bluuebunny/crossref_metadata_2025_split huggingface dataset:
In‐Vivo Knockdown of Cortactin Disrupts Basal Tubulobulbar Complexes and Results in the Presence of Redundant Junction Related Ectoplasmic Specializations
Authors: Adams Arlo , Palia Prunveer , Vogl Wayne 
Published: April 2020

In the mammalian testis, spermatogenic cells begin development in the germ cell niche or basal compartment of the seminiferous epithelium. They then translocate through massive junction complexes between neighboring Sertoli cells to enter the adluminal compartment and further differentiate. Removal of the junctions above and formation of new junctions below is necessary for the translocation of cells from one compartment to the other. Tubulobulbar complexes’ (TBCs) are Sertoli cell specific structures that are proposed to internalize intercellular junctions in the seminiferous epithelium. TBCs are actin cuffed endocytic structures that share many features in common with clathrin mediated endocytosis (CME). Both TBCs and CME vesicles begin with the association of a clathrin coated pit on the plasma membrane and the development of dendritic actin ‘cuffs’ on their neck regions. Rather than vesiculating, the necks of TBCs continue to lengthen into microns long structures, eventually developing an actin free ‘bulb’ region, which develops an ORP9 positive membrane contact site with an associated leaflet of endoplasmic reticulum. One of the proteins known to be highly specific for the dendritic actin around TBCs is the actin related protein, cortactin. It previously has been shown that knockdown of cortactin causes shortening and disorganization of those TBCs at the apex of the epithelium that are associated with late spermatids, and this is correlated with a delay in sperm release. However, it has not been shown if disruption of cortactin expression has similar effects on TBCs associated with the basal junction complexes and whether or not this has an effect on junction removal. Here we use electron microscopy (EM) and stimulated emission depletion (STED) super resolution microscopy to evaluate the phenotype of rat testes injected 5 times over a 9‐day period with siRNA targeting cortactin, and use western blots to confirm the knockdown. In vivo knockdown of cortactin in rat testes disrupts basal TBCs when sections labeled for ORP9 and cortactin/actin are evaluated by STED, and leads to the presence of redundant junction related material (ectoplasmic specializations) when evaluated by EM. These data are consistent with the conclusion that TBCs internalize basal junctions and are likely part of the mechanism of spermatocyte translocation from basal to adluminal compartments of the epithelium.Support or Funding InformationSupported by a Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grant to A Wayne Vogl (RGPIN‐2018‐03727).
CRP and Cra have Antagonistic Roles in the Regulation of the Fructose Operon in Vibrio cholerae
Authors: Beck Christina , Perry Sayde , Liu Jane 
Published: April 2020

Vibrio cholerae, the causative agent of cholera, survives in both the human small intestine, where pathogenesis occurs, and aquatic reservoirs by adapting to changes in carbon source availability. In order to metabolize fructose, one of the most prevalent sugars in freshwater environments, V. cholerae must selectively produce the necessary metabolic machinery. Here, in studying how V. cholerae regulates fructose metabolism at the genetic level, we found that Cra (catabolite repressor/activator) and CRP (cAMP receptor protein), two well‐described transcriptional regulators in Escherichia coli, have antagonistic roles in regulating the expression of fruB, a member of the fructose operon fruBKA. Using transcriptional assays, qRT‐PCR, and western blot analysis to evaluate changes in gene expression, we found Cra to be a repressor of fruB and CRP to be an activator of fruB. We also found CRP to be a repressor of Cra, suggesting that CRP and Cra interact to affect downstream fruB expression. In order to further elucidate the mechanisms by which Cra and CRP regulate fruB expression, we are currently focusing our efforts on mapping the fruB promoter region. Although more remains to be understood about the relationship between the two regulators, this work has elucidated previously unknown roles of Cra and CRP in regulating fructose metabolism at the genetic level in this facultative pathogen.Support or Funding InformationNational Institutes of Health, The Camille and Henry Dreyfus Foundation
Silk fibroin converts quiescent cardiomyocytes to pacemaker cells
Authors: HU YU-FENG 
Published: April 2020

IntroductionBiological pacemakers, generated by cell or gene delivery, have been developed as hardware‐free alternatives to electronic devices. However, the challenges associated with gene manipulation and immunogenicity complicate clinical applications.HypothesisThe biological pacemakers could be generated by the biomaterial, a gene and vector‐free strategy.MethodsKey features of pacemaker cells were assessed by patch clamp and calcium image in the rodent ventricular cardiomyocytes (VMs) after exposure to silk fibroin (SF) to characterize the conversion to pacemaker cells. The optical mapping was used to map in‐vivo biological pacemaker activity from SF‐implantation site. We used whole transcriptome analysis to explore the underlying mechanisms.ResultsSF‐induced pacemaker cells exhibited both functional and morphological features of genuine sinoatrial node (SAN) cardiomyocytes in vitro including long and slender morphology, HCN4 and Cx45 expressions, membrane (increased If and IK1 current) and calcium clock (diastolic calcium release). When implanted into the rat ventricle, SF induced biological pacemaker activity which functioned as a surrogate in situ SAN. Whole transcriptome analysis showed SF induced ectopic expression of vascular endothelial cadherin (Cdh5) on quiescent VMs. Cdh5 expression activated β‐catenin signaling and drove simultaneous expression of transcription factors during SAN embryogenesis, which converted quiescent VMs into pacemaker cells.ConclusionsThe SF‐based approach represented a new strategy to generate pacemaker cells for future clinical applications.Support or Funding InformationNHRI‐EX108‐10513SC
Suppression of histamine H1 receptor gene expression by antihistamines with inverse agonist activities
Authors: Fukui Hiroyuki , Mizuguchi Hiroyuki , Kitamura Yoshiaki , Takeda Noriaki 
Published: April 2020

Allergic rhinitis is an incurable disease whose prevalence rate is over 30% in Japan. Antihistamines are first‐choice drugs for the therapy of the disease. However, sole treatment of antihistamine is not enough to obtain satisfactory control. We elucidated histamine‐induced histamine H1 receptor gene expression as a pathological mechanism of allergic rhinitis. Treatment of epinastine, an antihistamine, to allergic rhinitis patients showed alleviation of nasal symptoms and correlated suppression of histamine H1 receptor mRNA elevation in nasal mucosa. We observed inverse agonist activities in some antihistamines. Suppression of inositol phosphates accumulation and suppression of histamine H1 receptor gene expression showed good correlation. These results suggested that antihistamines with inverse agonist activities have advantage for the therapy of allergic rhinitis.Support or Funding InformationResearch Grant from The Osaka Medical Research Foundation for Intractable Diseases
3D Visualization of Depression and Anxiety
Authors: Stalets Marissa , Rodgers Emily , Dufeau David 
Published: April 2020

We present a novel and technologically enhanced teaching tool geared towards both patients, students, and clinicians with the primary goal of educating these groups about the anatomical and biochemical implications of various syndromes involving depression and anxiety. Our secondary goal is to inform patients, students, and clinicians about the most common treatment methods used to treat these syndromes. We achieved this by mapping the mesolimbic system, otherwise known as the “reward pathway,” and other relevant neuroanatomical structures using a high‐resolution cryosection dataset made available by the Big Brain project at McGill University and constructing 3D visualized models using 3D analysis software tools. The results of this project were compiled into a series of three interactive educational modules serving each of our goals. The first module described the neuroanatomical structures involved in syndromes of depression and anxiety along with their functions. The second module describes various forms of clinical depression and treatment options, including the mechanism of action and how it applies to the underlying disease process. The third module did the same but described anxiety rather than depression. This tool is valuable to patients and care providers alike because it explains neuroanatomical and biochemical aspects of these syndromes in a straight‐forward and easy‐to‐understand visual manner. It also describes the context of the best pharmacological interventions for each syndrome, which is a substantial step to help engage patients in their personalized mental health treatment plan.Support or Funding InformationThank you to Dr. David Dufeau and Marian University College of Osteopathic Medicine for the resources and guidance throughout this research project.
Cannabidiol Effects on Post‐Seizure Neurogenesis
Authors: Victor Tanya R. , Elmes Matthew W. , Deutsch Dale G. , Tsirka Stella E. 
Published: April 2020

Epilepsy is a complex disorder characterized by a propensity to generate repetitive seizures with molecular and cognitive consequences. Many pharmacological treatments for epilepsy target neurotransmitters or ion channels, leaving nearly 30% of patients resistant to current medications. The recent approval of cannabidiol (CBD), a non‐psychotropic component of Cannabis sativa, has demonstrated the ability to lessen seizure severity and curb neuroinflammation in experimentally induced seizures in animal models. While CBD has been shown to effect multiple pathways and targets, its complete mechanism of action is still not understood.Current models of induced seizures mostly utilize treatments before or during epileptic events, though this practice does not normally occur in the clinic. Here we utilize a more translational model where CBD treatment is administered after the first seizure event. An inciting seizure increases the likelihood of secondary seizures due to epileptogenesis, where seizure susceptibility is increased due to molecular and structural changes within the central nervous system (CNS) parenchyma. The epileptogenic process includes changes in axonal sprouting, cell death, neurotransmitter release, and in the hippocampus aberrant granule neural network reorganization and neurogenesis. Microglia, the resident immune cells in the brain, are involved in hippocampal neurogenesis, controlling apoptosis of the newborn neurons and ultimately the numbers of granule cells. The objective of this study is to investigate whether CBD modulates microglial involvement in neurogenesis post seizure.MethodsFollowing seizure induction in 8–10 week old mice, mice are given BrdU for 7 days post seizure to monitor cell proliferation and survival. Newly generated neurons are assessed using NeuN and DCX, to identify differentiated neuroblasts and mature neurons, respectively. BrdU+ cells are used for quantification and assessment of differentiation and survival of new cells in the dentate gyrus. Neural stem cell (NSC) levels in the hippocampus are also compared using SOX2/GFAP co‐staining.ResultsOur data suggest that CBD affects the number of new neurons after seizure. Mechanisms contributing to neurogenesis will be explored.ConclusionsCBD contributes to microglia‐mediated changes in the numbers of granule cells and neurogenesis in a model of epilepsy.Support or Funding InformationThis work was partially supported by an NSF Predoctoral Fellowship and SBMS funds to TRV.
Novel Effect of Chitooligosaccharide on Cyst Growth Retardation in an in vitro Cyst Model of Polycystic Kidney Disease
Authors: Pathomthongtaweechai Nutthapoom , Muanprasat Chatchai 
Published: April 2020

Polycystic kidney disease (PKD) is a common inherited disorder characterized by multiple fluid‐filled cysts that replace normal renal parenchyma along the renal tubules leading to impaired renal function and end‐stage renal disease (ESRD). It has recently been reported that the activation of AMP‐activated protein kinase (AMPK) by metformin inhibits mammalian target of rapamycin (mTOR)‐mediated cell proliferation and cystic fibrosis transmembrane conductance regulator (CFTR)‐mediated chloride secretion in Madin‐Darby canine kidney (MDCK) cell, a cell line derived from distal nephron of dog kidneys. Therefore, AMPK activators are proposed to be a promising agent for PKD treatment. Recently, our research group has shown that chitooligosaccharide (COS) derived from chitin/chitosan activates AMPK in intestinal epithelial cells. This study was conducted based on the hypothesis that AMPK activation by COS may inhibit cyst growth in the in vitro cyst model of PKD. We found that COS at the concentrations of 10, 50, and 100 μg/mL decreased cyst growth in a concentration‐dependent manner without toxicity measured by MTT assays. Immunoblotting analysis showed that COS at 100 μg/mL activated AMPK and this effect was abolished by STO‐609, a calcium/calmodulin‐dependent kinase kinase beta (CaMKKβ) inhibitor. Moreover, COS could elevate the level of intracellular calcium. These results suggest that COS inhibits cyst progression by activation of AMPK and the increment of intracellular calcium level via CaMKKβ. Therefore, COS has novel potential application for the treatment of polycystic kidney disease.Support or Funding InformationFaculty of Medicine Ramathibodi Hospital, Mahidol University
Size Matters! Evaluating the Effect of Model Size on Anatomy Learning
Authors: Yang Jack (Xiaozhi) , DeYoung Veronica , Xue Yuanxin , Nehru Amit , Hildebrand Alexandra , Brewer-Deluce Danielle , Wainman Bruce 
Published: April 2020

Historically, learning anatomical specimens was limited to studying cadaveric materials, and by extension, specimens which are “life‐sized”. Recent technological advancements in 3D scanning and printing now allow for the production of inexpensive, durable anatomical replicas at virtually any size. This, however, creates a dilemma: what is the most effective model size to learn from? The goal of this project is to discover the appropriate size of an object to learn nominal anatomy and thus provide a critical step in improving anatomic education.We hypothesize that there is a curvilinear relationship between model size and learning, where a model too small or too big would not be the most conducive to learning and an ideal intermediate size can be determined. In this study, undergraduate students (n = 351) without prior anatomical training learned from four bones of varying normal anatomical size and features and were assessed on their ability to identify various landmarks. Thoracic vertebra (VE), hemipelvis (HE), sphenoid (SP), and scapula (SC) was 3D‐printed at four different scalar sizes. The VE and HE models were printed at 50%, 100%, 200%, and 400% scale, while SP and SC models were printed at 50%, 100%, 200%, and 300% scale. Each participant was randomly assigned to a group of two bone models (VE/HE or SP/SC) of a certain size, and randomized across the order in which they learned the models. They were then tested on the respective real bone specimens, followed by a qualitative survey reporting their experience with the 3D‐printed models, a Mental Rotations Test (MRT), and an Operation Span Test (OSPAN). Data collection for the 50% SP/SC group is still ongoing.Multiple regression suggested significant effects of Model Type, Model Size, MRT and OSPAN, (F(9, 596) = 17.96, p = 0.000, R2 = 0.2133). The most significant predictor of test score was MRT, which suggested a 10% increase in MRT score is associated with a ~3% increase in test score. The score variability independently accounted for by MRT and OSPAN was 14.6%, while the variability independently accounted for by model size and type was 7.8%. This means, that while test scores are primarily driven by participants’ mental rotation ability, model size remains an important feature that can be manipulated to improve learning. 3D printing allows for this in a cost‐effective way.Support or Funding InformationThis study was funded by the Education Program in Anatomy at McMaster University. Many thanks to the University of Buffalo for 3D printing the 400% VE and HE models.
Retinoic acid application transiently increases microglia and macrophages after optic nerve injury in frog Rana pipiens
Authors: De La Rosa-Reyes Valeria , Duprey-Díaz Mildred Vanessa , Blagburn Jonathan M , Blanco Rosa Esther 
Published: April 2020

Retinoic acid (RA) plays major roles during nervous system development, and during regeneration of the adult nervous system. We have previously shown that RA signaling pathway components are upregulated after optic nerve injury and that exogenous application of RA greatly increases the survival of axotomized retinal ganglion cells (RGCs). We have recently reported that the application of growth factors increases the number of macrophages in the regenerating optic nerve. The objective of the present study is to study in more the effects of RA application on the macrophages and microglia in the optic nerve after injury. We performed optic nerve crush and applied to the nerve either saline solution, retinoic acid or clodronate liposomes. We examined the optic nerves at one week, and two weeks, after axotomy with immunocytochemistry and electron microscopy. Our results show that application of RA to the optic nerve causes a significant increase in the number of macrophages and microglia one week after optic nerve injury. The macrophages present have considerable phagocytic activity and are consistently labeled with M2‐type macrophage markers. Numbers of activated microglia also increase near the injury site. Two weeks after axotomy and RA treatment there are fewer macrophages, and their ultrastructure shows less phagocytic activity. Clodronate liposome treatment at the time of axotomy apparently eliminates the macrophages present in the nerve at one week. We measured RGC survival at six weeks after axotomy and clodronate treatment and found no obvious effect of this treatment. In conclusion, the application of retinoic acid increases the number of macrophages and microglia present in the nerve after optic nerve injury. Further studies will explore if this effect plays a role in the success of optic nerve regeneration.Support or Funding InformationREB is supported by NIH grant GM116692, JMB by NIH grant NS081726 and VDR by NIGMS‐RISE R25 GM061838
Studies of specific yeast prion chimera to test hypotheses about prion propagation and Hsp104‐mediated prion elimination
Authors: MacElroy Andrew S. , Schock Sydney F. , Singh Atul K. 
Published: April 2020

In Saccharomyces cerevisiae, at least 10 proteins are known to form amyloid‐based prions which propagate in cell populations due to continuous fragmentation by molecular chaperones. Hsp104, Hsp70, and the Hsp40 Sis1 are all universally essential to this process; however, prions differ in their specific requirement for Sis1 activities, and some require additional Hsp40 chaperones. For reasons which are still being explored, only one prion, [PSI+], is eliminated when Hsp104 is overexpressed. We are utilizing prion‐chimeras originally developed by the Lindquist lab at MIT to test various hypotheses regarding prion propagation and Hsp104‐mediated elimination, including the role of amino acid content of the prion‐forming domain in determining prion‐Hsp40 requirements. Additionally, our investigations explore the hypothesis that the specificity for Hsp104‐mediated prion elimination is due to a unique protein region of the [PSI+]‐forming protein Sup35 known only as the “M” domain. Preliminary results indicate that the presence of the M domain is an important, if not essential, component in determining the ability of a prion to be eliminated by Hsp104 overexpression.Support or Funding InformationThis work was supported by the Lafayette College Chemistry Department, the EXCEL research scholarship program (
www.lafayette.edu), the National Institute of General Medical Sciences of the National Institutes of Health under Award Number R15GM110606, and the Henry Dreyfus Teacher Scholar Award. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or the Dreyfus Foundation.
Functional Platelet‐derived Mitochondria Induce the Release of Neutrophil Microparticles
Authors: Boudreau Luc H. , Foulem Robert D. , Léger Jacob L. 
Published: April 2020

INTRODUCTIONInflammation is an essential process of the host defense against infections, illness, or tissue injuries. However, unregulated inflammation has been associated with chronic inflammatory auto‐immune diseases, such as rheumatoid arthritis, multiple sclerosis and atherosclerosis. Polymorphonuclear neutrophils (PMNL) are amongst the immune cells involved in the acute inflammatory response used to fight bacterial infections. Once activated, PMNL release inflammatory mediators, enzymes and large quantities of microparticles in the extracellular milieu to recruit various immune cells required to fight the invading pathogens. Recent evidence also shows that platelets (PLTs), well known for their coagulation proprieties, are also implicated in the body’s inflammatory response. Interestingly, activated PLTs can release fully functional mitochondria in the extracellular milieu. Known as the powerhouse of the cell, the mitochondria share similar characteristics with bacteria. Therefore, we hypothesize that PLTs‐derived mitochondria present in the extracellular milieu, acting in a similar way as bacteria, induce a sterile inflammatory response that involves the PMNL.OBJECTIVESThe main objective of this study is to investigate the sterile inflammatory response of PMNL caused by the exposure of PLTs‐derived extracellular mitochondria.METHODSBlood was obtained from healthy consenting donors, then PMNL and PLTs‐derived mitochondria were isolated and purified. Following the co‐incubation of PMNL with various physiological doses of PLTs‐derived mitochondria, a characterization of the interaction between PMNL and PLTs‐derived mitochondria and an investigation of the inflammatory proprieties of PMNL was performed using flow cytometry, confocal microscopy, and high‐resolution respirometry.RESULTSOur data demonstrates that PLTs‐derived mitochondria associate with PMNL as quickly as 2 hours. This results in an increase of the PMNL intracellular calcium content and a 2.47‐fold mitochondrial‐dependent increase in oxygen consumption. Data also shows that PLTs‐derived mitochondria significantly induce, in a dose‐dependent manner, the release of PMNL microparticles.CONCLUSIONSThis study provides a new insight into the role of PLTs‐derived mitochondria in the context of sterile inflammation. Specifically, the characterization of its interaction with PMNL reveals a novel modulator in the fundamental inflammatory response. The knowledge gained from this study provides insight into the mechanism of sterile inflammation in auto‐immune diseases.Support or Funding InformationCanadian Institutes of Health Research, New Brunswick Health Research Foundation, New Brunswick Innovation Foundation