bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression profile of radiation‐responsive microRNAs in the human gastric cancer cell
Authors: Shen YanNan , Kim SangHoon
Published: June 2021

Ionizing radiation(IR) is generally used for cancer treatment. It induces cellular DNA damage through multiple mechanisms including ATM/ATR‐p53 pathways. MicroRNAs (miRNA) are small, well‐conserved, non‐coding RNA species that have recently been found to regulate protein translation by mRNA. Although miRNAs play important roles in the regulation of crucial processes such as the cell proliferation and apoptosis, the role of miRNA in tumor cells by IR is still unclear. In the present study, we determined the profile of microRNA expression in the SNU‐638 cells in response to IR. Firstly, we examined the radiosensitivity of SNU‐638 cells by the cell viability assay. After IR treatment, SNU‐638 cells showed high survival cell population than radiosensitive HCT116 cells. Thirteen miRNAs among the 200 miRNAs were selected, which the levels of their expression were changed more than two‐fold. The genes targeted by IR‐responsive miRNAs were predicted and yielded several genes, many of which are involved in the regulation of apoptosis, the cell cycle, and DNA repair. Taken together, these data may be useful for the regulation of genes involved cellular response to radiation.
Placenta Vitamin D Signaling in Preeclampsia
Authors: Wheeler Carol , Benoit Joseph N , Anderson Cindy M
Published: June 2021

National trends demonstrate that an overall increase in the prevalence of vitamin D insufficiency is associated with risk for preeclampsia (PE), particularly among those with reduced sun exposure. Previous studies in our lab demonstrated that 70% of women pregnant with their first baby living in the northern plains had pregnancies complicated by hypovitaminosis D and that 24% of the study participants experienced PE. As the placenta produces and utilizes vitamin D in a paracrine fashion, we investigated the differences in expression of key proteins involved in vitamin D signaling in placentas from women with normotensive pregnancy (NP) and in PE. Placenta samples were collected after delivery and frozen for analysis of protein expression by Western blot analysis. Lysates from placental homogenates were subjected to electrophoresis, transferred to a PVDF membrane and incubated with antibodies against 1á‐hydroxyase (CYP27B1), vitamin D (VDR) and retinoid X (RXR) receptors. There were no significant differences in expression of VDR or RXR between groups. Expression of CYP27B1 was significantly increased in PE compared to NP (p<0.05). Our findings indicate that placental conversion of the circulating form of vitamin D to the active form is increased in preeclampsia, increasing availability of vitamin D for utilization by the placental‐fetal unit. Supported by Robert Wood Johnson Foundation, APS.
Maternal Exercise Dose Response Effect on Fetal Heart Rate and Variability
Authors: May Linda E , Glaros Alan G , Gustafson Kathleen M , Suminski Richard R
Published: June 2021

We did a retrospective analysis to determine if there was a dose response relationship between maternal exercise and fetal heart rate (HR) and heart rate variability (HRV). We used hierarchal linear regression of 36 wk data from MCG recordings and activity questionnaires. Associations between maternal kilocalories per min (kcal/min) and total kcal expended (Tkcal) during exercise and fetal HR and HRV (ie. SDNN, RMSSD, VLF, LF, HF) were examined. We controlled for maternal resting HR, maternal weight, maternal kcal while sedentary, and fetal activity state. In univariate analysis, SDNN, VLF, and LF were positively correlated with Tkcal (p<0.05) and fetal HR was negatively correlated. SDNN and HF were positively correlated with kcal/min (p<0.05). After controlling for confounders, significant associations were found with Tkcal and SDNN (t=2.7) and between kcal/min and fetal HR (t = −2.3), RMSSD (t= 2.2), and HF (t=2.2). Maternal exercise variables explained significant portions of the variance in fetal measures: HR (17.8%), RMSSD (15.5%), SDNN (20.4%), and HF (15.7%). Maternal exercise intensity and energy expenditure has a dose response effect on the fetal heart consistent with the development origins hypothesis. These findings will guide future studies using exercise as an intervention for improving maternal and fetal outcomes in pregnancies complicated by gestational diabetes, obesity and hypertension.

Research supported by: KCUMB and Hoglund Brain Imaging Center funds

Grant Funding Source: KCUMB
Effective Strategies for Integration of Basic & Clinical Neuroscience
Authors: Norden Jeanette Jean
Published: June 2021

The Brain & Behavior Module at the Vanderbilt Medical School is a 13 week interdisciplinary module for second year medical students that encompasses both basic and clinical material (anatomy, pharmacology, radiology, pathology, and behavioral science) relevant to the Neurology and Psychiatry Clerkships. As Director of the Module, my role is to integrate basic and clinical neuroscience vertically (within a group of lectures) and horizontally (between lectures over the 13 weeks of immersion). There are inherent challenges to the success of such intense modular courses, for the students, the faculty, and for the module Director. Our experience has been that integration ‐ which goes to decreasing stress and neurophobia in students ‐ is best achieved by an interweaving of the basic and clinical sciences in multiple venues by all participating faculty, allowing for synergistic reinforcement of the knowledge to be acquired. Multiple examples of how basic science and clinical faculty cooperate in this endeavor will be discussed in the hope that some parts of our model may be helpful to others involved in such multidisciplinary courses/modules.
Effects of lifelong decreased IGF‐1 on physiological status in mice
Authors: Motch Susan Marie , Sell Christian , Graboski Robyn , Matzko Michelle , Stout Joseph Tobias , McCarter Roger
Published: June 2021

Caloric restriction (CR) and repression of the GH/IGF‐1/insulin signaling pathway increase lifespan, possibly via common genetic mechanisms of IGF‐1 regulation and lifespan. To elucidate effects of low plasma and tissue IGF‐1 on longevity, midi (IGF‐1 hypomorph) mice were compared to sex and age matched controls. Bone mineral density (BMD) was examined at 100±8 d (young, n=30) and 495±5 d (old, n=42) by dual energy x‐ray absorptiometry. Metabolic rate (MR), physical activity (PA) and core body temperature (CBT) were concurrently measured over 24 hrs. Data were analyzed by ANOVA with significance at p<0.05. BMD was lower in young midi mice vs. control, with no effect of sex. BMD was lower in old midi mice vs. control, with males exhibiting lower BMD than females. Length and weight were correlated in young and old mice. CBT did not vary by genotype or sex in young mice. CBT was affected by genotype and sex with a sex*genotype in old mice. CBT between young and old midi mice did not differ, though young and old control mice differed. Old midi mice displayed less PA than control with no sex effect. MR differed by genotype and sex with a sex*genotype. Lifelong depressed plasma IGF‐1 leads to some similarity between CR and midi mice (lower CBT, BMD) but not others (MR, PA) suggesting that lowered IGF‐1 is not the major factor involved in the life extending effects of CR. Supported by NIH Grant AG022443, PI C. Sell.
Effects of Daily Insulin Treatment on Bone in Rats Following Severe Burn and Disuse
Authors: Baer Lisa , Pidcoke Heather , Wu Xiaowu , Silliman David , Walters Thomas , Tou Janet , Wolf Steven , Wade Charles
Published: June 2021

Traumatic burn increases bone fractures in children and osteopenia in adults. Using our burn unloading model in rats (BU), we have previously observed a decrease in bone mineral density (BMD), content (BMC), and strength 14 days after burn. Insulin infusion is readily used for glucose control with burn patients and improves burn induced hypermetabolic responses, however the direct effect of insulin on bone is not well documented. In the present study we determined the effects of daily insulin treatment on BMD, BMC and strength using BU. Male rats were randomized into two groups: BU‐Vehicle (BUV; n=7) or BU‐Insulin‐Treated (BUI; n=6). Rats received a 40% total body surface area scald burn and following recovery were immediately hindlimb unloaded. Daily subcutaneous injections were either long‐acting insulin (5U/kg/day) or 0.9% saline. On day 14 after injury femurs were collected for DEXA scan and 3‐point breaking strength. No differences were observed in BMD. BUI showed a significant decrease in BMC, however no differences were observed in overall bone strength between the groups. Based on these results, insulin therapy at the current dose may not be effective for maintaining bone in burn patients. Funding: US Army MRMC & Juvenile Diabetes Foundation
Cost‐effective middle school summer camps build the science pipeline
Authors: Wyss J Michael , Radford David , Ginwright Shriley , Lockhart Jarrod , Williams Mary , Latner J. Bree , Chase Melissa , Steele Taylor , Wyss Steven A , Yang Jerry
Published: June 2021

To provide students (especially minority students) enrichment activities that will lead to an increased understanding of biology and interest in careers in biomedical science, we developed and evaluated CORD Summer Science Camps, which greatly enhance the interest and performance of participants in science education. Camps increased from 178 students (5 camps, 5 wks each; 2005) to nearly 500 students (6 camps, 5 wks each; 2009). All camps relate biological science lessons to the grade‐specific physical, biological and earth science the students encounter the next school year. In each 1‐wk (40h) camp, students conduct different hands‐on experiments and learn to analyze and present the results to their parents at camp's end. ~70% of campers are minority, urban student (50% live below the poverty line); 30% are from privileged schools. This integrates students across socio‐economic strata in science education. ~75% of area middle school teachers who gain professional training as Camp Facilitators use camp experiences in classrooms. Campers and teachers become more confident in hands‐on science and leaders in their classrooms. About 75% of the campers return for subsequent years and indicate increased interest in science careers (38% pre‐ vs. 76% post‐camp). We are assessing the impact of camps on standardized science scores. Camps are relatively cost effective, sustainable methods to build the biology pipeline.
Graduate students design neuroscience outreach educational activities for grades 3–5
Authors: Puder Barb
Published: June 2021

Graduate students in the Masters of Occupational therapy program at Samuel Merritt University selected a neuroscience topic area and created an informational poster, hands‐on learning activity to reinforce neuroscience concepts, and a short presentation describing the neuroscience topic. Students self selected to work with 3–4 other graduate students and submitted 3 possible neuroscience topics that they were interested in researching the relevant information. Requirements for the neuroscience educational outreach project included choosing a neuroscience topic that is interesting and relevant to children in grades 3–5, creating a poster that uses terminology and images that are age appropriate, and designing an interactive 10 minute presentation that includes a fun, hands‐on activity to engage the children in the neuroscience topic. Periodic assignments were given to keep each of the graduate students groups on pace for a successful completion of the project which was presented during the last week of the semester at a local elementary school. Graduate students were assessed on several categories regarding the poster, presentation, and the hands‐on activity. This score was part of their neuroscience grade for the semester. Graduate students also completed a survey regarding their perceptions and value of the project. Survey questions were based on the Likert scale.
FIRST: Fellowships in Research & Science Teaching: A differential approach to postdoctoral training
Authors: Mitrano Darlene A , Eisen Arri , Eaton Douglas C
Published: June 2021

As part of the IRACDA (Institutional Research and Career Development Awards, NIGMS) initiative, the FIRST program was developed in order to increase underrepresented minorities in postdoctoral fellowship positions in the life sciences, as well as to increase research oriented teaching at minority serving institutions (MSIs). This is accomplished through a collaborative effort, with the majority of the postdoctoral fellows conducting research at Emory University or Morehouse School of Medicine, while completing their teaching component at one of the historically black colleges in the Atlanta University Center (Morehouse College, Spelman College, or Clark‐Atlanta University). FIRST fellows, therefore, have a research‐intensive experience and at the same time have an opportunity to engage in teaching activities, an experience quite unusual compared to that of most traditionally trained postdocs. FIRST fellows are at least as successful by any measure as a traditionally trained postdoc. A study of 114 FIRST fellows compared to over 500 traditional postdoctoral fellows revealed that not only do FIRST fellows publish in scientific journals at the same rate as the comparison group, they obtain more academic jobs; receive NIH grants at a comparable rate; contribute to diversity at Emory; and to the faculty at the MSIs.
Legacy TRIPSEs (Tri‐Partite Problem‐Solving Exercises): Fostering Student Engagement and Learning in Large Classes
Authors: Rangachari Patangi K , Nastos Stash
Published: June 2021

The TRIPSE is a process‐oriented problem solving exercise that mimics the scientific process. Using limited data, students: 1) frame hypotheses; 2) design experimental tests; and 3) later with new information, reassess answers. Earlier we validated it as an evaluative exercise in a large class (The FASEB Journal. 2008; 22:767.1). We report here the use of TRIPSEs to promote student engagement and learning in a freshmen introductory biology class. In part 1, students learned about gene expression and cell signaling. In part 2, students (groups of 5) incorporated elements of what they had learned into designing novel problems that could be left as a “legacy” for future classes. Groups were told to: 1) write challenging problems based on published material; 2) state 4 plausible hypotheses and for each hypothesis indicate appropriate experimental tests; and 3) provide annotated references. Interim feedback was given. Groups were assessed on clarity, creativity, plausibility of hypotheses, relevance of the experiments proposed and corroboration for statements made. On an exit survey, a total of 322 students from 2 separate years rated this exercise the most valuable amongst several used.
Eukaryon: An undergraduate peer‐reviewed and managed life sciences scholarship journal
Authors: Fiske Michael , DebBurman Shubhik , Smith Pliny , Davis Shaun
Published: June 2021

Undergraduate research and inquiry‐based pedagogy are increasingly crucial components to college education in the U.S. Providing undergraduates the opportunity to publish their scholarship is a tool to increase scientific literacy and motivation for scientific careers. Spurred by an NSF grant in 2004, the Lake Forest College Biology Department developed a peer‐reviewed undergraduate research journal of life science scholarship entitled Eukaryon (http://www.lakeforest.edu/eukaryon). In each annual issue, Eukaryon publishes scholarship in the life sciences that students produced within undergraduate classrooms and faculty labs. We detail three aspects of the journal's development: 1) how it is governed, published, and financed by undergraduates; 2) how it establishes content and format; and 3) specific strategies that maintain selectivity, encourage inquiry‐based curriculum, and support a community of student scholars. Next, we discuss a survey‐based assessment of our journal's impact on curriculum and student community. We found that Eukaryon contributes significantly to the preparedness for future careers of its board members. Finally, we report an analysis of our online readership using data from Google Analytics. Future goals include strengthening community familiarity with Eukaryon and improving communication with faculty on submission and review progress.

(Supported by NSF‐CCLI)
Calcified Integumentary Structures in Anurans
Authors: Delorme Stephanie , Vickaryous Matthew Kenneth
Published: June 2021

Although often overlooked, the integument of many vertebrates is reinforced by skeletal elements. In anurans these calcified structures include osteoderms, bone‐rich elements, and the lamina calcarea, a calcified, acellular layer that lacks collagen. We investigated the histology of anuran integument with a special focus on taxa developing skeletal elements. When present, the lamina calcarea forms a thin but discrete horizon at the interface between the superficial and deep dermis. This layer is rich in glycosaminoglycans (GAG) and is structurally unlike bone and cartilage. In contrast, anuran osteoderms are composed of cellular bone with limited GAG. A partial ontogenetic series of the tree frog Phyllomedusa bicolor provides the first details of anuran osteoderms development. In P. bicolor osteoderm development begins as a cluster of small calcified centres localized over the head. With continued body growth, osteoderms become larger in size and become distributed across the dorsal and ventral surfaces of the body, and across the limbs. The leaf litter frog Brachytarsophyrys cariensis is unusual in developing both osteoderms (on the dorsal body surface) and a lamina calcarea (on the ventral body surface). Our data shows that anuran integument has the ability to form skeletal elements and suggests that these elements arose in multiple anuran lineages.

Grant Funding Source: NSERC
MicroRNA 200c Targets Noggin Expression: A Role for MicroRNAs in Dental Epithelial Cell Differentiation
Authors: Amendt Brad A , Cao Huojun
Published: June 2021

MicroRNAs are known to control gene expression patterns in other tissues and organs and in this report we demonstrate that craniofacial/tooth development is tightly controlled by microRNAs. To understand the role of microRNAs in tooth morphogenesis dental epithelial and mesenchyme tissue was screened for microRNA expression. There are specific sets of microRNAs expressed in the dental epithelium compared to the dental mesenchyme. Furthermore, specific microRNAs are expressed in incisors compared to molars revealing a code of microRNAs required for tooth development. Interestingly, bioinformatics analyses indicate only a small number of genes are regulated by microRNAs during late stages of tooth development. Conditional knockout (cKO) of Dicer1 (mature microRNAs) in the dental epithelium using the Pitx2‐Cre mouse results in multiple and branched enamel‐free incisors, a change in incisor patterning, and incisor and molar size and shape. Furthermore, deletion of oral epithelial microRNAs resulted in a cleft palate, revealing new mechanisms for their role in regulating palatogenesis. Epithelial microRNAs control incisor dental epithelial cell differentiation demonstrating a unique role for microRNAs in regulating in vivo dental stem cell biology. Noggin, a potent BMP inhibitor is up‐regulated in the Dicer1 knockout mouse, which prevents BMP signaling and cell differentiation. MicroRNA 200c targets Noggin to allow for normal tooth development and a mechanism to regulate cell differentiation. These results demonstrate a critical role for microRNAs in regulating tooth and craniofacial development through the dental stem cell niche and a mechanism involving microRNA 200c regulation of Noggin and BMP signaling. Support for this research was provided from grants DE13941 and DE18885 to BAA from the National Institute of Dental and Craniofacial Research.
Pannexin 3, a gap junction protein, Regulates Odontoblasts Differentiation
Authors: Iwamoto Tsutomu , Nakamura Takashi , Yamada Aya , Yamada Yoshihiko , Fukumoto Satoshi
Published: June 2021

Pannexin 3 (Panx3) has recently been identified as a new member of the pannexin family of gap junction proteins, however its expression and physiological function in tooth development have been unknown thus far. In this study, our objectives are to analyze the expression of Panx3 and to identify the role of Panx3 in tooth development. We found that Panx3 was strongly expressed in developing teeth. In situ hybridization revealed that Panx3 mRNA was expressed in the pre‐odontoblast layer, which represents an intermediate stage between proliferating dental mesenchymal cells and differentiated odontoblasts. Immunohistochemistry showed that the Panx3 protein was localized in cell membranes, especially in cell‐cell contact lesions as well as in cell‐matrix adhesion sites, where cells face the basement membrane and predentin. To analyze the function of Panx3, we used odontogenic cell line mDP6. Panx3 expression was induced during differentiation of mDP6 cells. Further, overexpression of Panx3 by transfection promoted mDP6 cell differentiation, while suppuration of endogenous Panx3 expression by siRNA inhibited mDP6 differentiation. Thus, our results suggest that Panx3 plays a role in the promotion of odontoblast differentiation. (Supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan and the Intramural Program of the NIDCR, NIH, USA.)
MICROANATOMY OF THE MANDIBULAR SYMPHYSIS IN LIZARDS
Authors: Holliday Casey , Gardner Nicholas , Paesani Sylvia , Douthitt Mallory , Ratliff Jack
Published: June 2021

Although the mandibular symphysis is a functionally and evolutionarily important feature of the vertebrate skull, little is known about the soft‐tissue morphology of the joint in squamate reptiles. Lizards evolved a diversity of skull shapes and feeding behaviors, thus it is expected that the morphology of the symphysis will correspond with functional patterns. Here we present new histological data illustrating the morphology of the joint in a number of lizard taxa. The symphyses of all taxa exhibit dorsal and ventral fibrous portions of the joints that possess an array of parallel and woven collagen fibers. The middle and ventral portions of the joints are complemented by contributions of Meckel's cartilage. Kinetic taxa have more loosely‐built symphyses with large domains of parallel‐oriented fibers whereas hard biting and akinetic taxa have symphyses primarily composed of dense, woven fibers. Whereas most taxa maintain unfused Meckel's cartilages, iguanids and geckos independently evolved fused Meckel's cartilages however the joint's morphologies suggest different developmental mechanisms. Fused Meckel's cartilages may be associated with the apomorphic lingual behaviors exhibited by iguanids (tongue translation) and geckoes (eye wiping). These morphological data shed new light on the functional, developmental, and evolutionary patterns displayed by the heads of lizards.

Grant Funding Source: West Virgina NASA Space Grant Consortium
Long faces and small eyes: Papionin models for the relationship between orbit size and facial prognathism
Authors: Yang David , Pagano Anthony Santino , Laitman Jeffrey T
Published: June 2021

The papionins are unique among anthropoid primates for their long faces and large canines. The genera Papio and Mandrillus are here used as natural experiments for facial elongation among microsmatic primates. It is hypothesized that the bony orbits of long faced papionins take on smaller dimensions so as not to encroach upon the elongated nasal cavity. Long faced Papio and Mandrillus are contrasted against relatively orthognathic Macaca, Cercocebus, and Lophocebus via 3D geometric morphometrics. Only adult males were used since marked sexual dimorphism was present among all species. 3D landmark coordinates were collected around the face and upper respiratory tract. Landmark coordinate data between individuals was optimally superimposed and scaled using the Procrustes method, allowing direct comparison of linear measures, areas, and volume. It was found that Papio and Mandrillus are distinguished from Macaca, Cercocebus, and Lophocebus in palate length, area of the piriform aperture, orbital volume, and width of the orbital rim (p<.05). There is a strong negative correlation (−.78, p<.0001) between orbital volume and palate length and a weaker correlation between orbital volume and palate width (.57601; p<.0001). The data supports a functional relationship between orbit size and nasal cavity length.

Grant Funding Source: Mount Sinai CAFM Student Research Support
Ephrins stimulate developmental bone growth
Authors: Atobajeun Lola , Fernandez Claudia , San Miguel Symone , La Jan , Opperman Lynne , Dechow Paul , Henkemeyer Mark , Benson M. Douglas
Published: June 2021

The Eph receptor tyrosine kinases and their ligands, the ephrins, control boundary formation during cranial suture development. They also regulate mineral homeostasis in the long bones. We hypothesized that ephrins regulate bone growth in the calvarium separate from pattern formation, and that exogenous activation of Eph signaling can be used to promote bone synthesis. We found strong expression of beta‐galactosidase in the periosteum and the dura mater of late embryonic ephrin‐B2/LacZ and EphB2/LacZ mice (in which a lacZ reporter gene was knocked in to the ephrin‐B2 or EphB2 locus, respectively) after suture patterning was complete, but during the period of vigorous bone growth. EphB2 was particularly concentrated in the osteogenic fronts of the frontal bones. Treatment of e17.5 mouse calvariae for five days with recombinant ephrin‐B2 ectodomain fused to human Fc dramatically increased bone volume without changing bone density or interfrontal/sagittal suture widths. We conclude that Eph signaling stimulates bone synthesis apart from its role in cranial patterning. Thus, recombinant ephrin may be useful for the repair of cranial defects. Ongoing studies are concentrating on the mechanism of ephrin action in osteoblast differentiation and gene expression.
Molecular Regulation of Osteoactivin Expression in Osteoblasts
Authors: Singh Maneet , Del Carpio‐Cano Fabiola , Monroy Alexandra , Popoff Steven N
Published: June 2021

Osteoblast (OB) commitment, proliferation and differentiation from mesenchymal stem cells are regulated by Bone Morphogenetic Protein2 (BMP2). BMP2 mediates these osteoinductive effects by recruiting Transcription Factors (TFs), namely Runx2, Homeodomain (HD), Smad1 and Smad4 proteins to the promoter of OB specific genes and induces their transcription. Our laboratory has previously shown that Osteoactivin (OA), OB‐related glycoprotein acts as a downstream mediator of BMP2 effects on OB differentiation and function. Furthermore, we showed that Smad1 signaling regulates BMP2 induced OA expression. The goal of our study is to characterize OB specific TFs that regulate BMP2 induced OA promoter activity. In order to study the effects of BMP2 on OA promoter activity, we cloned 1kb upstream of rat OA gene into a luciferase expression vector and have shown that BMP2 stimulates OA promoter activity in a time and dose‐dependent manner. Secondly, knockdown of Runx2, HD, Smad1 and Smad4 using specific siRNAs decreased BMP2 induced OA promoter activity. Similarly, loss of consensus binding sites of these TFs from OA promoter decreased OA promoter activity. Furthermore, ChiP assays at various stages of OB differentiation indicated that Runx2, Smad1, Smad4 and HD proteins associate differentially with OA promoter. Collectively, our results suggest that Runx2, Smad1, Smad4 and HD TFs differentially regulate BMP2 induced OA promoter activity during OB differentiation.

Grant Funding Source: NIAMS
Go/i Signaling is Involved in Facilitation of Neurite Outgrowth by Reelin
Authors: CHO SUNG‐KUK , YOON YEO‐CHO , KANG WOO‐YOUNG , SUH‐KIM HAEYOUNG , LEE YOUNG‐DON
Published: June 2021

The extracellular protein, reelin, which binds to ApoER2, VLDLR controls the positioning of radially migrating neurons and synaptic plasticity. During neurodevelopment activation of Go/i, a heterotrimeric G protein, facilitates the axon growth. Here we investigated the relationship between of Reelin signaling and Go/i during neurite formation of primary hippocampal neurons. In reeler, reelin‐deficient mice, neuritogenesis was significantly decreased. Inhibition of neuritogenesis in reeler was rescued by reelin treatment. The localization of Gαi1, Gαi3 and Gαo were obvious in the terminal portion of growing neurites in wild type, but not in reeler. In addition, treatment of pertussis toxin, Go/i inhibitor, suppressed the neurite formation by reelin. In study using F11 cells transfected Gαo mutant (Q205L), induction of neurite outgrowth by reelin could not be observed. To address the underlying mechanism, we investigated the downstream signaling pathway of reelin and Go/i using specific inhibitors. Reelin treatment induced the activation of MAPK‐JNK pathway, which is well‐known as a major pathway in neuritogenesis. Taken together, we suggest that Go/i signaling may be involved in induction of neuritogenesis by reelin.
Honokiol Regulates Phosphorylation of Tau via Inhibition of GSK3β
Authors: Le Nguyen Mai Khoa , Kim So‐Yeon , Lee Dalim , Kim Sung‐Soo , Suh‐Kim Haeyoung , Ryu Jae‐Ha , Lee Young‐Don
Published: June 2021

Abnormal phosphorylation of tau plays an important role in neurodegeneration and subsequent dementia in Alzheimer's desease (AD). Much effort has been dedicated to treat AD, including inhibits GSK3β which is major kinase responsible for hyperphosphorylated tau. Honokiol, one of the major components isolated from Magnolia obovata, has been reported to have variety of pharmacological activities including anti‐inflammation and neuroprotection.

In this study, the main objective was to determine whether Honokiol has inhibitory effects on phosphorylation of tau via modulation of GSK3β activities. Honokiol suppressed phosphorylation of tau in a dose‐dependent manner and inhibited GSK3β in 293T cells. Cholinergic neuroblastoma cell lines, LA‐N‐2 or SN56.B5.G4 were differentiated by CNTF and the cells were treated with oligomeric Aβ for an AD‐like pathogenic condition. The cells showed morphological changes such as long processes and branches, cell death and aggregation in the presence of CNTF or Aβ. Treatment with Honokiol not only recovered morphological changes but also decreased the phosphorylation of tau and the activity of GSK3β in vitro.

Although further experiments about CDK5 which is known as the other cause for tau phosphorylation are required, our results implies that Honokiol regulates the phosphorylation of tau and the phosphorylation of GSK3β.
Neuroinflammation in a Forceps Compression Model of Spinal Cord Injury
Authors: Vaughn Chloe N. , Henley Jessica B. , Iafrate Julia , Stevenson Edward , Jones T. Bucky
Published: June 2021

We sought to evaluate the neuroinflammatory response in a compression model of spinal cord injury (SCI). Because compression is a common mechanism of injury in human spinal cord trauma, such a model has clinical relevance. In this study we used forceps to induce lateral compression injury to the spinal cord, producing a moderate degree of injury as previously described in the contusion model (Basso et al., 1996).

Post‐injury inflammation has been implicated in secondary degeneration but has not been characterized in this model. We evaluated the inflammatory response in the injured spinal cord using immunohistochemistry. We found that neutrophils are present within the lesion site by 1 day post injury (dpi) but do not persist past 3 dpi. T‐cells were present by 1 dpi, mainly in gray matter at the epicenter. They peaked in number at 7 dpi, but persisted to 42 dpi. A similar time course was observed for macrophage activation, although the response was more widespread at the impact site and extended further rostrocaudally. Overall, we confirm that forceps compression of the spinal cord produces a neuroinflammatory response similar to what has been described in human spinal cord trauma and in contusion models of experimental spinal cord trauma. Thus, forceps compression represents a clinically relevant experimental model for evaluating secondary injury.

Grant Funding Source: MWU Biomedical Sciences program
Increased sensitivity of PACAP knockout mice to retinal ischemia
Authors: Reglodi D. , Szabadfi K. , Kiss P. , Tamas A. , Hashimoto H. , Baba A. , Shintani N. , Gabriel R. , Helyes Z. , Atlasz T.
Published: June 2021

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuroprotective peptide exerting protective effects in neuronal injuries. PACAP is protective in several models of retinal degeneration in vivo. Our previous studies have shown that PACAP treatment ameliorates the damaging effects of chronic hypoperfusion modeled by permanent bilateral carotid artery occlusion. The aim of the present study was to compare the degree of retinal damage in wild type and PACAP deficient mice in ischemic retinal insult. Mice underwent 10 min of carotid artery occlusion followed by 2‐week reperfusion. Retinas were then processed for histological analysis. It was found that PACAP deficient mice had significantly greater retinal damage, as shown by the thickness of the whole retina and the morphometric analysis of the individual retinal layers, such as inner and outer plexiform and inner and outer nuclear layers. The retinal damage in PACAP knockout animals could be attenuated by exogenous intravitreal PACAP. These results clearly show that endogenous PACAP reacts as a stress‐response peptide that is necessary for endogenous protection against different neuronal insults.
Histological validation of the diffusion tensor: feasibility in human brain tissue
Authors: Ortiz Jake , Hageman Nathan , Salin Ashley , Salin Megan , Dong Hong‐wei , Stark M. Elena , Vinters Harry V. , Toga Arthur W. , Wisco Jonathan J.
Published: June 2021

Taking advantage of anisotropic water movement caused by cytoarchitectural properties of brain tissue, Diffusion Tensor Imaging (DTI) is capable of representing approximate true nerve fiber orientations as a diffusion tensor at each voxel. The interpretation of DT images remains hampered in part by the error inherent in tensor calculations. This pilot study demonstrates the feasibility of histological validation of DTI in post‐mortem human brain tissue, with the eventual goal of constructing a probabilistic, ground‐truth atlas of fiber orientations specifically designed to reduce the uncertainty of tensor calculations. As a first step, we mapped nerve fibers of the corpus callosum, cingulum bundle and the intersecting region from images of histological sections nonlinearly registered to tensor maps. We confirmed that tensors accurately represented homogeneous but not necessarily heterogeneous fiber populations. Histological validation can provide a priori constraints for tensor calculations and will have a direct impact on the reconstruction of fiber tracts and help clinicians to identify and manage diseases involving white matter pathologies with greater sensitivity and specificity. This work was supported in part by NIH grants U54 RR021813, P41 RR013642, P50 AG16570, R21 MH083180, R90 DA023419‐01, T90 DA023419‐01 and GM08042.

Grant Funding Source: U54 RR021813, P41 RR013642, P50 AG16570, R21 MH083180, R90 DA023419‐01, T90 DA023419‐01 and GM08042
Phrenic nerve morphometry in male and female spontaneously hypertensive rats
Authors: Rodrigues Anaceres Ribeiro , Ferreira Renata Silva , Salgado Helio Cesar , Fazan Valeria Paula
Published: June 2021

Histological examination of the diaphragm showed gender differences in fiber composition but gender differences on phrenic nerve morphometry were not thoroughly addressed. Ventilatory differences were described between male and female rats, as well as among different rat strains. However, the morphology of the phrenic nerve was not investigated in spontaneously hypertensive (SHR) and normotensive Wistar‐Kyoto (WKY) rats. After arterial pressure and heart rate recordings, phrenic nerves of 20 week‐old male and female SHR and WKY were prepared for epoxy resin embedding and light microscopy. Morphometry was performed with the aid of computer software. Our results showed for male and female SHR remarkable longitudinal asymmetry on both sides and an asymmetry between sides on distal segments, which were not present on WKY nerves. Phrenic nerves were generally larger in males compared to females on both strains but larger on WKY compared to SHR for both genders. Myelinated fiber number and density were significantly larger on females of both groups and remarkably larger on WKY compared to SHR in both genders. Strain and gender differences on phrenic nerve myelinated fiber number is described for the first time in this experimental model of hypertension, pointing to the need for thorough functional studies of this nerve on male and female SHR.

Grant Funding Source: FAPESP
Alterations in atypical PKC expression and activity near the phrenic motor nucleus following cervical spinal injury
Authors: Guenther Courtney H. , Tubon Thomas C. , Windelborn James A. , Yin Jerry C.P. , Mitchell Gordon S.
Published: June 2021

Atypical protein kinase C (aPKC) is a kinase family critical for maintenance of hippocampal synaptic plasticity (Pastalkova and Serrano, et. al., 2006). aPKCs are expressed in phrenic motor neurons (Guenther and Mitchell, unpublished), a key site of respiratory plasticity (Mitchell, et. al., 2001), particularly following C2 cervical hemisection (C2HS). We tested the hypothesis that C2HS increases the expression and activity of atypical PKCs near the phrenic motor nucleus suggesting a contribution to injury‐induced respiratory plasticity. C2 laminectomy (sham) or C2HS were performed on adult, male Lewis rats. Ventral spinal segments C3‐5 were harvested 1, 3 or 28 days post‐surgery and prepared for enzyme activity assays and immunoblots to assess changes in aPKC activity and expression. Ventral cervical aPKC activity increased at 1 and 28 days (not 3) post‐C2HS (1 day: 63% vs shams ipsilateral to injury; p<0.05; 28 day: increased activity vs sham, but without difference ipsilateral and contralateral to injury; p<0.05). Total PKCζ protein expression was unchanged by C2HS, but total PKMζ (constitutively active PKCζ isoform) was increased vs shams ipsilateral to injury at 28 days post‐C2HS. Changes in aPKC expression near the phrenic motor nucleus following a C2HS suggest the possibility that aPKCs contribute to spinal respiratory plasticity following cervical spinal injury (NIH HL 80209 and HL69064).

Grant Funding Source: NIH
Controlled Expression of a Cystatin C‐Peptide as Therapy for Alzheimer's Disease
Authors: Shaw Phyllis A. , Luo Jian
Published: June 2021

Alzheimer's disease (AD) is an incurable, degenerative neurological disorder leading to memory loss, impaired cognitive abilities and changes in behavior and personality. Hallmarks of AD are neurofibrillary intracellular tangles and extracellular senile plaques, resulting from improperly processed amyloid precursor protein (APP) and aggregated amyloid β (Aβ) peptides. A major therapeutic effort in AD centers on prevention of Aβ aggregates/fibrils. The cysteine proteinase inhibitor, cystatin C (CysC), plays a prominent role in amyloidogenesis. The intact CysC molecule exerts both protective anti‐Aβ effect through its C‐terminal portion (amino acid 101–117) by preventing fibril formation and, a detrimental effect, in its role as a cathepsin inhibitor, preventing Aβ degradation. We showed that a similar cysteine proteinase inhibitor, cystatin S (CysS), is highly regulated in vivo by steroid hormones and the autonomic nervous system, and its 1.9kb promoter is active in vivo in the brain during postnatal development. We made a construct of the CysS promoter and the secreted CysC, 101–117 Aβ binding‐peptide, and are testing the best routes of construct delivery into TgCRND8 AD mice, determining the optimal concentration necessary for Aβ binding in neuroblastoma cells stably expressing APP695 and Aβ, and testing the expression of Aβ binding‐peptide in vivo in TgCRND8 mice.
Bifunctional protein for improved stent performance and endothelial regeneration
Authors: Baumer Yvonne , Leder Christoph , Schoenberger Tanja , Ziegler Melanie , Filzmayer Christina , Buehring Hans‐Joerg , Gawaz Mainrad , Schlosshauer Burkhard
Published: June 2021

Restenosis and thrombosis are undesired effects after stent implantation. Reduced or missing re‐endothelialization of injured coronary arteries shall be abolished by improving stem cell homing at the side of injury to support their differentiation into intact endothelium. We developed a bifunctional protein binding to collagen at the vessel lesion site and to stem/progenitor cells. ELISA studies demonstrated that this bifunctional protein binds to collagen type1 comparably strong as the control protein. Employing stem cell marker overexpressing cells and human CD34+ endothelial progenitor cells on a multi substrate array (MSA) spotted with 14 different extracellular matrix proteins demonstrated the bifunctionality and specificity of the protein. This specificity was substantiated in flow chamber adhesion experiments. Furthermore, we tested colony forming ability and by this endothelial like differentiation of CD34+‐cells on fusion protein coated surfaces. In conclusion, this bifunctional protein is a promising construct to support re‐endothelialization and therefore reducing restenosis and thrombosis risk after stent implantation.

Supported by the German Ministry of Education and Research (BMBF) # 01GU0727 and 01GU0726.
Surface modification by Gas Cluster Ion Beam (GCIB) as a novel drug delivery method for vascular stents
Authors: Khoury Joseph , Kodali Guru Charan , Tarrant Laurence JB , Svrluga Richard C , Kirkpatrick Sean R
Published: June 2021

Much discussion has focused on the possible inflammatory and thrombogenic influences of the polymer coating on drug‐eluting stents (DES) and the potential for late stent thrombosis. Biodegradable polymers and/or microporous materials are being evaluated by others. In this study, we have used Argon based GCIB to adhere the drug to the surface of a metal. GCIB bombardment results in surface modification of the top 100Å, allowing a thin coating of drug to adhere to the surface without polymer. Titanium foils cut into 1.5cm2 pieces were initially processed with GCIB at 5x1014 Ar clusters/cm2. 50μg of rapamycin (RAP) was placed on the surface; one group received a GCIB drug surface processing while another group did not. Pieces of foil were placed in plasma at 37oC on a rotator to simulate in vivo drug elution for up to 14 days (n=3). Pre‐ and post‐ eluted foils were then placed in plates and 104 mouse endothelial cells were seeded per foil, allowed to attach for 24 hours, and counts were measured by MTS assay. RAP on foils not receiving the second modification by GCIB eluted off within 24 hours and increased cell attachment from 576±403 cells (pre‐eluted) to 8876±1170 (24h post‐eluted, p<0.01). Foils that received a second GCIB modification caused the RAP to be eluted off more slowly, requiring 72 hours before cell attachment matched no‐drug controls (p<0.05). Therefore, GCIB could replace polymers as a drug delivery method for DES.
Insights on Rad51 Function from the Structure and Properties of a Filament Interface Mutant
Authors: Morrical Scott Walker , Chen Jianhong , Villanueva Nicolas , Rould Mark A.
Published: June 2021

Rad51 protein promotes homologous recombination in eukaryotes. Recombination activities are activated by Rad51 filament assembly on single‐stranded DNA. Previous studies of yeast Rad51 showed that His352 occupies an important position at the filament interface, where it could relay signals between subunits and active sites. Here we report the 2.5A X‐ray crystallographic structure of the H352Y mutant of yeast Rad51. Biochemical data indicate that H352Y is trapped in a high‐affinity ssDNA‐binding conformation with limited catalytic turnover. The structure of H352Y reveals a right‐handed helical filament in a high‐pitch (130Å) conformation with P61 symmetry. The H352Y structure closely resembles DNA‐bound structures of E. coli RecA protein. Tyrosine substitution at position 352 locks Phe187 from the adjacent subunit into a position that interferes with the gamma‐phosphate binding site of Walker A, potentially explaining the limited catalysis observed in H352Y. Comparison of Rad51 H352Y and other RecA/Rad51 structures reveals a correlation between DNA bound/unbound status and the trans/cis configuration of a conserved peptide bond near Walker B.

Funded by NIH grant number P01 CA098993.
Altering the substrate specificity of the Escherichia coli E1 Component of the 2‐Oxoglutarate Dehydrogenase Multienzyme Complex
Authors: Shim Da Jeong , Nemeria Natalia S. , Balakrishnan Anand , Jordan Frank , Farinas Edgardo T.
Published: June 2021

The substrate specificity of the 2‐oxoglutarate dehydrogenase multienzyme complex (OGDH) was engineered to accept unnatural substrates. OGDH catalyzes the rate limiting step in the citric acid cycle. It contains three components: a thiamine diphosphate dependent oxoglutarate decarboxylase (E1), a dihydrolipoylsuccinyl transferase (E2), and a dihydrolipoyl dehydrogenase (E3). The crystal structure, kinetic studies, and site directed mutagenesis analysis of the E1 component indicate that His260 and His298 are important for recognizing the distal carboxylate of 2‐oxoglutarate. Saturation mutagenesis libraries of His260, His298, and His260/His298 were constructed and screened for activity towards 2‐oxovalerate. Several E1 variants were isolated that are active towards 2‐oxovalerate, and the kinetic parameters were determined. In addition, circular dichroism investigations were performed that identified the formation of the pre‐decarboxylation thiamin‐bound intermediate via formation of the 1’, 4’‐imino tautomer of ThDP. The activity of the overall complex on reconstitution with the E2–E3 sub‐complex shows that substrate specificity is controlled at both the E1 and E2 levels.
Human DNA2 is a mitochondrial nuclease/helicase for efficient processing of DNA replication and repair intermediates and defective in myopathy
Authors: Shen Binghui , Zheng Li , Zhou Mian , Guo Zhigang , Lu Huiming , Qian Limin , Dai Huifang , Qiu Junzhuan , Yakubovskaya Elena , Bogenhagen Daniel F. , Demple Bruce
Published: June 2021

DNA2, a helicase/nuclease family member, plays versatile roles in processing DNA intermediates during DNA replication and repair. Yeast Dna2 (yDna2) is essential in RNA primer removal during nuclear DNA replication and is important in repairing UV damage, base damage, and double‐strand breaks. Our data demonstrate that hDNA2 migrates to the mitochondria, interacts with mitochondrial DNA polymerase γ, and significantly stimulates polymerase activity. We further demonstrate that hDNA2 and flap endonuclease 1 synergistically process intermediate 5’ flap structures occurring in DNA replication and long‐patch base excision repair (LP‐BER) in mitochondria. Depletion of hDNA2 from a mitochondrial extract reduces its efficiency in RNA primer removal and LP‐BER. In additon, we show that three point mutations of DNA2 gene identified in myopathy patients are defective in their biological and biochemical functions.