or hypercaloric (4.6 Kcal/g) diets for 20 weeks. Afterward, calorie intake, body weight, adiposity, glycemia, insulin, and leptin measurements were obtained. Cardiovascular analysis integrated systolic blood pressure, echocardiography, morphology, and myosin heavy chain composition. Canonical correlation analysis was used to evaluate the relationships between dietary macronutrients and metabolic plus cardiovascular parameters.

Results

Canonical scores for sucrose, saturated and unsaturated fatty acids were strongly correlated with adiposity, leptin and glycemia (r=0.9816; p<0.001). In cardiovascular context, canonical indexes were lower for unsaturated fatty acids, sucrose, and saturated lipids in the set of macronutrients; these scores were directly associated with the ratio between left atrium and aortic artery diameters (LA/AO), E‐wave, E wave‐to‐A wave ratio (E/A), and isovolumetric relaxation time (IVRT) (r=0.9975; p<0.001). Also, saturated and unsaturated lipids, followed by sucrose were directly correlated with morphological and molecular cardiac parameters (r=0.9271; p<0.001).

Conclusion

Despite of diet, sucrose, saturated and unsaturated fatty acids intakes were correlated with metabolic and cardiovascular disorders in SHR. Financial support: FAPESP
Low Dietary Intakes of Omega‐3 Fatty Acids and Niacin Are Associated with Schizophrenia in Korean Adults
Authors: Lim So Young , Kim Sung‐Wan , Yang Soo Jin
Published: June 2021

Increased prevalence of metabolic syndrome (MS) has been reported in subjects with schizophrenia. The underlying mechanisms have not yet clearly identified; however, medications and unhealthy lifestyle choices including physical inactivity and poor diet may be contributing factors for developing MS. In this study, we investigated whether dietary factors are associated with the prevalence of schizophrenia in Korean adults. Total 124 subjects were voluntarily participated: 1) control (CON, n = 67), and 2) schizophrenia (n = 57). Dietary habits and intakes from foods were assessed with modified questionnaires for dietary guideline adherence score and semi‐quantitative food frequency questionnaires. Dietary intakes of energy and nutrients were estimated using a computer software program based on the standard tables of food composition. Body mass index and total energy intakes were higher in subjects with schizophrenia compared with CON. Although dietary guideline adherence scores were not different among groups, male subjects with schizophrenia consumed lower levels of omega‐3 polyunsaturated fatty acids (PUFAs) (sum of eicosapentaenoic acid, docosahexaenoic acid, and docosapentaenoic acid, etc.), arachidonic acid, niacin, folate, and vitamin C. Moreover, odds ratios (OR) for schizophrenia were significantly attenuated in subjects with high dietary intakes of omega‐3 PUFAs and niacin (omega 3 PUFAs OR = 0.340, p = 0.037; niacin OR = 0.260, p = 0.010). Our results suggest the association between dietary factors and schizophrenia, and underline the necessity to develop appropriate dietary guideline for subjects with schizophrenia.
DMSO Represses Inflammatory Cytokine Production from Human Blood Cells and Reduces Autoimmune Arthritis
Authors: elisia ingrid , Nakamura Hisae , Cederberg Rachel , Lee Leorra , Lam Vivian , Adomat Hans , Guns Emma , Cait Jessica , Hughes Michael , McNagny Kelly , Samudio Ismael , Krystal Gerald
Published: June 2021

Dimethyl sulfoxide (DMSO) is currently being used by many people, both topically and orally, to treat inflammatory conditions as well as cancer. However, little is known about the efficacy, safety and mechanism of action of DMSO in humans. We therefore examined the effects of this polar solvent on whole human blood, stimulated or not, ex vivo, with E. coli. We found that, between 1 and 2%, DMSO markedly inhibited cytokine production without affecting cell viability. However, at greater than 5%, significant death of leukocytes was observed, suggesting that DMSO has only a narrow window of efficacy. The two in vivo generated metabolites of DMSO, dimethyl sulfide (DMS) and dimethylsulfone (DMSO2), required higher concentrations than DMSO to be anti‐inflammatory. Mechanism of action studies using purified human monocytes revealed that DMSO's anti‐inflammatory properties were due, at least in part, to inhibition of the ERK1/2, p38, PI3K and JNK pathways. In vivo mouse model studies revealed that topical administration of DMSO did not reduce the growth rate of subcutaneously implanted B16 melanoma cells, even though it did act in vitro to increase their differentiation. Importantly, however, topical administration of DMSO significantly ameliorated KBxN‐induced arthritis in mice and this was associated with lower levels of inflammatory cytokines in the joints. Taken together, our studies suggest that DMSO may be useful for some inflammatory conditions but care must be taken to avoid toxic effects.
Effect of the Complex Extracts of Lotus/Turmeric in accordance with Medical Nutrition Therapy in Subjects with Borderline Dyslipidemia
Authors: Lee Hansongyi , Lim Hyunjung
Published: June 2021

Aims

To investigate the effects of complex extracts of Lotus/Turmeric in accordance with medical nutrition therapy (MNT) in subjects with borderline dyslipidemia according to the Korea Society of Lipidology and Atherosclerosis (Total‐cholesterol: 200‐239 mg/dl or low density lipoprotein cholesterol: 130 ‐159 mg/dl or Triglyceride: 150 ‐199 mg/dl).

Methods

This 12‐week randomized, controlled clinical trial was carried out. Eighty‐six subjects divided 4 groups, PLACEBO (n=23), PLACEBO+MNT (n=19), ACTIVE (n=21), ACTIVE+MNT (n=23). Placebo or Active was ingestion of 4 tablets per daily for 3 month. Active groups were ingestion of the Lotus/Turmeric extracts, total 1,400 mg/daily. All subjects' anthropometrics, nutrient intakes, blood lipid parameters and anti‐oxidants were assessed baseline and after 12 weeks. The research protocol was approved by the institutional review board.

Results

In the ACTIVE group, HDL‐ cholesterol levels was significantly increased from the 58.9±11.2 mg/dl to 62.9±16.1 mg/dl (p=0.006) although TC, LDL‐cholesterol, and TG dose not changed. ACTIVE+MNT group, superoxide dismutase increased significantly after 12 weeks (changes: +0.9±1.4 U/mL) (P=0.006). Furthermore, the malondialdehyde also significantly reduced in the ACTIVE (change: ‐0.2±0.3 pmol/mg) (p=0.030) and ACTIVE+MNT (change: ‐0.1±0.2 pmol/mg) (p=0.048) groups.

Conclusion

The results suggest that the complex extracts of Lotus/Turmeric in accordance with MNT could be beneficial effect and might be useful for subject with dyslipidemia.
AdamTS‐A Developmental Function in Neuronal Development and Migration in Drosophila Melanogaster
Authors: Romero Selena , Zhu Yi , Wilson Beth , Skeath James
Published: June 2021

The Drosophila central nervous system (CNS) is an excellent model system in which to dissect the genetic and molecular processes that control the development of the nervous system. Through a forward genetic screen, we previously identified, a larval phenotype that leads to the formation of ectopic neural masses. These masses are composed of dozens to thousands of neuronal cell bodies at the junctures between the brain and the eyes, the ventral nerve cord and the leg imaginial discs, as well as within nerves. We determined that this mutant phenotype arises due to a transposable element insert within the 5' untranslated region of a disintegrin and metalloprotease with thrombospondin type 1 motif A (AdamTS‐A) gene. The AdamTS family of proteases are secreted proteins thought to play critical roles in extracullular matrix (ECM) formation and function, and their regulation has been linked to cancer progression. We have recently generated RNA and antibody probes to the AdamTS‐A and will report the RNA and protein expression of Adam TS‐A during larval CNS development.

Support from the NINDS/NIH grant number NS0365570 and the Opportunities in Genomics Research through the Genome Institute at Washington University in St. Louis, grant number R25H600668703.
Bioactive Micronutrients in Seven Species of Passiflora from Brazil
Authors: Camacho Márcio , Villamil Daniel , Mônaco Kamila , Corrêa Camila , Pereira Giuseppina , Chau Ming
Published: June 2021

Bioactive components in Passiflora vary according to their species. This study determined bioactive fat‐soluble components such as carotenoids and α and δ‐ tocopherols in seven varieties (P. paurifolia, P. guardeni, P. coccinea, P. edulis, P. alata, P. incarnate, P. mucionata) using a HPLC system. The highest amounts of lutein were observed in leaves of P. coccinea (3968, 81ug/g), P.gardeni (416, 66 µg/g), P.edulis (352, 58 µg/g), P.alata (332, 92µg/g), P. incarnata (284, 76 µg/g) and P. mucionata (109, 32 µg/g). P. paurifolia does not contain this carotenoid. Regarding cryptoxanthin it was found only in leaves of three species, P. mucionata (4, 04 µg/g), P. edulis (2, 89µg/g) e P.alata (1, 97µg/g). β‐carotene was the other carotenoid found in five species, P.gardeni (169,68µg/g), P.alata (138,10), P.coccínea, (112,12µg/g), P.edulis (62,61µg/g) and P. incarnata (17,43ug/g). α and δ‐tocopherol were not found in any species of Passiflora. Therefore the leaves of passion flower are rich in carotenoids and some of that can be useful for the dietary antioxidant strategy.