bluuebunny/crossref_metadata_2025 huggingface dataset:
Association between sleep, and dietary habits and exercise among adolescents (47.2)
Authors: Heskey Celine , Sirirat Rawiwan , Chen‐Chen Phang , Abigail Clarke , Siapco Gina , Sabate Joan
Published: June 2021

Inadequate sleep has been associated with increased adiposity and BMI among adolescents. Lack of sleep may influence dietary behaviors. The aim of this study is to determine the association between sleep, and dietary habits and exercise among adolescents. In this cross‐sectional study, 581 adolescents (47% males) ages 11‐19 years from 10 selected middle and high schools in Southern California and Michigan completed an online survey, which included a food frequency questionnaire and questions on lifestyle and sleep habits. Anthropometric measurements were taken during school visits. Data was analyzed using descriptives, t‐tests, ANOVA and χ2 with adjustment for relevant confounders. Physical activity was significantly associated with sleep duration (p=0.0431). Subjects who slept less than 8‐9 hours per night were more likely to exercise 蠅90 minutes per week compared to those who slept <8 or >9 hours. Sleep duration was significantly associated with breakfast consumption (p<0.0001). 74% of those who slept >9 hours always ate breakfast compared to 61% and 41% of those who slept for 8‐9 hours and <8 hours, respectively. Sleep duration was significantly associated with snack consumption after dinner in male subjects (p=0.035). Male subjects who slept 8‐9 hours were less likely to eat a snack after bedtime compared with those who slept <8 hours or >9 hours. Bedtime was significantly associated with type of lunch eaten (p=0.02). Students who went to bed after 9 pm were less likely to consume a convenience lunch (fast food or store‐bought) when compared to those who went to bed before nine. Sleeping patterns are associated with physical activity and dietary habits, which may have implications in weight management.
Conformational exchange in the mechanism of multidrug efflux by EmrE (470.3)
Authors: Henzler‐Wildman Katherine
Published: June 2021

EmrE is a small multidrug resistance efflux pump found in the inner membrane of E. coli. As a secondary active transporter, it imports two protons down their electrochemical gradient to drive the export of one polyaromatic cation substrate. By pumping polyaromatic cations out of the cell, it thus confers resistance to this broad class of substrates and contributes to antibiotic resistance.

As a multidrug transporter, EmrE transports a broad range of substrates with widely varying affinities. How does such a small transporter with a single small active site bind and transport this broad array of ligands? We have studied a series of tetrahedral compounds, as well as several planar substrates, to examine multidrug recognition and transport by EmrE. Using ZZ‐exchange NMR spectroscopy and lineshape analysis we find that the rate of conformational exchange between inward‐ and outward‐facing states varies over several orders of magnitude even within this limited set of substrates. Thus, the identity of the bound ligand controls the rate of this critical step in the transport process. The binding affinity also varies over a similar range and our data indicate that ligand off‐rate and conformational exchange rate are of similar magnitude for most ligands, indicating both processes may influence the rate of overall drug efflux. Our results suggest that tight binding ligands may stabilize the ground state sufficiently to block conformational exchange, and thus efflux. Assay in E. coli support this hypothesis, demonstrating that tight binding substrates reduce the rate of ethidium efflux by EmrE in its native environment.
Protein quality control in the endoplasmic reticulum (472.3)
Authors: Ploegh Hidde
Published: June 2021

In the context of our studies on host‐pathogen interactions we have developed a series of tools that are likely to be of broad utility. These tools include the development of a sortase transacylation‐based system for protein modification, which allows the stoichiometric and site‐specific installation of a wide variety of functionalities that cannot be provided in a template‐encoded manner. The method is particularly useful for site‐specific modification of type II membrane proteins on living cells. We have applied this method to labeling of the flu virus glycoproteins HA and N, both of which are refractory to tagging with GFP in the context of an infectious virus. We have developed a semi‐intact cell system, based on selective perforation of the plasma membrane with perfringolysin‐O. This preparation allows us to track the biogenesis of flu particles in detail. In this analysis we make use of single domain antibody fragments, derived from heavy chain‐only antibodies produced in camelids, to interfere with select steps in the virus assembly process. This analysis has revealed the participation of host factors that were not uncovered in any of the published genome‐wide shRNA screens. Finally, on the host side, we have used somatic cell nuclear transfer to generate new mouse models with which to study the interactions of flu with flu‐specific B cells. This approach uncovered the requirement for an internalizing receptor, in addition to surface‐disposed sialic acids to mediate entry of the virus.
The role of phosphorylation and acetylation in modulating mitochondrial metabolism (474.1)
Authors: Pagliarini Dave
Published: June 2021

Mitochondria are complex and dynamic organelles that are essential to the survival of nearly every eukaryotic cell. To generate a foundation for systematic investigations of mitochondrial function and adaptation, we recently established a protein compendium of these organelles across a wide range of tissues. This resource, termed MitoCarta, provides a robust, yet static view of the mitochondrial proteome. We are now applying MitoCarta as a framework for quantifying how mitochondrial proteins and post‐translational modifications (PTMs) change during metabolic perturbations, and to elucidate the roles of these changes in regulating mitochondrial activity. To do so, we blend large‐scale mass spectrometry‐based proteomics with focused biochemistry and molecular biology approaches.

In particular, we have recently taken this approach to capture the mitochondrial proteome dynamics during fasting, the onset of obesity, aging, caloric restriction and acute iron deprivation. Our analyses have revealed hundreds of dynamic phosphorylation and acetylation events and have enabled the production of quantitative, searchable maps of mitochondrial alterations across a spectrum of metabolic states. We have leveraged these data to demonstrate that key steps in ketogenesis and branched‐chain amino acid degradation are regulated by reversible PTMs. Moving forward, we plan to further elucidate the mitochondrial signaling network by identifying the regulatory enzymes responsible for managing mitochondrial PTMs, and to define the functions of additional mitochondrial PTMs that are altered in human disease.
Large‐scale development and implementation of highly sensitive, multiplex MRM‐based assays for quantification of proteins in biological settings (475.2)
Authors: Paulovich Amanda
Published: June 2021

Proteins carry out the biological functions of our cells and form the basis of diagnostic tests and treatments, yet over 95% of human proteins can’t be studied because we lack reliable laboratory methods (assays) for quantifying their abundances. This lack of standardized assays for quantifying the vast majority of human proteins has left the proteome clinically inaccessible, contributed to poor inter‐laboratory reproducibility of preclinical research, and indeed is the biggest impediment to translating novel protein diagnostics into clinical use, as well as a major impediment to systems biology studies needing to interrogate biologically relevant numbers of samples. Selected/multiple reaction monitoring mass spectrometry (S/MRM), especially coupled to the enrichment of target analytes, promises to relieve this limitation by offering a robust platform that enables efficient generation of highly sensitive, multiplex‐able assays that are standardized to support high reproducibility across laboratories and platforms. This seminar will review current data on the large‐scale generation, implementation, and performance of highly multiplex MRM‐based assays in biological settings relevant to cancer. En masse, these studies establish the feasibility and utility of scaling to develop analytically validated, MRM‐based assays to quantify all human proteins.
Single molecule sorting of the DNA repair machines (477.1)
Authors: Spies Maria
Published: June 2021

The functional diversity and dynamic nature of the eukaryotic proteome are highly enhanced by posttranslational modifications (PTMs). DNA repair enzymes are often controlled by reversible PTMs that modulate their activities and interactions with their targets to broaden the range of repairable DNA lesions. A fraction of a DNA repair protein carrying an activating or modifying PTM may greatly influence cellular fate. To expedite the quantification, analysis and comparison of distinct posttranslationally modified pools of a protein we developed a technique to simultaneously, but separately evaluate the activities of modified and unmodified proteins extracted from the human cells. I will describe the experimental strategy behind “single‐molecule sorting”, as well as its application to the analysis of several DNA repair helicases, which produce transient single‐stranded intermediates of DNA replication, recombination and repair. Individual protein molecules isolated from human cells with their native PTMs are analyzed using total internal reflection fluorescence microscopy (TIRFM). Separation of the activity trajectories originated from modified and unmodified enzymes allows us to quantify the fraction of the protein modified in the cell and to assess the effect of each modification or a combination of modifications on the target activity.
A SUMO Atlas shoulders the DNA damage response (477.3)
Authors: Zhao Xiaolan , Chung Inn , Sarangi Prabha , Wei Lei
Published: June 2021

The maintenance of genomic integrity requires multiple types of protein modification that are readily reversible and therefore capable of dynamic signaling and regulation. Our lab has been studying how protein modification by SUMO (small ubiquitin‐like modifier) contributes to the maintenance of the genome. Studies from our lab and others have shown that mutating sumoylation and desumoylation enzymes results in multiple defects in DNA repair and chromosomal segregation. For example, mutating the sumoylation E2 and the Mms21 E3 enzymes in yeast leads to the accumulation of toxic recombination intermediates when cells replicate over damaged templates. Some of these defects can be partly explained by the lack of modification of known DNA metabolism substrates, such as PCNA, Rad52, and TDG, sumoylation of which can modulate their DNA repair functions. However, majority of the phenotypes seen in mutants of the sumoylation enzymes cannot be explained by the lack of sumoylation of the previously known SUMO substrates. This indicates the existence of additional targets that contribute to the roles of sumoylation in genome maintenance. We have undertaken a biochemical approach to identify new sumoylation substrates and have examined how sumoylation contributes to the cellular response to DNA damage, both at a global level and at the individual substrate level. Our results indicate the existence of an extensive DNA damage‐induced sumoylation system that makes multiple contributions to genome stability and operates largely independent of the canonical DNA damage checkpoint kinase. Our progress to understand this new system and its effects will be reported and discussed.
Evolution of the role of Cdc48 and its interactions with the 20S peptidase (478.1)
Authors: Sauer Robert , Barthelme Dominik , Grabenstatter Jon , Jauregui Ruben , Chen James , Baker Tania
Published: June 2021

Proteasomes are ubiquitous ATP‐dependent proteases that function in eukarya, archaea, and actinobacteria. These proteolytic machines employ a barrel‐shaped 20S core peptidase and a hexameric AAA+ unfolding ring that unfolds and spools substrates into the chamber of the peptidase. Interestingly, the 20S peptidase can function with two very different types of AAA+ unfolding rings. For example, archaeal 20S can combine either with the double‐ring Cdc48 unfoldase or the single‐ring PAN unfoldase to execute ATP‐dependent proteolysis. Single‐ring AAA+ unfoldases that are homologous to PAN also function with the 20S in protein degradation in eukarya (the Rpt1‐6 ring of the 26S proteasome) and actinobacteria (Mpa). We will discuss differences between single‐ring and double‐ring AAA+ unfoldases, how the interactions between the 20S peptidase and its AAA+ partner are controlled, and possible mechanisms by which the N domain of Cdc48 controls rates of ATP hydrolysis, substrate unfolding, and affinity for the 20S peptidase.
Role of intestinal fibroblasts in inflammation across the intestinal epithelium (488.10)
Authors: Anderson Meghan , Smalls Anita , Reaves Titus
Published: June 2021

Fibroblasts are mesenchymal cells that produce extracellular matrix (ECM) proteins and play a major role in tissue repair (i.e., resolution of inflammation). Fibroblasts can affect cells in the local environment through the release of cytokines, growth and differentiation factors. CD36 is a membrane‐bound and scavenger receptor that is an indicator of inflammation and interacts with TLR‐2, 4, and 6. We investigated intestinal fibroblasts for expression of immune receptors and for a role of CD36 in intestinal inflammation using fibroblasts exposed to a series of cytokines. Analysis of supernatants revealed the presence of IL‐8 (potent chemoattractant for polymorphonuclear leukocytes, (PMN)) in IL‐6 treated normal and CD36‐deficient fibroblasts. However, results reveal a statistically significant reduction in PMN migration using IL‐6 treated CD36‐deficient cells. Kinase inhibitor experiments reveal that PI3 kinase pathway may be involved in the lL‐6 mediated release of IL‐8 from fibroblasts. Fibroblasts exposed to IL‐8 have been reported to display a myofibroblast‐like phenotype. Differences in cell adhesion and migration were also revealed using CD36‐deficient fibroblasts. Results show that fibroblasts can play a major role in the inflammatory responses in the intestine, PI3 kinase may be involved, and highlight CD36 as a target‐receptor that may modulate the inflammatory responses of fibroblasts.

Grant Funding Source: NSF‐RII, NIH/NIDDK 5T35 DK007431
WNT1‐inducible protein 1 regulates intestinal mucosal wound resealing (488.9)
Authors: Nishio Hikaru , Neumann Philipp , Leoni Giovanna , Quiros Miguel , Bernal Gabriela , Feng Mingli , Parkos Charles , Nusrat Asma
Published: June 2021

The intestinal epithelium has a remarkable capacity to migrate and proliferate to reseal wounds and re‐establish the epithelial barrier under conditions of inflammation. We report that a matricellular protein transcriptionally regulated by WNT/β‐catenin pathway referred to as WNT1‐inducible signaling protein 1 (WISP1) is upregulated in resealing colonic epithelial wounds in vitro and in vivo. Similarly, we observed increased WISP1 protein and mRNA expression in the intestinal epithelium of patients with active inflammatory bowel disease. Exogenous administration of WISP 1 promoted wound closure of model intestinal epithelial cell lines in vitro through increased epithelial proliferation and cell migration. The beneficial effects of WISP1 on wound closure were mediated by activation of focal adhesion kinase and c‐Src‐mediated signaling. These findings highlight a mechanism by which a WNT/β‐catenin target protein released by injured epithelial cells promotes mucosal wound closure.
Concept mapping in the physiology lab to promote student learning (530.7)
Authors: Karnatz Matthew , Ackerman Ralph
Published: June 2021

We report on a two year study to determine the effectiveness of concept mapping to promote student learning through conceptualization of physiological knowledge. Students were enrolled in a 300 level physiology lecture with a lab component. We worked only in the lab for concept mapping, where, during the first year we started halfway through the semester and had several full labs on concept mapping covering respiratory, blood pressure, and fluid balance control. We use concept mapping to focus more on conceptualization and application instead of rote memorization (however, students need to know the proper terms to create a good map). The second year we integrated concept mapping into the normal labs as pre‐labs with an initial lab on “how to create a concept map”. This would be typical of an instructor initially adding concept mapping to their existing labs. We discuss how to implement concept mapping into a lab using both years as case studies. The first year results focus on student opinion and logistics within the lab. The second year results focus on streamlining the procedures that worked and experimentally testing the effect of using concept maps as pre labs on student performance. Students report higher learning on concept map labs as well as feeling they better prepare them for the future.
PORTAAL: a practical observation rubric to assess active learning in the college science classroom (531.9)
Authors: Wenderoth Mary Pat , Eddy Sarah , Converse Mercedes
Published: June 2021

Active learning is defined and implemented in a variety of ways. Yet how activities are implemented in the classroom may be key to whether student learn. Recognizing the cognitive science underpinning the activity is therefore an important first step to successfully implementation. Unfortunately, there are few easy to use observation tools to assess implementation of active learning. We created our tool/rubric by first reviewing the literature and identifying four key dimensions shown to increase student learning: 1) Ericsson’s “deliberate practice” (1998); 2) encouraging meaningful participation through accountability addresses the need for attention in learning (Bjork 2002) & Chi’s Active‐Constructive‐Interactive framework (Chi 2009); 3) logic development is an opportunity for students to generate, interpret and elaborate on their understanding (deWinstanley & Bjork 2002) and 4) reducing student apprehension which reinforces Dweck’s mindset work (1999). We deconstructed each dimension into 28 different elements observable in a classroom and used PORTAAL: a Practical Observation Rubric to Assess Active Learning to observe 27 different instructors across two years of introductory Biology. PORTAAL was able to successfully differentiate a wide range of active learning behaviors and future work will correlate student academic performance in class with PORTAAL results.

Grant Funding Source: NSF DUE 1118890
Prospecting the use of quick response codes in the gross anatomy laboratory (533.1)
Authors: Traser Courtney , Hoffman Leslie , Seifert Mark , Wilson Adam
Published: June 2021

Background & Purpose The use of quick response (QR) codes in institutions of higher education is becoming increasingly popular, yet literature concerning their use in medical education is scant. This study examined student perceptions of the usefulness of QR codes as study‐aids in a Medical Gross Anatomy course and statistically analyzed whether this resource impacted student performance.

Methods Question prompts and QR codes tagged on cadaveric specimens and models were made available for four weeks as a learning resource to medical (n=155) and doctor of physical therapy (n=39) students. Each QR code provided answers to posed questions in the form of embedded text or external links. Students’ perceptions were gathered using a formative questionnaire and practical exam scores were used to assess potential gains in student achievement.

Results Overall, participants responded positively to the idea of using QR codes in the laboratory. Majority of respondents (64/73) used the QR codes at least once, 89% (57/64) of students agreed the codes augmented their learning of anatomy, and 56% (36/64) found the codes to be more useful than traditional laboratory study‐aids. The students’ most noticeable objection to using QR codes was the reluctance to bring their smartphones into the lab. A repeated measures analysis of covariance revealed no statistical difference (p=0.108) between unit exam scores on practical exam 1 (pre‐intervention) and practical exam 2 (post‐intervention).

Conclusion This study suggests that learners welcome the innovative implementation of QR code technology in the gross anatomy laboratory and such an intervention has no negative effects on practical exam performance.
improving performance on gross anatomy practical examination through weekly quizzes (533.7)
Authors: Harper Noah , Morton David
Published: June 2021

A standard to assess first year medical students’ knowledge of gross anatomy is a timed practical examination. Research indicates weekly quizzes are associated with better academic success in an anatomy course, and they strengthen academic performance, lower exam anxiety, and support student‐teacher communication. Previously, this research team investigated if students who had taken an anatomy course prior to medical school performed better in anatomy than students who had not. Results indicated that students who took anatomy with cadavers earned higher grades on the final cadaver practical exam (in a statistically significant manner) than students that had either no anatomy or anatomy without cadavers. Based on this information, the questions we now address are: 1) Does the frequency of administration of practical exams have an effect on final practicum performance?, and 2) Will weekly practical quizzes effect the performance disparities between students who have had cadaver‐based anatomy as premedical students and those who have not? To address these questions, weekly practical quizzes consisting of 10 questions related to material covered in the previous week's lab were administered at the beginning of each lab. Final gross anatomy practical scores were then compared to those of the previous four years. We expect results and conclusions to be available late January, 2014.
Development of a pre‐medical embryology course using backward course design (534.1)
Authors: Cassidy Keely
Published: June 2021

Medical students often bemoan a lack of previous experience with embryology, to their detriment in the early years of medical school. To fulfill this need, an upper‐level pre‐medical embryology course was developed using backward course design in which the instructor first identified the desired results and determined acceptable evidence, then planned learning experiences and instruction upon that framework. It structured student learning on assessments intended to provide evidence that students have achieved course goals: mastery of embryological processes and structural relationships leading to complex adult anatomy and analysis of birth defects. Enrollment was 21 students, most in nursing or pre‐medical programs, and their career goals shaped the final assessment: write a plan for a hypothetical family expecting a child with a certain birth defect, focusing on the embryological basis, prevention, and treatment. Course content decisions were based on topic density and overlap with other courses, and lectures, exams, adult donors, and active learning (TBL, journal article analyses, metacognitive blogs) were included in the course curriculum.
The Pre‐Medical Sciences Master’s program at Texas Tech University Health Sciences Center: training better medical students (534.9)
Authors: Schneider Brandt , Lee Vaughan
Published: June 2021

The TTUHSC Graduate School of Biomedical Sciences has developed a 2‐year non‐thesis Pre‐Medical Sciences Master’s program (http://www.ttuhsc.edu/gsbs/mspremedsci.aspx) designed for students who need additional preparation for medical school or whose goal is anatomical education. Students are indistinguishably immersed with incoming medical students and participate alongside medical students in class, lab, and small group activities in the first year curriculum. During the second year, students participate as teaching assistants and take an Advanced Cell Biology course with PhD students. In addition, students design and carry out an education project including either the development of self‐directed learning units or the creation of novel educational materials. This program has exceeded expectations. It greatly promotes inter‐professional teamwork, stimulates independent learning, helps meet the teaching mission in light of reduced numbers of anatomical educators, and provides a better educational environment for our medical students. Historically, after graduation, all of our students who applied have been admitted to and excel in medical school. This program improves the educational training of our current medical students and provides our school with excellent future medical students.

Grant Funding Source: This project was funded by the Graduate School of Biomedical Sciences at TTUHSC.
Learner characteristics involved in visuospatial anatomy comprehension (535.1)
Authors: Glena Katlyn , Johnson Marjorie , Allman Brian , Zibrowski Elaine , Elizabeth Hampson , Nguyen Ngan
Published: June 2021

The practical aspect of human anatomy instruction is becoming less dissection based and increasingly more reliant on computerized, three‐dimensional (3D) teaching tools. Some students find it difficult to learn from these tools and it is important not to alienate this group of learners. Previous studies have demonstrated that the spatial relations subcomponent of spatial ability is predictive of success in 3D anatomy comprehension, but the effect of other spatial ability subcomponents has not been examined. This study explored the role of three subcomponents of spatial ability and the effect of prior domain knowledge in comprehending a 3D teaching tool. By adding more spatial ability subcomponents and a knowledge component the results may provide a broader picture of the relevant learner characteristics involved in comprehending these tools. 60 students from Western University were recruited. Participants performed an anatomy knowledge test (AKT), three standardized spatial tests, and a pre and post spatial anatomy task (SAT) with a 3D teaching tool as an intervention. Preliminary data analysis (N=24) suggests that participants’ scores on the AKT and the three spatial ability tests are predictive of learning from the 3D teaching tool (p<0.01). These results may have implications for the appropriate design and implementation of computerized 3D instructional animations.

Grant Funding Source: Lippincott Williams Wilkins/AAA Education Research Scholarship
CANmeds physician competencies inadvertently realized in the gross anatomy lab (535.4)
Authors: Farias Anna , Awuku Mark
Published: June 2021

Introduction: The cadaveric gross anatomy lab is a learning environment in which the students learn multiple skills and tasks. In the process of learning the intricacies of the human body, the students learn, demonstrate and acquire competencies such as professionalism, collaboration, communication and scholarly learning. This study was based on student evaluations to measure the degree to which the listed competencies are realised in the Gross Anatomy Lab.

Observations:The first lab session starts with a memorial service to honour and respect the generous “gift” by the donors. This sets the tone for gratitude, respect and responsibility with which the learners treat their ‘first patient’ whenever they are in the lab. They approach the learning as a group and share the information and participate in the discussions among themselves as well as the rest of the class when there is an unusual finding. The learning environment calls for collaboration and communication within the specific group and also with the rest of the class.

Learners’ feedback: At the end of each course the evaluations polled on the professionalism, collaboration and scholarly learning were rated at 4.3/5 and 4.4/5 among year one and two respectively.

Conclusion: The learning environment in the gross anatomy lab can effectively nurture competencies such as professionalism, collaboration, communication and scholarly learning in addition to medical expert. The anonymous evaluation of the learning environment, by the learners, also provided some comments regarding the strong collaborative environment and team work. The scores as well as the comments are strong indicators that the learners are realizing other competencies such as professionalism, communication and collaboration to the same level as that of medical expert. Medical schools therefore should be aware of the competencies realized in the gross anatomy lab during the process curriculum design.
Rovsing’s sign in appendicitis ő an example for the effect of erroneous translation on medical teaching (536.10)
Authors: Prosenz Julian , Hirtler Lena
Published: June 2021

Objective: To investigate whether Rovsing’s sign in the diagnosis of appendicitis is described and applied inaccurately in current literature, scientific papers and books, quantify the problem and investigate the cause of this error.

Background: The way Rovsing’s sign is elicited is not described uniformly throughout the literature.

Methods: A Pubmed search was conducted and 1069 individual studies were searched for the use of Rovsing’s sign. 53 studies were included in this study, of these 14 described the way the sign was executed. Additionally 3 current English (text)books on surgery, 3 German (text)books on surgery and 2 German books on anatomy were analyzed. The descriptions in studies and books were compared to the original publication by Niels Thorkild Rovsing in 1907.

Results: No included study which described the way Rovsing’s sign is elicited gave a correct description. Of the 3 English (text)books on surgery none described it accurately, of the 5 German (text)books all yielded a correct description of this sign.

Conclusion: Rovsing’s sign is used erroneously in clinical practice and medical research and most likely already taught incorrectly to most medical students. All statistical data available on this sign must be questioned and reinvestigated in future well‐designed studies to assess the actual value of Rovsing’s sign in the diagnosis of appendicitis.
Revisiting the first patient: review of craniofacial anatomy and its role in dermatological practice (536.3)
Authors: Adar Tony , Swerdloff Daniel , Pagano Anthony , Marquez Samuel
Published: June 2021

Dermatology is a branch of medicine that focuses on diagnosis of over 3,000 skin diseases and disorders. The unique nature of this medical discipline requires the physician to be proficient in delicate surgical procedures, pathology, and pharmacologic practice, thus dermatologists are highly skilled specialists. An anatomical workshop for residents of the department of dermatology was held at SUNY Downstate to review and augment foundational knowledge of underlying craniofacial anatomy with the goal of facilitating and translating basic principles of cutaneous surgical procedures. Intimate spatial relationships of dermal, subcutaneous and fascial layers, muscle, nerves, and bone were “brought to life” for residents rarely afforded the opportunity to perform human dissection. Important concepts reviewed included significance of the mid‐pupillary reference line, utilized in location of supraorbital, infraorbital and mental foramina for safe injection of Onabotulinum toxinA (Otx), a potent neuromuscular inhibitor. Review of muscular anatomy with particular attention to the overlapping zygomaticus minor (Zm) and orbicularis occuli (Oo) demonstrated the manner in which inadvertent delivery of Otx to the Zm can induce “relaxation” (paralysis) of Oo when injected beyond the ‘safe‐zone’ (2 mm lateral to mid‐pupillary line). Mastery of craniofacial anatomy remains indispensable for proper and safe cutaneous and cosmetic procedures.
Changing myoglobin’s paradigm: a novel link between lipids and myoglobin (537.4)
Authors: Schlater Amber , Kanatous Shane
Published: June 2021

During endurance exercise, terrestrial mammals rely on erythrocytic oxygen to fuel aerobic metabolism in working muscle. Physiological changes associated with endurance training elicit responses that increase muscular blood flow and subsequent oxygen delivery . Intramuscular oxygen stores, alternatively, appear to bear little significance in sustaining aerobic metabolism during endurance exercise, evident by the inability to appreciably release intra‐muscular stored oxygen during normoxic exercise; yet, terrestrial endurance athletes have more myoglobin (Mb) than their sedentary counterparts. Accordingly, Mb’s traditional functional paradigm pertaining to oxygen storage and transport does not appear to be fully applicable to terrestrial mammals in vivo. Here, we provide data offering alternative paradigm, where increases in Mb expression were associated with increases in lipid supplementation. C2C12 cells were cultured in normoxic or hypoxic conditions with glucose or 5% lipid. Mb assays and western blots revealed lipid hypoxic cells had higher Mb than glucose hypoxic cells and both lines of normoxic cells. Normoxic cells were also compared to soleus tissue from normoxic rats fed a high fat diet; both normoxic high fat conditions showed no change in carbon monoxide (CO)‐binding Mb, but increased total Mb protein, suggesting an increase in non‐CO‐binding Mb. Moreover, Mb increases did not parallel CN increases, but did, however, parallel increases in lipid supplementation and oxidative stress markers, suggesting an alternative pathway for Mb regulation associated with lipid and independent of CN. In light of these novel data, and in conjunction with the inability of terrestrial mammals to appreciably utilize Mb oxygen stores, we propose an alternative paradigm, whereby in the context of lipid metabolism, Mb as an oxygen store appears less relevant than Mb as scavenger of reactive oxygen species or potential transporter.
Cloning, expression and purification of recombinant Slx4 from Saccharomyces cerevisiae (539.3)
Authors: Diaz Martinez Angel , Cussiol Jose Renato , Smolka Marcus
Published: June 2021

Cells are constantly exposed to exogenous and endogenous sources of DNA damage. In the presence of genotoxic stress, the DNA damage checkpoint signaling (DDC) pathway contributes to genomic maintenance by promoting a transient cell cycle arrest and recruiting numerous repair proteins to sites of lesions, thus maintaining genomic integrity. Slx4 is an evolutionary conserved scaffold protein that is important for the cellular response to genotoxic stress. Slx4 interacts with several DNA repair factors and also with components of the DDC. Recently, our group showed that Slx4 in Saccharomyces cerevisiae is capable to interact with a DNA replication initiation protein called Dpb11, therefore dampening the checkpoint by counteracting the Rad9 adaptor protein. This competition based mechanism is important to avoid hyperactivation of the DDC which can be deleterious to the cells. In order to better understand the role of Slx4 during DDC, we decided to purify the protein and to elucidate its structure by X‐ray diffraction. Our hypothesis was that by employing the method of metal affinity chromatography using cobalt a resin we would be able to purify our Slx4 protein. Here, we cloned, expressed and purified recombinant Slx4 by heterologous expression in Escherichia coli. We cloned a truncated form of the Slx4 gene (361aa to the end) into an expression vector pET28a. We transformed the product into an E. coli expression strain Bl21 (DE3) that contains the T7 RNA polymerase to express the Slx4 gene through the T7 promoter from the pET28a vector. To induce the expression of the Slx4 gene, we added IPTG to further purify the protein. In conclusion we were able to express Slx4 (361aa‐end) in the soluble fraction of the cell and purify in its native form through metal affinity chromatography using cobalt resin.

Grant Funding Source: R25GM096955
The discovery of the pulmonary circulation by Ibn al Nafis during the 13th century: an anatomical approach (543.9)
Authors: Le Floch‐Prigent Patrice , Delaval Dominique
Published: June 2021

Harvey is usually credited of the discovery of the circulation. However Ibn‐Al‐Nafis is the real inventor of the pulmonary circulation in the late 13th century. The medical thesis of Al‐Tatawi, Frankfurt in 1924, the papers of Meyerhoff , 1935, the M.edical thesis of Chehaddé, 1955, the book of Chadli and al. demonstrated clearly this fact. Ibn Al Nafis as a great physician in Cairo published his results in the “commentaries of the anatomy of the canon of Avicenne”. We analyzed the text concerning the pulmonary circulation and concluded that not only clinical, physiological and pathological results were used but that Ibn‐Al‐Nafis performed himself several human dissections.
Less is more: optimal myoendothelial communication entails less gap junctions (546.7)
Authors: Hald Bjørn , Jacobsen Jens Christian , Welsh Donald
Published: June 2021

The myoendothelial junction (MEJ) has been assigned an integral role in the regulation of small resistance arteries. Efficient and proper information transfer between the endothelium and the smooth muscle cell (SMC) layer is thought to depend on a high density of MEJ and direct coupling facilitated by gap junctions (MEGJs) herein.

In this study, we study how heterogeneity in distribution and number of MEGJs affects the electrical communication between cell layers. We redesigned a detailed computational model of an arteriole to allow for the introduction of heterogeneity in both the number and spatial distribution of MEGJs.

In our first simulations, we maintained a contant total conductance of MEGJs across the arteriole, but reduced the number of MEGJs. If less than 20% of SMCs harbours an MEGJ, an increase in conduction strength is observed. However, reducing MEGJs also reduces the spatial control of membrane potential in the SMC layer. This may to some extent be ameliorated by an increase in SMC‐SMC coupling.

Moreover, an decrease in electrical conductance between the cell layers increased electrical spread in the endothelium but limited the radial current flow into SMCs. Thus, reducing the number and conductance of MEGJs provides for a scenario where the SMC layer across a finite distance along the vessel receives a specific quanta of charge, i.e. no decay of signal is observed in the SMC layer. This may underlie the observation of non‐decaying conduction along small arterioles.

We conclude that in contrast to common belief, a low MEGJ density enhances electrical spread along the vascular wall. Furthermore, low MEGJ density may underlie the observation of augmented conduction.
Exercise‐induced erythrocyte dysfunction in individuals with sickle cell trail (547.1)
Authors: Mukherjee Shyamali , Hardy Robert , Hinds Joseph , Das Salil
Published: June 2021

Irreversibly sickled cells (ISC) remain unaggregated in the presence of oxygen and yet fail to function normally. We reported earlier that sickle cell trait (SCT) individuals are potentially at risk with microvascular complications due to physical exercise. We also reported that human RBC membrane is heterogeneous and during hemolysis of RBCs, the RBC membrane is sliced into two populations, dense and light. The objective of this study was to characterize whether physical stress causes changes in specific biochemical and physical properties of SCT erythrocytes under conditions of (a) physical stress or (b) deoxygenation. Fractionations of RBC demonstrated differences in the density or age profiles of RBC between AA, AS and SS thus indicating a higher clearance and shortened survival rate. Our studies indicated that the exposure of SCT subjects to treadmill resulted in an increase not only in the susceptibility of red blood cells to oxidation but also in the density of RBC membrane and loss of deformability and increased membrane rigidity in AS than AA. The increased susceptibility of red blood cells to oxidation as a consequence of exercise in AS was associated with relatively excessive production of H2O2 and lipid peroxidation, and increased ATPase’s activities. The damaging consequences on their performance in these conditions could be due to tissue oxygen delivery limitations.
Rapid induction of hypothermia by the Thermosuit system profoundly reduces infarct size and anatomic zone of no‐reflow following ischemia/reperfusion in rabbit and rat hearts (547.12)
Authors: Herring Michael , Dai Wangde , Hale Sharon , Kloner Robert
Published: June 2021

Background: While hypothermia reduces myocardial infarct size, non‐invasive and rapid cooling methods are lacking. This study tests the effectiveness of a novel cooling apparatus on myocardial infarct size and no‐reflow area in two models of coronary artery occlusion (CAO).

Methods and Results: Anesthetized rabbits (protocol 1) and anesthetized rats (protocol 2) were randomized to Normothermic (N) or Hypothermic (H) groups after isolation of a proximal coronary artery. Animals were subjected to 30 minutes CAO and 3 hours reperfusion. In protocol 1, hypothermia was initiated, using the Thermosuit apparatus (convective‐immersion cooling), 5 minutes after the onset of CAO to a goal temperature of ~32°C. In protocol 2, hypothermia was initiated 2 minutes after the onset of CAO to a goal temperature of ~30°C. Goal temperature was reached in ~20 minutes. In protocol 1, hypothermia caused an 82% reduction of infarct size as a percentage of the ischemic risk zone (N, 44±5%; H; 8±2%, P<0.001) and an 89% reduction of the no‐reflow area (N, 44±4%; H, 5±1%, P<0.001). In protocol 2, hypothermia caused a 73% infarct size reduction (N, 51%±5%; H, 14±6%, P<0.001) and a 99% reduction of the no‐reflow area (N, 33±5%; H, 0.4±0.3%, P<0.001).

Conclusion: The Thermosuit device induced rapid hypothermia and limited infarct size and no‐reflow to the greatest extent ever observed in this laboratory with a single intervention.
Characterization of the nurA (DR_0836) and herA (DR_0837) genes from Deinococcus radiodurans (549.3)
Authors: Mueller Charles , Levi Jonathan , Julin Douglas
Published: June 2021

HerA proteins are members of the FtsK/HerA superfamily of ATP‐dependent DNA translocases. FtsK is a bacterial protein that translocates double‐stranded DNA during cell division. The herA gene is found in an operon with the gene for its nuclease partner NurA. In archaea, herA and nurA are essential genes that encode enzymes capable of processing a variety of DNA substrates. Evidence obtained from studying these enzymes in vitro in P. furiosus and in vivo in S. acidocaldarius indicates that they have a critical role in homologous recombination in archaea. Recent in vitro work in S. solfataricus has demonstrated a dsDNA translocase activity for the NurA/HerA complex, an activity associated with FtsK from E. coli. This result and the phylogenetic relationship between FtsK and HerA suggest HerA has a similar role in archaea as FtsK in bacteria.

Homologs of herA and nurA are found in a few bacterial genomes. In most cases, these bacteria lack a ftsK homolog. The functions of NurA and HerA in bacteria are not known. We chose to investigate the roles of NurA and HerA in Deinococcus radiodurans, typically studied for its extreme resistance to double‐strand DNA breaks. The D. radiodurans genome has homologs of nurA and herA in an operon, and it also has a ftsK gene. We made strains with deletions of either herA or nurA and characterized their basic growth properties and sensitivity to DNA damaging agents. Our results indicate that neither gene is essential in D. radiodurans, and deletion of the genes do not cause significant sensitivity to DNA damaging agents. The herA deletion strain displayed a distinct phenotype consisting of slower growth and larger cell types, which may indicate an FtsK‐like function for HerA in D. radiodurans. The HerA protein has been expressed and purified from E. coli and basic characterization is underway.

Grant Funding Source: None
Elucidation of the substrate recognition mechanism of VcCry1 (550.2)
Authors: Ahmad Rohe , Gindt Yvonne
Published: June 2021

Isothermal titration calorimetry will be used to study the temperature dependence of DNA binding to Vibrio cholerae cryptochrome 1 (VcCry1), a DNA repair enzyme. VcCry1 uses light‐driven electron transfer to repair UV damaged single strand DNA but not double strand DNA. The enzyme is a good model to study substrate binding, since the repair process requires absorption of light and is separated from the damage recognition process. VcCry1 has an active site FAD which is required for substrate binding and repair. The thermodynamics along with the quantification of the electrostatic and nonelectrostatic interactions important in the VcCry1 substrate binding will be examined and compared to DNA photolyase, a related DNA repair enzyme that binds double stranded DNA.

Grant Funding Source: Supported by The Margaret and Herman Sokol Institute for Pharmaceutical Life Sciences
Pulling a ligase out of a “HAT” (555.13)
Authors: Morgan Julie , Greer Susanna
Published: June 2021

Major histocompatibility class II (MHC II) molecules present pathogenic and tumorigenic antigens to CD4+T cells and play vital roles in regulating adaptive immune responses. MHC II is controlled at the level of transcription by the Class II Transactivator (CIITA) which is in turn regulated through a series of sequential posttranslational modification. Acetylation of CIITA by the histone acetyltransferase (HAT) pCAF, results in CIITA nuclear localization. Once nuclear, CIITA is ubiquitinated and binds the MHC II promoter. In addition to its HAT activity, pCAF was recently identified as an E3 ligase for the nuclear phospho protein Mdm2. We show here pCAF increases CIITA transactivation and ubiquitination; suggesting pCAF also drives CIITA ubiquitination. We have previously linked CIITA phosphorylation with subsequent mono‐ubiquitination and increased CIITA transactivation. We now further identify K63 linked ubiquitination of CIITA and provide novel links between acetylation, phosphorylation and ubiquitination of this critical inflammatory protein. Our data suggests particular posttranslational modifications of CIITA result in very different functional outcomes, and may enable the identification of biomarkers for immune diseases associated with dysregulated antigen presentation.
DHHC acyltransferases are regulated by palmitoylation at multiple cysteine residues (555.14)
Authors: Gottlieb Colin , Linder Maurine
Published: June 2021

DHHC acyltransferases catalyze the addition of palmitate to proteins at cysteine residues. In vitro, the enzyme forms an acyl‐enzyme intermediate prior to transferring the acyl group onto substrate proteins. The DHHC cysteine is necessary for both autoacylation and transfer. Both wild type DHHC3 and the inactive DHHS3 (C157S) mutant enzyme are palmitoylated in cells, suggesting that cysteine residues other than C157 are modified. DHHC proteins are known to oligomerize, providing a mechanism for intermolecular palmitoylation and potentially accounting for palmitoylation of the DHHS3 mutant in cells. Using a custom mass spectrometry method, we directly identified palmitate modifying C146 in DHHC3. C146 is in the conserved cysteine rich domain (CRD) of DHHC proteins. Surprisingly, metabolic labeling studies demonstrated that mutation of C146 or other cysteine residues in the CRD resulted in increased palmitoylation of the enzyme in cells, while reducing autoacylation and transfer activity in vitro. Our working model proposes that DHHC3 autoacylates at the DHHC cysteine and distributes palmitate to other sites in the CRD through both cis and trans‐palmitoylation. We hypothesize that the mutation of palmitoylation sites in the CRD perturbs secondary structure, resulting in increased accessibility of cysteines for intramolecular transfer.

Grant Funding Source: GM051466