bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of Learning Paradigms on the Retention of Anatomical Knowledge in Medical Students
Authors: Ahmed Mohammed , Dykowski Sara , Tooley Trevor , Helland Timothy , Barremkala Malli
Published: November 2023

Introduction

Practicing clinicians today, especially surgical specialists, feel that medical students' anatomical knowledge is inadequate and does not meet the minimum threshold for safe practice. Culprits for this loss of knowledge are not exactly understood, however, key contributors include inadequacies during initial learning, lack of reinforcement in later phases, and non‐adaptive teaching methods for a growingly diverse population of learners. The primary goal of this study is to describe or suggest patterns of learning, learning tools, and teaching methods that promote retention of learned anatomical knowledge in medical students. A secondary goal is to compare and assess perceptions of students on anatomy learning and retention in pre‐clerkship years (i.e. years 1 and 2), and during clerkships (i.e. years 3 and 4).

Methods

An online survey was sent to all medical students (i.e. years 1 to 4) at the Oakland University William Beaumont School of Medicine (OUWB), approximately 450 students. Information collected included demographics, learning style preferences, learning tool preferences, perceived level of anatomical knowledge retained, and opinions on curricular structure.

Results

Ninety‐five students (52.1% male; 73.7% from years 1 and 2) responded to the survey. Fifty‐one students (54.8%) stated having no prior experience with anatomy, or having taken an anatomy class, while 42.9% of respondents reported utilizing anatomical principles less than 50% of the time. Seven respondents (10.6%) stated having retained 20% anatomical knowledge learned in first‐year medical school, 22 (33.3%) stated 21–39% retention, 29 (43.9%) stated 40–59% retention, and 8 (12.1%) stated greater than 60% retention. Each category of students was asked to provide a rank order of utilized studying resources. Students who reported <60% retention utilized dissection labs and professor‐written notes as their main resources. Students with >60% reported primarily using also the latter but with less frequency, atlases, and multiple‐choice questions. When re‐prompted to select resources for optimal retention, in both groups (<60% and >60% retention), there was an increase in frequency of students selecting dissection labs most pronounced in the >60% group. There was also an increase in frequency of students selecting atlases in the 21–39% and >60% groups. Across all groups, professor‐written notes were selected less frequently.

Conclusion

Anatomy education and retention relies on a major visual and kinesthetic (experiential) component, as seen in the preferences and perceived retention levels of students at OUWB. The use of written notes may be initially important for establishing context, but does not seem to be a significant contributor to long‐term retention. As more data is collected, these selections may show concrete trends in the way students are retaining anatomical information.
The Use of Radiological Imaging to Enhance Gross Anatomical Learning: Thoughts and Practices
Authors: Inglis Stuart Donald , Delbalso Angelo M , Dannenhoffer Raymond P
Published: November 2023

The integration of basic radiology into gross anatomy courses is becoming increasing prevalent in modern medical curricula. Anatomy faculty at the University at Buffalo are actively developing methods to increase pre‐clinical exposure to and interest in medical imaging during the medical gross anatomy course. In the most recent academic year, bodies selected for the medical and dental dissection courses underwent full body CT scans, with the data stored in DICOM stacks. Files were reviewed by a panel containing both anatomists and radiologists for anomalies and pathological conditions. Cases relevant to each dissection lab were preloaded onto a Sectra Visualization Table for viewing prior to the start of each session. Students were educated in using the visualization software to manipulate the images and encouraged to visit the dissection table to correlate the images with the active dissection. Students were also equipped with copies of the DICOM stack for their respective tables and taught to use software to identify structures on radiological images for their dissection reports. Despite initial issues in working with the software, students found the incorporation of radiological images to be both interesting and beneficial to their anatomical education.
Everything Old Is New Again: An Alternate Approach to Human Hip Dissection Inspired by a Classic 19th Century Experiment
Authors: D'Abarno Anthony , Singleton Michelle
Published: November 2023

Disarticulation of the human hip is among the most arduous joint dissections routinely performed in gross anatomy laboratories. The force required to distract the femoral head can fracture the femur or acetabulum, especially when applied by inexperienced student dissectors to a geriatric donor. We conducted a literature review to better understand the factors that contribute to passive stabilization of the hip in hopes of minimizing use of force and averting hard and soft tissue damage during hip dissection. We found that early experiments by Weber and Weber (1837) first demonstrated the role of atmospheric pressure differentials in resisting distraction of the femoral head. Subsequent experimental research has clarified the relationship among three factors that contribute to this phenomenon: 1) the cross‐sectional area of the femoral head; 2) the saturated vapor pressure of synovial fluid at body temperature; and 3) the integrity of the acetabular labrum and the zona orbicularis of the joint capsule, the latter being necessary to maintain intra‐articular pressure levels. The force F required to distract the hip is given by F = (πD2/4) * (Pa − Pi), where D is the diameter of the femoral head (m), Pa is atmospheric pressure (mBar), and Pi is the saturated vapor pressure of synovial fluid at body temperature (mBar). Thus, equilibration of atmospheric and intra‐articular pressures should minimize or eliminate the force required for hip distraction. As a practical application to anatomical education, we have performed multiple standard hip dissections (Grant's Dissector, 16th ed.) with the additional step—following the example of Weber and Weber (1837)—of drilling a 3 mm hole in the acetabulum ~2 cm superior to the acetabular notch. Using this procedure, the femoral head slides easily from the socket with no need for the complex maneuvers described in many dissection manuals. Adoption of this technique should provide human anatomy students with a less stressful dissection experience, as well as the opportunity to directly observe the interplay of factors contributing to normal human hip function.

Support or Funding Information

This research was supported by the Midwestern University Department of Anatomy, and did not require IRB approval.
The Anatomy Laboratory as a Fulcrum of Basic and Clinical‐Science Integration
Authors: Nausheen Fauzia , Al‐Eyd Ghaith , Ettarh Rajunor , Hassan Sherif , Yakub Mohsin , Suskind Robert , Tenore Alfred
Published: November 2023

Introduction

At California University of Science and Medicine, School of Medicine (CalMed‐SOM) we have designed an active learning, competency‐based and Clinical Presentation (CP) driven curriculum which avails itself of current, established, best pedagogical learning methodologies collated from the world's most advanced educational institutions, all guided by adult learning strategies. With the introduction of this active‐learning, system‐based and team‐driven curriculum (tightly integrated with “Clinical Presentations” [CP]), there is a need to create laboratory sessions that support content integration with the active learning methods used. With this in mind, CalMed‐SoM has designed the anatomy lab experience as a hands‐on multidisciplinary learning adventure which emphasizes the prompt exposure of pre‐clinical students to clinical conditions integrating, from day 1 of medical school, structure and function with pathology.

Methods and Results

During the course “Transport and Delivery of Life's Elements” (i.e., Cardiovascular‐Pulmonary System), a lab session on thoracentesis has been designed. The laboratory session is aligned with the week's CP (dyspnea), clinical case (pleural effusion), and clinical skill session. The utilization of the appropriate learning venue containing spirometers and its interpretation software, multimedia applications, ultrasonographs, manikins and human cadavers, along with the use of student volunteers, will allow to integrate the planned learning objectives of anatomy, physiology and pathology for this clinical application session of thoracentesis. The session is designed to provide students with the opportunity to apply and integrate knowledge across disciplines.

The overall learning outcomes of this particular lab session, include:

To describe the ultra‐sonographic anatomical appearance of thoracic wall, thoracic/abdominal viscera and the identification of fluid in pleural cavity.

To demonstrate the procedural skills of thoracentesis and the use of bedside ultrasonography.

To explain the principle of spirometry and estimates of different lung volumes and capacities.

To relate the effects of pneumothorax and pleural effusion on pulmonary function.

Conclusions

Multidisciplinary integrated medical education using anatomy lab will be an effective tool to improve knowledge and skills of medical students early in the curriculum to better prepare them to meet the challenging diagnostic and safe practice requirements needed in the clinical years. In addition, performance of bedside procedures, will promote spiral integration of the basic and clinical sciences.
Building Bridges Between Brazil and the US: Anatomical Sciences and Medical Education
Authors: Curcio Daniella Franco , Laitman Jeffrey T.
Published: November 2023

Brazil and the US represent the two most populous countries in the western hemisphere with the greatest number of medical schools and students in training. Over the past decade there has been considerable cross‐fertilization among anatomists in the two countries particularly in the American Association of Anatomists / Experimental Biology Meetings. Medical schools and universities in the US generally count on excellent infrastructure; high quality, motivated faculty; and cutting edge technology. The interplay of these assets results in flexible strategies that address the needs of millennial students and a modern way of learning that is interactive, multimodal and stimuli‐dependent. In Brazil, different educational settings have particular difficulties and needs. Building Bridges is an international initiative in medical education to foster collaborations and exchange experiences in anatomical sciences among students, researchers, and educators in Brazil and the US. Here we report the results from the first endeavor of our ongoing initiative. We organized three meetings that were held in Brazil, in November 2015, hosted by private, state and federal medical universities, respectively: Santa Casa School of Medical Sciences in Sao Paulo, University of Sao Paulo ‐ Ribeirão Preto, and Federal University of Rio de Janeiro. Each meeting consisted of lectures on both current research and educational topics in anatomy by faculty from the Icahn School of Medicine at Mount Sinai and local professors. In addition, there were discussions on medical curricula, and specific challenges faced by each school. Students representing academic organizations (e.g. “Anatomy Leagues”) had the opportunity to introduce their work, ask questions, and discuss subjects of particular interest, for example availability of cadaver dissection and the use of imaging technology in the gross anatomy lab. The active participation of over 300 students and professors showed a clear interest for this program. There were also meetings with the Deans and Course Coordinators in the different settings to exchange experiences, discuss current issues in medical curricula and specific difficulties in teaching anatomy from the different educational standpoints. These opportunities allowed for us to better understand specific needs of medical students in each type medical school in Brazil. Lastly, we met with Brazilian Government Officials in both Brazil and the US to give feedback. We are currently planning on student workshops visiting programs and meetings in both the US and Brazil as part of the ongoing Building Bridges initiatives.

Support or Funding Information

Support Sources: National Counsel of Technological Development – CNPq; Santa Casa School of Medicine of Sao Paulo; University of Sao Paulo ‐ Ribeirao Preto; Federal University of Rio de Janeiro; The Anatomical Record.
Physiology Understanding Week (PhUn Week), First Time in Kingsville, Texas: The Immune System
Authors: Johnson Rachel Ann
Published: November 2023

In November, approximately 100 seventh grade students came to the Texas A&M University ‐ Kingsville campus to learn about the human immune system. Students were introduced to three main aspects of the immune system: antibodies, antigens, and B‐cells. When discussing the concept of immune responses, a superhero theme was used. Therefore, Anti‐Man was a representation of the antibody, Director B was a B‐cell, and the Germinator was an antigen. Technology was used to explain the individual role each of these play in an immunological reaction. After actively participating in the presentation, students' knowledge was tested in a modified game of rock‐paper‐scissors. In this activity, the scissors represented antibodies, rock was called B‐cell, and paper became antigen. Students were then divided into two groups, each group had to agree on being either antibodies, B‐cells, or antigens. The two teams showed their agreed signs, and then had to decide which team won. For the purposes of this activity, antibodies were beat by B‐cells, which were beat by antigens, which were beat by antibodies. This game allowed students the opportunity to further understand the chain effect of the antibody, B‐cell, and antigen interactions. The wonder upon each of the children's faces as they came to comprehend the complex interactions of the human body gave proof to their grasp of the topic.
Medical Education Across Borders–Are You GAAAME?
Authors: Ziermann Janine M. , Curcio Daniella Franco
Published: November 2023

A core tenet of the American Association of Anatomists's (AAA) mission is to strive for equality, with a continuous effort on supporting the professional development of underrepresented minorities (URMs). We believe that by fostering new and existing communication channels we can enhance the self‐confidence of young URM students and inspire them to take actions in pursuit of their professional dreams. In America, the US Department of Education lists 415 Hispanic‐Serving Institutions (HSIs), 58 Predominantly Black Institutions (PBIs), and 105 recognized historically black colleges and universities (HBCUs). Between Brazil and the US, ‘Building Bridges’ is our latest international initiative: it involves the Icahn School of Medicine at Mount Sinai in New York and three major Brazilian Universities, actively promoting access to knowledge and strengthening liaisons between the AAA and the SBA (Sociedade Brasileira de Anatomia – Brazilian Society of Anatomy). Connecting American and Brazilian schools, colleges, and universities in an inclusive program would benefit thousands of students, who would have the opportunity to experience human anatomy through the AAA, obtain information and attend meetings, participate in publications and potentially become members of the AAA. Here, we present an interactive web‐based quiz‐game with the acronym GAAAME for ‘Go American Association of Anatomists for Minority Education’. Our strategy comprises the following steps to be carried on for Brazil and the US: 1) Identify target‐institutions: schools, colleges, and universities; 2) Promote the AAA and the SBA by sharing digital and print advertising material on AAA journals and activities; 3) Present the GAAAME concepts; 4) Obtain students' feedback and ideas, thus developing the initiative in solidarity with their different ethnicities and cultures; and 5) Form groups of minority students, faculty, and SBA and AAA members who will development and fundraising teams. Our vision is that this project will engage students in creating and solving challenging quizzes, foster communications with more advanced educators and researchers, facilitate interaction with peers, and contribute to increasing diversity in the AAA community.

Support or Funding Information

Grant Funding Source: National Counsel of Technological Development – CNPq
Segmenting the Data Stream: Harnessing contrast‐enhanced imaging from widely available, anonymized patient data to Generate 3‐D Training Tools for Clinical Gross Anatomy
Authors: Todd Jarred T , Gignac Paul M
Published: November 2023

Understanding the organization of the human body—particularly through cadaveric dissection— is the foundation of modern medical education. A hands‐on appreciation for the remarkable three‐dimensional (3‐D) complexity and integration of nerves, blood vessels, muscle, sinew, and bone allows students to acquire the detailed knowledge of human anatomical structures necessary to build their own conceptual maps of how the body is organized. To forward these goals, educational applications for 3‐D technology are progressively expanding and becoming entrenched in medical and non‐medical classrooms as well as in doctors' offices and surgery bays. In healthcare, 3‐D technology is expected to revolutionize several areas of medicine by providing new methods for enabling mock diagnosis of normal and pathological states during student training, practicing clinical procedures in silico and physically with 3‐D printed models, and facilitating the development of patient‐customized prosthesis for faster recoveries with fewer complications. These initiatives have resulted in a vast amount of preexisting, anonymized 3‐D radiographic datasets, such computed tomography (CT) scans or magnetic resonance imaging (MRI), that can be readily reconstructed into 3‐D models. Here we describe a pipeline to access already available contrast‐enhanced CT and MRI datasets to develop 3‐D anatomical atlases (abdominal, cardiac, and cerebral vasculature using Avizo) for augmenting classroom learning initiatives through digital hubs such as Radiopaedia.org and DiceCT.com. Granted that the datasets have sufficient contrast differences allowing for differentiation of tissues, thick‐section scans allow for rapid reconstruction but may compromise anatomical accuracy, while thin‐section scans provide greater anatomical detail but also require extensive segmentation. We discuss how decimation tools and resampling approaches can be applied to smoothing the resulting 3‐D models. In addition, we illustrate steps that must be taken when choosing both the type of scan and the amount of detail desired in the final product. As new modalities for 3‐D technology arise, teaching styles and training materials that can evolve to embrace these new tools will see expanded applications spanning from student to patient. We anticipate that physicians‐in‐training will take such applications to even loftier heights by sourcing exemplar datasets from digital hubs that provide greater opportunities to share 3‐D models and provide more resources to a wider scope of students and clinicians.

Support or Funding Information

Funding was provided to Paul Gignac through the National Science Foundation (grants no. 1450850 and 1457180) and the Oklahoma State University Center for Health Sciences.
Visualization of Cardiac Anatomy: New Approaches for Medical Education
Authors: Whitman Rachel Marie , Dufeau David
Published: November 2023

3D Visualization is becoming an increasingly utilized tool in medical education and patient interaction. It is used for training, diagnosis, intervention design, and patient education. Many physicians and medical students can envision an anatomically correct heart, in addition to the anatomical conformation of congenital heart defects. The ability to medical students and medical professionals to visualize these structures can be greatly improved by augmenting diagnostic imaging modalities (e.g. CT and MRI) with 3D renderings assembled from these data. We present here methods for the recovery of 3D structures and anatomical associations from CT Data. These data, acquired from the St. Vincent Indianapolis Cardiology/Radiology department and the National Institute of Health Cancer Imaging Archive, were analyzed using the 3D analytical software FEI Amira at the Marian University College of Osteopathic Medicine 3D Visualization Laboratory. Relevant anatomical structures, landmarks, and anomalies were assembled to create 3D prints, interactive digital models, and YouTube videos. We assembled models from two date sets: one of an anatomically correct heart, and the other of a heart after corrective surgery for the Tetralogy of Fallot congenital anomalies. Both projects have been replicated via 3D printing, and both have an associated YouTube video that enables the clinicians, students, and patients to view the structures in three‐dimension. Developing 3D Visualization techniques gives us the capability to create more accurate, interactive, and detailed images of cardiac anatomy. Our 3D Visualizations show great potential in advancing medical education and in improving communication and interactions between patients and physicians.
An authentic link to the clinic: Implementation of an integrated anatomy‐radiology curriculum
Authors: Darras Kathryn , Reeves Janine , Roston Alexandra , Hu Jeff , Nicolaou Savvas , Forster Bruce , Krebs Claudia
Published: November 2023

Since 2011, the University of British Columbia (UBC) medical curriculum has taught anatomy in conjunction with radiology in order to give students a greater understanding of anatomical relationships and their application to clinical practice. Based on student preference, we are preparing a learning resource which encompass highly‐interactive computed tomography (CT) studies using an anatomy visualization table (Sectra, Linkoping, Sweden). Real patient CT scans are loaded into an anatomy visualization table and students can virtually cut and dissect these datasets to expose the anatomy in two and three dimensions. We would like to present a model for the integration of digital dissection with cadaveric dissection in a first year curriculum of a large medical school, with the aim of providing a direct link from the lab to the clinical application.

Methods

Normal CT studies of the spine, thorax, heart, abdomen and pelvis were identified from the online case library affiliated with the anatomy visualization table and labeled to reflect specific learning objectives for first year UBC medical students. The radiological and gross anatomy objectives are closely linked. The students were provided with lab guides that specify UBC‐labeled files for study, learning objectives and relevant anatomy/clinical review questions pertinent to each lab. Students viewed both the two and three dimensional images during the lab.

Results

This anatomy visualization table is a highly valuable learning tool as it allows students to better visualize and relate anatomical structures they learn in dissection lab with the same structures on two dimensional and three dimensional CT images. The table provides improved appreciation for anatomical relationships by allowing students to zoom, rotate and dissect 3D CT images with simple swiping hand gestures. This abstract will present lessons learned from this integration including the highlights and the challenges.

Conclusions

For medical students, who will be primarily looking at anatomy on radiology images throughout their careers, this technology allows them to become more familiar with radiological imaging early on in their training. Our radiological anatomy‐gross anatomy integrated curriculum allows students to link the clinical application of the anatomy to their dissection experience, thereby providing an authentic link to the clinic where they will be applying their knowledge throughout their careers.
Widespread Growth Factor Expression in the Reptilian Epidermis Indicates Roles During Homeostasis and Wound Healing
Authors: Subramaniam Noeline , Petrik James , Vickaryous Matt
Published: November 2023

The skin is the primary interface between an organism and its external environment. As such, it participates in the maintenance of homeostasis, as well as functions as a physical barrier, protecting against physical trauma, solar radiation, and pathogens. These functions are mediated, at least in part, by various growth factors, including members of the vascular endothelial growth factor (VEGF), fibroblast growth factor (FGF), and transforming growth factor β (TGFβ) families. Although growth factors have long been documented in the dermis, particularly during wound healing, less is known about their expression in the overlying epidermis. Here we performed a spatio‐temporal investigation of growth factor expression within the epidermis of a representative reptile, the leopard gecko (Eublepharis macularius). Recent work has demonstrated that leopard gecko skin has an intrinsic capacity to heal scar‐free. Therefore, we investigated the pattern of growth factor expression before, during, and after scar‐free wound healing. We demonstrate that the gecko epidermis expresses a diverse panel of growth factor ligands and receptors, including VEGF, VEGF receptors 1 (VEGFR1) and 2 (VEGFR2), FGF‐2 and FGFR1, and TGFβ1 and activin βA. Curiously, only VEGFR1 and FGFR1 were dynamically expressed, and only during the earliest phases of re‐epithelization; otherwise all the proteins of interest were constitutively present. Using double‐immunofluorescence, we observed co‐localization and/or overlapping expression patterns of VEGF/VEGFR2/VEGFR1 and FGF‐2/FGFR1. These findings indicate that homeostatic and injury‐mediated functions of the epidermis may involve both paracrine and autocrine signalling. Furthermore, they provide compelling evidence that keratinocytes expressing these growth factors may participate in non‐angiogenic roles during homeostasis. We propose that constitutive growth factor expression by keratinocytes is associated with functions specific to reptilian biology, including body‐wide skin shedding, protection against ultraviolet photodamage, and photobiosynthesis of vitamin D.

Support or Funding Information

Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grants 400358 (to MV) and 401013 (to JP), and Ontario Veterinary College Scholarship (to NS)
Sexual Dimorphism of Human Vallate Papillae – A Human Cadaveric Study of Normative Morphology
Authors: Rickards Allen A , Holmes J Scott , Ganoe Maria R , Bliss Kaitlyn N , Russell Michelle L , Lynch Hannah L , Zdilla Matthew J. , Lambert H Wayne
Published: November 2023

The vallate papillae, or circumvallate papillae, are located at the posterior dorsum of the tongue, which form a V‐shaped row immediately anterior to the sulcus terminalis. Normally, the vallate papillae are surrounded by a vallum, a small mound of tissues, which creates a sulcus (or trench) around the papilla. The inner surface of the vallum houses approximately half of the taste buds located within the tongue, which plays an important role in bitter taste perception and biased food selection. Biased food selection and differences in eating behavior, in general, have been linked to sex differences, and numerous studies have noted differences in taste perception between males and females, especially with regard to bitter taste. Previous research within our laboratory demonstrated that females (aged 18–26) possess a higher number of vallate papillae than their male counterparts, based upon endoscopic examination of the tongues of 79 individuals (40 females, 39 males). Furthermore, males were found to have a larger mean vallate papillae diameter than their female peers, demonstrating that vallate papillae are sexually dimorphic at the gross anatomical level. To expand upon this in vivo study, 103 human cadaveric tongues were dissected at West Virginia University, with approval of the West Virginia Anatomical Board, in order to identify, photograph, count, and measure the perimeter of the 1,069 individual vallate papillae present. The resulting data has implications for the understanding of biased food selection among sexes.

Support or Funding Information

WV Research Challenge Fund [HEPC.dsr.14.13]; WVU Initiation to Research Opportunities (INTRO) Summer Research program
Characterization of a non‐canonical role for Tcf3 in skin tumorigenesis
Authors: Ku Amy Tsu , Nguyen Hoang
Published: November 2023

Skin serves as the first line of defense against pathogen and solar radiation. As DNA mutations accumulate from chronic UVB exposure, signaling network crucial for skin homeostasis is disrupted, leading to abnormal cell growth and the development of skin cancer such as cutaneous squamous cell carcinoma (cSCC). The transcription factor TCF3 (or TCF7L1) participates in canonical Wnt signaling by binding to nuclear β‐catenin and regulates the expression of Wnt‐responsive genes. Understanding how TCF3 contributes to skin tumorigenesis will help reveal the practical value of strategies targeting Wnt signaling pathway. Our lab previously presented TCF3 as a tumor promoter from the chemical‐induced model of murine cSCC. In this work, I evaluate the oncogenic potential of TCF3 in the xenograft model of human cSCC and determine how TCF3 impacts on different stages of skin tumorigenesis. To dissect TCF3's functionality, I apply separation‐of‐function mutants to TCF3‐altered cellular processes, including oncogene‐induced senescence (OIS) and cell migration. The results suggest that TCF3 utilizes alternative pathways to β‐catenin‐mediated signaling in TCF3's pro‐tumorigenic activities. Therefore, I proceed with in‐depth transcriptomic analyses to identify downstream effectors and signaling involved in TCF3‐mediated tumor growth. In conclusion, TCF3‐altered cellular processes converge into increased oncogenic potential, sustained tumor growth and enhanced invasiveness. Advanced studies will warrant the identification of novel biomarkers and targets in cSCC treatments.

Support or Funding Information

Support from 5T32GM088129‐03, 2T32HL092332‐11 and 5T32HL92332‐12 to A.K.
Stereocilin Paralogs Have Distinct Roles in Zebrafish Inner Ear Development
Authors: Kramer Kenneth L , Inserra Christopher J
Published: November 2023

In the vertebrate inner ear, sound and movement are transduced into electrical signals by specialized bundles of cilia on the apical surface of sensory hair cells. Each hair bundle is composed of a single Tubulin‐filled kinocilium attached to multiple rows of Actin‐packed stereocilia. Cilia are linked to each other and to an overlying membrane by a complex of proteins that includes Stereocilin (STRC), a putative GPI‐linked protein that is expressed in both stereocilia and kinocilia in mouse auditory and vestibular hair cells. Since loss of STRC in mouse results in defects in the attachments both between neighboring cilia as well as between cilia and the overlying tectorial membrane, we sought to determine if STRC is similarly required in zebrafish inner ear development. We generated antibodies that specifically distinguish the two zebrafish STRC paralogs and found that STRCa and STRCb are both expressed during hair cell formation but with slightly different expression patterns. STRCb is found on the distal tip of kinocilia when the hair cells first start tethering to the seeding otoliths, while STRCa is found a few hours later in a few punctate locations along the distal half of the kinocilium. Knockdown of strcb by injecting antisense morpholinos or eliminating strcb in CRISPR/Cas9‐generated mutant embryos blocked the ability of kinocilia to tether otoliths to hair cells. In contrast, otolith attachment appeared to be normal in strca morphants and mutant embryos. Kinocilia in strcb morphant and mutant embryos were normal in length, yet proteins in the unattached otolith were disorganized compared to otoliths in normal and strca‐deficient zebrafish. We conclude that STRCb is uniquely required for zebrafish otolith attachment and organization.

Support or Funding Information

The work was supported by grants from the National Institutes of Health (8P20GM103471) and the Creighton Health Science Strategic Investment Fund.
Cell‐Specific Relations between Vasopressin and Renin‐Angiotensin‐Aldosterone Systems
Authors: Giesecke Yvonne , Giesecke Torsten , Kawahara Katsumasa , Koshimizu Taka Aki , Gimber Niclas , Schmoranzer Jan , Bachmann Sebastian , Mutig Kerim
Published: November 2023

Vasopressin (AVP) is crucial for body water and electrolyte homeostasis. Its antidiuretic effects are mediated chiefly via renal V2 vasopressin receptors (V2R). Parallel activation of V1a and V1b receptors (V1aR and V1bR) may modulate the renin‐angiotensin‐aldosterone system (RAAS). Despite a large body of functional evidence for a crosstalk between AVP and RAAS, little is known about distribution of AVP receptors in RAAS‐related signalling. In this study we have comparatively analyzed V1aR, V1bR, and V2R distribution in the kidney, adrenal glands, and brain with focus on their potential role in modulation of RAAS.

Custom antibodies to individual AVP receptors were generated and characterized using respective mouse knockout tissues. Distribution of the receptors was studied by immunofluorescence, immunocytochemistry and super resolution imaging using rodent and human tissues.

The anti‐V1aR antibody produced signal in macula densa cells of the kidney, all three cortical zones of the adrenal glands, and suprachiasmatic and supraoptic nuclei of the hypothalamus. V1bR was detected in the mark of adrenal gland and hypothalamic nuclei but not in the kidney. V2R signal was observed along the distal nephron and principal cells of collecting duct cells of the kidney, but not in the adrenal glands or hypothalamus.

In summary, our results document presence of V1aR and V1bR in hypothalamic nuclei and adrenal cortex which suggest their involvement in the release of adrenocorticotropic hormone and aldosterone. In addition, expression of V1aR in macula densa cells may affect renin release in a paracrine way.
Primary Sertoli Cell Cultures from 30‐Day Old Rats Form Cords with a Morphologically Differentiated Phenotype
Authors: Sriram Aarati , Huynh Hong Doan Nghi , Shadarevian John , Djaksigulova Diana , Vogl A. Wayne
Published: November 2023

The seminiferous epithelium lines the seminiferous tubules of the mammalian testes and is comprised of Sertoli cells and germ cells. Sertoli cells are tall columnar cells that extend from the base of the seminiferous epithelium to the tubule lumen. They play a key role in spermatogenesis by providing support and nourishment for the developing spermatogenic cells. We have been working to establish a primary culture system for studying junction turnover in morphologically differentiated Sertoli cells. We have found that when the age of the rats used for isolating Sertoli cells are older than 20 days, the cells respond much differently to the same culture conditions than cells from younger animals. The purpose of this study is to characterize the differences in growth and morphology between these two populations of cells. Sertoli cells were enzymatically dissociated from the testes of 20‐day or older rats and were plated at a density of 1.3×106 cells /0.2 mL on Transwell culture inserts coated with undiluted Matrigel. Sertoli cells were cultured in serum‐free defined medium (SFDM) containing hormones (FSH and testosterone), vitamins and antibiotics and incubated at 34°C in 5% CO2 for either 9 days or 2 weeks. The cultured cells were treated with a brief hypotonic wash after two days to lyse any contaminating germ cells. Finally, the cells were fixed and processed for immunofluorescence microscopy or transmission electron microscopy. Sertoli cells isolated from 20‐day old rats formed a confluent layer within 24–48 hours with cells having a cuboidal to columnar shape. Cells formed a monolayer interrupted by areas where they formed focal clumps. The cells generally were polarized with nuclei and junction networks basally located. Sertoli cells isolated from 25–30 day old rats rapidly reorganized to form a condensed mass from which cylindrical cords of cells projected into the surrounding remodeled Matrigel. Sertoli cells in the cords were columnar and their irregularly shaped nuclei were located basally around the periphery of the cords. Intercellular junctions appeared better formed at the base of Sertoli cells than in cultures from 20‐day old animals. Claudin‐11 staining confirmed the presence of numerous tight junctions within ectoplasmic specializations. All the primary cultures were predominantly comprised of Sertoli cells with few contaminating germ and myoid cells. This study presents an improved primary Sertoli cell culture system that more closely resembles the in vivo phenotype and may be a more effective model for studying junction turnover.

Support or Funding Information

Supported by a NSERC Discovery Grant (#155397‐2013) to AWV.
Expansion of Hepatic Progenitor Cell Population in Liver Fibrosis of Elderly Cadavers
Authors: Chiu Sophia , Mak Ki M.
Published: November 2023

Human hepatic progenitor cells (HPC) normally reside in a quiescent state in the canals of Hering located at the interface between the portal tract and the periportal parenchyma of the liver lobule. In chronic liver disease such as alcoholic liver disease, nonalcoholic fatty liver disease, viral hepatitis, chronic cholestasis, hereditary liver disorders, and neoplasia, HPC become activated and proliferate. Being bipotential progenitors, HPC can differentiate into cholangiocytes or hepatocytes. Expansion of HPC population in the periportal parenchyma, which constitutes the HPC compartment, is termed ductular reaction or HPC response. Preliminary studies revealed ductular reaction associated with liver fibrosis of elderly cadavers with diverse causes of death (FASEB 2016; 30:1044.4; 1044.5). The present study assessed fibrosis‐associated HPC expansion in progressive stages of fibrosis of the aged liver. Paraffin liver sections of embalmed cadavers were immunoperoxidase stained using monoclonal cytokeratin 7 (CK7) antibody. Sections were post‐stained with Sirius red stain for assessment of fibrosis. HPC are small oval‐shaped cells containing an ovoid nucleus and scant cytoplasm that stained positively for CK7, a phenotypic marker of HPC. In livers with minimal fibrotic changes, CK7 + HPC lined the canals of Hering, formed reactive ductules in the form of anastomotic strings of cuboidal cells in the periportal HPC compartment, and presented as isolated cells in the liver parenchyma at variable distance from the portal tract. In septal fibrosis, in addition to the abovementioned lobular distribution, CK7 + HPC were observed at the septal‐parenchymal border of developing septa and bridging septa, and around perisinusoidal fibrotic lesions. Counts of HPC from 11 liver samples in each stage of fibrosis showed that the number of cells per HPC compartment increased from 7.93 ± 1.32 in minimum fibrosis to 17.18 ± 2.96 (P = 0.01) in septal fibrosis, and 21.83 ± 3.69 (P = 0.002) in bridging fibrosis. In the mid‐pericentral lobular areas, HPC counts increased from 2.72 ± 0.96 cells/field in minimum fibrosis to 6.92 ± 1.39 cells/field (P = 0.02) in septal fibrosis, and 11.15 ± 1.99 cells/field (P = 0.001) in bridging fibrosis.

Conclusions

Expansion of HPC population in the periportal HPC compartment in advanced stages of fibrosis represents a HPC response to liver fibrogenesis. Increased intralobular distribution of HPC likely resulted from periportal to lobular parenchyma migration, possibly relating to hepatic regeneration consequent to liver injury caused by fibrosis. Elderly cadavers provide a valuable source of liver specimens for assessing HPC expansion associated with liver fibrosis progression in the aged liver.
Cell Migration and Palatal Fusion
Authors: Logan Shaun , Benson Douglas
Published: November 2023

Cleft palate, one of the most common birth defects, occurs as a result of the developing secondary palatal shelves' failure to grow to midline, degrade the midline epithelial seam (MES), and fuse to leave a confluent shelf. The mechanism of MES degradation is largely unknown, but the role of ephrin signaling in palatal fusion suggests an induction of epithelial to mesenchymal transition (EMT). Our goal is to determine the role of ephrin signaling in MES degradation. We dissected palatal shelves from embryonic day 14.5 Ephrin‐B2/GFP mice, in which a nuclear‐localized green fluorescent protein (GFP) under control of the ephrin‐B2 (EB2) gene labels 100% of medial edge epithelia (MEE). The adhered shelves were placed oral‐side down in a glass‐bottomed dish, encased in low‐melt agarose, and submerged in culture medium. Palates were imaged same day for a total of 48 hours, using a Zeiss 880 laser scanning confocal with a humidified chamber. Displacement of Ephrin‐B2/GFP labeled cells was tracked using Imaris Software. After imaging, palates were sectioned and stained with hematoxylin and eosin. Fluorescently labeled MEE cells moved as an aggregate/sheet and not individually. Histology confirmed that the imaged palates fused. Cell expression of EB2/GFP was visible throughout the 48‐hour experiment. The data indicate that MES degradation involves MEE cell migration and suggest that elements of the EMT program are a part of palatal fusion. Continued expression of EB2/GFP throughout fusion suggests that MEE cells do not undergo apoptosis prior to seam degradation.

Support or Funding Information

NIDCR R01DE022804
In Vivo Inhibition of microRNA (miR) Using a Novel miR Inhibitor System Reveals a Role for miR‐17‐92 in Palate Elevation and/or Extension
Authors: Ries Ryan J. , Yu Wenjie , Eliason Steven L , Cao Huojun , Amendt Brad A.
Published: November 2023

MicroRNAs (miRs) are short non‐coding RNA molecules approximately 22 nucleotides long. miRs bind to complementary targets on the 3′ untranslated region (UTR) of messenger RNAs (mRNAs), attenuating mRNA translation via either mRNA strand degradation or sequestration. Given their redundancy throughout the genome, conventional knockout strategies are difficult to employ when studying miRs. To circumvent this difficulty, we have developed a novel method of miR inhibition, the Plasmid‐Based miRNA Inhibition System (PMIS), to allow for the simultaneous knockdown of homologous miRs. Short hairpin structures that flank the anti‐sense miR target site on the PMIS inhibitor complex (PMIS‐IC) dramatically increase miR inhibition by associating with RISC complex factors. The PMIS‐IC is composed of native, unmodified nucleic acids and can be stably integrated into the genome. In this study, we employ this strategy to analyze the effects of inhibition of mature miRs in the miR‐17‐92 cluster. Here we show how the PMIS system can be used to inhibit miRs from the miR‐17‐92 cluster and associated family members in stable cell lines and transgenic mice. We also show how inhibition of the miR‐17‐92 cluster can lead to differences in the point of palate formation arrest in mice carrying PMIS‐ICs for miR‐17‐92. These results demonstrate the effectiveness of this novel miR inhibition strategy and indicate new roles for miR‐17‐92 in palatogenesis.

Support or Funding Information

University of Iowa Carver College of Medicine

University of Iowa College of Dentistry

University of Iowa Research Foundation

National Institutes of Health (National Institute for Dental and Craniofacial Research)
Gene expression of angiogenesis markers in mouse mandibular condyle during development
Authors: Sato Iwao , Maeda Yuuki , Miwa Yoko , Sunohara Masataka
Published: November 2023

Angiogenesis and osteogenesis markers are an important factor during development in relation between mandibular condyle and Meckel's cartilage. However, it has many functions in addition to the formation of mandible. We need to clearly the relationship between mandibular condyle and Meckel's cartilage to angiogenesis and osteogenesis markers during formation in mouse fetal and early postnatal stages. Therefore we try to investigate the correlation between these markers during the development of the mouse mandible. We analyzed the expression and localization of VEGF by in‐situ hybridization in the developing mouse mandibular condyle from embryonic days 12.5 to days 5. We also detected the mRNA abundance of mandible with various markers from embryonic and early postnatal stages by real‐time RT‐PCR. Our results was performed for mouse mandible in different postnatal stages and after birth and the expression of the following some mRNA markers (CD31, LYVE‐1, VEGF, collagen I, II, osteopontin, and osteocalcin). The antisense probe for VEGF was detected in mesenchymal cells surrounding around MC and bone matrices area from E17.5, and clearly localization was seen at E18.5 in the embryonic mouse mandible by in‐situ hybridization. These data suggested that angiogenesis markers were effected by development stages after or before birth in morphogenesis of mandible. VEGF also affects the other morphological markers in these specific stages terminating the differentiation in mouse mandibular condyle development.

Support or Funding Information

None
A Sox2‐Lef‐1 Protein Interaction Inhibits Lef‐1 Transcriptional Activity and WNT Signaling During Odontogenesis
Authors: Sweat Mason E , Yu Wenjie , Eliason Steve , Amendt Brad A
Published: November 2023

Tooth development proceeds through a series of steps wherein epithelial and mesenchymal tissue layers cooperate to form a tooth bud, and cells in the tooth bud are organized and differentiated into a mature tooth. Throughout this process, the changes in gene expression that are required for tooth formation are prompted by transcription factors. Lef‐1 is a transcription factor and a major effector of WNT signaling, and the loss of Lef‐1 expression or an ectopic increase in Lef‐1 results in dramatic dental anomalies in mice. The transcriptional mechanisms responsible for maintaining appropriate levels of Lef‐1 expression and downstream targets are not well understood. We show that Sox2, a major regulator of dental stem cells inhibits Lef‐1 transcriptional activation. Endogenous Sox2 directly binds to Lef‐1 and we have identified specific motifs in the Sox2 and Lef‐1 proteins regulating this interaction. By immobilizing different GST‐labeled Sox2 domains we show that the HMG domain of Sox2 is required for the Sox2‐Lef1 protein interaction. Functionally, Sox2 is capable of disrupting the association of Lef‐1 protein with its consensus DNA binding sequence. Taken together, these data suggest a novel mechanism wherein the Sox2 protein functions to negatively regulate the expression of Lef‐1 and Lef‐1 target genes during odontogenisis. These new mechanisms may also regulate other epithelial organs during development.

Support or Funding Information

Iowa Institute for Oral Health Research, College of Dentistry, The University of Iowa, Iowa City, IA.