bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamic Regulation of TET2 by Phosphorylation and Dephosphorylation at Ser99
Authors: Kundu Anirban , Shelar Sandeep , Ghosh Arindam P. , Ballestas Marry P. , Kirkman Richard P. , Nam Hye-Young , Brinkley Garrett J. , Karki Suman , Mobley James A. , Varambally Sooryanarayana , Sudarshan Sunil
Published: April 2020

The ten‐eleven translocation enzyme was discovered when an 8‐year‐old boy diagnosed with acute myeloid leukemia (AML) containing the t(10;11)(q22;q23) in 2003. Three members, TET1, TET2 and TET3, belong to this dioxygenase family of enzymes. TET2 is widely implicated in cancer and in aging due to its role as a global epigenetic modifier catalyzing oxidation of 5‐methylcytosine (5mC) to 5‐hydroxymethylcytosine (5hmC), 5‐formylcytosine (5fC) and then 5‐carboxylcytosine (5caC) in 2‐oxogluterate (2‐OG) dependent manner. TET2 mutations are prevalent in malignancies including acute AML. Notably, most TET2 mutations in AML are monoallelic. Given the presence of a remaining wild type allele in these malignancies, characterizing the mechanism that regulate TET2 protein could have therapeutic significance.

Human TET2 is 2002 amino acids in length. The catalytic domain is located in the C‐terminal portion of the protein between amino acids 1129–1936. The C‐terminal catalytic domain comprises a cysteine rich domain (1129–1312 amino acids) followed by a double stranded β‐helix domain (DSBH) which make a compact fold to execute the catalytic activity. The C‐terminal part of TET2 has been reported capable of oxidizing 5‐mC to 5‐hmC independent of the N‐terminus. As such, the function of the N‐terminus of this large protein remains poorly characterized.

Here, we identify that several isoforms of the 14‐3‐3 family of proteins bind TET2. 14‐3‐3s bind TET2 when phosphorylated at serine 99 (S99). AMPK‐mediated phosphorylation at S99 promotes TET2 stability and increases global DNA 5‐hydroxymethylcytosine. 14‐3‐3s’ interaction with TET2 serves to protect S99 phosphorylation. Disruption of this interaction leads to both reduced TET2 phosphorylation and decreased protein stability. Furthermore, we identify that the protein phosphatase 2A (PP2A) can interact with TET2 and dephosphorylates S99. siRNA knockdown of PP2A (B‐alpha subunit) or pharmacologic inhibition of PP2A was able to significantly increase TET2 phosphorylation and protein level as well. Collectively, our study provides novel insights into the role of the N‐terminal domain in TET2 regulation. Moreover, they demonstrate the dynamic nature of TET2 protein regulation that could have therapeutic implications for disease states resulting from reduced TET2 levels and/or activity.

Support or Funding Information

The research reported in this article was supported by R01CA200653, I01BX002930, and a Mike Slive Foundation Grant (to S.S.) and in part by the UAB Comprehensive Cancer Center (P30CA013148).
Deletion of Kir5.1 Abolished the Inhibitory Effect of High Sodium (HS) Intake on the Basolateral K Channels in the DCT and Thiazide‐Sensitive Na‐Cl Cotransport (NCC)
Authors: Lin Daohong , Wu Peng , Zhang Dandan , Gao Zhong-Xiuzi , Duan Xinpeng , Xiao Yu
Published: April 2020

Background

We have previously demonstrated that dietary sodium intake regulated the basolateral Kir4.1/Kir5.1 channel activity in the distal convoluted tubule (DCT) such that low sodium (LS) intake stimulated whereas HS intake inhibited the Kir4.1/Kir5.1 in the DCT. The HS‐induced inhibition of basolateral Kir4.1/Kir5.1 in the DCT was essential for the HS‐induced inhibition of NCC because HS failed to inhibit NCC in the kidney‐specific Kir4.1 knockout mice. While Kir4.1 is responsible for permeating the basolateral K conductance, Kir5.1 is an important regulatory subunit of the basolateral K channel of the DCT. The aim of the present study is to explore the role of Kir5.1 in mediating the effect of HS on the basolateral K conductance of the DCT and NCC expression/activity.

Methods

We have used the whole‐cell recording technique to measure the basolateral K (Kir4.1) conductance and DCT membrane potential, employed immunoblotting to examine the expression of phosphor‐NCC (pNCC) and total NCC (tNCC), and conducted renal clearance method to measure thiazide‐sensitive renal sodium excretion in Kcnj16+/+ (WT) mice and Kcnj16−/− mice (Kir5.1 KO).

Results

We confirmed the previous finding that HS decreased the basolateral K conductance of the DCT, depolarized the DCT membrane, and inhibited the expression of pNCC and tNCC in WT mice. In contrast, LS intake increased the basolateral K conductance, hyperpolarized DCT membrane, and augmented expression of pNCC and tNCC in WT mice. The deletion of Kir5.1 not only increased the basolateral K (Kir4.1) conductance in the DCT and also abolished the inhibitory effect of HS on the basolateral Kir4.1 activity. Also, LS intake did not further stimulate the basolateral Kir4.1 of the DCT. The expression of pNCC and tNCC was upregulated in Kir5.1 KO mice in comparison to WT. Furthermore, neither HS nor LS had an effect on NCC expression in Kir5.1‐deficient mice, suggesting the role of Kir5.1 in mediating the inhibitory effect of HS on NCC expression. Renal clearance experiments also showed that the deletion of Kir5.1 not only augmented the thiazide‐induced natriuresis but also abolished the inhibitory effect of HS on the thiazide‐induced natriuresis in Kir5.1 KO mice.

Conclusion

Kir5.1 plays an important role in mediating the inhibitory effect of HS intake on the basolateral K channel activity in the DCT and NCC activity/expression.

Support or Funding Information

NIH‐DK115366
The development of peptide‐coated nanoparticles for targeting and drug delivery to cancer cells
Authors: Poursharifi Mina , dragulska Sylwia Anna , Chen Ying , Martignetti John , Mieszawska Aneta
Published: April 2020

Ovarian Cancer (OC) is a most lethal female malignancy worldwide, mostly due to the late‐stage diagnosis and high recurrence rate. Survival statistics remain stuck at ~30% for the past four decades highlighting the necessity to improve the early stage detection and therapeutic outcomes. Nanotherapy in OC may provide significant and unique benefits for drug efficacy as well as for reducing the therapeutically limiting adverse effects associated with current treatment regimens. Herein, we report a unique nanoparticle (NP)‐based platform of peptide‐stabilized poly (lactic‐co‐glycolic acid) (PLGA). The ultimate goal is NP‐mediated targeted delivery of anticancer therapy and contrast agents. Morphology and small size (~100 nm) of NPs provide them an ideal candidate for mechanistic studies. We present detailed imaging studies of NPs uptake by OC cells as well as NPs distribution within cellular organelles obtained with high resolution live cell imaging. The results show a preferential accumulation of NPs in mitochondria. In addition, the in vitro studies include the NP‐based delivery of anticancer agents, such as Pt(II) and mitochondria‐targeting lonidamine. This project demonstrates potential of peptide‐stabilized nanocarrier as an effective nanomedicine against cancer. The PLGA is FDA approved and the amino acids of the peptides have Generally Regarded as Safe (GRAS) status. The NP platform is completely biodegradable, and as we demonstrate biologically active.
The Oligomerization Status of IRE1α Kinase/RNase Determines Its Signaling Outputs Under Endoplasmic Reticulum Stress
Authors: Oakes Scott A.
Published: April 2020

Accumulation of unfolded proteins in the endoplasmic reticulum (ER) triggers signaling pathways called the unfolded protein response (UPR). Under remediable levels of ER stress, the adaptive UPR (A‐UPR) activates transcriptional and translational changes that restore homeostasis. However, under irremediably high ER stress, these adaptive measures fail and the signaling pathways instead trigger programmed cell death—referred to as the terminal UPR (T‐UPR). We discovered that IRE1α—an ER transmembrane bifunctional kinase/endoribonuclease (RNase)—is a critical life‐death switch that employs kinase auto‐phosphorylation levels as a mechanism to control its filamentous status and toggle between an A‐UPR or a T‐UPR depending on the level of upstream ER stress. Remediable ER stress causes low‐level kinase auto‐phosphorylation and dimerization that restricts IRE1α’s RNase activity to excising an intron in XBP1 mRNA, which upon re‐ligation and translation of spliced XBP1 produces the homeostatic transcription factor XBP1s (s=spliced) that then upregulates genes encoding ER protein‐folding and quality control components. However, sustained ER stress induces high‐level kinase autophosphorylation to promote higher‐order IRE1α oligomerization; under these conditions, IRE1α RNase hyperactivation leads to massive degradation of ER‐localized mRNA and T‐UPR events including: (1) loss of differentiated cell identity, (2) local sterile inflammation, and (3) programmed cell death through pyroptosis and apoptosis. We have recent insights into how the oligomerization status and signaling outputs of IRE1α contribute to cancer.

Support or Funding Information

The work was supported by grants: American Cancer Society Research Scholar Award; Harrington Discovery Institute Scholar‐Innovator Award; UCSF and Onyx Oncology Innovation Alliance; American Association of Cancer Research; R01CA219815; R01EY027810.
Pembrolizumab treatment increases chemotaxis of cytotoxic T cells in head and neck cancer patients within the adenosine‐rich tumor microenvironment
Authors: Newton Hannah S. , Chimote Ameet A. , Gawali Vaibhavkumar S. , Palackdharry Sarah , Hildeman David , Janssen Edith , Wise-Draper Trisha , Conforti Laura
Published: April 2020

Head and neck cancer (HNC) is the sixth most common cancer worldwide with a five‐year survival rate of 50%. Immunotherapy has been emerging as a promising treatment modality as conventional treatment strategies (surgery, chemotherapy and radiation) have limited efficacy and cause debilitating co‐morbidities. Immunotherapy harnesses the patient’s own immune system to fight disease. Targeting the immune system is crucial because immune cell (specifically cytotoxic T cell) infiltration into the tumor microenvironment (TME) and subsequent cytotoxicity is critical for tumor elimination. Pembrolizumab, an anti‐PD‐1 (programmed cell death‐1 receptor) antibody, is an immunotherapy currently approved for metastatic or recurrent unresectable HNC and is in clinical trials for curative intent. While there have been promising long‐lasting clinical responses to Pembrolizumab, many patients still do not respond. Thus, it is critical to understand the mechanism of action of Pembrolizumab in order to reduce treatment resistance. Moreover, it has been shown that Pembrolizumab increases cytotoxic T cell density in the TME; nevertheless, the underlying mechanism is not fully understood. However, it is known that there is an increased accumulation of adenosine (Ado), an anti‐inflammatory purine nucleoside, in the TME of HNC patients, which inhibits T cell chemotaxis by blocking the function of the potassium ion channel, KCa3.1. Therefore, to elucidate possible effects of Pembrolizumab on T cell chemotaxis, we studied KCa3.1 activity and subsequent chemotaxis in CD8+ peripheral blood T cells (PBTs) of naïve (never treated) HNC patients before/after Pembrolizumab treatment (n=25) via electrophysiological and 3D chemotaxis experiments. Furthermore, Pembrolizumab treated patients were categorized as responders (R) or non‐responders (NR) based on treatment response (i.e., reduction in tumor viability with a cutoff of ≥20% for R). We observed that Pembrolizumab increased the KCa3.1 activity in post‐treatment patient PBTs by 29.8% (p=0.026). Because we observed an increase in KCa3.1 activity in treated patients, we next assessed whether there was an effect on the chemotaxis of PBTs in presence of Ado. While Ado inhibited the chemotactic ability in treatment naïve PBTs by 53.9% (p<0.001, n=22), we determined that post‐treatment R PBTs were not inhibited by Ado (p = 0.142, n=11) whereas post‐treatment NR PBTs were inhibited by Ado (p = 0.005, n=10). These data support a role of ion channels and chemotaxis in patient response to Pembrolizumab and could lead to better understanding of the effect of Pembrolizumab in immune cell infiltration into the tumor and, ultimately, its mechanism of action/resistance.

Support or Funding Information

DoD CA160714; T32CA117846; 2R01 CA95286
Pharmacologic Effects of a Synthetic Macamide Analogue (ZYPC) on A549 and HeLa Cells
Authors: Lam Melanie , Rondón-Ortiz Alejandro Nico , Martínez-Málaga Jimena Alexandra , Gonzales-Urday Ana Lucia , Pino-Figueroa Alejandro
Published: April 2020

Lepidium meyenii L., known as the Maca plant, is acclaimed by the scientific community for its various health benefits in traditional medicine, including prevention of gastritis, depression, and cancer, as well as increasing fertility and improving sexual performance. Macamides (N‐benzylamides) are secondary metabolites derived from Maca that have demonstrated neuroprotective, anti‐oxidative, anti‐inflammatory, endocannabinoid system modulatory, and mild anti‐diabetic properties. The present study investigates the pharmacologic effect of N‐(8Z‐heptadecen‐1‐yl)‐O‐(3‐pyridylmethyl) carbamate (ZYPC), a synthetic macamide analog, on A549 non‐small cell lung cancer and HeLa cervical cancer cells. Both cell lines were treated with different concentrations of ZYPC (1–100 μM). After 24 h, cell viability was measured using a 3‐(4, 5‐dimethylthiazol‐2‐yl)‐5‐(3‐carboxymethoxyphenyl)‐2‐(4‐sulfophenyl)‐2H‐tetrazolium (MTS) colorimetric assay. ZYPC was found to significantly reduce cell viability in a dose‐dependent manner. The half maximal inhibitory concentration was calculated, using dose‐response non‐linear regression, as 38.94 μM for A549 cells and 38.35 μM for HeLa cells. Subsequently, a caspase 3 colorimetric assay was executed to study ZYPC‐induced reduction of cell viability through activation of apoptosis. HeLa cells were treated with 0, 30, 50, and 100 μM ZYPC and cell lysates were collected after 5 h incubation. A reagent containing DEVD‐pNA, a substrate for caspase 3, was added to the lysates, and incubated for 2 h at 37°C. Unexpectedly, ZYPC was found to have no effect on caspase 3 activity. Then, the involvement of a pro‐survival mechanism was assessed through AKT and phospho‐AKT (p‐AKT) chemiluminescent western blotting. Results obtained from treatment with 0, 10, 30, and 50 μM ZYPC for 24 h were inconclusive for total AKT and p‐AKT levels in both cell lines. Preliminary western blots of poly (ADP‐ribose) polymerase (PARP), a DNA repair enzyme, and its inactive cleaved form indicated constant expression levels of both proteins in A549 cells. Interestingly, there was an apparent increase in cleaved PARP in HeLa cells, which may suggest late‐stage apoptosis. However, further studies are needed to better elucidate the mechanism of anti‐cancer activity exhibited by ZYPC.

Support or Funding Information

MCPHS University Summer Undergraduate Research Fellowship Program
Balancing Parts: Integrating Uncertainty/Ambiguity into Anatomy Education
Authors: Lazarus Michelle D. , Rees Charlotte , Stephens Georgina
Published: April 2020

Tolerance of Uncertainty/Ambiguity (ToU/A) is a construct describing individuals’ responses to ambiguous stimuli. ToA is recognised as impacting healthcare outcomes, however, little is known about whether education may impact students’ ToA, and if so, how we can develop pedagogy to improve students’ ToU/A. Framework analysis of longitudinal qualitative data, collected via online discussions and semi‐structured interviews from 207 undergraduate medical students across three semesters of anatomy, identified anatomy pedagogy which either fostered or hindered students’ ToU/A. Anatomy curricula which integrates opportunities for students to distribute responsibility for knowledge (i.e. working in teams) and/or provide anonymity for those conveying knowledge (i.e. online anonymous discussion forums) appear to foster students’ ToU/A, while didactic educator approaches (suggesting that knowledge is finite and knowable) appear to hinder students’ ToU/A. Thematic analysis across time‐points suggest that active learning approaches can be effective stimuli for students’ ToU/A development. Active learning approaches within our study balanced communication of anatomy discipline knowledge with clinical applications, so that the context of knowledge became the stimulus of uncertainty. Exemplar anatomy curricula fostering ToU/A include “choose your own adventure” chest pain team‐based learning, where students elect which patient information they receive and in which order, engaging anatomical knowledge through the “adventure”. Another approach is using short clinical case scenarios wherein more than one underlying anatomical deficit may explain symptoms described in the scenario. Despite evidence that students develop ToA/U in their cognition and behaviour (e.g. through acknowledgement of the clinical importance of ToU/ToA, and perseverance when faced with ambiguous/unknown knowledge), student talk in this study suggests that emotionally, students remain negative about ToU/A (e.g. feeling overwhelmed, fearing errors). Together our data and examples illustrate that foundational science knowledge education can be balanced with effective student development of ToU/A, but student feedback may remain negative; an important consideration in evaluating effective curricula. Future work will focus on whether students’ negative emotions change with time and clinical exposure.

Support or Funding Information

Research supported by Monash Education Academy, Faculty of Medicine, Nursing and Health Sciences (Monash University) and a Medical Education Travel Fellowship
Correlation of Parkinson’s diseases with Vitamin D
Authors: PATEL MEET
Published: April 2020

Parkinson's disease (PD) is the neurodegenerative disorder which leads to progressive deterioration of motor function due to loss of dopamine‐producing brain cells in elderly people. Exposure to non‐genomic and genomic factors play a major role in the etiology and progression of Parkinson’s Disease. Genetic research has led to the identification of several monogenic forms of the disorder and non‐genetics studies examining polymorphism of the vitamin D receptor show high risk of Parkinson’s disease. An important relation between low serum vitamin D and Parkinson’s disease has been revealed, which suggest that high level of vitamin D may decrease a person’s risk of developing Parkinson’s disease. Calcitriol (1,25‐dihydroxyvitamin D3, a type of Vitamin D is highly active metabolite with an appropriate receptor in the central nervous system that can be involved in decreasing neurodegenerative disorder. In this review, the authors attempt relates the indirect relation of vitamin D with Parkinson’s disease through genomic and non‐genomic factors.
Whitening Effect of Hydrothermal Extracts of Phragmites Communis and its Active Compound, p‐coumaric acid
Authors: So Gyeongseop , Ha Chang Woo , Jang Sohee , Kim Sung Hyeok , Kim Youn Kyu , Lim Hyosun , Sohn Eun-Hwa
Published: April 2020

Reed (Phragmites communis) is a perennial grass forming extensive colonies inwetland. The study aimed to compare the antioxidant and whitening effects of roots of P. communis extracts (RPC) by three different extraction processes, using 100% EtOH, 70% EtOH and hydrothermal fluid methods. The amounts of total phenolic compound(TPC), total flavonoid contents(TFC), and p‐coumaric acid (pCA), a major active constituent, were evaluated. Melanogenesis‐related factors such as tyrosinase, L‐DOPA oxidase, and DPPH radical scavenging activities by all three RPC were also measured. Among them, hydrothermal method of RPC(h‐RPC) showed the best effect in the contents of TPC and TFC, DPPH scavenging activity, tyrosinase‐ and L‐DOPA oxidase inhibitory activities. Highest pCA content was also detected from h‐RPC (3.05 ppm). Finally, we confirmed that the color of the cell pellet and the amount of melanin synthesis were significantly reduced in the presence of h‐RPC and pCA in α‐melanin cell stimulating hormoneinduced B16F10 cells. The anti‐melanogenic potential of h‐RPC and pCA will provide an insight into developing a new skin whitening product.

Support or Funding Information

This research was a part of the project entitled "Development of multi‐functional cosmetic materials derived from halophyte", funded by the Ministry of Oceans and Fisheries, Korea.
Comparing in vitro and in vivo Bicistronic Design Protein Production
Authors: Gomez Paul , McKlin Sam , Chen Queena , Mazariegos Christopher , Zhou John , Seligmann Haley , Bashaw Benjamin , Peterson Andy , Shildknecht Kaitlyn , Tauchen Ali , Tittor Raechel , Poet Jeffrey L. , Eckdahl Todd T. , Heyer Laurie J. , Campbell A. Malcolm
Published: April 2020

Synthetic biology takes an engineering approach to biological systems for the construction of parts, devices and systems that address challenges associated with medicine, biofuels, the environment, and more. Although the production of natural and engineered proteins is fundamental to this effort, a persistent challenge to protein production has been unreliable translation directed by a given ribosomal binding site (RBS) depending on which gene of interest is downstream of it. Based on many observations, investigators have hypothesized that the 5’ UTR containing an RBS and early codons can bind to downstream coding mRNA to form base paired secondary structures that decrease the efficiency of translation initiation. In 2013, Drew Endy and colleagues developed bicistronic designs (BCDs) for predictable and reliable in vivo translation. BCDs contain two RBSs and two protein‐encoding cistrons. The first RBS leads to the production of a 16 amino acid leader polypeptide with no functionality, whereas the second RBS directs the production of a protein encoded by a gene of interest. A key feature of BCDs is that the stop codon for the first cistron (TAA) shares its terminal adenine with the first base of the start codon (ATG) for the second cistron. In other words, TAATG is both a stop codon in one reading frame of cistron 1 and a start codon in a shifted reading frame for cistron 2. BCDs were found to be better at producing predictable protein levels in vivo than simple RBSs, thereby improving the ability of BCDs to function reliably and predictably as classic synthetic biology parts in bacterial cells. We wanted to learn if BCDs worked equally well for in vitro protein production during cell‐free protein synthesis (CFPS). CFPS employs transcriptional and translational machinery extracted from cells producing the viral T7 RNA polymerase. We built GFP expression cassettes using 19 BCDs and tested them with CFPS. We compared the rank order of the 19 BCDs in terms of in vitro CFPS GFP production with the in vivo results published by the Endy group. We calculated a rank correlation of 0.88 between the in vitro and in vivo data, showing that BCD control of protein production is comparable in both contexts. The significance of our work is that synthetic biologists using CFPS are well advised to employ BCDs so they can produce predictable amounts of protein for widespread applications in vitro.

Support or Funding Information

NSF RUI MCB‐1613281 to Missouri Western State University; NSF RUI MCB‐1613203 to Davidson College
Fundamentals of Teaching in the Basic Sciences
Authors: Goodman Barbara E.
Published: April 2020

Graduate students in the School of Medicine have had an elective course on how to teach offered either annually or every other year since 2007. The initial course design was based on suggestions from other educator colleagues (especially Dr. Dee Silverthorn). The course is offered in 90 minute to 2 hour time blocks once a week throughout a 15‐week semester. The course is discussion‐based with online reading material provided through the course management system and various assignments during the course (mostly involving having each student design aspects of a hypothetical course that they may teach in their future careers). Topics for the discussions include: the three‐legged stool, importance of learning objectives, planning a course, designing a course, learning preferences, teaching styles, visual aids and active learning, assessing student learning, exams and grading, evaluating a course, teaching portfolios and other resources. The professors provide feedback for specific assignments during the course which include: a minute paper on a course that the student liked and why, how to choose a textbook, writing learning objectives, writing a syllabus, writing a report on a class observation, writing a short review paper on an evidence‐based teaching pedagogy, writing 5 active learning ideas for their course, a short videotaped teaching session (as a preliminary to the final microteach session), finding 3 different online resources for their course, and a peer and self‐evaluated 10‐minute videotaped microteach session that acts as the final examination for the course. The short review paper was added recently when this course became a required course for the non‐thesis option master’s program in basic biomedical sciences so that the students would have experience with scientific writing. The culminating videotaped microteach session was felt to be so valuable for the students that a short videotaped session was added earlier so that students could make improvements after their initial attempt. An interesting recent complication in the course is that although the course was initially designed for preparing future faculty, now the majority of the students in the course are pre‐professional students hoping for acceptance into health professional programs where classroom teaching may not be a high priority. Thus, the emphases of the discussions and assignments have changed to address relevance to these students. However, student evaluations following the two classes with predominantly master’s students have still shown appreciation of the discussions and assignments for future health practitioners.

Support or Funding Information

Dr. Goodman was partially supported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM103443.
The relationship between Alzheimer's disease and vascular risk factors
Authors: Chen Xianghui , Zhang Weidong , Qiao Xiaohong , Li Youliang , Duan Hubin , Sun Pinglian
Published: April 2020

Objective

To study the relationship between Alzheimer's disease(AD) and vascular risk factors.

Methods

53 patients with AD were selected as the case group and 40 elderly patients with normal cognitive function as the control group. The distribution of hypertension, diabetes, abnormal lipid metabolism, cerebral infarction, carotid artery stenosis and plaque, carotid artery intimal thickening, intracranial blood flow velocity slowing, smoking and drinking history were analyzed. Logistic regression model was used to analyze the relationship between above factors and AD.

Results

Hypertension, diabetes, blood lipid metabolism, cerebral infarction, neck vascular stenosis and plaque, carotid artery intimal thickening, slow cerebral artery blood flow between AD and the control group with significant difference (P < 0.05), while there was no significant differences in moking and drinking alcohol history between AD case group and the control group (P > 0.05).

Conclusion

AD is closely related to vascular risk factors. Hypertension, diabetes, abnormal lipid metabolism, cerebral infarction, carotid artery stenosis and plaque, carotid artery intimal thickening and intracranial blood flow velocity are independent risk factors for AD.

Support or Funding Information

This study was supported by Lvliang City Science and Technology Fundation Grant, Lvliang people‘s hospital Fundation Grant, This study was supported by the National Natural Science Foundation Youth Fund (30600637), China Postdoctral Science Foundation Grant (2019T120195), China Postdoctral Science Foundation Grant (2014M561207), the Shanxi Province Basic Research Program ( 2010021034‐4, 201601D011119), the Shanxi Scholarship Council of China (2011‐096 and Key Projects No.4), Department of Human Resources and Social Security of Shanxi province, The science and technology research projects of health department of Shanxi Province(2016D011119), The Teaching Reform Program of Shanxi Higher Education, The Education Reform Project of Postgraduate Students in Shanxi Province, The Education Reform Project of Shanxi Medical University.
Genome‐wide Quantitative Trait Loci (QTL) Mapping of Metabolites in Human Cerebrospinal Fluid
Authors: Zeltser Nicole , Elting Rachel , Boltz Toni , Loohuis Loes Olde , Ophoff Roel
Published: April 2020

Genome‐wide association studies (GWAS) are used to identify genetic loci that are associated with a trait of interest in order to decipher underlying biological mechanisms. Metabolite measures in cerebrospinal fluid (CSF), the fluid that surrounds the brain, provides insight into brain function that may be relevant for neurobehavioral traits and neuropsychiatric disorders. We performed a GWAS of 1,000 metabolites in a sample of 600 human subjects, the largest set of CSF data used in a GWAS to date. We applied standard quality control of genetic data including missingness, minor allele frequency cut‐off, and population stratification. Phenotype data were checked for outliers, and non‐normal data were transformed using inverse rank normalization. A linear association, including age and sex as additional covariates, was performed using the PLINK whole genome association analysis toolset. Significant SNP associations were found in 45 metabolites. Analysis via functional mapping and annotation (FUMA) downstream GWAS analysis tool found that many of these SNPs have been associated with other traits by other GWAS. Linked traits include blood metabolite levels and brain related traits such as cognitive performance. Results show that a GWAS can be successfully performed with CSF metabolite data, and indicate a relationship between CSF and blood metabolite levels. Further steps include analysis of other CSF metabolites and the use of dosage genotype data for potentially more detailed results.
Inhibition of leucine aminopeptidase affects myocyte proliferation and differentiation
Authors: Osana Shion , Murayama Kazutaka , Suzuki Naoki , Takada Hiroaki , Kubota Takahiro , Nagatomi Ryoichi
Published: April 2020

Background

Loss of skeletal muscle mass in the elderly, namely sarcopenia, is a rapidly growing global health problem. Understanding the mechanism of muscle mass regulation is essential to confront this problem. The balance of muscle protein synthesis and degradation is thought to be one of the key issues. Previous reports showed that muscle‐specific deletion of a crucial proteasomal gene, Rpt3 (also known as Psmc4), resulted in severe skeletal muscle atrophy (Kitajima Y et al., 2014). Although proteasomal proteolysis plays an essential role in the maintenance of muscle mass, the underlying mechanism remains unknown. Proteasome generates oligopeptides in the cytoplasm, which are further degraded quickly to amino acids depending on several aminopeptidases (Hershko A et al., 1992, Saric T et al., 2004). Although aminopeptidases may have a significant role in the maintenance of muscle mass, not much is known. Microarray analysis of transcripts in Rpt3 knockdown myocytes revealed an upregulated leucine aminopeptidase related genes. In this study, therefore, we examined the contribution of leucine aminopeptidase in the proliferation and differentiation of cultured myocytes.

Methods

We examined the consequence of leucine aminopeptidase inhibition on C2C12 myocytes under different culture conditions. Leucine aminopeptidase was inhibited by bestatin methyl ester (Bes‐ME; 100 μM), an aminopeptidase inhibitor. Proliferation was evaluated by counting the number of Ki67 (proliferation marker) positive cells. Differentiation was evaluated by myosin heavy chain (MyHC) staining and fusion index. Fusion index was calculated as the ratio of the nuclei number in myocytes with two or more nuclei versus the total number of nuclei. Additionally, phosphorylation of mammalian (or mechanistic) target of rapamycin (mTOR) and eukaryotic translation initiation factor 4E‐binding protein (4EBP1), were evaluated by Western Blotting because they are key factors for muscle protein synthesis.

Results

Leucine aminopeptidase inhibition by Bes‐ME significantly suppressed the activity of leucine aminopeptidase. The proliferation of C2C12 myoblasts was significantly reduced and the differentiation of C2C12 myoblasts was also markedly restricted after Bes‐ME administration. The protein expression levels of MyHC, Myogenin, and MyoD responsible for myodifferentiation were also significantly reduced. Furthermore, we also discovered that the phosphorylation of mTOR was significantly suppressed after leucine aminopeptidase inhibition.

Conclusions

Leucine aminopeptidase is essential for myoblast proliferation and differentiation; however, further studies are required to clarify the role of aminopeptidase in skeletal muscle growth.

Support or Funding Information

Acknowledgements

This work was supported by JSPS KAKENHI Grant Number 18H04080.
Siglec‐E Retards Atherosclerosis by Inhibiting CD36‐Mediated Foam Cell Formation
Authors: CHAU Lee-Young , Hsu Yaw-Wen , Chiang Ming-Tsai , Hsu Fu-Fei , Angata Takashi , Crocker Paul R.
Published: April 2020

The accumulation of lipid‐laden macrophages, foam cells, within sub‐endothelial intima is a key feature of early atherosclerosis. Siglec‐E is a member of sialic acid binding lectin predominantly expressed on myeloid cells to transduce inhibitory signal upon interacting with its ligands. Whether Siglec‐E expression on macrophages impacts foam cell formation and atherosclerosis remains to be established. To this end, both apoE‐deficient and apoE/Siglec‐E‐double deficient mice were placed on high fat diet for 3 months and their lipid profiles and severities of atherosclerosis were then assessed. The results showed that Siglec‐E deficiency accelerated atherosclerosis without affecting lipid profile in apoE deficient mice. In vitro experiments demonstrated that Siglec‐E deletion facilitated the uptake of acetylated or oxidized low density lipoprotein (LDL) and augmented foam cell formation in macrophages. By performing proximity labeling and proteomic analysis, we identified CD36 as a cell surface protein interacting with Siglec‐E. Notably, the interaction between Siglec‐E and CD36 was not affected by the sialylation status of CD36. Further experiments demonstrated that oxidized LDL induced transient Siglec‐E phosphorylation and recruitment of SHP‐1 in macrophages. VAV, a downstream effector implicated in CD36‐mediated oxidized LDL uptake, was shown to interact with SHP‐1 following oxidized LDL treatment. Moreover, Siglec‐E deficiency enhanced VAV phosphorylation induced by oxidized LDL. Collectively, these data demonstrate that Siglec‐E attenuates atherosclerosis in apoE‐deficient mice through suppressing CD36‐mediated signaling responsible for modified LDL uptake and foam cell formation in macrophages.

Support or Funding Information

This work was supported by a grant from Ministry of Science and Technology of Taiwan (MOST‐106‐2320‐B‐001‐020‐MY3).
Biochemical Composition and Antimicrobial Activities of Seed Extracts of Avocado (Persea americana)
Authors: Akalazu Jacinta Ngozi , Uchegbu Rosemary Izunwanne
Published: April 2020

Evaluation of the bioactive compounds present in plant extracts play an important role in developing effective antimicrobial agents that can combat resistant plant and animal fungal pathogens. The objectives of the study were to evaluate the biochemical composition and antifungal activities of Avocado (Persea americana) seed extracts. The fungal pathogens for the study were isolated from yam tuber. They were Botryodiploidia theobromae, Rhizopus stolonifer, Aspergillus flavus, Fusarium oxysporium and Geothricum candidum. Pathogencity tests showed that all the fungal isolates were pathogenic on the yam tuber. The results of the antifungal activity of Avacado pear seed using paper disc diffusion technique against the pathogenic fungi showed that it had varied degree of inhibitions on all the fungal pathogens, Botryodiploidia theobromae (13.20), Rhizopus stolonifera (14.02) A. flavus (12.10), F. oxysporium (9.30), G. candidum (7.00).Identification of antifungal compounds using GCMS revealed the presence of 119 compounds. The compounds with highest in abundance are 2‐n‐Octylfuran,this was followed by 13‐tetradecenal and 9,12‐Octadecadienoic acid and finally 2,15‐Octadecedien‐1‐ol acetate. It is suggested that Avacado pear seed be used on the treatment of plant and animal fungal diseases.

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Similarities Between Stem Cell Niches in Glioblastoma and Bone Marrow: Rays of Hope for Novel Treatment Strategies
Authors: Hira Vashendriya , Breznik Barbara , de Jong Annique Loncq , Khurshed Mohammed , Molenaar Remco , Lah Tamara , van Noorden Cornelis
Published: April 2020

Glioblastomais the most aggressive primary brain tumor. Slowly dividing and therapy‐resistant glioblastoma stem cells (GSCs) reside in protective peri‐arteriolar niches and are held responsible for GSC maintenance and glioblastoma recurrence. Recently, we showed similarities between GSC niches and hematopoietic stem cell (HSC) niches in bone marrow. Acute myeloid leukemia(AML) cells hijack HSC niches and are transformed into slowly‐dividing and therapy‐resistant leukemic stem cells (LSCs). Current clinical trials are focussed on removal of LSCs out of HSC niches to differentiate and to become sensitized to chemotherapy. In the present study, we elaborated further on these similarities by immunohistochemical analyses and fluorescence microscopy of17 biomarkers in paraffin sections of human glioblastoma and human healthy bone marrow. We found all 17 biomarkers to be expressed both in hypoxic peri‐arteriolar HSC niches in bone marrow and hypoxic peri‐arteriolar GSC niches in glioblastoma. Our findings implicate that GSC niches are being formed in glioblastoma as a copy of HSC niches in bone marrow. These similarities between HSC niches and GSC niches provide a theoretic basis for the development of novel strategies to force GSCs out of their niches, in a similar manner as in AML, to induce GSC differentiation and proliferation to render them more sensitive to anti‐glioblastoma therapies.

Support or Funding Information

This study was financially supported by the Dutch Cancer Society (KWF; UVA 2014‐6839 and UVA 2016.1‐10460), the European Program of Cross‐Border Cooperation for Slovenia‐Italy Interreg TRANS‐GLIOMA (Program 2017), the IVY Interreg Fellowship, the Slovenian Research Agency (Program P10245) and by the Fondation pour la Recherche Nuovo‐Soldati 2019.
Effective Antihypertensive Treatment with Epoxyeicosatrienoic Acid Analog (EET‐A) and 20‐HETE Antagonist (AAA) of Spontaneously Hypertensive Rats (SHR)
Authors: Gawrys Olga , Walkowska Agnieszka , Kompanowska-Jezierska Elzbieta , Husková Zuzana , Honetschlägerová Zuzana , Falck John , Imig John , Červenka Luděk
Published: April 2020

Introduction

Despite decades of research the death toll of hypertension and associated disorders increases significantly each year. This impose the need to further determine the mechanisms underlying the development of hypertension and to implement more efficient therapeutic strategies. Growing number of research suggest the important role of cytochrome P‐450 dependent metabolites of arachidonic acid (AA) in blood pressure regulation. The epoxyeicosatrienoic acids (EETs) exhibit vasodilatory activity, they decrease sodium reabsorption and possess renoprotective and anti‐inflammatory properties. Considering the beneficial effects of EETs, we decided to test the efficiency of stable analog of 14,15‐EET [disodium (S)‐2‐(13‐(3‐entyl)ureido)‐tridec‐8(Z)‐enamido) succinate] called EET‐A, together with AAA [N‐disodium succinate‐20‐hydroxyeicosa‐6(Z),15(Z)‐diencarboxamide], a novel receptor antagonist of 20‐HETE which is another potent AA metabolite with both pro‐ and antihypertensive activity. To test the antihypertensive potential of both substances we employed spontaneously hypertensive rats (SHR), which are currently considered the best model to study human essential hypertension.

Methods

Male SHR in the established stage of hypertension (16 week old) were treated for four weeks with EET‐A (n=8), AAA (n=6), as well as the combination of EET‐A and AAA (n=7) and compared to age‐matched untreated SHR (n=10) and WKY (n=5). Both tested substances were administered in drinking water in the dose of 10 mg/kg/day each. Systolic blood pressure (SBP) was measured by telemetry. Once a week observations in metabolic cages (food and water intake, feces, diuresis) were performed; urine, blood and tissue samples were collected for further analysis.

Results

EET‐A or AAA given alone had no significant effect on blood pressure of SHR. However, the combined treatment with AAA + EET‐A was significantly antihypertensive (161±5 vs 180±3 mmHg, p<0.05). Additionally, combined AAA and EET‐A attenuated cardiac hypertrophy expressed as left ventricle weight to tibia length (22.95±0.61 vs 24.75±0.35 mg/mm in untreated SHR, p<0.05), decreased kidney ANG II level, increased natriuresis and increased the excretion of nitric oxide metabolites (7.6±1.4 vs 4.8±0.7 μmol/24h in untreated SHR, p<0.05).

Taking into account all the beneficial impact of the combined treatment with EET‐A and AAA on cardiovascular and renal function of adult SHR we suggest that it constitute a very promising novel antihypertensive strategy.

Support or Funding Information

Supported by National Science Centre Poland (grant no 2017/26/M/NZ5/00367)and MH CZ ‐ DRO („Institute for Clinical and Experimental Medicine – IKEM, IN 00023001“)
Neuroanatomical Approaches to Improving Student Learning in Science Education
Authors: Azzam Mohammad , Easteal Ronald
Published: April 2020

Students are usually led to believe that rote memorization is the best technique, and perhaps the only technique, to learn factual information. In disciplines such as Anatomy, and in science education generally, this belief is largely observed because factual information is constantly taught. This review challenges the said notion. First, the different stages of memory and its pathways are explained to provide the basis for the several teaching and learning paradigms which, if employed in the classroom, can improve student learning in science education. Next, the teaching and learning paradigms are described. These paradigms include cognitive load theory, dual encoding theory, the spiral syllabus, bridging and chunking during lectures, sleep consolidation, and retrieval practice. Although such paradigms are especially important in science education, they can be utilized in the teaching and learning of any discipline.

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N/A
Comparative Effects of Taurine, Vitamin E and Nuclea latifolia (Stem‐Bark) Extract on Object Regonition and Anxiety on Wistar Rats Subjected to Water Immersion Restraint Stress
Authors: Iliya Ezekiel , Eze Ejike Daniel , Jiya Peter Babatsu
Published: April 2020

Stress is the feeling of emotional or physical tension, that may arise from any event or thought that makes person become frustrated, angry, or nervous. Anxiety and cognitive impairment can be normal during stressful situations. The comparative effect of Taurine, Vitamin E, and Nuclea Latifolia (Stem back) extract object recognition and anxiety in wistar rats subjected to water immersion restraint stress was carried out. Total of twenty four rats weighing between 100g – 120 g were used for this study. The animals were divided into four groups before the commencement of the experiment as follows: Group A: Serves as the control and received 1ml/kg distilled water for three weeks. Group B: Received 0.2 ml/kg vitamin E for three weeks. Group C: Received 200 mg/kg taurine (TAU) for three weeks and Group D: Received 200 mg/kg of Nauclea latifolia (NL) (stem bark) extract for three weeks. At the end of the three weeks, animals were starved for 24 hours before the commencement of the stress procedure. They were anesthetized using chloroform vapour after which they were restraint on a wooden plank (25 by 19 cm) with the four arms anchored horizontally and were immersed into water up to the xiphoid level for two hours. After the stress procedure, object recognition task and anxiety behaviours were assessed. The result of the training phase on object recognition showed that, TAU group explored the novel object more (64.92 %) followed by Vit. E group (63.90 %). The NL group has the least percentage (52.75 %) in exploring the novel object that was presented to the animals. During the test phase, it was noticed that, the TAU together with Vit. E groups showed similar behaviour with that of the training phase as they explored the Novel object presented to them more (71.52 %) and (70.05 %) respectively. On the anxiety model, while the Vit. E group shows significant increase in the number of entries into the open arm, the Tau group revealed significant decrease in the number of entries into the closed arm. Vit. E, Tau and NL groups showed significant increase in time spent by the animals in the open arm. The researched therefore, showed that Taurine and Vitamin E improved object recognition in the rats and also inhibited anxiety‐like behavior in Wistar rats that were subjected to water immersion restraint stress.

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Personal Funding
The Prevalence of Exercise Prescription‐related Course Offerings in U.S. Medical School Curricula: Exercise is Medicine®
Authors: Dirks-Naylor Amie J. , Crump Lauren Hartsell , Le Nam
Published: April 2020

Exercise training has proven to be beneficial in the prevention of disease. In addition, exercise can improve the pathogenesis and symptoms associated with a variety of chronic disease states and can attenuate drug‐induced adverse effects. Exercise is a drug‐free polypill. Exercise is Medicine®, a joint initiative between the American Medical Association and the American College of Sports Medicine, was launched in 2007 to call on all health care providers to counsel patients and prescribe exercise in the prevention and treatment of chronic disease states. However, according to the Behavioral Risk Factor Surveillance System data, 48% of Americans still do not meet the national physical activity guidelines. The cause behind the high prevalence is unknown and likely multifactorial. One factor may be under‐education of medical professionals on how to council patients and develop exercise prescriptions. Therefore, the aim of the study was to determine the prevalence of course offerings in U.S. medical school curricula regarding training in exercise prescription. School websites were accessed for information regarding course offerings in MD and OD programs. Only those programs whose course descriptions of their full curriculum were accessible online were included in the analysis. The search terms used were exercise prescription, exercise, fitness, and physical activity. 61% (86/141) and 69% (24/35) of MD and OD granting programs were included in the analysis, respectively. Four programs offered courses with the term “exercise prescription” in the course title or description. Only 4 programs offered courses with the terms “exercise, fitness, or physical activity” in the course title. 36% of programs offered courses with the terms “exercise prescription, exercise, fitness, or physical activity” in the course description or course topics, with only 17% of those programs offering required courses. The data suggests that medical students may be under‐educated in the area of exercise prescription, which may contribute to the high prevalence of inactivity.