bluuebunny/crossref_metadata_2025 huggingface dataset:
Deposition of collagen perivascular by Angiotensin II action in the aorta independent of hemodynamic effects
Authors: da Silva Viegas Katia Ap. , Souza-Oliveira Thais Cristina , Rabechi Nara Baptistella , Paiva Aline , Alves Izabel , Machado Raquel Cristina Campos , de Jesus Piva Camila , Martins Alvaro Batista , Lacchini Silvia
Published: June 2021

The vascular lesions through trophic stimulation of your wall are an important mechanism of action of Angiotensin II (AngII) in extracellular matrix (ECM) by modulation of synthesis x degradation collagen. The aim was to assess whether AngII can act in the ECM by modifying the deposition perivascular collagen (COL) without hemodynamic effects. Mice were treated for 7 and 14 days with subpressor doses of AngII(100ng/kg/min); Losartan(LOS)‐AT1 receptor blocker(20mg/kg/day); Los+Ang; Control (CONT). Measured systolic blood pressure (SBP) and heart rate (HR) by tail plethysmography. Histological sections of aorta were stained with picrosirius to quantify the perivascular COL analyzed in brightfield and polarized light. Results were compared by 2‐way ANOVA p≤0.05. No changes in SBP and HR between groups and nor significant differences of perivascular COL fibers thick (type I‐polarized light) in treated for 7 days but there was a slight increase these in 14 days with AngII(0.56±0.13au) and Los+AngII (0.55±0.14au) compared to CONT(0.49±0.13au) and LOS(0.38±0.04au) significantly reduced the presence of these fibers. In thin COL fibers (type III) there was no difference. The results suggest that the AngII can act on deposition of perivascular COL fibers thick acting in ECM independent of hemodynamic factor through the activation of the AT1 receptor and through other ways for combined treatment.

Grant Funding Source : FAPESP ‐ Research Foundation of the State of Sao Paulo
Phenanthrene affects cardiovascular function in the developing African clawed frog (Xenopus laevis)
Authors: Hausken Krist N. , Layman Kimberly A. , Whittemore Susan L.
Published: June 2021

The incomplete combustion of fossil fuels results in deposition of the tricyclic aromatic hydrocarbon phenanthrene into air, soil, and water. This common and persistent contaminant is also present in tobacco smoke, coal tar sealant, and weathered crude oil. Surprisingly, little is known about its toxicity. To gain a better understanding of potential risks to humans, we characterized the effects of phenanthrene exposure using embryos of X. laevis, an important model for vertebrate development. Heart rates and evidence of arrhythmia were recorded during various stages of heart development using an inverted dissecting microscope, Leica™ camera, and the video recording and editing software Camtasia.™ In stage 42 embryos, phenanthrene exposure (5 ppm, 28.5 μM) led to bradycardia with a decrease to 79% of control values at 6 hrs followed by a further reduction to 69% by 10 hrs. After 5 hrs of depuration, heart rates returned to control levels. Arrhythmia was also evident in phenanthrene‐exposed embryos. A 42% increase in interbeat variability was observed following a 10‐hr exposure to 5 ppm. These phenanthrene‐induced effects may be stage‐specific as cardiac function in stage 47 animals, with a mature morphology, and in stage 37 animals was less affected. These findings underscore the need for further characterization of the cardiotoxic effects of this common pollutant.

(Supported by NH‐INBRE grant 1P20RR030360‐01)
Structural Composition of Myocardial Infarction Scar Does Not Differ Between Male and Female Middle‐Aged Rats
Authors: Bogatyryov Yevgen , Kelly Molly , Christensen Lance P. , Tomanek Robert J. , Dedkov Eduard I.
Published: June 2021

A number of studies indicate that females have a better reparative response in earlier stages of myocardial infarction (MI) compared to males. Nevertheless, little is known concerning the sex‐related differences in a later stage of MI healing (i.e. scar formation). Accordingly, we designed our study to determine whether the composition of the scar is affected by the biological sex of post‐MI animals. A large MI was induced in 12‐month‐old male (M‐MI) and female (F‐MI) Sprague‐Dawley rats by ligation of the left coronary artery. Four weeks after the MI, rats with transmural infarctions, greater than 50% of the left ventricular free wall, were evaluated. The fractional volume of fibrillar collagen (FC), cardiac myocytes (CM), myofibroblasts (MF) and vascular smooth muscle cells (VSMC) was determined in each scar. The mean scar size and thickness were comparable between F‐MI and M‐MI rats (61.3±3.9% vs. 63.8±2.5% and 0.66±0.04mm vs. 0.73±0.05mm, respectively). Although, there was a great degree of heterogeneity in spatial distribution of the analyzed structural components within each scar, their mean content showed no significant differences between F‐MI and M‐MI rats (FC: 53.3±6.6% vs. 61.6±4.5%; CM: 3.1±0.4% vs. 3.3±0.4%; MF: 9.6±1.2% vs. 9.6±1.7%; VSMC: 3.4±0.5 vs. 2.3±0.5%). Our data are the first to demonstrate that biological sex does not influence the structural composition of the MI scar in middle‐aged rats.

Grant Funding Source: Internal Award from the NYCOM Office of Research (E.I. Dedkov) and NIH RO1‐HL62587 (R.J. Tomanek)
Undergraduate Dissection Techniques: Unique Views of Internal Structures
Authors: Coniglio Lacy Nichole , Saenz Zoe Marie
Published: June 2021

Cadaver dissection is the most efficient means of disseminating anatomical knowledge, inviting anatomy educators to expand and simplify dissections in order to maximize the student experience. Undergraduates rarely have the opportunity to produce meaningful dissections of the human head and neck, prompting our group to streamline techniques and methods for head and neck dissection. Incorporation of an in class sagittal section of the head and neck, or hemi‐section, proved invaluable to students in improving their understanding of the respiratory, digestive, and nervous systems (A. Koba 2010). In order to improve dissection technique and the dissemination of anatomical knowledge, coronal and sagittal sections were made by various techniques, and assessed for ease of integration and educational value. The hemi‐section was produced by the classic method and compared to a hemi‐section made by removing a supero‐lateral quadrant from the neurocranium, addressing concerns of simplicity and time management. The coronal section of the head and neck was made posterior to the foramen magnum, preserving the brain stem and provided new perspectives of intact structures. This research simplified advanced dissection techniques in a cost effective manner, engaging students in their learning.

Grant Funding Source : n/a
Dissection of a female perineum: a colored anatomical model in plaster (Sciences e Pedagogie, Paris) of the middle of the 20th century
Authors: Le Floch-Prigent Patrice Pierre
Published: June 2021

The aim of the study was to analyze the anatomical veracity of an anatomical model of the dissection of a female perineum, realized in plaster by the factory: “Sciences and Medecine”, in Paris, in the middle of the 20th century. It was a colored representation of a dissection which was different on each side. The teaching model belonged to the collections of the department of anatomy, 45 rue des Saints Pères, Paris 6ème. We determined its technics of presentation and fabrication. The sample was observed after cleaning and measuring. At least we performed photographies with a numerical camera under several angles of view. The anatomical veracity was good and this types of teaching models could be still useful in the departments of anatomy.

Grant Funding Source: DEPARTMENT OF ANATOMY
Head and neck dissection: a non colored anatomical model in plaster of the Nicolas‐Augier‐Roux collection
Authors: Le Floch-Prigent Patrice Pierre
Published: June 2021

The aim of the study was to appreciate the anatomical veracity of a non‐colored, plaster sample of the Nicolas‐Augier‐Roux (beginning of the 20th century in Paris). The sample belonged to the teaching collection of the department of anatomy, 45 rue des Saints Pères, Paris 6ème. It was studied and photographed to determine its technics of presentation and of fabrication. We observed the sample after cleaning and measuring. It was similar to another model belonging to the department of anatomy in Brussels (ULB) which was CT‐scanned by its head: Pr LOURYAN. This cheap models were teaching tools which can still be useful in the departments of anatomy as they are very anatomically accurate.
The Use of Medical Images in Teaching Human Anatomy
Authors: Rae Guenevere , Skelton Michele , Bevers Emma
Published: June 2021

The purpose of this study is to determine whether the use of medical images increases student performance in learning human anatomy. 36 undergraduate students enrolled in basic human anatomy and physiology course were recruited and divided into two groups. A pretest was given to both groups to assess knowledge base on the human skeletal system prior to beginning the study. The control group was taught the anatomy of the skeletal system by lecture using cadaveric images, models, and artistic illustrations. The experimental group was taught the same material using medical images, cadaveric images, models and artistic illustrations. The lecture time and content were standardized. The study consisted of three sessions on the following topics: the general anatomy of the human skeleton; the bone markers of the axial skeleton; the bone markers of the appendicular skeleton. Student performance was assessed via a posttest after each learning session. After conclusion of the study, a survey was given asking students to rate how well each material (cadaveric images, medical images, artistic illustrations, and models) helped them learn and enjoy the lessons. Results of the study indicate that no differences exist between group posttest scores, even though differences do exist between the student opinions of the materials’ contribution. Medical imaging appeared to lower student satisfaction in the lesson, regardless of the lack of difference between posttest scores.

Grant Funding Source : N/A
The Use of 3‐D Stereoscopic and 2‐D Videos as Pre‐Lab Tool for Anatomy Dissection
Authors: Alkhalili Sereen Mahmoud , Hannon Kevin , Coppoc Gordon L
Published: June 2021

The hypothesis was that viewing a 2‐D or 3‐D stereoscopic pre‐lab video containing interspersed instructions and short multiple‐choice questions would improve students’ ability to efficiently perform dissections and more quickly identify required structures. First‐year D.V.M. class was divided into 21 dissection teams of four students each. Teams were randomly assigned to one of three groups, A, B and C. Preparation for all groups included reading the lab manual. Video groups interacted with a 30‐minute video. All groups were allowed 45 minutes to dissect the thorax of a dog and find a list of assigned structures followed by a quiz.

Preliminary results are as follows: There was no significant difference (p > 0.05) among the groups on the post‐dissection quiz nor on the number of structures the groups identified during the dissection period (p > 0.05).

In conclusion, the first phase of this cross‐over design experiment that will include dissection of abdomen and pelvis did not support the hypothesis that video pre‐dissection laboratory preparation would lead to greater speed in identifying required anatomical structures or performance on a post‐dissection quiz. However, for aspects that might reflect on the need for laboratory teaching assistance, there was a trend that supported the hypothesis that video preparation would be superior to lab manual preparation.

Grant Funding Source: School of Veterinary Medicine, Purdue University,
The John Martin Rare Book Room as a learning resource
Authors: Heidger Paul M. , Hirst Donna L.
Published: June 2021

The John Martin Rare Book Room (RBR) houses over 5,000 rare books relating to the biomedical and health sciences. The most notable works in the collection were a bequest to the University Libraries from Dr. John Martin (1904–1996), an anatomist and neurosurgeon, who collected the books over a period of many decades. The RBR offers numerous services promoting learning, including publishing a monthly newsletter, offering a monthly lecture series during the school year, and creating regular exhibits from the resources of the collection. The Curator of the RBR, Donna Hirst, MPH, MLS, gives all first‐year medical students an anatomy lecture focusing on the social and historical ambivalence toward the body and human dissection, together with the legal and social history of dissection. The Sparks Essay Contest, coordinated through the RBR, is offered annually to all medical students. Significant monetary prizes recognize the best essays that examine a timely issue in medicine incorporating historical, ethical, and cultural perspectives. Tours and open houses of the room are offered to groups upon request, often featuring specific aspects of the collection. Among courses utilizing the RBR have been graduate courses in Anatomy and Cell Biology, and a course in Classics on Galen and Human Dissection. The John Martin Rare Book Room serves the University as a vital resource in anatomical instruction and historical research.
Clinical utility of PA anatomy education
Authors: Lane Richard , Gardner April , Bennett-Clarke Carol , Hogue Patricia , Hankin Mark
Published: June 2021

The University of Toledo Physician Assistant Studies (UT PA) program matriculated its first class in 1996. Students are drawn primarily from Ohio and Michigan and the typical graduating class performs well on the Physician Assistant National Certification Examination. UT PA students that graduated between 2006–2010 (N=145) had a first‐time mean pass rate of 97.8% (± 2.0) compared to the national mean of 93.0% (± 1.0). The UT PA human anatomy course is a traditional medical gross anatomy and neuroanatomy course taught by Anatomy faculty in the College of Medicine. It consists of 120 hours of classroom instruction that include lectures and labs. In addition to providing a broad foundation in human anatomy, this course emphasizes the practical application that anatomical knowledge has in the clinical setting. The goal of this study is to understand the value of this anatomy education for recent UT PA graduates. Recent UT PA graduates will be surveyed to assess their level of appreciation, usefulness, and applicability of the anatomy education they received and the specific anatomical subjects they have found to be most useful. The data will be sorted by the clinical specialty of the responders. This information will be used to evaluate the current course and revise it accordingly to better fit the needs of our future graduating students.
What motivates medical students to learn anatomy?
Authors: Selby Keegan , Singh Supriya , Ratner Shemer , Johnson Marjorie
Published: June 2021

Independent e‐learning modules (IL's) using sound pedagogy and cognitive learning theories are often used to replace the lost lecture or lab hours once given to anatomy. This change should reflect the documented characteristics of the “net” generation. However, participation in and/or completion of the IL's is low. Our initial hypothesis is that motivation to learn is not tied solely to formal evaluation or comfort with e‐learning. An adjunct to learning anatomy through IL's was offered for Years 1 and 2 medical students. The students organized a surgical oriented anatomy club (SOA) to help them understand the relevant anatomy in a clinical context. This study compares the motivation of students to participate in anatomy taught through the surgery club with that of IL's. The SOA club arranged for surgeons to demonstrate several common surgical procedures. After the demonstration students were asked to review the relevant anatomy on cadavers with the assistance of an anatomist and the surgeon. Participation for the events was voluntary and was full beyond capacity within minutes of posting the session information. Participation rates will be compared to that of IL's. In addition, survey data will gauge student motivation for attending the SOA sessions or completing IL modules. Preliminary results suggest more student initiatives designed with clinical relevancy should be considered part of future medical curricula.

Grant Funding Source : None
Undergraduate study approaches to anatomy, using surface or deep approaches
Authors: Dunham Stacey , Notebaert Andrew J
Published: June 2021

Anatomy courses require students to learn many new terms and structures and so they may approach learning anatomy in a style contrary to how they approach learning in other courses. The purpose of this study was to examine how undergraduate anatomy students approach learning in anatomy and how their learning approach affects course grades. Students enrolled in an undergraduate anatomy course completed an initial survey which asked for demographics, GPA, and exam sores up to the midterm. Students also completed a modified study process questionnaire resulting in a deep approach score (likely to study for understanding) and surface approach score (likely to use rote memorization). Initial surveys were completed by 105 students. Average age was 20.0 years and average GPA was 3.39/4.00. On average, deep approach score was 31.0 ± 6.4 and surface approach score was 24.7 ± 5.9. Current class percentage averaged 77.9% ± 19.94. Correlations between both approach scores and grade were not significant (deep approach and current grade r=.005, p=.967 and surface approach and current grade r=.107, p=.340). Initial results find that students trend toward deeper approaches to learning, but current analysis indicates low correlations between study styles and midterm grades. This may indicate that approaches to learning do not account for the variance in course achievement. This study was not funded.

Grant Funding Source : N/A
Integrating basic science and clinical subject material into a clinically relevant skin‐musculoskeletal course for first‐year medical students
Authors: Severson Arlen R , Repesh Lillian A , Westra Ruth E , Johns Alan M , Hoffman Richard G
Published: June 2021

A clinically relevant integrated skin‐musculoskeletal systems course with the appropriate topics from the basic science disciplines and the clinical physical exam were incorporated into a 7‐week course for first‐year medical students. The course included cadaver dissection of the back and extremities, and the histology, microbiology, physiology, pathology and pharmacology related to skin, muscle, bone and joints. Clinical faculty presented an overview of the most common clinical problems and diseases affecting these anatomical structures. Delivery of the educational content involved several curricular pedagogies: lectures, cadaver laboratory, peer teaching, self‐study, clinical case studies, and physical exam skill sessions. The microbiology component was delivered via self‐study with clinical cases presented in advance of group discussion. The course material was divided into two sections: the first 4 weeks covered skin, basic immunology, laboratory dissection of the back and upper extremities, and physical exam skills related to these areas. The last 3 weeks covered dissection of the lower extremities, physical exam skills, physiology and pharmacology of the neuromuscular junction, and pathology and clinical aspects of muscle, bones and joints. Assessment of the course revealed a high degree of satisfaction for the overall course, with requests for only additional formative assessments.
Near‐peer teaching in an anatomy course
Authors: Omaña Rodrigo Enrique Elizondo , López Santos Guzmán , Bahena Eduardo Navarro , Durán Claudia Pámanes
Published: June 2021

The near‐peer teaching has been used in anatomy labs to utilize the tutor's experience to supervise students’ activities and to act as the instructor in the laboratory. The goal of the present study was to describe the design and implementation of an anatomy course program with a large number of registered students that utilizes near‐peer teaching as a strategic solution, as well as to evaluate students’ perceptions of this program. Due to the large number of students who enroll in the Anatomy course each semester, near‐peer teaching was implemented as a strategy in which instructors (e.g., senior students) participated in the theoretical or practical activities. A total of 700 students were registered for the anatomy course, of which 558 (79.7%) agreed to participate in this study. In general, the practical section (e.g., the clinical hour, image anatomy, and laboratory anatomy) of the course was viewed most favorably (54.8%, n=306), with the practice of dissection/prosection in the laboratory as the highest valued (34.9%, n=195). Near‐peer teaching is a viable option that satisfies the demands of modern curricula using small groups to stimulate learning, despite a large number of students.
Medical Students in the Microscopic Anatomy Course: Active Learning as an Evolving Response to the Adult Learner's Needs
Authors: Jurjus Rosalyn A , Krum Janette , Goldman Ellen
Published: June 2021

The microscopic anatomy lab experience in the U. S. is evolving from using light microscopes to employing virtual microscopy using digital technology. There is a view that active learning principles and laboratory‐acquired skills have also been marginalized. However, we believe this actually provides a better opportunity for active learning because computer monitors can be readily shared. Accordingly we developed lab sessions that apply principles of adult learning and instructional design. These include linking, hooking, engagement, transfer and teaching to different learning styles and preferences. We will test the hypothesis that these principles will improve the medical students’ performance and satisfaction. Preliminary data based on student surveys of lab activity shows positive responses.

This is a comparative study with a quasi‐control group. Two years of student performance and course evaluation data will be compared. Student exam scores, course evaluations, surveys of lab activity and faculty surveys of students’ participation will be collected. The survey data will be analyzed by tabulating the ratings for each question and survey written comments will be analyzed for themes. This approach also favors development of teamwork skills that will not be directly assessed in this study, but could be implied from comments of participants.

Grant Funding Source: Pending ‐ SMHS Medical Education Grant
A qualitative exploration of how students self‐assess anatomy performance
Authors: Finn Gabrielle Maria , Sawdon Marina A
Published: June 2021

This qualitative study explores how students self‐assess their performance on anatomy examinations. 84 first and second year medical students completed an online questionnaire following summative anatomy examinations. Students entered a numerical predication of their grades for the anatomy practical exam and written exam. In addition, students provided free‐text justifications for their predications. Justifications were analysed using a grounded‐theory based approach. When self‐assessing, students employ a multi‐faceted approach. The major considerations offered by our students included; their exam technique, their knowledge of course content and the standards required to pass, their performance in relation to that of their peers’ and their previous experience, education and exam performance. Further analysis is underway to determine whether the strategies employed differ across students’ self‐assessment abilities and demographics. Our data provides a unique insight into how students’ self‐assessments are complex and often informed by their experiences prior to medical school. This presents a challenge for faculty as factors out with the immediate learning context appear to impact on students’ ability to reflect on their performance. Further insight into students’ self‐assessment techniques will enable faculty to provide targeted support, particularly within anatomy.

Grant Funding Source: Internal funding
Creating Integrated Histology Flash Cards with Clinical Correlations in Teaching Histology
Authors: Cui Dongmei , Daley William P , Yang Gongchao
Published: June 2021

Well designed histology flash cards are a useful tool for learning histology. Integrating pathological images and clinical cases makes flash cards especially helpful in enhancing students’ understanding of how the normal tissue could develop into abnormal tissue, and what pathological condition and clinical disease may be associated with this change. The integrated flash cards provide students a good platform to maximize their exposure to clinical information in their early stage of medical training. Because of the portability of the flash cards, they serve as convenient learning tools for quick review, self‐assessment, and preparation for exams. We created customized histology flash cards at four levels: (1) Introductory cards that help students to quickly review basic organization of the materials; (2) Cards with normal and pathological images to help students to appreciate their differences; (3) Cards with key point information to strengthen their knowledge base, and (4) Cards with practical questions and clinical vignette in USMLE style for students to prepare for various exams. In brief, Histology Flash Cards with Clinical Correlations serve as an efficient, convenient and flexible tool for students to learn histology, as well as strengthen their ability to integrate histology and related clinical information.

Grant Funding Source: University of Mississippi Medical Center
Identification and characterization of small molecules that inhibit binding of the third plant homeodomain finger of JARID1A to histone H3
Authors: Wagner Elise Katherine , Flemming Rod , Nath Nidhi , Denu John
Published: June 2021

A number of histone‐binding domains, such as plant homeodomain (PHD) fingers, are implicated in cancer pathogenesis through improper binding of chromatin. For example, the third PHD finger (PHD3) of the histone demethylase JARID1A, which binds histone H3 trimethylated at lysine 4 (H3K4me3), has been identified as a nucleoporin 98 fusion partner in acute myeloid leukemia. Because of its relevance in disease, we sought to identify small molecule inhibitors of JARID1A PHD3 binding to H3K4me3. We utilized HaloTag technology to develop a novel biochemical assay for high throughput screening of JARID1A PHD3. Following screening of a clinical library, we identified three small molecules that inhibit JARID1A PHD3 binding to H3K4me3 peptides, which was validated using independent biochemical methods. We also characterized the specificity of these compounds towards other PHD fingers. Our data suggest two independent mechanisms of inhibition by these compounds. One compound irreversibly modifies JARID1A PHD3 to cause structural destabilization. The other two compounds appear to inhibit H3K4me3‐binding PHD fingers but not those PHD fingers that bind unmodified H3K4, suggesting a competitive mode of binding inhibition.
Human skeletal muscle histone modifications in response to six weeks of endurance training
Authors: Ydfors Mia , Lindholm Maléne , Sundberg Carl Johan
Published: June 2021

Differences in training responsiveness between individuals can to a large degree be explained by genetic differences. Whether epigenetic modifications, which are known to change over the life span and to be partly related to physical activity levels, are altered with training has been studied sparsely. The differences in responsiveness to training could, among other things, be due to the pre‐existing epigenetic pattern or to epigenetic changes with training.

Purpose

To investigate the histone modification pattern following a period of endurance training.

Methods

Sedentary males (n=16) performed 6 weeks of endurance training. Muscle biopsies were taken before and after the training period. One high‐responder (n=8) and one low‐responder (n=8) group were identified on the basis of their improvement in physiological parameters. Protein content was analyzed with Western blot.

Results

There was no significant change in histone modification pattern (H3ac, H3K9ac, H3K9me1 and H3K9me3) with training in either of the groups or when pooling the groups. There was a tendency (p=0.09) of increased mono‐methylated lysine 9 in histone H3 (H3K9me1) after training in the high‐responder group.

Conclusion

H3K9me1 is associated with transcriptional repression and the tendency to higher H3K9me1 may implicate a role in down‐regulation of genes following exercise.
Epigenetic regulation of Dual‐specificity phosphatase 5 as a functional tumor suppressor and biomarker for gastric cancer
Authors: Shin So-Hyun , Kang Gyeong Hoon
Published: June 2021

Dual‐specificity phosphatase 5 (DUSP5) regulates the duration and magnitude of phospho‐activated ERK1/2 of the MAPK cascades and is recently proposed as a tumor suppressor. However, the epigenetic regulation of DUSP5 and its critical roles in gastric cancer (GC) remain unknown. Here, we compared differential RNA expression profiles of GC cell lines with or without treatment of 5‐aza‐2¡̄‐deoxycytidine and we found that the expression of DUSP5 was dramatically decreased by DNA methylation. Hypermethylation of DUSP5 promoter was detected in GC tissue samples, but not in normal healthy gastric mucosa tissue samples. Restoring DUSP5 expression in DUSP5‐silenced GC cell lines decreased their growth and colony formation ability by causing arrest in G1 to S cell cycle phase through the dephosphorylation of phosphor‐ERK1/2 in nucleus. Moreover, in a set of surgically resected GC cases (n=179), GCs with hypermethylation of DUSP5 promoter region (30.2%) showed significantly shortened survival compared with GCs without DUSP5 methylation (p=0.009). These results implicate that silencing of DUSP5 by hypermethylation of its promoter causes longer maintenance of phosphorylated ERK1/2, which drives cell proliferation, by which DUSP methylation contributes to gastric carcinogenesis. Furthermore, DUSP5 methylation might serve as a prognostic marker for GC, which needs to be validated in a large large of GC samples.
Transcriptomic and Epigenomic Characterization of Mouse Models Simulating Features of Post‐Traumatic Stress Disorder
Authors: Muhie Seid , Hammamieh Rasha , Miller Stacy-Ann , Chakraborty Nabarun , Jett Marti
Published: June 2021

Social defeat mouse models were used to simulate human post‐traumatic stress disorder (PTSD). C57B6 and BALB/c mice exposed to SJL aggressor mice exhibited behaviors accepted as PTSD‐in‐mouse phenotype. Transcripts in hemi‐brain, blood and heart of mice were assayed using Agilent's genome‐wide arrays, and meta‐analyzed with human PTSD data from gene expression omnibus (GEO). DNA methylation changes in hemi‐brains of mice were assessed using Agilent's tiling arrays, and compared with human DNA methylation data from GEO.

About 200 transcripts were differentially regulated in both mice and human data. Induced transcripts were involved in axonogenesis, neuroblast proliferation, response to steroid hormone, apoptosis, inflammation, histone modification and nucleotide biosynthesis. Suppressed transcripts were involved in adaptive immunity, dopamine signaling, DNA methylation, synapse organization, matrix adhesion, nerve impulse transmission, T cell commitment and neural crest cell migration. Hypo‐ & hyper‐methylated genes were associated with immune response, neurogenesis, synaptic transmission, anxiety, and chromatin and protein modifications.

Integrative analyses of (mice and human) transcriptomic and DNA methylation data were indicative of validity of social defeat mice as human PTSD models in search of biomarkers of PTSD that are accessible in human blood.

Funding: Office of Army Surgeon General
Integrin α6β4 regulates promoter demethylation of EGFR ligands
Authors: Carpenter Brittany , Chen Min , O'Connor Kathleen
Published: June 2021

Pancreatic cancer is the 4th leading cause of cancer death in the US due to late detection and lack of effective treatments. The integrin α6β4 is overexpressed in most pancreatic adenocarcinomas where it contributes to tumor invasion. We find that integrin α6β4 overexpression leads to upregulation of amphiregulin and epiregulin, ligands of the epidermal growth factor receptor. The purpose of this study is to address how integrin α6β4 upregulates these ligands. Pancreatic cancer cells expressing low levels of α6β4 were treated with DNA methyltransferase inhibitor 5‐aza‐2′deoxycitidine. Using Q‐PCR, our results show that inhibition of DNA methyltransferase increases amphiregulin and epiregulin expression at the mRNA level. In contrast, when cells with high levels of integrin α6β4 were treated with the hypermethylating agent, S‐adenosylmethionine, it leads to a decrease in amphiregulin and epiregulin expression. Using ELISA, we also provide evidence that integrin α6β4 promotes increased autocrine secretion of amphiregulin after promoter demethylation. This study indicates that overexpression of integrin α6β4 in pancreatic cancer is associated with the hypomethylation of amphiregulin and epiregulin promoters, which may play an important role in the development of malignancy. This study was supported by the Markey Cancer Center.
Effect of chronic hypoxia on pregnancy‐mediated transcriptional regulation of ERα in ovine uterine arteries
Authors: DASGUPTA CHIRANJIB , Zhang Haitao , Chen Man , Zhang Lubo
Published: June 2021

Objective

Chronic hypoxia attenuates pregnancy‐induced increase in uterine artery blood flow. Previous studies demonstrated that ERα was downregulated in uterine arteries by chronic hypoxia during gestation, leading to heightened uterine arterial myogenic tone. This study examined transcriptional mechanisms of ERα promoter in ovine uterine arteries.

Methods

We cloned a 2 kb ovine ERα promoter region, which is about 97% homologous to bovine ERα promoter.

Results

The ERα promoter contains SP1, PR‐AB, and a non‐canonical USF binding sites, each having CpG dinucleotides. Authenticity of the transcription factors recognizing these sites were confirmed by EMSA and super shift experiments. Deletion of each site significantly attenuated ERα promoter reporter gene activity. SP1, but not USF and PR‐AB, sites, were hypomethylated in pregnant as compared with nonpregnant uterine arteries. In contrast, chronic hypoxia during gestation resulted in hypermethylation of SP1 and USF, but not PR‐AB, binding sites in the uterine arteries. Additionally, CpG methylation inhibited the binding of transcription factors to ERα promoter.

Conclusions

The results demonstrate a novel mechanism of promoter methylation in transcriptional regulation of ERα gene expression patterns in the uterine arteries in response to pregnancy and hypoxia.

(Supported in part by NIH grants HL89012 and HL110125)
5‐azacytidine influences ATP synthase gene expression and ATP levels in Escherichia coli
Authors: Cantatore Sarah J , Militello Kevin T
Published: June 2021

5‐azacytidine is a nucleoside analog that once incorporated into DNA, traps cytosine‐5 DNA methyltransferases and blocks further DNA methylation. Thus, 5‐azacytidine is commonly used as a DNA demethylating agent. We have performed gene expression DNA microarray analysis of E. coli cells treated with 5‐azacytidine to determine if cytosine DNA methylation influences gene expression. To our surprise, expression of all eight ATP synthase genes was up‐regulated in response to 5‐azacytidine. The effect was observed at early stationary phase, and not observed in logarithmic phase. To determine if cells treated with 5‐azacytidine have altered levels of ATP, we used a luciferase assay to measure ATP levels in untreated and 5‐azacytidine‐treated cells. The change in ATP levels in response to 5‐azacytidine is complex. At logarithmic phase, there are increased levels of ATP in 5‐ azacytidine‐treated cells, but this increase subsides as cells reach stationary phase. The increase in ATP levels in 5‐azacytidine‐treated cells at logarithmic phase was observed in a cytosine DNA methylation deficient strain (dcm knockout). These data indicate the model of DNA methyltransferase trapping cannot explain the changes in ATP levels, as this strain lacks a cytosine DNA methyltransferase. These data also indicate that 5‐azacytidine does not cause its effect on ATP metabolism through a loss of DNA methylation, and may occur via a novel mechanism. This work was supported by the Geneseo Foundation.
Enhancing Directed Double Strand Breaks with the Ref and RecA Protein
Authors: Won Sang Joon , Cox Michael M.
Published: June 2021

The Ref protein derived from bacteriophage P1 enhances RecA dependent DNA recombination via an unusual mechanism in vivo. Ref, acting in concert with the RecA protein, is a universal restriction enzyme that promotes DNA cleavage at any desired sequence in vitro. Ref is a RecA dependent nuclease that only cleaves DNA in the presence of an active RecA filament. More interestingly, DNA cleavage can be restricted to the sequence within a D‐loop formed by RecA‐mediated homologous pairing. The mechanism of such a remarkable reaction is still unknown and the yield of the double strand DNA (dsDNA) break is relatively low. Briefly, in this in vitro assay, a RecA filament is first formed on an oligonucleotide. Once the RecA filament is active, ldsDNA and Ref are added to catalyze the cleavage of dsDNA within the D‐loop. I report that a relatively high pH (pH 8.9), the use of RecA variants such as RecA E38K, and a high magnesium concentration (15 mM), can significantly enhance the reaction. The targeted DNA cleavage reaction is dependent on variables that contribute to the stability of D‐loops and RecA filaments. Optimization of the reaction could possibly yield a new method for gene‐function analysis, gene therapy, and construction of artificial chromosomes. This work was supported by a grant from the National Institutes of Health (GM32335, to MMC), a Hilldale Undergraduate Research Fellowship, and a Mary Shine Peterson Scholarship.
Regulation of MSH2 activity by acetylation and ubiquitylation
Authors: Tong Dan , Zhang Mu , Xiang Shengyan , Gu Liya , Zhang Xiaohong , Li Guo-min , Huang Jian
Published: June 2021

DNA mismatch repair (MMR) plays an important role in maintaining genome stability. Defects in MMR lead to certain types of cancer syndrome, including hereditary non‐polyposis colorectal cancer. MSH2 is a key component in MMR, and it interacts with MSH6 and MSH3 to form the MutSα and MutSβ heterodimers. Recent studies have shown that the half‐life of MSH2 is regulated by HDAC6‐catalyzed acetylation or ubiquitylation on lysine (K) residues 249, 332, 819 and 892. To determine the effect of HDAC6 on MMR function, nuclear extracts derived from mouse embryotic fibroblasts(MEFs) that are proficient and deficient in HDAC6 were examined for MMR activity using a functional in vitro assay. We demonstrate here that HDAC6‐deficient MEFs exhibit an MMR activity that is more active than that in HDAC6‐proficient MEFs, suggesting that HDAC6 negatively regulates MMR function. To distinguish if the reduced MMR activity by HDAC6 is due to acetylation or ubiquitylation of MSH2, K249, K332, K819 and K892(4K) of MSH2 were substituted for glutamines(4KQ) or arginines(4KR), which mimic acetylation or mimic deacetylation status as well as making the protein unubiquitinated, respectively. The resulting MSH2‐4KQ and MSH2‐4KR were allowed to form MutSα‐4KQ and MutSα‐4KR, and tested for their biochemical activities, including mismatch binding, ATP/ADP binding, ATPase, and sliding along the DNA helix. Our results show that in comparison with wild type MutSα, both MutSα‐4KQ and MutSα‐4KR show reduced biochemical activities. However, MutSα‐4KQ appears to be more active than MutSα‐4KR in each activity examined. These results suggest that HDAC6 negatively regulates MSH2 function, probably through ubiquitylation, which either leads to MSH2 degradation via proteasome or directly inhibits MSH2 activity.
The Functional Role of the Mre11 Dimeric Interface
Authors: Albrecht Dustin , Herdendorf Timothy , Nelson Scott
Published: June 2021

The Mre11/Rad50 (MR) complex is a central player in DNA repair and is implicated in the processing of DNA ends caused by damage‐induced double strand breaks. Rad50 is a member of the ABC protein family, which are dimeric enzymes that bind and hydrolyze ATP. Mre11 is a ssDNA endonuclease and a dsDNA exonuclease that forms a heterotetrameric complex (M2/R2) with Rad50. Recent structures indicate that the MR complex undergoes extensive conformational rearrangements during ATP hydrolysis and nucleotide excision. Notably, two forms of the Mre11 dimer interface have been observed, denoted here as ‘open’ and closed’. In order to understand the significance of these two states, we have generated and characterized a bacteriophage T4 Mre11 mutant (Mre11L101D) that is expected to disrupt the closed form of the Mre11 dimer. The variety of functional assays suggest that the Mre11 dimer interface undergoes ATP‐driven conformational rearrangements (i.e., an opening and closing) during the nuclease reaction cycle. Productive complex assembly and the nucleolytic reaction occur when Mre11 is in the open state, whereas translocation of the MR along the DNA substrate requires formation of the closed state.
Mechanism and Regulation of the Helicase‐driven Path of DNA End Resection in Saccharomyces cerevisiae
Authors: Niu Hengyao , Chen Xuefeng , Kwon Youngho , Chung Woo-Hyun , Zhu Zhu , Ira Grzegorz , Sung Patrick
Published: June 2021

The ends of DNA double‐strand breaks (DSBs) are resected in a cell‐cycle dependent fashion, to yield 3′ overhanging single‐stranded DNA (ssDNA) for the recruitment of factors critical for DNA damage checkpoint activation and repair by homologous recombination. As defined in studies in the budding yeast Saccharomyces cerevisiae, the DNA end resection process involves multiple, redundant pathways, one of which entails the Sgs1 helicase and its partners Rmi1‐Top3, Dna2 helicase/nuclease, Mre11‐Rad50‐Xrs2 (MRX) complex, and the heterotrimeric ssDNA‐binding protein RPA. We have reconstituted the DNA end resection reaction using the aforementioned factors. To gain further insight into how Cdk1 kinase activity regulates the motor‐driven resection pathway, we have purified the Cdk1‐Clb2 complex and examined its effects both in our reconstituted system and in conjunction with genetic analysis. Our studies indicate that one means of Cdk1 regulation is via direct phosphorylation of Dna2 at three consensus sites located within its amino‐terminus, which promotes efficient recruitment of Dna2 to DSBs. However, phospho‐mimic mutations of Dna2 cannot fully bypass the requirement of Cdk1 activity in vivo, raising the possibility that Cdk1 also targets other factor(s), an idea that is being examined using our reconstituted system. The biochemical and genetic systems that we have established should serve as a useful model for delineating the mechanistic intricacy of the DNA break resection process in eukaryotes.
Enhanced apoptosis and altered DNA repair underlie improved outcomes in HPV‐positive head and neck cancer
Authors: Kimple Randall J , Torres Alexandra D , Blitzer Grace C , Smith Molly A , Armstrong Eric A , Lambert Paul F , Harari Paul M
Published: June 2021

PURPOSE

To investigate molecular mechanisms underlying improved clinical outcome in patients with human papillomavirus (HPV)‐positive head and neck cancers (HNC).

METHODS

Radiation (RT) and chemotherapy response was assessed by clonogenic survival and proliferation assays. Cell line xenografts were used to assess response to chemoradiation with either cisplatin or cetuximab. Cell cycle distribution, apoptosis, and DNA damage response were assessed by flow cytometry, immunohistochemistry, and western blot.

RESULTS

HPV‐positive cells showed greater sensitivity to RT than HPV‐negative cells (mean SF2: 0.27 vs. 0.64, p<0.0001), but similar sensitivity to chemotherapy. Compared to RT alone (mean time to quadruple = 19 d), the concurrent use of either cisplatin (33 d, p=0.0011) or cetuximab (83 d, p<0.0001) induced a significant delay in tumor xenograft growth. RT induced a greater G2 arrest in HPV‐positive compared with HPV‐negative cells (mean increase 36% vs. 15%, p=0.0035). Following RT, p53 expression was increased and significant induction of apoptosis was seen at 24 and 48 hours. Between 4 and 24 hours post‐RT, significantly more γ‐H2AX foci were observed (42% vs. 80%, p=0.017).

CONCLUSIONS

Improved tumor response of HPV‐positive HNC may reflect increased sensitivity to radiation and chemoradiation mediated through G2 arrest, apoptosis, and altered DNA repair kinetics.
The Anti‐oxidant Cysteamine Confers a Protective Effect Against Oxidative Damage in Saccharomyces cerevisiae in the Absence of Endogenous Repair Machinery
Authors: Steele Allison M , Chikwana Edward , Mordan-McCombs Sarah
Published: June 2021

In vitro chemical studies have shown that natural anti‐oxidants are beneficial compounds that prevent and may help repair DNA damage caused by intracellular reactive oxygen species (ROS), such as hydrogen peroxide. Thiols (RSH), naturally‐occurring, electron‐rich, biological compounds, donate electrons to neutralize free radicals, removing them from the body. Here we seek to elucidate the biochemical pathway by which the thiol cysteamine protects against oxidation by hydrogen peroxide in the absence of endogenous oxidative stress or DNA repair machinery. Previous investigations have shown that Saccharomyces cerevisiae gene deletion strains lacking the ctt1 gene exhibit increased growth in the presence of cysteamine as compared to their untreated counterparts when assaulted with hydrogen peroxide. The current study investigates the potential protective effects of cysteamine in other S. cerevisiae knock‐out strains. Western Blot analysis is used to test protein levels in the oxidative response and DNA repair pathways in the presence and absence of cysteamine.
FANCA has intrinsic affinity to nucleic acids with preference for single‐stranded forms
Authors: Yuan Fenghua , Qian Liangyue , Zhao Xinliang , Liu Jesse Y , Song Limin , D'Urso Gennaro , Jain Chaitanya , Zhang Yanbin
Published: June 2021

Fanconi anemia complementation group A (FANCA) gene is one of the 15 disease‐causing genes and has been found to be mutated in ~60% of Fanconi anemia patients. Using purified protein, we report that human FANCA has intrinsic affinity to nucleic acids. FANCA binds to both single‐stranded and double‐stranded DNA, however, its affinity to single stranded DNA is significantly higher than to double‐stranded DNA in an electrophoretic mobility shift assay. FANCA also binds to RNA with an intriguingly higher affinity than its DNA counterpart. FANCA requires a certain length of nucleic acids for optimal binding. Using DNA and RNA ladders, we determined that the minimum numbers of nucleotides required for FANCA recognition are ~30 for both DNA and RNA. By testing the affinity between FANCA and a variety of DNA structures, we found that a 5′ flap or tail on DNA facilitates its interaction with FANCA. A patient‐derived FANCA truncation mutant, Q772X, has diminished affinity to both DNA and RNA. On the contrary, the complementing C‐terminal fragment of Q772X, C772‐1455, retains the differentiated nucleic acids binding activity (RNA>ssDNA>dsDNA), indicating that the nucleic acids binding domain of FANCA is primarily located at its C‐terminus where most disease causing mutations are found.