bluuebunny/crossref_metadata_2025 huggingface dataset:
Aldehyde Dehydrogenase‐2 (ALDH2) Ameliorates Chronic Alcohol Ingestion‐Induced Hepatic Steatosis and Inflammation
Authors: Guo Rui , Ren Jun
Published: June 2021

Objectives

Mitochondrial aldehyde dehydrogenase (ALDH2) plays a critical role in the detoxification of the ethanol metabolite acetaldehyde. Chronic alcohol intake has been shown to result in alcoholic liver disease although the precise mechanism of action is still elusive. This study was designed to examine the impact of global ALDH2 overexpression on chronic alcohol ingestion‐induced hepatic steatosis and inflammation.

Methods

FVB and ALDH2 mice were placed on a 4% alcohol or control diet for 12 weeks. Serum levels of AST/ALT and total bilirubin, levels of hepatic triglyceride, hepatic histology on steatosis, fat metabolism‐related proteins and inflammatory factors were examined.

Results

Chronic alcohol intake led to enhanced levels of total bilirubin and hepatic triglycerides, increased fat deposition as evidenced by H&E and Oil‐Red‐O stainings, elevated fatty acid synthase, stearoyl‐CoA desaturase 1, phospho‐acetyl‐CoA carboxylase‐1, PPAR γ, PGC‐1α, IL‐6 and phosphorylation of AMPK and Stat3, decreased phosphorylation of mTOR as well as unchanged levels of G6Pase‐α, TNF‐α and IL‐1β, the effects of which were overtly attenuated or ablated by ALDH2, with the exception of fatty acid synthase, G6Pase‐α, TNF‐α and IL‐1β expression.

Conclusions

In summary, our results revealed that ALDH2 may effectively ameliorate chronic alcohol ingestion‐induced hepatic steatosis and inflammation.
Tobacco components activate the acetylcholine signaling pathway in bronchioalveolar carcinoma
Authors: Lau Jamie K. , Brown Kathleen C. , Crabtree Clayton M. , Dom Aaron M. , Buckley Adam W. , Harman Jarrod C. , Dasgupta Piyali
Published: June 2021

Nicotine, the addictive component of cigarettes, accelerates cell proliferation and angiogenesis through nicotinic acetylcholine receptors (nAChRs). We show for the first time that acetylcholine (ACh) stimulates proliferation of A549 and H358 human bronchioalveolar carcinoma (BAC) cells. The maximal proliferation of human BACs was observed at 2 μmol. Furthermore, we observed that nicotine induced the production of ACh in a concentration and time‐dependent manner. The pro‐secretory activity of nicotine was mediated via α7‐ and β3‐containing‐nAChRs. Western blotting and ELISA analysis indicates that proteins involved in ACh signaling pathway, vesicular acetylcholine transporter (VAChT), choline transporter 1 (CHT1) and acetylecholinesterase (AChE), are expressed on A549 and H358 human BACs. Finally, VAChT and CHT1 inhibitors (vesamicol and hemicholium respectively) caused robust apoptosis of A549 and H358 cells and had little effect on normal lung cells. Our observations raise the possibility that tobacco components like nicotine promote the growth of human BACs by stimulating ACh production. Further, the disruption of this process may have applications in BAC therapy.
Nicotine induces the up‐regulation of alpha‐7‐nicotinic receptors (α7‐nAChRs) in human squamous carcinoma of the lung via transcriptional mechanisms
Authors: Dasgupta Piyali , Lau Jamie K. , Brown Kathleen C. , Luo Haitao , Chen Yi Charlie
Published: June 2021

Nicotine, the addictive component of cigarettes, promotes the proliferation of lung cancers via nicotinic acetylcholine receptors (nAChRs), specifically the α7‐nAChR subtype. We explore the effect of nicotine treatment on α7‐nAChR levels in squamous cell carcinomas of the lung (SCC‐L) in vitro and in vivo. Nicotine increased α7‐nAChR levels in three human SCC‐L cell lines. Nicotine‐induced up‐regulation of α7‐nAChR was dependent on α7‐nAChRs and the protein kinase C (PKC) pathway. The up‐regulation of α7‐nAChR upon nicotine treatment was confirmed in H520 tumors xenotransplanted on chicken chorioallantoic membranes (CAM). Real‐time PCR and luciferase assays showed that nicotine increased α7‐nAChR levels by transcriptional mechanisms in SCC‐L cells and CAM tumors. ChIP showed that nicotine induced the binding of GATA4 or GATA6 to Sp1 on the α7‐nAChR‐promoter, thereby increasing its levels in human SCC‐L. Long‐term exposure to nicotine could up‐regulate α7‐nAChRs on SCC‐L, thereby promoting its progression and metastasis. Our data would also be relevant to SCC‐L patients exposed to nicotine via second‐hand smoke or nicotine patches/gums used to quit smoking.
Hands on Physiology Activity Improves Learning of Special Senses
Authors: Carvalho Helena , Collins Kristy M.
Published: June 2021

Variety of methodologies have been employed to teach the sciences with different levels of success since students have different learning styles such as auditory, visual and Kinesthetic. Kids’ Tech University is an annual event at Virginia Tech University where middle school students attend a series of lectures in sciences and also actively participate in a hands‐on activity. This study evaluates the efficacy and improvement of understanding of physiology when the passive lecture is followed by a related practical activity. 250 students took a survey before and after a lecture on physiology of special senses. The same survey was offered to the students that participated in a practical exercise where they experienced the special senses with guidance from medical students from Virginia Tech Carilion School of Medicine and students from Jefferson College physician assistant program. The basic knowledge about the special senses was observed in 75% students before the lecture. Only 23% of students could respond the question about similarities between the special senses. 38% correctly responded to how the signal reached the brain. After the lecture the proportion of correct answer increased significantly (p<0.001) to 84%, 63% and 60% respectively, but a misconception regarding how the signal reached the brain increased (6% before to 16% after the lecture). After participating in the hands‐on activity the number of correct responses increased significantly (p<0.001) and the misconception was reduced (5%). In conclusion the lecture deepened the students understanding about the special senses but the hands‐on activity was more efficient to correct misconceptions. To better teach for a life longing learning and to prevent misconception it is necessary to apply interactive methodologies that reach a large number of students learning styles.
Computer‐driven Mechanical Model Enables Investigation of Complex Cardiovascular Concepts
Authors: Sweeney Terrence E
Published: June 2021

In further development of a mechanical model designed for teaching cardiovascular physiology (FASEB J. 23:633.4, 2009), we show the ability of the model to clarify to the student complex cardiovascular concepts. The fluid‐pumping apparatus is comprised of components that model cardiac and vascular elements and is controlled by a PowerLab (ADInstruments) system. Students can vary heart rate, systolic contraction strength and duration, arterial and venous compliance, pre‐ and post‐capillary resistance, the skeletal and thoracic pumps and vascular volume. Cardiac output and vascular pressures are measured directly and displayed continuously. The system enables data overlay and rapid parameter analysis from the simple to the complex. The model can both isolate individual control elements that are difficult to isolate in vivo, and integrate control elements that may be difficult to integrate ex vivo. For example, model data will show that the user can demonstrate that venular dilation during hyperemia is required to moderate increases in capillary pressure and improve skeletal muscle pump performance. Additionally, through visualization of the cardiac element and analysis of system parameters, the student can compare the consistently positive impact of the skeletal muscle pump on cardiac filling and performance to the cyclically reversing effect of the thoracic pump. Supported by The University of Scranton.
Role of cAMP‐PKA/CREB pathway in regulation of AQP 5 production in rat nasal epithelium
Authors: Zheng Ming , Wang Weiwei
Published: June 2021

Objectives

Aquaporin 5 (AQP5) is a water‐specific channel protein. In this study, we investigated the possible role of the cyclic adenosine monophosphate‐protein kinase A/cyclic adenosine monophosphate response element binding protein (cAMP‐PKA/CREB) pathway in the regulation of AQP5 in nasal epithelial cells.

Methods

Rat nasal epithelial cells were cultured and treated with the PKA inhibitor H89 or cAMP inducing medicine forskolin for 12 or 24 hours in vitro. AQP5 and phosphorylated CREB (p‐CREB) at serine133 (Ser133) were detected by immunocytochemistry, Western blotting or real‐time PCR. Experiments were repeated 10 times.

Results

After treatment with H89 for 12 or 24 hours, the number of cells positive for AQP5 and p‐CREB (Ser133) were decreased, p‐CREB (Ser133) and AQP5 protein decreased, and AQP5 mRNA decreased. After treatment with forskolin for 12 or 24 hours, the number of p‐CREB (Ser133) and AQP5 positive cells increased, p‐CREB (Ser133) and AQP5 protein increased, and AQP5 mRNA was increased.

Conclusion

Both H89 (PKA inhibitor) and forskolin (cAMP inducing medicine) regulate AQP5 production through the cAMP‐PKA/CREB pathway, which could influence the secretary function of the submucosal glands in nasal epithelium.
Association between TaqI vitamin D receptor (VDR) gene polymorphism and LDL‐C in Cuban Americans
Authors: Shaban Lamya , Zarini Gustavo G. , Exebio Joel C. , Shin Dong-Ho , Huffman Fatma G.
Published: June 2021

The study investigated the association between TaqI vitamin D receptor (VDR) polymorphism (rs 731236) and LDL cholesterol (LDL‐C) in Cuban Americans (CA). This cross‐sectional study included CA (n=253) with (n=111) and without (n=142) type 2 diabetes (T2D). Participants were recruited from multiple sources from Miami‐Dade and Broward Counties, Florida. TaqI VDR polymorphism was genotyped by the RFLP‐PCR method. Plasma LDL‐C was assayed by an enzymatic method. Analysis included descriptive statistics and analysis of variance (ANOVA). Participants without T2D had significantly higher LDL‐C levels than those with T2D (P=.015). ANOVA showed that after controlling for confounding variables, participants without T2D carrying the tt genotype had significantly higher LDL‐C levels compared to those carrying the genotypes TT (P=.006) and Tt (P=.028). Findings provide a better understanding of the relationship between TaqI polymorphism and LDL‐C in this minority group. A need for further exploration as to how TaqI VDR polymorphism (rs 731236) is associated with cardiovascular disease risk factors and diabetes status is recommended. Funding for this research was provided through an NIH/NIDDK sponsored grant.
Analysis of Study Logs in an Anatomy Learning Skills Course
Authors: Schutte Audra
Published: June 2021

At Indiana University, MSCI M100: Improving Learning Skills in Anatomy is a 1‐credit hour course which supplements Anatomy A215: Basic Human Anatomy. MSCI M100 was developed in 2010 to help Anat A215 students by teaching them effective approaches to learning anatomy and becoming better regulators of their learning process.

While time management is one component of regulation of learning many Anat A215 students utilize their study time inefficiently, as evidenced by the author. One MSCI M100 assignment is the study log (adapted from Classroom Assessment Techniques). Students complete a daily study log, three days prior to each of the four Anat A215 exams. Students record when and where they study, and rate their productivity during that time on a scale ranging from one to four (1=not productive, 4=very productive).

Analysis of the study logs was performed to see if students’ time management changed while enrolled in MSCI M100. Preliminary results indicate there is an improvement in students’ time management between studying for the first and second Anat A215 exams. Specifically, 2nd exam study logs showed students studied, fewer hours overall (p<.000), but logged significantly more productive hours (p<.000). Student feedback (via weekly blogs for M100) indicates study logs raised student awareness of how they utilize their time. These results also support the efficacy of MSCI M100 in improving student learning skills.

Grant Funding Source : NA
Epigenetic Regulation of the Class II Transactivator (CIITA) by the Polycomb Repressive Complex 2 (PRC2)
Authors: Boyd Nathaniel , Greer Susanna
Published: June 2021

Major Histocompatibility Class II (MHC II) molecules present extracellular peptides to T helper cells and are thus responsible for initiating, regulating, and terminating immune responses to foreign agents and tumors. Expression of MHC II is induced by the inflammatory cytokine interferon gamma (IFN‐γ) which induces transcription of the Class II Transactivator (CIITA), the master regulator of MHC II genes. Transcription of CIITA through the IFN‐γ inducible CIITA promoter IV is frequently suppressed in tumor cells through specific trimethylation of histone H3 lysine 27 (H3K27me3) by the histone methyltransferase Enhancer of Zeste Homolog 2 (EZH2). EZH2 is the catalytic subunit of the Polycomb Repressive Complex 2 (PRC2); however the mechanisms of recruitment of PRC2 to mammalian promoters remains unknown as proteins in the PRC2 complex contain no DNA binding domains. We identify here two EZH2 binding partners which bind DNA and regulate PRC2 recruitment to CIITApIV. We demonstrate that Yin Yang 1 (YY1), the homolog of the Drosophila PRC2 recruiting protein Pleiohomoeotic (Pho), and Jumonji domain containing protein 2 (JARID2), a member of histone demethylase family, both interact with EZH2. Upon IFN‐γ stimulation, YY1 dissociates from CIITA pIV while Jarid2 CIITA pIV binding increases, suggesting novel roles for each of these proteins in regulating inducible gene promoter dynamics. Research supported by grants from the American Cancer Society, the Georgia Cancer Coalition, and the Georgia Research Alliance (to S.F. Greer).
Structural insights into the function of FANCM‐mediated complexes
Authors: Saro Dorina , Sachpatzidis Aristidis , Zheng Xiao-Feng , Singh Thiyam Ramsing , Meetei Amom Ruhikanta , Xiong Yong , Sung Patrick
Published: June 2021

FANCM is a Fanconi anemia nuclear core complex protein required for the functional integrity of the FA‐BRCA pathway of DNA damage response and repair. FANCM is a highly conserved protein that belongs to the SF2 superfamily of helicases/translocases. Previously, we have reported the isolation and characterization of two novel histone‐fold‐containing FANCM‐associated proteins, MHF1 and MHF2. These proteins form a complex that is important for DNA damage‐induced monoubiquitination and foci formation of FANCD2, as well as chromatin localization of FA nuclear core complex proteins. Depletion of MHF complex resulted in elevated MMC‐induced chromosome aberrations, and sensitivity of cells to MMC and camptothecin. Biochemical characterization of the MHF complex showed that it assembles into a heterodimer that binds double stranded DNA and most importantly enhances the DNA branch migration activity of FANCM. We have carried out structural work to characterize the MHF complex by X‐ray crystallography. Based on the high resolution crystal structure of the MHF complex we have designed and tested mutants of MHF complex that are defective in DNA binding and FANCM interaction and showed that both protein‐protein interactions and DNA binding of MHF complex are needed for stimulation of FANCM activity.
Regulator of G Protein Signaling 6 (RGS6), A Novel Modulator of Doxorubicin‐induced Cardiotoxicity and Apoptosis
Authors: Maity Biswanath , Yang Jianqi , Huang Jie , Fisher Rory A
Published: June 2021

Doxorubicin (Dox) is an anthracycline chemotherapeutic agent that is among the most effective and widely used drugs for treatment of human cancers. Unfortunately, Dox induces life‐threatening cardiotoxicity, which is the major limitation to its use. The cellular mechanisms that underlie cardiac damage upon Dox treatment remain unclear, but are known to involve p53‐mediated apoptosis, loss of ventricular function and sinus tachycardia. Here, we show that RGS6, a protein indispensable for parasympathetic control of automaticity, is dramatically induced by Dox in atria and ventricles of mice and mediates subsequent activation of ATM, p53, and apoptosis. Indeed, hearts or isolated ventricular mycocytes from RGS6−/− mice are protected from Dox‐induced p53/ATM phosphorylation, Bax upregulation and cleavage of caspase‐3. RGS6 is required for Dox‐mediated reactive oxygen species production, essential for subsequent activation of DNA damage signaling. Participation of RGS6 in this pro‐apoptotic signaling cascade is independent of its canonical role as a G protein regulator. Unsurprisingly, mice lacking RGS6 are protected from Dox‐induced heart damage. These results effectively identify RGS6 as a critical mediator of Dox‐induced cardiotoxicity in vivo. Thus, RGS6 inhibition might represent a viable means to protect against heart damage in patients undergoing Dox chemotherapy.
Autophagy Stimulation Manipulates Burkholderia cenocepacia Infection in a Cystic Fibrosis Mouse Model
Authors: abdulrahman basant ahmed , khweek Arwa Abu , caution kyle , akhter anwari , Tazi Mia , kopp benjamin , amer amal
Published: June 2021

Cystic fibrosis (CF) is the most common inherited lethal disease of Caucasians. It is caused by mutations in the cystic fibrosis transmembrane conductance regulator which results in multi organ dysfunction. The most common mutation is the ΔF508. Infection by Burkholderia cenocepacia (BC) is a particularly lethal threat to CF patients because it causes severe and persistent lung inflammation. Autophagy is a cell survival mechanism in which, an autophagosome, engulfs non‐functional organelles then delivers them to the lysosome for degradation. Our Immunofluorescence (IF) data demonstrated that autophagy is defective in ΔF508 macrophages. The ubiquitin binding adaptor protein p62 is needed to deliver several types of ubiquitinated cargoes to the autophagosomes. Our IF and Colony Forming Unite assay in wild type (WT) macrophages showed that when P62 is down regulated using siRNA, the bacterial survival increased. Interestingly, when we over expressed p62, we observed a significant decrease in the bacterial survival. The ΔF508 macrophages did not respond similarly to WT showing a problem in the delivery of BC to the autophagosomes. Here, we demonstrate that an autophagy inducer, Rapamycin, markedly decreases BC infection by enhancing the clearance of BC. Therefore, this study reveals that autophagy is a novel target for new drug development for CF patients to control BC infection and accompanying inflammation.
Cloning and characterization of EF‐Tu and EF‐Ts from Pseudomonas aeruginosa
Authors: Palmer Stephanie , Rangle Edna , Montalvo Albert , Mugica Diego , Bullard James
Published: June 2021

Introduction

Elongation factors (EF) facilitate many steps in the process of the biosynthesis of proteins. EF‐Tu in particular plays a central role in the GTP‐dependent placement of aminoacylated tRNA at the A site on the ribosome. EF‐Ts acts by catalyzing the change of EF‐Tu from a GDP‐bound inactive state to a GTP‐bound active state.

Results

Analysis of both P. aeruginosa EF‐Tu and EF‐Ts showed they are 84% and 55% identical to their E. coli counterparts, respectively. Both P. aeruginosa EF‐Tu and EF‐Ts were cloned and over‐expressed in E. coli and purified to greater than 95% homogeneity. EF‐Tu was shown to be active when assayed for activity using the GDP exchange assay. Kinetic parameters for the interaction of EF‐Tu with its substrate GDP were calculated to be: KM = 33 μM, Vmax = 0.125 μM/min, kcat = 1.65 × 10−3 sec−1. The specific activity of EF‐Tu was calculated to be 45 μM min−1 per mg of EF‐Tu. At a 1:20 ratio of EF‐Ts:EF‐Tu, EF‐Ts stimulated the exchange of GDP by EF‐Tu 10‐fold.

Conclusion

EF‐Tu and EF‐Ts identified in P. aeruginosa were cloned, expressed and purified and shown to be functional in GDP exchange assays suggesting that each is functional in protein synthesis.
Inhibition of PC1/3 and PC2 by 2,5‐dideoxystreptamine derivatives
Authors: Vivoli Mirella , Caulfield Thomas R. , Martínez-Mayorga Karina , Johnson Alan T. , Jiao Guan-Sheng , Lindberg Iris
Published: June 2021

The prohormone convertases PC1/3 and PC2 are eukaryotic serine proteases involved in proteolytic maturation of peptide hormone precursors and implicated in a variety of pathological conditions, including obesity, diabetes, and neurodegenerative diseases. In this work, we screened 45 compounds obtained by derivatization of a 2,5‐dideoxystreptamine scaffold. We found four promising PC1/3 competitive inhibitors and three PC2 inhibitors exhibiting various inhibition mechanisms with sub‐ and low micromolar inhibitory potency against a fluorogenic substrate. Low micromolar concentrations of certain compounds blocked the processing of the physiological substrate proglucagon. Interestingly, we identified compounds that were able to stimulate both 87 kDa PC1/3 and PC2 activity. Molecular modeling studies revealed interactions of the PC1/3 inhibitors with the active site that suggest structural modifications to further enhance potency. In support of experimental data suggesting that PC2 inhibition likely occurs via an allosteric mechanism, we identified several possible allosteric binding sites using computational searches.

This work was supported by DA050854 to Iris Lindberg.
Palmitate and ceramide induce human monocytic cell toxicity towards neuronal cells
Authors: Little Jonathan Peter , Madeira Jocelyn M , Klegeris Andis
Published: June 2021

Obesity is associated with increased risk of Alzheimer's disease (AD) but the mechanisms linking these conditions are poorly understood. Levels of saturated fatty acids and their metabolites (e.g., ceramides) are elevated in obesity and can induce inflammation in peripheral immune cells. Brain inflammation, which is mediated by activation of glial cells, is increased in AD. Our aim was to determine if elevated levels of saturated fatty acids or ceramides could induce inflammation and cause neurotoxin secretion from glial cells. Human THP‐1 monocytes and U‐ 373MG astrocytoma cell lines were used to model microglia and astroctyes, respectively. Incubation of THP‐1 cells, but not U‐ 373MG cells, with 125–250 μM palmitate for 48 h increased secretion of the pro‐inflammatory cytokines MCP‐1 and IL‐8 (p<0.001). Transfer of conditioned media from palmitate‐treated THP‐1 cells to SH‐SY5Y neuroblastoma cells caused a significant increase in neuronal cell death (p<0.001). Palmitate had no effect on U‐373 MG cell‐mediated neurotoxicity (p>0.05). Similar to palmitate, treatment of THP‐1 cells with C2 ceramide significantly increased neurotoxicity (p<0.001). These results indicate that increased circulating levels of palmitate or ceramides in obesity may contribute to neurodegeneration through inflammatory pathways involving microglia. Supported by the NSERC
Deficiency of S‐nitrosoglutathione reductase compromises expression of the ER stress‐related proteins in LPS‐treated liver
Authors: Ozawa Kentaro , Tsumoto Hiroki , Tsujimoto Gozoh
Published: June 2021

Nitric oxide (NO) promotes tumor formation by inflicting damage to DNA, which could lead to mutations or other genomic rearrangements. Recently, we discovered that S‐nitrosoglutathione reductase (GSNOR) deficient mice (GSNOR KO) are prone to spontaneous tumor formation, suggesting that S‐nitrosylation might be involved in tumorigenesis. To clarify the mechanism of tumorigenesis in GSNOR KO, we compared the protein expression profiles of wild‐type and GSNOR KO following LPS‐treatment. We identified 19 up‐regulated and 19 down‐regulated proteins in the GSNOR KO using Two‐dimensional difference gel electrophoresis analysis and following mass spectrometry. Further immunoblot analysis revealed that several members of the protein disulfide‐isomerase (PDI) family were expressed at lower levels in the GSNOR KO and that there were also differences in the expression patterns of the ER chaperone isoforms. These results suggest that S‐nitrosylation could potentially regulate the expression of many proteins, including those of the chaperones and folding catalysts in the ER, under inflammatory conditions some of which may promote carcinogenesis induced by NO.
Estrogen Receptor Activation of CaM Kinase I and ERK
Authors: Todd Mikayla S. , Schmitt John M.
Published: June 2021

Cell growth and development is regulated by the cell signaling second messenger, calcium, that regulates key enzymes and genes in cells including the Calcium/Calmodulin‐dependent protein kinases (CaM Ks) and their downstream target ERK. In particular, CaM KK and its substrate CaM KI promote ERK and Elk‐1 activation in MCF‐7 breast cancer cells. Estrogen (E2) may utilize CaM KK and ERK to promote breast cancer cell proliferation, however it is unclear which E2 receptor promotes cell proliferation through CaM Ks. Estrogen is a ligand for estrogen receptors (ER) of the alpha(α) and beta(β) forms, as well as G protein‐coupled receptor 30 (GPR30). E2 has recently been shown to activate ERα and GPR30 leading to ERK activation however transgenic mice and cells that express only a membrane localized form of ERα are capable of activating ERK. The precise ER that is responsible for activating CaM KI and ERK in MCF‐7 breast cancer cells remains unknown. Our goal was to evaluate which estrogen receptor controls the calcium‐mediated activation of CaM KI and ERK in MCF‐7 cells. E2 stimulation of MCF‐7 cells potently stimulated both CaM KI and ERK phosphorylation, an effect that was blocked by the selective ERα inhibitor, MPP. MPP did not block epidermal growth factor activation of ERK. In contrast, the GPR30 antagonist, G15, did not block E2 stimulation of CaM KI or ERK. These results suggest that E2 is capable of activating CaM KI and ERK through ERα but not GPR30 in MCF‐7 cells.
Expression profiles of genes coding for oil biosynthesis in developing Tung seeds
Authors: Cao Heping , Shockey Jay M. , Klasson Kjell T.
Published: June 2021

Understanding plant oil biosynthesis will help to create new oilseed crops with value‐added properties. Our project focuses on engineering oilseed crops to produce industrial oils, such as tung (Vernicia fordii) oil, which are rich in novel fatty acids that impart valuable properties to their respective oils. We have identified a number of genes for tung oil biosynthesis, but less is known about the expression patterns of the genes. In this study, qPCR approach was used to explore the expression profiles of > 40 genes potentially related to oil biosynthesis in developing tung seeds. Total RNAs were extracted from 11 stages of three tung trees. RNA concentrations and integrity were determined before the cDNAs were synthesized from the total RNAs. qPCR transcript profiling during tung seed development revealed that most genes studied fell into one of two classes. The first set of genes were significantly upregulated during the middle to late stages of seed development; whereas the second set of genes were transcribed at higher levels in younger seeds, and declined as development progressed. These results suggest that genes in the first group potentially play important roles in tung oil biosynthesis and that those in the second group are probably more important in the synthesis of lipids other than triacylglycerols. This study should provide useful information for selecting targets for oil improvement in transgenic plants.
Development of Analytical Methods for Determination of Peptide Concentration
Authors: Lang Ewa , Rupprecht Kevin , Fishpaugh Jeffrey
Published: June 2021

Measuring the concentration of peptides can be challenging. Amino acid sequence, solubility and the folding of the peptide can influence measurement. A280 measurement requires that chromophores trytophan and tyrosine be in the sequence. Lyophilized peptides contain water and salts so concentration determined by weight alone is imprecise. Amino acid analysis is the default method for measuring peptide concentration, but it is very labor intensive. We evaluated two microtiter plate methods for potential use in peptide quantitation. Pierce BCA Protein Assay (bicinchoninic acid) combines the reduction of Cu2+ to Cu1+ by protein in an alkaline medium with the selective colorimetric detection of the (Cu1+) by bicinchoninic acid. Peptides of three or more residues form the reaction, which is measured at 562 nm. LavaPep™ Protein & Peptide quantification kit depends on a small, naturally‐occurring fluorescent compound, epicocconone, that reversibly binds to lysine, arginine, and histidine residues to yield an intensely red‐fluorescent product (Ex/Em: 540/620 nm). Various peptides were used to generate standard curves to characterize the affect of standards used on the concentration. Results presented will demonstrate the relative robustness, accuracy and precision of the two methods.
TopoCell – An image analysis tool to study intracellular topography
Authors: Fachada Vasco Rui Carvalho Pereira Fernandes , Fachada Nuno , Turpeinen Tuomas , Rahkila Paavo , Rosa Agostinho , Kainulainen Heikki
Published: June 2021

We aim to build an open source bioimage informatics tool capable of analyzing 2D and 3D topography (absolute and relative localization, shape, size and intensity) of distinct types of intracellular particles (such as organelles, proteins, lipids and other detectable biomolecules) in different types of cells in a high‐throughput fashion.

To achieve this, TopoCell is primarily designed to run fluorescence microscope images, where all possible channels imaged are used to semi‐automatically detect different particles, define cell type and segment cells. To each detected particle, a coordinate, intensity, volume, and sphericity value are attributed. Cells are detected through a plasmalemma marker, where volume and coordinate values are generated after the segmentation step. Numerous cell types can be defined by the user accordingly to the density of given intracellular particles. By controlling a dedicated filter, the user can select which cells and particles to consider in the analysis according to their detected values, including relative distance between two or more types of particles. Visualization of results and statistical analysis are also available.

TopoCell allows us to comprehensively measure in still microscope images, the distribution of intracellular phenomena which otherwise would be negligible to the naked eye, as for instance translocation or particle association.
Investigations of nanoparticle toxicity and uptake of Cerium oxide and Titanium dioxide in Arabidopsis thaliana (L.)
Authors: TUMBURU LAXMINATH , Andersen Christian , Betts Julia N , Johnson Mark G , King George A , Plocher Milt , Reichman Jay R , Rygiewicz Paul T
Published: June 2021

The emergence of nanotechnology and incorporation of nanoparticles in consumer products necessitates risk assessment from an environmental and health safety standpoint. To date, very few studies have examined nanoparticle effects on terrestrial species, especially plants. In this study, we examined responses at the molecular level when A. thaliana was exposed to two environmentally relevant nanoparticles (NPs)‐i) titanium dioxide (TiO2), and ii) cerium oxide (CeO2) during different life stages. Particle suspensions were determined to be in the <100 nm diameter range. A. thaliana plants were exposed to NPs (concentration: 500 mg/L) starting in the seed stage, to weekly suspensions of these mixtures. Following exposure, three different tissues (cotyledons, roots, leaves at different developmental stages were harvested, and RNA was extracted. With the use of Affymetrix Arabidopsis ATH1 Genechip, a comparison of RNA expression profiles was made between control and NP‐treated A. thaliana tissues. The results varied with developmental stage, and showed differences in gene up‐ and down‐regulation. Uptake studies involving transmission electron microscopy (TEM) observations are underway. The findings of this research will help to elucidate mechanisms of action associated with NP‐induced toxicity and may provide primary screening information for terrestrial ecological risk assessments.
Evidence for the Presence of Phosphate Groups on the Two Major Proteins of the Hamster Sperm Acrosomal Matrix and Their Potential Role in Acrosomal Hydrolase Binding Activities
Authors: Alvarez George , Buchanan Teresa L , Raychoudhury Samir , Nagdas Subir K
Published: June 2021

We previously isolated an acrosomal fraction, termed ALM, from hamster cauda epididymal spermatozoa which contains specific domains of the acrosomal matrix and an adherent detergent‐insoluble complex, termed the acrosomal lamina; which is derived from the outer acrosomal membrane. By SDS‐PAGE, the ALM fraction exhibited two major polypeptides of Mr=29,000 (ALM29) and Mr=20,000 (ALM 20). ALM complex binds both acrosin and N‐acetylglucosaminidase (NAGA) in a dose‐dependent manner. The present study was undertaken to examine the presence of phosphate groups on both ALM29 and ALM20 polypeptides and to identify the binding competency of dephosphorylated ALM complex to acrosomal hydrolases. Our diagonal gel electrophoresis data reveals that both ALM29 and ALM20 polypeptides are not joined by disulfide bridge(s). Both ALM29 and ALM20 polypeptides showed a reduction in size (~3 kDa) by alkaline phosphatase treatment. A sedimentation assay was employed to determine whether alkaline phosphate treated ALM possesses acrosin and NAGA binding sites. There was a ~50% reduction in binding between the dephosphorylated ALM complex and acrosin/NAGA. Our studies conclude that both ALM29 and ALM20 polypeptides are phosphorylated and that the ALM complex‐acrosomal hydrolase interactions require the phosphorylation of ALM polypeptides. Supported by NIH/NIGMS/1SC3GM096875‐01/FSU McNair and Graduate School Program.
New Cell Biology Assays Using a Stabilized Luciferase Exhibiting Red Luminescence
Authors: Jiang Ying , Naleway John Joseph
Published: June 2021

Bioluminescence is the light produced in certain organisms as a result of luciferase enzyme mediated oxidation of a luciferin substrate. This oxidation is linearly dependent on the presence of adenosine tri‐phosphate (ATP). We have recently developed a codon‐optimized luciferase based upon the gene sequence of the natural luciferase isolated from Luciola cruciata (Japanese firefly). This new gene codes for a modified amino acid sequence which exhibits improved expression levels in mammalian cells, red‐shifted emission wavelengths (560 to 619 nm) as well as improved thermal stability, while still utilizing D‐luciferin and ATP as substrates.

This new recombinant enzyme was purified to 98% homogeneity from both transfected E. coli bacteria and HEK 293 mammalian cells using Ni+2‐affinity chromatography as the His6‐tagged protein. Enzymes isolated from either source were found to have equivalent activity in ATP dependent assays. The purified recombinant luciferase was used to develop a number of ultrasensitive biological assays useful in measuring cell number, cell viability as well as the cytotoxicity effects of several chemotherapeutic drugs (Etoposide and Doxorubicin) when added to murine NIH3T3 cells in culture. We also utilized the purified enzyme to sensitively detect bacterial cell number down to 1 × 102 cells and adapted the methods for use in high‐throughput antibiotic screening protocols. By cloning the luciferase downstream of TFkB responsive elements, the activation of TF‐κB (with stimulation by TNFα) in human HEK 293 cells could be quantitated by directly monitoring the increase in luminescence from activation of luciferase expression. Finally, since this new luciferase has a red‐shift in light emission, it was utilized in multiplexed assay conditions with other green (firefly) or blue‐emitting (Renilla) luciferases in a number of coupled biological assays. Moreover, application of this new luciferase for use in in vivo imaging was explored.
Community‐based design and national testing of an assessment tool to measure student understanding of enzyme kinetics in undergraduate biochemistry
Authors: Loertscher Jennifer , Minderhout Vicky , Lewis Jennifer E , Villafane Sachel M
Published: June 2021

Most biochemists would agree that students should understand the fundamentals of enzyme kinetics and inhibition by the end of an introductory undergraduate biochemistry course. Assessment instruments can be used to determine whether students understand this important concept at the end of a course and can help guide changes in instruction to improve student learning in this area. As part of an NSF‐funded project to test and disseminate active learning materials for biochemistry on the national level, we designed a multi‐part question to test student understanding of enzyme kinetics through application of the concepts in a new context. The new context, a saturable binding event, was used in order to ensure that students demonstrated meaningful understanding of key concepts rather than memorization. A community of over thirty expert biochemistry instructors from a variety of institutions participated in choosing the question topic, developing and revising the question and designing a grading rubric. The question consists of eight parts, three of which allow for student free response. The experts agreed that five of the eight question parts assessed skills in the higher levels of Bloom's taxonomy including analysis, synthesis and evaluation. Results from use of the instrument at several institutions nationwide will be reported.
Galactooligosaccharides: effects on calcium absorption and gut microflora in young premenarcheal girls
Authors: Whisner Corrie Marie , Martin Berdine R , Clavijo Andrea , Nakatsu Cindy H , McCabe George P , McCabe Linda D , van den Heuvel Ellen GHM , Schoterman Margriet HC , Weaver Connie M
Published: June 2021

Adolescence presents an opportunity to influence peak bone mass with prebiotic agents like galactooligosaccharides (GOS) that increase calcium absorption in the large intestine in animal models and adults. The objectives of this study were 1) to investigate the dose response of GOS supplementation on calcium absorption during growth and 2) to assess total colonic microbiota and bifidobacteria to better understand the mechanism of action. Using a randomized, double‐blind, crossover design, thirty‐one 10–13 year old girls consumed smoothie drinks enriched with 0, 2.5 or 5 g GOS twice a day for 3 weeks. The 3‐week periods were separated by 2‐week washout periods. Fractional calcium absorption was determined, using dual stable isotope methods, as the ratio of oral and intravenous tracers in urine over 48 hours. DNA was extracted from fecal samples to quantify total bacteria and bifidobacteria using qPCR. Preliminary results show that GOS increased fractional calcium absorption. Bifidobacteria as a proportion of total bacteria was also significantly greater after consumption of GOS. Results suggest that GOS consumption increases fractional calcium absorption in young girls through possible mechanisms in the large intestine.

Grant Funding Source : FrieslandCampina Domo
Individualized counseling for a Healthy People 2010 diet results in surpassed fruit and vegetable intakes
Authors: Sidahmed Elkhansa , Cornellier Maria , Askew Leah , McCoy Alexandra , Rapai Mary , Sen Ananda , Djuric Zora
Published: June 2021

The objective of the study was to evaluate adherence to two different diets that were implemented in a colon cancer prevention study. A total of 120 subjects were randomized to 6 months of individualized counseling for a Healthy Eating or a Mediterranean diet. The counseling was implemented starting with one face to face meeting with a registered dietitian followed by telephone counseling at the same intervals in both arms. A checklist was developed for assistance in meeting goals in both arms. The percentage of goals met in the two groups was similar with 87% of goals met in the Healthy Arm and 82% in the Mediterranean arm. Although the fruit and vegetable goals were lower in the Healthy arm, these goals were surpassed resulting in statistically similar fruit and vegetable intakes in the two groups (7.8 vs 8.1 servings/day). Carotenoid intakes and blood carotenoid levels also increased to similar extents in the two arms. These results indicate that individualized counseling for meeting Healthy People 2010 goals can result in better than expected compliance, resulting in carotenoid intakes similar to what can be achieved with a Mediterranean diet. Supported by grant NIH CA120381

Grant Funding Source : NCI
Don't walk this talk: public health discourse that disempowers
Authors: Porter Christine M.
Published: June 2021

How many of us have “targeted low‐income audiences,” “served at‐risk populations,” or “empowered consumers?” Using critical discourse analysis, this paper examines how public health messages position the people and “communities” with whom we work. Using examples from obesity prevention, I demonstrate how some language used in public health discursively disengages communities from creating their own health and reinforces existing marginalizations, particularly by discounting citizen agency and expertise. This paper does not propose “politically correct” speaking, or a list of words to avoid; it surfaces the power relations and values that our language reflects and reinforces. The paper closes with alternative and additional frames for public health nutrition that center the margins. For example, describing people as negotiators, actors, or citizens denotes much more power than reducing them to clients, consumers, or audiences. The talk we walk in public health nutrition should cast the public not as (for example) recipients of our services or targets of our interventions, but as powerful actors in creating individual and community health. No funding to report.
Nutritional knowledge and overweight or obesity in Mexican urban women of low socioeconomic level
Authors: Galindo-Gómez Carlos , Juárez-Martínez Liliana , Shamah-Levy Teresa , García-Guerra Armando , Ávila-Curiel Abelardo , Quiroz-Aguilar Marco Antonio
Published: June 2021

The objective was to identify the association between the knowledge about nutrition with obesity or overweight in 1,245 women from low income households in Mexico City. Data was obtained of an Urban Food and Nutrition Survey 2002 in Mexico City. An ordinal logistic regression model was used to estimate the association between overweight and obesity (OWOB), obesity (OB) and nutrition knowledge and select variables. The variables that related to OWBO were: correct (OR=2.00, p=0.017) or regular nutrition knowledge (OR=1.54, p=0.035), age of 30 yr old (OR=3.00, p<0.001), to be illiterate or have not finished the elementary school (OR=2.00, P=0.016), elementary and junior high school completed (OR=1.61, p=0.034), belonging to a medium ‐ low socioeconomic status (OR=1.90, p=0.003), and high fat consumption (OR=1.65, p=0.024). For OB were: age of 30 yr old (OR=2.42, p<0.001) and high fat consumption (OR=1.67, p=0.029). Approach to intervention strategies in the control and prevention of overweight and obesity should be considering the level of knowledge of nutrition of the population.
Dietary patterns and risk factors for glucose abnormalities in Korean adult population
Authors: Song SuJin , Paik Hee Young , Song YoonJu
Published: June 2021

The aim of this study was to identify dietary patterns associated with risk of insulin resistance (IR), impaired fasting glucose (IFG), and diabetes (DM) in Korean adult population. We analyzed data of 3,871 subjects from the 2007–2008 Korea National Health and Nutrition Examination Survey from 30 to 65 years of age with no treatment history of diabetes, hypertension, or dyslipidemia. IR was defined as the highest quintile of the homeostasis model assessment of insulin resistance (HOMA‐IR) scores. IFG was defined as fasting blood glucose of 100–125 mg/dL and DM as fasting blood glucose ≥ 126 mg/dL. Using factor analysis based on data from food‐frequency questionnaire, we identified five dietary patterns: 1) the many foods pattern, 2) the western pattern, 3) the mixed rice & beans pattern, 4) the white rice & kimchi pattern, and 5) the noodle & beverage pattern. HOMA‐IR significantly decreased across quintiles of mixed rice & beans pattern scores and noodle & beverage pattern scores were associated with higher fasting glucose after adjustment for all potential confounders. In multivariate logistic regression analysis, subjects in the highest quintile of mixed rice & beans pattern scores had lower odds ratios for IFG (OR: 0.83) and IR (OR: 0.77). Noodle & beverage pattern had a positive association with IFG. In conclusion, mixed rice & beans pattern was associated with lower risk of glucose abnormalities among Korean adults.
Korean middle school students’ recognition and consumption of fast food
Authors: Chang Kyung Ja , Kim Sung-Eun
Published: June 2021

The purpose of this study is to investigate recognition and consumption of fast food in Korean middle school students. This cross‐sectional study was conducted using a questionnaire and the subjects were 503 middle school students in Incheon metropolitan city, Korea. Data were analyzed with SPSS 12.0 program. In case of mother having a job, students were frequently eating more hamburgers (42.6%), drinking more carbonated beverages (72.0%) and consuming fast food 1–2 times a week (72.0%) compared to students whose mother did not have a job. As for the food items that are most frequently being consumed at fast food restaurants, hamburgers were being consumed most frequently by males (38.5%) and females (33.1%) according to gender. In case of French fries, females (27.5%) were consuming them significantly more frequently compared to males (17.9%). The reason for consuming fast food was in order of ‘good taste’ (31.6%), ‘quickness in consumption’ (23.1%), and the reason of ‘good taste’ was the highest for both genders. As for the frequency of consumption, 3–4 times a week (58.4%) was the highest followed by less than once a month (28.2%). As for price of fast food, the response of ‘reasonable’ (55.5%) and ‘expensive’ (39.8%) was high. As for sanitation condition, both males and females responded ‘average’ and as for services of fast food restaurants, females answered significantly higher service score compared to males(p<0.01). As for nutrition knowledge on fast food according to gender, females got significantly higher score (9.4) compared to males (7.5)(p<0.01). The students that frequently ate fast food had unbalanced diets and did not frequently eat vegetables and fruits. Therefore, nutrition education is necessary for those with high frequency of fast food consumption to practice balanced diets.
Nutrient intakes of Canadian children consuming breakfast and ready‐to‐eat cereal compared with non‐consumers
Authors: DiFrancesco Loretta , Barr Susan I. , Fulgoni Victor L.
Published: June 2021

Higher nutrient intakes are reported in breakfast consumers vs. non‐consumers, and in those who include ready‐to‐eat cereal (RTEC) at breakfast vs. those who do not. We examined these differences using Day 1 data from a nationally‐representative sample of children (age 4–18; n = 12,303) participating in the Canadian Community Health Survey Cycle 2.2 (2004). Overall, 90% of Canadian children ate breakfast, but consumption decreased with age, from 98% (age 4–8) to 82% (age 14–18). Breakfast consumers had higher (P ≤ 0.01) energy intakes than non‐consumers (mean ± SE: 2243 ± 15 vs. 2026 ± 44 kcal/d), but total fat intakes were lower (mean ± SE: 77.9 ± 0.3 vs. 83.6 ± 1.0 g/d). They also had higher energy‐adjusted intakes of carbohydrate (CHO), fibre, calcium (Ca), phosphorus (P), magnesium (Mg), iron (Fe), potassium (K) and vitamins A, B1, B2, B6, folate, C and D. Among breakfast consumers, about 37% included RTEC. Energy intakes of RTEC consumers and non‐consumers did not differ (mean ± SE: 2239 ± 26 vs. 2245 ± 19 kcal/d). RTEC consumers had lower total fat and cholesterol intakes, and higher intakes of CHO, fibre, Ca, P, Mg, Fe, K and vitamins B1, B2 and B6. Breakfast and RTEC breakfasts can make positive contributions to the nutrient intakes of Canadian children. (Supported by Kellogg Canada Inc.)