bluuebunny/crossref_metadata_2025 huggingface dataset:
miRNA‐21 and miRNA‐133b are decreased in hyperthyroidism‐induced cardiac hypertrophy – Critical role of Type 1 Angiotensin II receptor (AT1R)
Authors: Diniz Gabriela Placoná , Takano Ana Paula C , Barreto‐Chaves Maria Luiza Morais
Published: June 2021

Objective

To investigate the miRNA‐21 and miRNA‐133b levels in T4‐induced cardiac hypertrophy, as well as the potential influence exerted by AT1R in this response. Hyperthyroidism was induced by daily intraperitoneal injections of 0.100 mg/kg of T4 for 14 days. The T4 group was then subdivided into two groups: T4 and T4 plus Losartan, which was administrated in the drinking water once a day for 14 days (200mg/L). The analysis of miRNA expression was assessed by MicroRNA Stem Loop RT‐PCR.

Results

Cardiac hypertrophy in T4 group confirmed by the higher heart weight, heart weight to tibia length ratio, as well as by the increase of ANF and skeletal‐alpha actin levels, was prevented by Losartan. In addition, T4‐induced cardiac hypertrophy was accompanied by decreased miRNA‐21 and miRNA‐133b levels (P<0.01, vs control). However, the AT1R inhibitor blocked the reduction of miRNA‐21, as well as attenuated the reduction of miRNA‐133b promoted by T4.

Conclusions

Considering that increased miRNA‐133b was able to prevent the cardiac hypertrophy development, and that miRNA‐21 inhibition promoted cardiac growth, it is possible that the modulation of these miRNAs may exert an important role to the cardiac hypertrophy development induced by T4, as well as to the prevention of the cardiac growth evidenced by the AT1R blockade. Research support: Supported by Foundation for the Support of Research in the State of São Paulo.
Testing is a superior method for learning anatomy and physiology information.
Authors: Dobson John
Published: June 2021

The testing effect observes that tests actually strengthen learning by prompting the retrieval of information. Although the testing effect is among the most robust findings in experimental psychology, much of its empirical support is from laboratory studies and it has been virtually ignored as a strategy for enhancing learning in the classroom. The purpose of this study was to investigate the testing effect in a university anatomy and physiology course. Students repeatedly studied three sets of passages that described structures and concepts pertaining to 1) cardiac electrophysiology, 2) ventilation and 3) endocrinology. Each student was randomly assigned to study one of those three passage sets by reading it three consecutive times (RRR), another by reading and then rereading it while taking notes (RRN) and the third by reading it, recalling as much as possible (i.e., testing) and then rereading it (RTR). Retention assessed after one week was significantly greater (ANOVA; F = 3.73, P = 0.02) following RTR (53.95 ± 1.72) compared to RRR (48.04 ± 1.83) and RRN (48.31 ± 1.78). In conclusion, testing is an effective means of improving learning of anatomy and physiology class.
Students’ satisfaction with their 1st year: cross‐sectional study at the new Medical Studies in English Program at the University of Split School of Medicine
Authors: Valic Zoran , Malicki Mario , Bradaric Kristina , Bozic Josko , Ljutic Dragan , Valic Maja
Published: June 2021

Students’ satisfaction with their studies grants institutional prestige, attracts prospective students, and helps the institutions set and improve their educational goals. Aim of this study was to assess students’ perceptions of the first year at the new Medical Studies in English Program at the USSM. We conducted a designed student satisfaction survey and a Course section of the Quality of Life Enjoyment and Satisfaction Questionnaire (QLESQ) to students at the beginning of their second year. In total 16 out of 19 students (9 male and 7 female), aged 19 to 24 (M=21±1.5) and with a GPA of M=3.4±0.5 returned the completed questionnaires. Most students (n=9, 60%) rated their overall satisfaction with the USSM as above average, with 12 (75%) students giving that same grade to the quality of their first year teaching. Lowest rated item was opportunities for extracurricular activities with 9 (56.3%) students rating the item poor. Students overall score on the course section of the QLESQ survey was 35.3±3.5. Using two different satisfaction questionnaires we showed that students which enrolled at the new Medical Studies in English Program where highly satisfied with their first year of study. As the most negative aspects of the Program were extracurricular activities, the USSM included obligatory physical education course and language activities for students.

This study was supported by University of Split School of Medicine.
A Practical Class to Teach Masticatory Efficiency for Dental Students
Authors: Rocha Maria Jose Alves , Silva Mauro Ferreira , Garcia Dorival , Mestriner‐Júnior Wilson
Published: June 2021

The aim was to set up a practical class to teach the students about the several factors that can affect masticatory efficiency. Before starting the test we asked that the student volunteers to report whether or not they have any masticatory problem and to masticate a piece of parafilm or a sugar‐free chewing gum for 20 seconds to remove any resting muscular memory. Subsequently, they are instructed to masticate the way they usually do a Me Mastig® capsule made of polyvinyl acetate filled with beads containing a calcium chlorid fuchsin dye. After 20 seconds, the beads (fragmented or not) are collected and the content is dissolved in 5 ml of water and agitated for 30 seconds. The solution is then filtered through filter paper and the fuchsin dye quantified by spectrophotometry. The masticatory force of each volunteer is then calculated by regression analysis of the extracted fuchsin concentration against a standard curve generated from a force‐controlled masticatory apparatus. As future dentists, the students are instructed to discuss the problems that the volunteer students reported and discuss the possible factors affecting the respective masticatory efficiency.
The use of a Prematriculation Medical Physiology course to help at‐risk students develop appropriate study plans
Authors: Miller Cynthia Jayne
Published: June 2021

Medical schools frequently have trouble recruiting students from rural areas and ethnic minorities, and often these students are at high risk for academic challenges. A Prematriculation Program was created to assist at‐risk students in preparing for demanding medical school didactic schedules. Students were exposed to core content, as well as strategies for time management, study skills, etc. On pre‐surveys, students predicted an increase in their number of study hrs/ lecture hr, from 6.2 hrs in undergraduate coursework to 8.4 hrs in medical school coursework (n=17). Students reported to revise their study methods by increasing the use of instructors’ notes, multimedia resources, flow charts and diagrams, and group study strategies. A discussion illustrated the infeasibility of 8.4 hrs of study time with the content load, so students were exposed to active learning strategies and study skills to improve their ability to master Physiology efficiently. At the end of the course, a post‐survey was given, and students reported similar plans for their study methods. However, there was a reduction in the planned hours of study to a more realistic 3.9 hrs of study time/hr of lecture.
The vascular basis of perforator flaps of the upper abdominal wall: an anatomical study
Authors: Duscher Dominik , Tinhofer Ines , Huemer Georg M. , Schmidt Manfred
Published: June 2021

Introduction

Pedicled perforator flaps in the trunk such as the DIEAP (deep inferior epigastric artery perforator) flap have increasingly been used for reconstructive purposes. However perforator flaps of the upper abdominal wall derived from the SEA (superior epigastric artery) and DIEA have not been widely reported. The aim of this study was to investigate the vascular basis of perforator flaps of the upper abdominal wall based on the epigastric vascular axis and to describe the location and size of the individual flaps.

Material and Methods

The superior and deep inferior epigastric artery perforators of the supraumbilical abdominal wall of ten fresh cadavers were injected with methylene blue solution or india ink. The location and size of the labelled skin area was observed. The arterial perforators were dissected and the length, diameter and the distance of the midline, xiphoid process and umbilicus to the perforation point was recorded.

Results

The SEAPs and DIEAPs supplied the ventromedial skin of the lower thoracic and supraumbilical abdominal wall in a sequential order. The mean size of all injected skin areas was 65,8 ± 33,7 cm2 (9,9 × 8,8 cm). A mean number of 9,8 ± 4,2 perforators was identified. The mean external diameter of the dissected perforators was 0,8 ± 0,3 mm (range 0,5 – 1,9 mm). The perforator length until arborisation averaged 3,6 ± 1,1 cm (range 2 – 7 cm). Most Perforators were located in an area 4–6 cm from the midline and 5–10 cm below the xiphoid process.

Conclusions

A reliable anatomy of SEAP and DIEAP flaps of the upper abdominal wall could be demonstrated. Especially subcostal SEAP flaps are of clinical interest. These flaps may be rotated cranially for lower chest wall reconstruction or deflected caudally for abdominal wall reconstructive purposes. The harvest site could be closed directly or via a reverse abdominoplasty procedure.

Grant Funding Source: AAA
The effects of resveratrol on bone and growth plate cartilage in leptin‐deficient mice
Authors: Cooley Joseph , Broderick Thomas L , Al‐Nakkash Layla , Plochocki Jeffrey H
Published: June 2021

In this study, we tested the effects of resveratrol treatment on bone and cartilage tissue in wild‐type and ob/ob mice. Ob/ob mice are leptin‐deficient and serve as an animal model for obesity and diabetes and exhibit reduced bone mass relative to controls. Resveratrol is a phytoestrogen that has been shown to maintain bone mass by inhibiting osteoclastogenesis and stimulating osteoblast activity. Our findings indicate ob/ob mice have reduced bone strength relative to controls. However, ob/ob mice treated with resveratrol exhibited significantly greater cortical area and resistance to torsion than untreated ob/ob mice (p < 0.05). A similar pattern was observed with resveratrol treatment of wild‐type control mice. In both wild‐type and ob/ob mice, resveratrol treatment reduced the thickness of the calcified layer of the proximal tibia growth plate (p < 0.05), but the overall thickness of the growth plate remained unchanged. Our results suggest resveratrol treatment affects bone and cartilage tissue in the mouse model for obesity and diabetes.

Grant Funding Source: AZCOM Summer Research Fellowship
The effect of alliin on oxidative stress‐induced bone‐like cells (UMR 106–01 BSP)
Authors: Joseph Eugene E , Gibbons Meliseanna , Molina Sonia
Published: June 2021

Osteoporosis is a disease that is characterized by bone mass depletion with subsequent bone fractures. Antioxidants are reported to play a role in the prevention of bone loss and reduction of oxidative stress. There is ongoing research on how alliin, an antioxidant found in garlic, prevents or mitigates the progression of many disease processes. However, there is still much research to be done on the protective effects of alliin against bone loss. In this study, it was hypothesized that alliin mitigates the effect of the oxidative stressor, buthionine sulfoximine (BSO), on bone‐like cell cultures (UMR 106–01 BSP) by lowering the levels of the apoptotic marker, caspase‐3. UMR cells were routinely passaged, plated (1.0 × 106 cells/well in six‐well culture plates), then incubated (37 degrees C, 5% CO2) for four hours. The cells were then co‐treated with different concentrations of alliin (10, 50, 75, 100 μM) and 75 μM of BSO then re‐incubated. After 84 hours, caspase‐3 activity was analyzed using cell flow cytometry (Accuri C6) to assess cell viability and apoptotic response to induced oxidative stress. The results showed that BSO induced oxidative stress in UMR cells in a concentration‐dependent manner. Treatment with BSO correlated with higher levels of caspase‐3 activity. Co‐treatment with alliin and BSO, exhibited a concentration‐dependent correlation with a decrease in caspase‐3 activity. These findings suggest that alliin protects bone cells from oxidative stress apoptosis by inhibiting caspase‐3 activity. These findings may help in the elucidation of the potential benefits of garlic (alliin) on bone health. (Supported by the Dean's Office of the College of Arts and Sciences of La Sierra University, Riverside, CA).
Absent Right Cranial Vena Cava and Persistent Left Cranial Vena Cava in a Goat
Authors: Freeman Larry E.
Published: June 2021

An atypical cranial vena cava in a goat cadaver was discovered during student dissection lab. The malformation consisted of absence of the cranial vena cava on the right side (RCVC) and a persistent left cranial vena cava (LCVC) located on the left of the other great vessels in the cranial mediastinum. This LCVC continued over the left aspect of the base of the heart, received the left azygos vein, and joined with the great cardiac vein to unite with the coronary sinus that, in turn, emptied into the right atrium. Drainage from most right side tributary veins converged to form a vein that joined with the LCVC. Right costocervical‐vertebral and azygos veins emptied directly into the right atrium. Both a LCVC and a RCVC are normal for some mammals species, and cases of a persistent LCVC in addition to a normal RCVC have been reported for man, dog, horse, and domestic ruminant, often as incidental finding. Rare cases in man of persistent LCVC with absent RCVC in which the LCVC empties into the left atrium result in clinical problems. We believe this to be a unique caprine case of LCVC without RCVC and may have presented no or minor clinical issues as the LCVC emptied into the right atrium.
Important variations of the axillary arch muscle that every clinician should know
Authors: Turner Benjamin Paul , Tubbs R Shane , Loukas Marios
Published: June 2021

The axillary arch (AA) has been recognized as the most common variant muscle of the axilla since the early 1800s. It has been called various names by different authors such as: “axillopectoral muscle,” “axillodorsal muscle,” “achselbogen muskell,” “arcus axillaris,” and the “pectodorsal muscle;” but it is widely and most commonly known as “Langer's axillary arch”. It presents in varied morphological variants both in unilateral and bilateral fashion and has no apparent physiological function. Much disagreement still exists about its nerve supply and embryological origin. With the advent of medical imaging and upper limb surgery the clinical importance of this muscle has been re emphasized. The aim of this study was to conduct a meta‐analysis on AA muscle with special emphasis on its anatomy, embryology, clinical implications and potential iatrogenic complications. The embryological origin of the AA is most likely to be a remnant of the Panniculus carnosus— highly developed pectoral muscle group in some lower animals, but in humans has regressed throughout evolution. Incidence of the AA ranges from 3.0% to 19.0% in cadaveric studies, and 0.25% to 37.5% in clinical observations. Knowledge of this important muscular anomaly and familiarity with the AA in the clinical setting, especially in surgical procedures of the axilla (i.e. axillary lymphadenectomy and sentinel node biopsy), or in any procedure requiring dissection of the axilla where the neurovascular structures could be compromised, is important for establishing differential diagnoses of other pathologies in the axilla, as well as crucial for proper lymph node staging.
Anterior and posterior reactive postural responses in healthy elderly and Parkinson disease
Authors: Smith Lorinda K. , Singer Madeline L. , Foreman K. Bo , Dibble Leland E.
Published: June 2021

Postural instability in Parkinson disease (PD) results in coordination deficits that contribute to a high incidence of falls (>; 70% fall each year, 13% fall >; once per week). To investigate the characteristics of reactive postural responses in PD, we examined lower extremity biomechanical variables during induced anterior and posterior falls. Ten males with mild to moderate PD (mean age=68.7±6.9 y) on medications, and 10 healthy male controls (mean age =66.8± 6.3 y) underwent motion analysis laboratory measurement of the following kinematic variables: reaction time, step length, step time, peak center of mass (COM) velocity, and recovery time. Anteriorly and posteriorly directed reactive responses were provoked through falls generated with an electromagnetic tether‐release system. Between group comparisons were performed using separate Mann‐Whitney U tests (p≤0.01). During anterior falls, PD subjects demonstrated decreased step length (p<.01) and decreased peak COM velocity (p<.01). No significant differences were found for posterior falls. In this study, PD subjects demonstrated hypokinetic and bradykinetic (decreased and slowed) reactive responses (anterior >; posterior) during falls compared to controls. Future research is necessary to clarify the effects of therapeutic interventions (rehabilitation, medication, surgery) on reactive responses and falls in PD.

Grant Funding Source: Univ. of Utah College of Health
Description of the microscopic anatomy of sural nerves in newborn rats.
Authors: Sanada Luciana Sayuri , Carmo Elisabete Cássia , Machado Nathália Leilane Berto , Nessler Randy Alan , Fazan Valéria Paula Sassoli
Published: June 2021

The morphology of the sural nerve is widely study, however, little is known about the morphometry of this nerve in newborns. We describe the morphological and morphometric aspects of different segments of the sural nerve in female Wistar rats (N=6) with 15 days of life. The animals were anesthetized, perfused and the right and left sural nerves were removed and prepared for epoxy resin embedding and light microscopy. Morphometric analysis was performed with the aid of computer software, and took into consideration the fascicle area and diameter, as well as myelinated fiber number, density, area and diameter. Comparisons between segments and sides were performed and significant differences were considered when p<0.05. There were no differences on the morphology and morphometry between the segments and sides of sural nerve in female Wistar rats with 15 days of life. Comparing the present results with previous data from our laboratory [1], we conclude that the newborns have similar fascicular parameters to those of 30 days‐old animals leading to the conclusion that the sural nerve is fully developed at birth and will grow following the animals’ body weight increase with age.
A Study of the Origin, Length, and Prevalence of the Meniscofemoral Ligaments in Comparison to the Cruciate Ligaments
Authors: Callier Dylan Michael , Zacher Joseph Edward , Tren Vinh , Steinkoenig Colleen , Bruzzini Kristen
Published: June 2021

Research regarding the data of the meniscofemoral ligaments has been poorly described; although it is probable that they have functional and clinical significance. This study investigated the prevalence, placement, and size of the meniscofemoral ligaments and their relationship with the cruciate ligaments. The measurements were made using the traditional digital caliper method on cadaveric specimens with formaldehyde preservation. All measurements were recorded by multiple observers and averaged for consistency and accuracy.

Grant Funding Source: None
Histological classification of the lateral patellofemoral ligament
Authors: McIntyre Michael B , Marberry Kevin , Kondrashov Peter
Published: June 2021

The knee joint capsule is a complex structure comprised of connective tissue, ligaments, vasculature, and nerves. The histological structure of the knee joint capsule has not been well documented. The aim of this study was to classify the histological structure of the lateral patellofemoral ligament (LPFL), which is the thickest portion of the lateral patellofemoral retinaculum. We then compared the LPFL to the anterior cruciate ligament (ACL), and the quadriceps tendons (QT). The ACL served as an example of a “true” ligament. The LPFL was compared to the QT to determine its validity as an autograft in LPFL reconstruction. Samples of the LPFL, ACL, and QT were harvested from six cadaveric knees, embedded in paraffin, sectioned, and stained with H&E, Masson's trichrome, or orcein. Representative samples were obtained throughout the entire LPFL and analyzed using light microscopy. Our data shows that the LPFL is composed of multiple elements including dense connective tissue, loose connective tissue, vasculature, elastin, and nerve fibers. Presence of these elements differentiates LPFL from the ACL and QT. Unlike the ACL and QT, the LPFL is not uniformly composed of dense regular connective tissue. Documenting the histological structure of the LPFL may contribute to the understanding of the pathoetiology of patellofemoral pain syndrome.

Grant Funding Source: N/A
THE EFFECT OF LIFE‐LONG ENDURANCE EXERCISE ON COLLAGEN CROSS‐LINKING OF THE HUMAN PATELLAR TENDON
Authors: Couppé Christian , Svensson René Brüggebusch , Grosset Jean‐Francois , Kovanen Vuokko , Karlsen Anders , Nielsen Rie Harboe , Skovgaard Dorthe , Kjaer Michael , Magnusson S. Peter
Published: June 2021

Introduction

It remains unknown if life‐long habitual endurance exercise influences the accumulation of Advanced Glycation Endproducts (AGE) cross‐links that are closely associated with aging and disease in connective tissue.

Purpose

To examine the effect of aging and life‐long habitual endurance exercise on the collagen cross‐linking of the human patellar tendon.

Methods

We investigated 13 healthy injury free master athletes (old trained men, OT; age 59–75 years, running distance of 49±3 km/wk over 29±3 yrs (mean±SEM)), 12 old untrained controls (OC; matched to OT for BMI and age) and 10 young men matched for current running distance (young trained,YT; age 21–34, 48±4 km/week) and 12 young untrained controls (YC; matched to YT for BMI and age). Percutaneous tendon biopsies were obtained and analyzed for enzymatic (HP&LP) cross‐links, pentosidine, and collagen concent from the patellar tendon. A 2‐way ANOVA was used for statistical analysis.

Results

Pentosidine increased with age (P<0.001)(Pentosidine: OT, 39±4; OC, 48±3; YT, 11±2 and YC, 9±1 mmol/mol collagen). There was an interaction between age and training (P=0.019), such that master athletes (OT) had a lower tendon pentosidine than OC (P = 0.006).

Conclusion

These are the first data to demonstrate AGE cross‐links in tendon in male master athletes are lower compared to old untrained controls. The results suggest that life‐long habitual endurance exercise can counteract the aging process in tendon and thereby possibly reducing the risk of injury. Research support: Danish Association of Rheumatism.
Expression of Calbindin‐D9k and Vitamin D Receptors in the External Genitalia of She‐Camels (Camelus dromedarius)
Authors: Inpanbutr Nongnuch , Emam Mahmoud A. , El‐Zoghby Ihab , Bolon Brod
Published: June 2021

To investigate the possible role of calcium metabolism in modulating external genitalia function of she‐camels, the localization and distribution of calbindin‐D9k (CaBP9k) and vitamin D3 receptors (VDR) were examined immunohistochemically in samples of vestibule, vulva, and clitoris acquired during breeding and non‐breeding seasons. Animals (n=6 per season) were healthy adults (5 to 10 years) with no history of reproductive tract or systemic disease. CaBP9k was expressed in the cytoplasm of epithelia, but not the stroma or muscles of all three regions examined. Moreover, CaBP9k was expressed in the superficial squamous cell layer of the vulva, and in the basal and parabasal layers of the vestibule. VDR was expressed in the nuclei of epithelia of all three regions, vestibule, vulva, and clitoris. The chondrocytes of the clitoris also expressed VDR. The expression of CaBP9k and VDR was higher during the breeding season than the non‐breeding season, suggesting that external genitalia function may be dependant in part on elevated calcium metabolism during this season.

This paper was supported by grant from the government of Egypt (MAE and EME) and OSU Research Fund (NI).
Characterization of Segment Defect Regeneration in the Axolotl Fibula
Authors: Chen Xiaoping , Stocum David
Published: June 2021

We determined the critical size defect (CSD) for regeneration of the axolotl fibula by surgically removing bone segments comprising 10, 20, 40, and 50% of the length of the fibula in 15–20 cm axolotls. The limbs were first subjected to X‐ray and micro‐CT imaging. Half the specimens were stained with methylene blue/alizarin red for cartilage/bone and the other half sectioned and stained with H&E. H&E staining revealed initiation of cartilage formation in 10% and 20% defects by one month; cartilage and bone were regenerated by three months. None of the 40% defects regenerated cartilage by two months post‐operation, and 7/8 (87.5%) failed to regenerate cartilage at three months. None of the 50% defects regenerated cartilage after three months. H&E staining showed that the 40% and 50% defects filled in with fibrous soft tissue. We concluded that the CSD is slightly less than 40% of the length of the fibula. A pig small intestine submucosa (SIS) scaffold did not promote regeneration across a 50% defect. The scaffold degraded by two months after implantation while the defect was filled by connective tissue and disorganized muscle fibers. Our aim is to use the axolotl fibula as a new, convenient model to screen for the optimum combination of growth factors/scaffold that promotes cartilage formation in a CSD, thus mimicking the first step of normal endochondral bone development and fracture repair.
The effect of tension on the growth of keratinocytes in a skin equivalent culture model
Authors: Meek Bill D , Vaughan Melville B , Edgar Natalie
Published: June 2021

The skin organotypic culture environment (Vaughan, et al 2009) was used to study the effect of tension forces on the fine structural features of the epithelium. Two different keratinocyte cell cultures were used in these experiments, Ker‐CT‐Ras and the HaCaT cell line. Both cell types grew to 7–15 layers of keratinocytes with a distinct basal layer present on a collagenous matrix composed of fibroblasts. The skin explants were harvested and prepared by standard methods for both light and transmission electron microscopy. Keratin intermediate filaments were not as dense and tonofibrils appeared reduced in the no‐tension model. In the tension model, desmosomes showed distinct features with evident attachment plaques and a dense radial band. Keratin filaments interacted with plaques of the desmosomes. In apical layers, early quantitative assessment showed 3 desmosomes/5μm of cell surface as compared to 8 desmosomes/5μm of cell surface in the no‐tension vs. tension model, respectively. Hopefully, these results will help to serve as a further means of understanding keratinocyte interaction and keratinocyte‐dermal interaction in wound healing, aging, and skin development.
Evolution of the colon in paddlefish, Polyodon
Authors: Wechter Todd , Theodosiou Nicole
Published: June 2021

Evolution of the colon in paddlefish, Polyodon

During the vertebrate transition from water to land, the colon played a vital role in water retention, thus allowing for survival in new terrestrial environments. While the colon was previously thought to have evolved following the vertebrate transition onto land, research showing that cartilaginous fish contain a colon‐like organ suggests pre‐adaptation to the terrestrial movement. To determine if the rudimentary colon is unique to cartilaginous fish or was retained throughout the vertebrate lineage, I am examining the paddlefish for a rudimentary colon. The paddlefish, Polyodon is a basal ray‐finned fish, and can provide insight into the evolution of the colon in the vertebrate lineage. Polyodon digestive tracts were examined for the presence of acid mucins, a histological marker for water absorption in the colon. In addition, I will report isolation and expression of the colon patterning genes Hoxd13 and Hoxa13. The possible existence of a region in the paddlefish digestive tract containing acid mucins and Hoxd13 and Hoxa13 expression would suggest that vertebrates were pre‐adapted to transition to land. Furthermore, this data would imply that the colon‐like organ found in cartilaginous fish was not the result of convergent evolution, but was rather a homologous organ.
Damaged DNA induced UV‐damaged DNA‐binding protein (UV‐DDB) dimerization and its roles in chromatinized DNA repair
Authors: Yeh Joanne I , Levine Arthur S. , Du Shoucheng
Published: June 2021

UV light‐induced photoproducts are recognized and removed by the nucleotide‐excision repair (NER) pathway. In humans, the UVdamaged DNA‐binding protein (UV‐DDB) is part of a ubiquitin E3 ligase complex (DDB1‐CUL4ADDB2) that initiates NER by recognizing damaged chromatin with concomitant ubiquitination of core histones at the lesion. We report the X‐ray crystal structure of the human UV‐DDB in a dimeric complex with two molecules of damaged DNA. The functional significance of the dimeric UVDDB, in a complex with damaged DNA, is validated by electron microscopy, atomic force microscopy, solution biophysical, and functional analyses. We propose that the binding of damaged DNA induces domain folding in the context of the bound UV‐damaged DNA substrate, resulting in NT‐helical ordering of DDB2, which in turn promotes dimerization. The temporal and spatial interplay between domain ordering and dimerization provides an elegant molecular rationale for the unprecedented binding affinities and selectivities exhibited by UV‐DDB for UV‐damaged DNA. Financial contributions: NIH AI76121, GM82251, GM66466, ES019566, Commonwealth of PA, DOH and Pitt Fdn.
The use of DNA repair as a tool in the study of factors involved in pathogenicity of Corynebacterium genus and characterization of a DNA repair gene present in pathogenic species.
Authors: de Oliveira Lopes Debora , Resende Bruno Carvalho , Carvalho Vasco Azevedo , Miyoshi Anderson , Machado Carlos Renato , Dos Santos Luciana Lara
Published: June 2021

The DNA repair pathway is an important mechanism in the maintenance of the genomic stability and the vulnerability of this system makes it an interesting target in search on the control of infectious organisms. The goal of this work was to identify and characterize DNA repair pathways of eight species of Corynebacterium, pathogenic or not, and select and characterize an enzyme, present only in pathogenic species, through in vitro assays to confirm its involvement in DNA repair. Was verified that the pathways of DNA repair in general are well conserved and NER and recombination pathways showed having virtually all conserved genes. The enzyme selected, uracil DNA glycosilase, is important to remove uracil from DNA with can cause transversion C/G into T/A during a replication event. The mug gene sequence of C.pseudotuberculosis, obtained from CoryneRegNet databank, was amplified by PCR and cloned in expression vector pET21 to be sequenced. To confirm expression in E. coli BL21 strain was carried out a western blot assay. The protein was purified and is currently being tested in in vitro experiments to verify its ability to recognize uracil in oligos sintehtyzed. An understanding of the precise molecular functions of the enzymes participating in DNA metabolism and in the maintenance of pathogen genome integrity can be the key to search for targets to be used in therapeutic intervention. Financial support: FAPEMIG
Regulation of mismatch repair protein MutSα functions by its Walker A and Walker B motifs
Authors: LEE SANGHEE , GU LIYA , LI GUO‐MIN
Published: June 2021

The mismatch recognition protein MutSα is an MSH2‐MSH6 heterodimer. Mismatch binding by MutSα triggers a series of downstream mismatch repair (MMR) reactions, including interacting and communicating with other MMR proteins by sliding along the DNA helixes. These MutSα biochemical functions rely on its ATP/ADP binding and ATPase activities. Like many ATPases, both MSH2 and MSH6 subunits contain a Walker A motif and a Walker B motif. However, how these Walker motifs contribute to the MutSα functions is not fully understood. To address this issue, a series of MutSα mutants that contain a lysine (K) to alanine substitution in Walker A motif and a glutamate (E) to K substitution in Walker B motif of either MSH2 or MSH6, or both were analyzed for their MMR activities. We show here that MSH2 mutations in Walker A or B motif block MutSα ATP hydrolysis, suggesting that the MSH2 Walker motifs regulate MutSα ATPase activity. Interestingly, the E‐K substitutions in the Walker B motif produce a mutant MutSα that, instead of sliding off, undergoes direct dissociation from a mismatch‐containing DNA molecule regardless of the presence of magnesium. This indicates that the E residue in the Walker B motif of either MSH2 or MSH6 is required for magnesium binding, and is indispensable for MutSα conformational change to form a sliding clamp. Our work provides new insights into how the Walker motifs contribute to MutSα activities in MMR.
Mobilization of the actin cytoskeleton by microbial pathogens
Authors: Welch Matthew , Reed Shawna , Lamason Rebecca
Published: June 2021

Diverse bacterial pathogens hijack the actin cytoskeleton of host cells to facilitate attachment, internalization, or spread. Our focus is on the human pathogen Rickettsia parkeri, which undergoes actin‐based motility in the cell cytosol to promote cell‐to‐cell spread during infection. R. parkeri and other Rickettsiae are unusual in expressing two bacterial proteins that activate actin polymerization: RickA activates the host actin‐nucleating Arp2/3 complex while Sca2 directly nucleates actin by mimicking host formins. By following Rickettsia throughout infection, we discovered two temporally and mechanistically distinct phases of motility. Early motility is slow, paths of movement are curved, and actin polymerization requires the host Arp2/3 complex. In contrast, late motility is rapid, paths of movement are straighter, and actin polymerization is independent of the Arp2/3 complex. Rickettsia with a transposon insertion in RickA are defective only in early motility, whereas bacteria with a transposon insertion in Sca2 fail to undergo late motility. Thus, RickA activates the Arp2/3 complex during early actin‐based motility, while Sca2 independently nucleates actin during late motility. Rickettsia are unusual in their ability to promote two temporally and mechanistically distinct phases of motility during infection, and each phase may perform a unique function in the bacterial life cycle.
PprA: A Key Protein in Extreme Radioresistance of Deinococcus
Authors: Czerwinski Matthew , Junop Murray
Published: June 2021

DNA double‐strand breaks (DSBs) represent the most severe form of chromosomal damage. In higher organisms, one DSB represents a lethal event. This severity is broadly conserved with sensitivities ranging from one to only a few DSBs. Bacteria of the genus Deinococcus represent an unusual exception and are able to withstand extremely high levels of DSBs caused by ionizing radiation, UV radiation, and genotoxic chemicals. The extraordinary radioresistance of Deinococcus results from the capacity of the organism to efficiently repair DSBs. The repair mechanism responsible for this remarkable damage tolerance is currently poorly understood. Several genes have been identified as being both unique and essential for repair within Deinococcus species. Of these, a defect in ppra has been demonstrated to be particularly important for damage recovery. The goal of this work was to better understand the repair mechanism through which Deinococcus is able to recover from DNA damage through biochemical characterization of PprA. Here we characterize the interaction of PprA with DNA. EMSA experiments were done to determine the length‐dependence, as well as DNA end preference of PprA binding with DNA. PprA was also shown to form bundles of long fibrous filaments by transmission electron microscopy. Implications for how these filaments function in DNA repair are discussed. Characterizing the filament formation and DNA binding activity of PprA is essential to clarify its proposed function in bridging and aligning DNA in Deinococcus genome repair.
Creation of a Physical 3D Homo sapiens Single‐cell Model using Confocal Microscopy Data via Rapid Prototyping
Authors: Paliwal Vipin , Hausch Paul , Padmanabhan Vakeel , Ramachandran Ramani
Published: June 2021

Physical three‐dimensional (3D) cells models are used to enhance understanding of cellular processes. Previously 3D models were artist's renditions of how cells were thought to look like and were not exact. The purpose of this study was to develop a novel process of converting a z‐axis stack of two‐dimensional confocal microscopy images into physical three‐dimensional cell models. Confocal microscope images of a single Homo sapiens endothelial cell were captured depicting the various cell stages of mitosis/cytokinesis or cell division. The high resolution of these images allowed for a single cell model to be produced, which has never been done before via rapid prototyping process. We investigated novel methods to create the most accurate and visually informative 3D cell models possible. Images were further enhanced in the medical imaging program Mimics, resized and oriented in the modeling program Freeform, and converted to the .STL file format prior to production on the rapid prototyping machines. Physical cell models were created using both the Z Corporation's Spectrum Z™ 510 printer and the 3D SystemsTM 250 SLA printer.

These new cell models created from microscopic images could benefit research scientists as well as classroom teachers, who can also now explain cellular processes like mitosis kinesthetically by providing hand size models to their students.
DNA repair regulates chromatin structure and key transcription factors
Authors: Snow Benjamiin , Estabrook Todd , Pei De‐Sheng , Strauss Phyllis R
Published: June 2021

Oxidative damage to DNA and its repair are ongoing in all cells. The key enzyme is AP endonuclease1 (Apex1), loss of which is lethal. We study Apex1 in zebrafish to follow events during early embryogenesis. Complete Apex1 knockdown leads to embryonic death as differentiation begins. Partial knockdown causes defective brain and heart. Here we explore initial events that occur upon knockdown of Apex1. Loss of Apex1 protein after knockdown begins 2.5 hours later, although apex1 message does not change for 24 hours. Loss is accompanied by loss of Creb1 protein and CrebÕs binding partners. Subsequently, DNA repair polymerase § fails to appear, probably because transcription requires Creb1. The first detectable change in transcription of any species is a striking increase in histone H2B mRNA without alteration of polyadenylated H2B mRNA, with increasing ubiquitinylated H2B (H2BK120ub1) protein. While transcription of other histones remains unchanged, transcription of many genes usually activated later is increased. Simultaneously, transcription of several ribosomal proteins but not that of myc is diminished. Since H2BK120ub1 is correlated with chromatin activation, we conclude that Apex1 is involved in maintaining correct chromatin structure with broad ramifications for the study of birth defects and cancer development. (Supported by Northeastern University and the G. Harold and Leila Y. Mathers Foundation.)
miR‐126 Dependent Decrease of Skeletal Muscle Microcirculation Contribute to Exercise Intolerance in Pulmonary Arterial Hypertension.
Authors: Potus francois , Malenfant simon , Breuils‐Bonnet Sandra , Margaillan guillaume , Bonnet sébastien , Provencher steeve
Published: June 2021

Pulmonary arterial hypertension (PAH) is characterized by a significant exercise intolerance which does not correlates with the cardiac output at rest and during exercise. Interestingly a close relationship between muscle circulation, exercise capacity and miR‐126 (pro‐angiogenic miRNA) had been shown. We hypothesized that miR‐126 is downregulated in peripheral muscles of PAH patients, decreasing microcirculation and contributing to exercise intolerance. Compared to control patients matched by both age and sex, PAH patients displayed dramatic decrease in exercise tolerance. This decrease correlates with a diminution in quadriceps microcirculation (CD31 immunofluorescence) and associated with a significant downregulation of miR‐126 expression. In vitro, using isolated CD31+ cells from human quadriceps biopsies, we demonstrated that the downregulation of miR‐126 triggered the activation of SPRED‐1 in PAH impairing the angiogenic response. All these abnormalities were reversed by treating the cells with miR126 mimic; while inhibition of miR126 in healthy cells mimic the PAH phenotype. In vivo, intraquadriceps injection of miR‐126 antagomir significantly reduced exercise tolerance compared to rats control. We demonstrated for the first time that exercise intolerance in PAH is associated with the decrease of skeletal muscle angiogenesis secondary to the downregulation of miR‐126.
Sumoylation of Sall4B, an essential stem cell transcription factor, regulates its stability, subcellular localization and transcriptional activity
Authors: Yang Feikun , Ma Yupo , Dai Wei
Published: June 2021

Sall4 is a transcription factor that plays a key role in the maintenance and self‐renewal of embryonic stem cells. However, little is known about its regulation. Sall4B, a major isoform of Sall4, was primarily ubiquitinated when ectopically expressed in HEK293T cells, and a significant fraction was further modified by sumoylation. Constitutive SUMO‐modification of Sall4B was also detected in Tera‐1 cell line. Sall4B sumoylation was independent of ubiquitination and lysine residues 156, 316, 374, and 401 were essential for sumoylation. Chromatin fraction contained more SUMO‐deficient Sall4B than wild type Sall4B. Despite a shorter half‐life than the wild‐type counterpart, SUMO‐deficient Sall4B interacted with Oct4 more efficiently. RNAi‐mediated silencing of SALL4 expression in Tera‐1 cells caused significant down‐regulation of both Oct4 and Sox2, which was rescued by ectopic expression of wild type SALL4B but not by SUMO‐deficient mutant. Significantly, compared with the wild‐type Sall4B, SUMO‐deficient mutant exhibited compromised trans‐activation or trans‐repression activities in reporter gene assays. Combined, our studies reveal sumoylation as a novel form of post‐translational modification for regulating the stability, subcellular localization and transcriptional activity of Sall4.
Mapping the ubiquitination sites of the transcriptional repressor ICER
Authors: Woldeamanuel Fanaye Bezunesh , Crow Marni , Sebasco Ethan , Molina Carlos A.
Published: June 2021

Inducible cAMP Early Repressor (ICER) is a dominant negative transcriptional repressor that expresses tumor suppressor activity. In cancer cells, ICER is abnormally expressed and becomes modified by ubiquitination. Consequentially, ICER is degraded or re‐localized from the nucleus to the cytosol where it is non‐functional. When ICER is reintroduced into cancer cells, it inhibits the transformed phenotype of cancer cells. The goal of this project is to map the specific ubiquitinated lysines on ICER to define the functional consequences of ICER ubiquitination and cytosolic localization. Eleven mutant forms of ICER were constructed using site‐directed mutagenesis. All lysine residues were substituted to arginine to create an ICER molecule without lysines. Another 10 independent mutants were generated where all but one specific lysine was substituted by arginine. All clones were subcloned in a mammalian expression vector. By using these altered forms of ICER, we have found that ICER is ubiquitinated in several lysines. This data suggested alternative modes of regulating ICER degradation and subcellular localization. The long‐term goal of this project is to identify ways of inhibiting ICER degradation in cancer cells as a new mode for cancer treatment. This project was supported by a grant from the Sokol Institute of Pharmaceutical Sciences, College of Science and Mathematics, Montclair State University.
Possible role for the adaptor protein complexes in sorting of human papillomavirus
Authors: Schlegel Angela M , Chapman Janice A. , Campos Samuel K.
Published: June 2021

Human papillomaviruses (HPVs) infect genital and mucosal epithelium and persistent infection with high‐risk strains like HPV16 are associated with cervical, anogenital, and nasopharyngeal cancers. HPV16 particles consist of two structural proteins: the major capsid protein L1 and the minor capsid protein L2. In a previous yeast two hybrid screen using L2 as bait, we identified a potential interaction with a truncated μ chain (μ2) of the adaptor protein complex‐2 (AP2). The AP complex family directs vesicular sorting of cargo via interactions between the AP complex μ subunits and YxxΦ motifs within the cytoplasmic tails of transmembrane receptors. Interestingly L2 contains a number of YxxΦ motifs with variable conservation among different HPV types. The goal of this project is to determine which AP complexes L2 interacts with, how these interactions occur, and the functions of these interactions during infection. Yeast two‐hybrid screens indicate that full‐length L2 interacts with all AP complex μ chains but smaller L2‐YxxΦ peptides have preference for specific μ chains. Infections with YxxΦ‐mutant viruses show decreased infectivity, strongly supporting a role for AP complexes in infection. siRNA single knockdowns of AP complex μ chains show varying effects on infectivity. Current work seeks to understand the nature of these defects and to assess specific roles for L2‐AP complex interactions during infection.

Support for this research comes from the Undergraduate Biology Research Program (U. of Arizona UBRP) and the American Cancer Society Research Scholar Grant RSG‐117469.
Phosphorylation of p57/coronin‐1 regulates phagocytosis through the modulation of actin‐binding activity
Authors: Oku Teruaki , Ando Yusuke , Tsuiji Makoto , Toyoshima Satoshi , Tsuji Tsutomu
Published: June 2021

The actin‐binding protein p57/coronin‐1, a member of the coronin protein family, is selectively expressed in hematopoietic cells and plays crucial roles in the immune response through reorganization of the actin cytoskeleton. We have studied on the intracellular distribution of p57/coronin‐1 in differentiated HL60 cells during phagocytosis, and found that p57/coronin‐1 and F‐actin were transiently accumulated on phagosomes and then dissociated from phagosomes before phagosome‐lysosome fusion. When the cells were treated with inhibitors of protein kinase C (PKC), p57/coronin‐1 and F‐actin remained on phagosomes without dissociation. This phenomenon seems to be similar to that observed after phagocytosis of Mycobacterium bovis BCG. Since we found that p57/coronin‐1 was phosphorylated by PKC, we assumed that the phosphorylation of p57/coronin‐1 triggered its dissociation from phagosomes and is an essential step for phagosome‐lysosome fusion. Recently, we reported that phosphorylation at Thr‐412 of p57/coronin‐1 regulated its actin‐binding activity and increased during phagocytosis. These studies provide mechanistic insights into roles of p57/coronin‐1 in phagosome‐lysosome fusion and bacterial parasitism.