bluuebunny/crossref_metadata_2025_split huggingface dataset:
Spectrin is a crucial component of EPEC pedestals
Authors: Ruetz Tyson Joel , Guttman Julian 
Published: June 2021

Enteropathogenic E.coli (EPEC) is one of the main pathogens contributing to roughly 2 million diarrhea related child deaths worldwide each year. Upon ingestion of contaminated food or water, EPEC colonize the intestinal tracts of their hosts, attach to the surface of enterocytes and generate characteristic attaching and effacing (A/E) lesions. These lesions involve the collapse of host microvilli and the generation of pedestal‐like structures atop infected cells. Pedestal formation and disease progression is induced by bacterial proteins (effectors) that are translocated into the host cell cytoplasm by a syringe‐like type‐three secretion system (T3SS). The three major components of the host cell cytoskeleton; actin, microtubules and intermediate filaments, are all altered during EPEC pathogenesis. We have identified another major host cytoskeletal network that is involved in EPEC disease, the spectrin cytoskeleton. Here we demonstrate that spectrin is recruited to the base of EPEC pedestals and through the use of EPEC effector mutants, we show that this localization is mediated by the translocated intimin receptor (Tir). Spectrin knockdowns using siRNA have revealed that spectrin is necessary for pedestal formation to occur. Our findings provide the first evidence of spectrin's involvement in EPEC pathogenesis and identify it as a crucial protein involved in pedestal formation and disease progression.Grant Funding Source: CIHR, NSERC
Antioxidant Effect on Myofibroblast Differentiation
Authors: Palmer Tiffany Robin , Vaughan Melville B. 
Published: June 2021

Antioxidants have been shown to decrease various fibrotic cells in culture, however little is known about their effects on dermal fibrotic cells. In this study we will investigate the effects of several different antioxidants on dermal myofibroblast differentiation. This will be accomplished by measuring the percent of myofibroblasts and cell number in differing treatment groups. The antioxidants chosen for our study include: carcinine, epigallocatechin gallate, and lycopene. Immortalized human fibroblasts treated with TGF‐β are plated at a concentration of 0.2 x 105 cells per coverslip. After incubating 72 hours, the coverslips are fixed, stained with immunofluorescence, and photographed. The preliminary results suggest that myofibroblast presence is somewhat reduced by carcinine; however this antioxidant had little effect on the cell number. Experiments with epigallocatechin gallate and lycopene are underway. The antioxidants that affect myofibroblast differentiation in the coverslips will then be tested in stress‐relaxed collagen lattices. These studies may be useful to develop future therapies for Dupuytren's contracture and other fibrotic tissue pathologies. Funded by a UCO student RCSA Award.Grant Funding Source: UCO RCSA Award
Upregulation proliferation and senescence markers in hyperplasia induced in mouse skin by sulfur mustard exposure
Authors: Chang Yoke‐Chen , Wang James D. , Soriano Melannie , Gordon Marion K. , Gerecke Donald R. 
Published: June 2021

Chemical vesicants such as sulfur mustard (SM) can produce severe tissue damage. A time‐course study using mouse ear skin exposed to a low dose (0.08 mg) of SM resulted in excessive keratinocyte hyperplasia about three days after exposure. The abnormal cell proliferation was reminiscent of that observed in several types of cancer and potential oncogenic hyperproliferative signal molecules were examined to determine if the two systems were comparable. Immunohistochemical, Western blotting and RT‐PCR studies were performed and markers of skin injury and repair were examined for various time‐points. The protein, p16‐INK4a, which triggers either growth arrest or apoptosis was hyperexpressed in the hyperplasia areas of injured skin. The p16‐INK4a co‐expressed with laminin γ2, a polypeptide that promotes epithelial sheet migration over a wound bed. The endoplasmic reticulum stress‐regulated gene, GADD153/CHOP had a similar expression pattern to that of p16‐INK4a. In contrast, both keratin 5, an intermediate filament protein of basal epithelial cells and Ki67, a marker of cells in the growth stage of the cell cycle had a much more restricted distribution. Understanding the basic mechanism of action of vesicant‐induced skin injury and repair should help to identify new targets of potential medical countermeasures against chemical vesicants. This work is supported by ES005022, EY09056, and AR055073.Grant Funding Source: ES005022, EY09056, and AR055073.
Efficient myogenic commitment of hESCs and iPSC‐derived cells on biomimetic materials replicating myoblast topography
Authors: Hwang Nathaniel Suk‐yeon , Langer Robert S 
Published: June 2021

Recent development in stem cell technology has demonstrated human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) as possible cells for cell‐based therapy and studies of particular differentiation program. However, spontaneous and heterogeneous differentiation of these cells may limit the potential use. Here, we describe the efficient generation of mesenchymal cell population from both hESCs and iPSCs having non‐turmoigenic potentianl and their commitment into myoblast. In skeletal development, increased cell‐cell contact plays critical initial steps for myogenic commitment. We demonstrate that iPSCs and hESCs‐derived cells can undergo efficient myogenic commitment by topographical cues present in their environment. We have created substrates from biomimetic materials that can replicate the mico‐ and nanoscale topography of fully differentiated skeletal myoblast. When hESCs and iPSCs‐derived mesenchymal cells were cultured on biomimetic pattern, mesenchymal cells followed the underying myoblast pattern. Furthermore, gene expression and cell fusion index showed enhanced myogenic commitment on these substrates. These results demonstrates that myogenic potential of hESCs and iPSCs‐derived cells are highly dependent on the micro‐ and nanoscale topographical cues provide by skeletal myoblast.Grant Funding Source: NIDCR (NIH)
Student interaction with a novel computer interface in a gross anatomy lab
Authors: Goldman Evan Michael 
Published: June 2021

Results. Philadelphia University retrofitted its on‐campus gross anatomy lab with a customized computer system and interface. The student use of the technology has revolutionized the way the course material is learned. The computers provide a host of interactive tools to instantly correlate the anatomy of the cadaver with radiographs, 3‐dimensional rotatable views, animations, photographs, and illustrations. Instant access is also available to the course lecture notes, study guides, muscle reference sheets, and online resources. The computer interface developed for the lab computers provides intuitive and reliable access to these materials.Conclusion. The goal of this project was to create an intuitive user‐friendly interface that students would freely choose to integrate into their dissection and studying routines. As evidenced by student feedback and instructors observations, students receive tremendous benefits from the computer access. Students are continually observed referencing online atlases, dissectors, and lecture notes. When asked questions there is greater prevalence of students providing correct answers as compared to previous years. Instructors also receive fewer basic questions from students and more questions indicative of a greater comprehension of the anatomy.
The Clinical Relevance of Teaching the Medial line Theme of the Lower Limb to Future Healthcare Professionals
Authors: Benninger Brion 
Published: June 2021

ObjectiveThis project investigated the current common overview approaches to lower limb anatomy teaching and suggests an alternative technique. Introduction. The overview of the lower limb is often described as thigh, leg and foot regions, and subdivided into anterior, posterior, medial and lateral compartments. This method guarantees all structures of the lower limb are taught. Virtually all healthcare institutions have had to distill anatomy into accelerated courses.MethodsLiterature search was conducted regarding teaching techniques of the lower limb and morphology of 60 dissected embalmed cadavers was observed to assess patterns which correlate with common clinical conditions and techniques.ResultsLiterature review revealed no specific teaching techniques. However, observations commonly revealed compartment and/or systems layout. Dissections revealed an overwhelming number of clinically important structures along the medial line of the lower limb. Reinforcement or stability structures, venous access, arterial supply, and nerves were emphasized greater along the medial line versus the lateral line.DiscussionAnatomy courses have resulted in less student‐instructor contact time. Specific techniques would aid the efficiency of learning. This project demonstrates that emphasizing lower limb structures along the medial line would be a useful technique teaching future clinicians.
Death is only the Beginning: Cadaver Use at Indiana University
Authors: Husmann Polly 
Published: June 2021

Cadaver shortages have plagued some areas of the country, yet other places have no problems fulfilling their cadaveric need. The state of Indiana has had few issues maintaining a supply of donated remains and has begun providing cadavers to other states in 2009. One possible reason for this abundance is that there is only one medical school the state is attempting to accommodate. However, it is one of the largest medical schools in the nation and thus requires many cadavers. Other possibilities include high levels of knowledge of the anatomical donation program or less restrictive criteria for donation, though none of these have been studied. This demographic study examines the origin and use of cadavers at the Indiana University School of Medicine (IUSM). De‐identified donor data from IUSM was examined including age at death, sex, cause of death, last location of residence, year of death, ethnicity, and final use of donation. Descriptive statistics were run to assess trends among demographic data and final use. Analysis revealed that most donors are Caucasian, die of chronic medical conditions, and tend to come from large cities. These data support the hypothesis that those with chronic diseases are more likely to donate their remains for research and education out of respect for and/or knowledge of health care. The study also discusses the uses of donated cadavers and the proportion of donations that are used for each purpose. Utilization of the donors typically begins within six months to one year of death and the largest numbers of cadavers are used for medical education. This information may be valuable for those deciding whether or not to donate. This study represents the first step in a larger project to compare these data with data from areas combating cadaver shortages to establish ways that these shortages may be resolved.Grant Funding Source: None
The logistics of facilitating a dissection based anatomy curriculum in a distributed medical program
Authors: Fraser Jennifer , Mogerman Emma , Krebs Claudia 
Published: June 2021

When the University of British Columbia (UBC) medical school began a distributed model for medical undergraduate education in 2004, the delivery of gross anatomy teaching needed to adapt to these new circumstances. The pedagogical decision to continue with a dissection – based anatomy curriculum, supplemented with selected prosections was made. Core gross anatomy teaching was front‐loaded into the curriculum during the first term when all students are based at the Vancouver site. Thereafter, all cadaveric material is shipped to the satellite sites so that students can continue with their anatomy education remotely. As the Body Donation Program at UBC's Vancouver campus is the only program of its kind in British Columbia, all cadavers for the distributed program are processed at UBC. The distributed medical education program has been operational for five years, and while successful, there have been some logistical and operational challenges:
The acquisition of cadaveric material.
The complexities of tracking and monitoring the anatomical material between all three sites.
The processing and maintenance of anatomical material.
The set up and monitoring of technical audio‐visual (AV) equipment in the lab.
Evaluating quality of medical students’ dissections in the gross anatomy laboratory
Authors: Kumar Neha , Hassinger James Peyton , Scoville Elizabeth A , Christensen Kevin , Nyauncho Clive , Lachman Nirusha , Pawlina Wojciech 
Published: June 2021

Gross anatomy laboratory dissection experience continues to be an integral part of the majority of gross anatomy courses in the United States. Student‐dissected specimens, typically used for practical exams administered during the course, vary in quality. In most anatomy courses the quality of cadaveric dissection is not counted toward the final grade, therefore students are not overly concerned with completing their daily laboratory assignments. Often gross anatomy teaching faculty is challenged to find good examples of dissected structures to be used for testing purposes, frequently spending additional dissecting time to secure adequate material for examinations. Dissection at Mayo Medical School is performed as a collaborative effort of a four‐member dissection team. A new method was implemented to evaluate dissection and provide the students with weekly formative feedback regarding the quality of their dissection. Team performance was graded and constituted 5% of the total grade for the course. A ten‐point evaluation tool was used that incorporated both objective and subjective measures for determining whether students had met learning objectives. Implementation of dissection evaluation resulted in the improvement of dissection quality. The dissection evaluation proved to be a valuable assessment tool in our anatomy curriculum and one that students found beneficial.Grant Funding Source: Department of Anatomy
Neuropeptides and Colon Cancer; Another Notch in the Complexity of Tumorigenesis
Authors: Light Naomi G , Benton Keith D , Hermann Rebecca J , Failing Jarrett , Dorsam Glenn P 
Published: June 2021

In 2009 an estimated 50,000 deaths in the United States will be attributed to colon cancer. Dysregulation of tumor suppressors and oncogenes are a major cause of uncontrolled cellular proliferation, leading to tumerogenesis. The proto‐oncogene, Notch, is associated with colon cancer. It has been previously shown that cleavage of membrane tethered Notch releases the intracellular domain fragment (NICD) which can enter the nucleus. Manifestation of tumerogenesis is caused by elevated NICD entering the nucleus and activating gene transcription. A proliferative G‐protein coupled receptor, vasoactive intestinal peptide receptor 1 (VPAC1), is highly expressed in colon cancer cells and contains numerous Notch putative consensus sequences within its promoter. We hypothesize that the reason why colon cancer cells, HT‐29, have elevated VPAC1 expression levels is due to enhanced Notch signaling. To this end, Dibenzazepine, a γ_secretase inhibitor which prevents the cleavage of Notch was utilized to determine whether VPAC1 expression was altered. By SYBR green qPCR, VPAC1 mRNA was found downregulated by Dibenzazepine in a concentration dependent manner over 48 hours. This observation is the first to link VPAC1 regulation to Notch signaling and therefore further elucidation of this mechanism will allow for greater insight into the etiology of colon cancer.Funding: NIH‐K01 1K01DK064828 and COBRE 2P20RR05566.
Activation of muscarinic receptor 2 stimulates proteasome function in cardiomyocytes
Authors: Ranek Mark John , Wang Xuejun 
Published: June 2021

Ubiquitin‐proteasome system (UPS)‐mediated proteolysis plays a central role in protein quality control and degradation in the cell. UPS dysfunction was observed in various forms of heart disease and might play an important role in the development of congestive heart failure (CHF).The parasympathetic nervous system (PNS) activity is decreased during CHF. As an initial step to investigate if and how PNS regulates the UPS in the heart, we employed neonatal rat ventricular myocytes (NRVM) culture. To better monitor UPS activity, a validated UPS surrogate substrate (a modified Green Fluorescent Protein, GFPu and GFPdgn) and a red fluorescent protein (RFP, not a specific substrate of the UPS) were co‐expressed in cultured NRVMs via adenoviral gene delivery.We found that stimulation of the muscarinic 2 receptor with pilocarpine significantly reduced the GFPu/RFP ratio, which is reversed by inhibiting protein kinase G (PKG). We have further found that the phosphorylation status of the Rpt6 subunit of the 19S proteasome and the beta 5 subunit of the 20S proteasome was markedly altered by stimulating this receptor. These preliminary findings suggest that PNS may positively regulate cardiac proteasome through activating PKG. Supported by NIH grants HL072166; HL085629, and HL068936 and American Heart Association grant 0740025N.
Compartmentalization of eicosanoid substrates, signaling lipids, and enzymes into nanoparticles in human cerebrospinal fluid
Authors: Fonteh Alfred Nji , Oborina Elena , Harrington Michael 
Published: June 2021

ObjectiveTo find out whether lipid substrates and metabolizing enzymes are compartmentalized into discrete particles in human cerebrospinal fluid (CSF).MethodsLumbar CSF was fractionated by ultracentrifugation to obtain a 17,000 g pellet (P2), a 200,000 g pellet (P3), and supernatant fluid (S3). We used transmission electron microscopy (TEM) to examine the morphology of particles, and liquid chromatography tandem mass spectrometry (LCMS) to obtain lipid composition of all fractions. Colorimetric or fluorometric assays, and shotgun sequencing were used to measure enzyme activities and identify proteins, respectively.ResultsTEM revealed particles of different sizes (30–200 nM) and structures in P3 fractions. Organic extraction showed P3 fractions contained lipids. LCMS identified the major phospholipid classes in P3. Positive or negative parent ion scanning revealed enrichment of polyunsaturated fatty acids, phosphatidylinositol and sphingomyelin in P3 compared with S3 fractions. Shotgun sequencing and enzyme activity assay identified proteins and enzymes of the prostaglandin biosynthetic pathway in P3.ConclusionsThere are discrete membrane particles within CSF that are compositionally distinct from bulk CSF. Compartmentalization of enzymes on these particles can play important roles in brain function by controlling the metabolism of eicosanoid and other lipid signaling pathways.
Arachidonoyl substrate specificity of diacylglycerol kinases
Authors: Shulga Yulia , Topham Matthew K. , Epand Richard M. 
Published: June 2021

Diacylglycerol kinases (DGKs) are an important class of lipid signaling enzymes. DGKε is unique among DGK isoforms in having specificity for DAG substrates with an arachidonate moiety. This contributes to the enrichment of lipid intermediates in the PI‐cycle with arachidonate. The importance of DGKε in neuronal function has been demonstrated in studies with knockout mice. We have shown that removal of 58 residues from the amino terminus of this protein, including the membrane‐inserting segment, had a negligible effect on substrate specificity. It is intriguing that the domain, responsible for DGKε specificity for DAG substrates with an arachidonate moiety, still is unknown.We have now identified a region of DGKε containing four conserved residues, very similar to those found responsible for recognition of arachidonic acid in lipoxygenases. We demonstrated that several mutations within this region of DGKε significantly impact enzyme activity, decreasing it to less than 2% of the activity of wild‐type DGKε. Moreover, the mutants showed a lower ratio of the enzyme activity with a DAG substrate containing an arachidonate moiety to substrates without an arachidonate moiety. The relationship of this finding to the substrate specificity of other isoforms of DGK is also explored. (Supported by the Natural Sciences and Engineering Research Council of Canada, grant 9848)
Measuring arrestin binding to G Protein Coupled Receptors: BRET and recruitment assays tell different stories.
Authors: Gimenez Luis E , Digby Gregory J , Weaver Charles D. , Conn P. Jefffrey , Gurevich Vsevolod V. 
Published: June 2021

In mammals, visual (Arr1) and cone arrestin (Arr4) target activated photoreceptors, whereas ubiquitous arrestin2 and 3 (Arr2 and Arr3) regulate non‐visual GPCRs. We explored the interaction of Arr1, 2 and 3 with the M1–M2 muscarinic acetylcholine receptors (M1/M2AChR) and the β2‐adrenergic receptor (β2AR) by direct visualization of arrestin recruitment and bioluminescence resonance energy transfer (BRET). Venus‐tagged arrestins were expressed in HEK‐293 cells and imaged after agonist stimulation of co‐transfected M1AChR, M2AChR or β2AR. The formation of puncta corresponding to the receptor‐arrestin complexes recruited to coated pits was observed for M1AChR and β2AR with Arr3, but not Arr1. For BRET assays, the same receptors fused with Renilla luciferase 8 (RLuc8) were co‐expressed in COS‐7 cells with increasing amounts of Venus‐tagged arrestins. In contrast to recruitment assays, all three arrestins yielded saturable BRET signals with near‐identical profiles. Considering that measured affinity of Arr2 and Arr3 for M2AChR or β2AR is much higher than that of Arr1, our results suggest that BRET is less sensitive to differences in receptor preference of arrestins than recruitment assays.NIH grants GM077561, GM081756 (VVG), MH073676 (PJC), and F32 MH087039 (GJD)
Structure of D‐AKAP2‐PKA RI isoform complex: Insights into AKAP specificity and selectivity
Authors: Sarma Ganapathy N , Kinderman Francis S , von Daake Sventja , Taylor Susan S 
Published: June 2021

cAMP‐dependent protein kinase (PKA) regulates a variety of biological activities by phosphorylating a broad range of target proteins. Spatial and temporal specificity of PKA activity is, in turn, achieved by a family of scaffolding proteins called as the A‐kinase anchoring proteins (AKAPs). One of these, dual‐specific AKAP 2 (D‐AKAP2) binds to both the Regulatory (R) isoforms (RI and RII) of PKA. Specifically, the dimerization and docking (D/D) domain at the N‐terminus of the R subunits forms a four‐helix bundle that provides an interface for AKAP binding.In order to understand the basis of D‐AKAP2 binding to the RIα subunit and understand the molecular determinants that make D‐AKAP2 dual‐specific, we solved the structures of the RIα D/D domain with and without the D‐AKAP2 peptide. The structure of the apo crystal form was solved using the anomalous signal from inherently present sulfur atoms. The structures highlight the importance of redox‐sensitive disulfides in affecting AKAP binding. An unexpected shift in the helical register of D‐AKAP2 compared to RIIα:D‐AKAP2 complex structure makes the mode of binding to RIα, novel. Further, the comparison allows us to deduce a molecular explanation for the sequence and spatial determinants of AKAP specificity. This work was supported by HHMI, American Heart Association Postdoctoral Fellowship to G.N.S, and NIH grants GM34921 and DK54441 to S.S.T.