bluuebunny/crossref_metadata_2025_split huggingface dataset:
Aberrant Retinal Angiogenesis, Structure, and Synaptic Morphology in an in vivo Mouse Model of ROP
Authors: Mezu‐Ndubuisi Olachi Joy , Macke Erica , Kalavacherla Raja , Nwaba Amanda , Ikeda Akhiro 
Published: June 2021

ObjectiveRetinopathy of prematurity (ROP) is a condition of aberrant retinal vascularization in premature infants. Vascular endothelial growth factor (VEGF) is a major angiogenic factor in both physiologic and pathologic retinal angiogenesis including ROP. However, cellular changes that take place in ROP are not fully understood. The aim of this study is to understand changes in retinal morphology, vasculature, and structure in mice with oxygen‐induced retinopathy (OIR), a model for ROP.MethodsTwenty‐two mice underwent hyperoxia exposure at 77% to induce OIR, while twenty‐three age‐matched control mice were raised in room air (RA). After anesthesia, pupils were dilated and fluorescein was injected intraperitoneally. After in vivo imaging at P19, P24, P32, and P47, eyes were isolated and fixed in Bouin's fixative for morphological analysis by H&E staining, and paraformaldehyde for immunohistochemistry. For immunohistochemistry, markers for photoreceptor presynapses (PSD95), bipolar cells (PKCalpha), microglia (iba1), and vascularization (VEGF) were used to investigate synaptic abnormalities, inflammation, and vascularization. To detect apoptosis, ApopTag Fluorescein In Situ Apoptosis Detection Kit (Millipore) was used.ResultsOIR mice had a vascular‐avascular retinal phenotype, with more dilated veins and more tortuous arteries than RA mice in early ages. The OIR retinas showed disorganized synaptic interactions between photoreceptor cells and bipolar cells, with significant thinning in the ganglion cell, inner nuclear, inner plexiform, and outer plexiform layers. Retinal cross‐sections showed altered structure in OIR mice, with dilated blood vessels and neovascularization in the inner retina. There was significantly increased microglia activity in ganglion cell, inner nuclear, and inner plexiform layers. There was increased apoptosis in OIR mice, which decreased over time, but persisted until P47. There was increased VEGF expression in bipolar cells. TUNEL staining showed increased apoptosis in OIR mice than RA mice at P19 (P<0.001), P24 (P<0.001), P32 (P<0.001), and P47 (P<0.001). Apoptosis decreased over time with developmental maturity of the OIR mice, but persisted at P47.Conclusion: Our results demonstrate patchy vascular–avascular phenotype, defective synaptic interactions and persistent apoptosis in the OIR retina, which affected retinal vascularization, and structure in the long term. Our study aids the understanding of the cellular and molecular complexities of the retina following oxidative stress, and could lead to potential new therapeutic targets.Support or Funding InformationDepartment of Pediatrics, University of Wisconsin, Madison (OJM), National Institutes of Health (R01 EY022086) (AI), and Timothy William Trout Professorship in Eye Research (AI)
VEGF is required for the initiation of Cerebral Cavernous Malformations
Authors: Glading Angela , DiStefano Peter , Chen Tiehua , Su Hua , Whitehead Kevin 
Published: June 2021

There is a narrow developmental window of susceptibility shortly after birth in which genetic deletion of CCM genes is sufficient to lead to spontaneous vascular malformations. Using injection of viral vectors, we show that CCM lesion formation not only requires loss of CCM genes but also active angiogenesis. Furthermore, VEGFR2 activation is increased following the depletion of KRIT1, CCM2 or CCM3 gene expression in endothelial cells (although increased VEGFa production is only seen with the loss of KRIT1). Subsequently, we show that inhibition of VEGF with the kinase inhibitor SU5416 reduces CCM lesion formation and vascular permeability in Krit1ecKO mice, with a decreased likelihood of lesion hemorrhage as shown by peri‐lesion iron deposits. This mechanism is not specific to KRIT1, as inhibition of VEGF using bevacizumab also reduces lesion formation in Ccm3ecKO mice.Support or Funding InformationThis work was supported by NIH grant HL117885 to A.J.G, NIH grant NS075168 to K.J.W., and AHA pre‐doctoral fellowship 14PRE2048009 to P.V.D.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Creating a Unique Graduate Training Program for Educator Scholars in the Anatomical Sciences
Authors: Notebaert Andrew 
Published: June 2021

Healthcare education has made large leaps to catch up to new educational theories and best practices. Teaching roles in healthcare have traditionally fell to basic science researchers and clinicians with content area expertise but possibly little educational training. Recently, there has been a push to educate future teachers through graduate programs, most notably in anatomical education. While most of these education track PhD programs devote a large amount of training in the anatomical sciences and pedagogy skills, other educational facets are taught sparingly. Using other graduate programs in both science education and anatomy education as curricular models, the University of Mississippi Medical Center has restructured its PhD in Clinical Anatomy which itself was a restructuring of a traditional PhD in Anatomy. The original Clinical Anatomy PhD had students taking the medical anatomy courses (gross anatomy, histology, developmental, and neuroscience) and supplementing with four total credit hours of education training over three courses. Additionally, students would serve as a teaching assistant for each of the medical anatomy classes, earning additional credit hours before progressing to the dissertation phase of the program. The new Clinical Anatomy curriculum was designed to focus on three equal aspects of training; anatomical content, educational content, and educational research. The program gives equal focus to each of these important aspects as well as the integration of these areas with each other to train educator scholars capable of contributing to the education and research mission of healthcare educational programs. The anatomical content courses were restructured to focus on a foundational level of graduate anatomy content rather than limiting the training to only medical anatomy. The educational courses were expanded and focus on three areas; learning theory from both an educational and neuroscience perspective, pedagogical theory and methods, and curricular development and design. The research experiences were restructured so that the students get early exposure to research projects and statistical analysis, leading into abstract and manuscript work much earlier in their training that in previous renditions of the program. However, as with any graduate program, the graduates have to be able to fill positions for which they were trained upon graduation. Based on a recent pilot analysis of job openings seeking teachers of the various anatomy disciplines, there are a variety of open anatomy education vacancies but it is unknown if these vacancies would value students trained under these new graduate programs over the traditionally trained anatomists and basic science researchers.Support or Funding InformationThis project was not fundedThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Integration, Evolvability, and Constraint within the Primate Functional Shoulder Complex
Authors: Agosto Elizabeth R. , Auerbach Benjamin M. 
Published: June 2021

In vertebrates, especially primates, the morphology of the scapula is closely associated with locomotion, and individual traits of the shoulder are hypothesized to be under directional selection to meet these behavioral demands. By ascribing individual traits to variation in locomotor behaviors, however, researchers implicitly assume traits evolve independently. Traits that share mechanical function may covary genetically, and could therefore evolve through correlated responses to selection. The scapula, for example, is part of a complex of functionally related traits, linked both developmentally and mechanically, which includes dimensions of the clavicle, humerus, and basicranium. This study investigates whether the functional relationships among these skeletal regions relate to evolutionary covariance by comparing patterns of integration and evolvability among homologous traits in humans and tamarins.Three‐dimensional landmarks were taken from the scapula, humerus, clavicle and basicranium of 109 humans (Homo sapiens) and 100 tamarins (Saguinus sp.). Twelve interlandmark distances were calculated from these landmarks for analysis, representing a complex of traits relating to mechanical function from both the basicranium and the shoulder girdle. Analyses aimed at examining evolutionary covariance were applied using both raw and data scaled by the geometric mean. The strength of integration was quantified using the analysis of eigenvalues and a measure of trait variational autonomy. Evolvability was estimated by applying 1000 simulated random selection gradients to the phenotypic variance/covariance matrices and comparing evolutionary responses among taxa. Statistical significance of differences among taxa was determined by recomputing each value of integration and evolvability for 1000 iterations of the bootstrapped sample. A t‐test was used for pairwise comparisons of the bootstrapped values among taxa (α = 0.05).Results indicate when traits from the basicranium and shoulder girdle are evaluated as a complex, the ability of these regions to respond to directional selection is decreased compared to their responses were they independent. Evolutionary independence among these traits is not supported, and analyses instead indicate evolutionary constraint among these regions. Morphological differences in the nuchal region and shoulder complex between humans and tamarins further reflect differences in the ability to respond to directional selection in these two genera, which may have had functional implications for evolution of the shoulder in apes and monkeys. Namely, the traits in humans are more tightly integrated, resulting in weaker evolvability.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Warburg Effect and Lactate Signaling Augment Fgf Signaling to Promote Sensory‐neural Development in the Otic Vesicle
Authors: Riley Bruce , Kantarci Husniye 
Published: June 2021

In a forward ENU screen in zebrafish, we recovered a novel mutation that specifically impairs development of sensory hair cells and neurons in the otic vesicle. Whole‐genome sequencing identified a null mutation in the gene encoding the glycolytic enzyme Phosphoglycerate Kinase‐1 (Pgk1). Targeted disruption of Pgk1 using CRISPR/Cas9 yields an identical phenotype. Surprisingly, analysis of genetic mosaics showed that Pgk1 acts non‐autonomously to promote efficient Fgf signaling at a distance, as required for specification of neurons and hair cells of the inner ear. Expression of known Fgf ligands appears normal but Fgf‐reporters Pea3 and Erm are dramatically under‐expressed in mutant embryos. Moreover, transient overexpression of Fgf8 from an inducible transgene does not rescue the phenotype, suggesting a problem with signal transduction. These and other data led us to the discovery that otic cells require metabolic reprogramming to enhance glycolysis and “aerobic fermentation” – similar to the “Warburg Effect” exhibited by tumor cells – to promote synthesis and secretion of lactate. Wild‐type embryos treated with drugs that block synthesis or secretion of lactate mimic the phenotype of Pgk1 mutants, whereas Pgk1 mutants are fully rescued by treatment with exogenous lactate. Lactate treatment of wild‐type embryos elevates expression of Pea3 and Erm even when Fgf signaling is blocked. Thus, by raising steady‐state levels of Pea3 and Erm (critical effectors of the Fgf‐MAPK pathway), lactate signaling primes the pathway and renders cells more responsive to dynamic changes in Fgf signaling required for sensory‐neural development in the inner ear.Support or Funding InformationNIH‐NIDCD grant R01‐DC03806This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
SMRT‐Cappable‐seq reveals complex operon variants in bacteria
Authors: Ettwiller Laurence , Yan Bo 
Published: June 2021

Gene expression in bacteria is organized into operons, functional units of genomic DNA containing clusters of genes under the control of single promoters. The definition of an operon relies on the precise identification of the start of transcription (TSS), the end of transcription and (TTS) and the relation between the TSS and the TTS for each transcript molecule. Yet current experimental methods for sequencing RNA in bacteria rely on the necessary fragmentation of transcripts for short read sequencing. Consequently the larger context by which genes are expressed and the phasing between the start and the end of primary transcripts are lost. As a result, and despite overwhelming mass of transcriptome information, accurate operon structure have so far been solved only for a handful of cases. Here we describe SMRT‐cappable‐seq as the first experimental methodology to identify operons genome‐wide in prokaryotes. It combines the isolation of full‐length primary transcripts with long read SMRT sequencing. Applied to E.coli, SMRT‐cappable‐seq identifies a total of 2300 operons from which around 900 are novel. Importantly, our result reveals a pervasive read‐through of previous experimentally validated transcription termination sites. Termination read‐through represents a powerful strategy to control gene expression and redefines operons with thousand of new operons containing more genes than previously thought. Taken together this data provides a first glance at the complexity of the ‘operome’ in bacteria and presents an invaluable resource for understanding gene regulation and function in bacteria.Support or Funding InformationNew England Biolabs Inc. and Pacific Biosciences of California, Inc supported this research.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Describing the molecular mechanism by which the A2 protein improves survival in mice with endotoxemia.
Authors: Martinez Marina , Da Qi , Fisipe Titilope A. , Cruz Miguel A. 
Published: June 2021

Von Willebrand factor (VWF) is an essential protein that mediates the platelet adhesion to the exposed subendothelium surface and significantly contributes to arterial thrombosis and stroke. Secreted VWF from the stimulated endothelium as long strings remain anchored to the surface of the endothelial cells and interact with circulating platelets, leading to vessel occlusion. We have shown that the isolated A2 domain of VWF (A2 protein) contains bind activity for the A1 domain, vimentin, and fibrin(ogen). In fact, as a result of those interactions, the A2 protein impairs platelet adhesion to VWF and fibrin(ogen), formation of VWF strings on the endothelium, and fibrin polymerization. The A2 protein was effective in preventing fibrin‐rich microvascular thrombosis and improving survival (100%) of mice with endotoxemia (LPS)‐induced disseminated intravascular coagulation (DIC). Thus, a goal was to identify the binding sites of A1 domain, vimentin, and fibrin in the A2 protein and dissect the mode of action by which the A2 protein recuperates the sick animals. To determine the amino acid residues that are important in the recognition for vimentin, A1 and fibrin, I used the crystal structure of the A2 domain as a guide for point mutations. Two mutations, E1523A and E1567A were introduced in the A2 protein. The purified mutants were tested for their binding capacity for the ligands using ELISA. Moreover, the A2 mutants were tested in mice with endotoxemia (LPS). We are now demonstrating that E1523A mutation had a reduced binding affinity for Vimentin and A1 domain. Strikingly, E1567A and E1523A mutations significantly abolished the binding to fibrin(ogen). Importantly, both mutations failed to replicate the beneficial effect of the wild type A2 protein in improving survival of mice with LPS‐induced DIC. Together, these findings suggest that the A2 protein impairs microvascular thrombosis in mice with endotoxemia mainly via its interaction with fibrin.Support or Funding InformationThis research was supported by funding provided by the following NIH Diversity Supplement: R01GM112806‐0351.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The metabolomic signature of Giardia sp.
Authors: Allain Thibault , Groves Ryan , Lewis Ian A. , Buret Andre G. 
Published: June 2021

BACKGROUNDGiardia duodenalis is a protozoan parasite responsible for giardiasis, a parasitic disease characterized by intestinal malabsorption, diarrhea, weight loss and abdominal pain. The pathophysiological processes occurring during giardiasis involve epithelial abnormalities, mucus depletion and microbiota disruption, however the mechanisms are poorly understood. Upon infection, Giardia, which has a minimal biosynthetic capacity, compete locally with the commensal microbiome for nutrients and ecological niches in the duodenum.AIMIn this context, we performed a metabolomic study to characterize the nutritional requirements and the secretome of several Giardia strains, isolated from human patients (assemblages A and B), outbreaks, as well as a variety of mammals.METHODSThe metabolomic profiles of Giardia sp. isolates were determined using mass spectrometry. Trophozoites were grown to confluence in MTYI media and supernatants were collected at different time points (3, 6, 9, 12 and 24 hours) and mixed with ice cold methanol. Methanolic extracts were analyzed by Ultra High Performance Liquid Chromatography mass spectrometry and resolved by Hydrophilic interaction liquid chromatography column.RESULTSWe identified a multitude of produced and consumed compounds such as amino acids, nucleic acid precursors, protein catabolism precursors, as well as carbohydrates and energy metabolites. Interestingly, nutrition requirements of Giardia trophozoites are significantly different between isolates, both intra and inter‐assemblages. For instance, isolate WB6 (assemblage A) is more auxotrophic for arginine than GSM strain (assemblage B), while GSM strain requires more inosine, uridine and as paragine than other isolates.CONCLUSIONThis study gives insight into how Giardia survives the host's intestinal tract and shows strain specific metabolomic signatures. A metabolomic approach may further help to understand the impact of nutritional modulation by Giardia trophozoites on the gut microbiome and host physiology.Support or Funding InformationGrant support: University of Calgary Eyes High postdoctoral fellowshipThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Shp2/Ptpn11 Deletion Suppresses Liver Tumorigenesis Driven by MET, β‐Catenin and PIK3CA
Authors: LIU Jacey 
Published: June 2021

Most recent experimental data demonstrated a liver tumor‐suppressing effect for Shp2/Ptpn11, as ablating Shp2 in hepatocytes triggered spontaneous development of hepatocellular adenoma (HCA) in aged mice and also aggravated hepatocellular carcinoma (HCC) induced by chemical carcinogen or Pten deletion. Herein we report surprisingly that despite the induction of hepatic injuries, inflammation, and fibrosis, Shp2 deletion in hepatocytes suppressed liver tumorigenesis driven by overexpression of oncogenic proteins MET and ΔN90‐β‐catenin (MET/CAT), or MET and PIK3CAH1047R (MET/PIK). Shp2 loss inhibited proliferative signaling from MET, Wnt/β‐catenin, Erk and PI3K/Akt pathways, but induced cell senescence following exogenous expression of the oncogenes. These results clearly distinguish a requirement for Shp2 in mediating and amplifying the oncogenic signals of these oncoproteins from a tumor‐promoting hepatic microenvironment resulting from Shp2 deficiency. Based on these data, we propose a new molecular targeted therapy by inhibiting Shp2 for HCCs driven by oncogenic signals emanating from RTKs and other upstream molecules. However, to achieve optimal and lasting therapeutic effect, it is necessary to suppress the tumor‐enhancing environmental factors produced secondary to Shp2 inhibition. This simultaneous targeting approach may be developed into a widely used clinical recipe, as long as the cell‐intrinsic drivers and the stromal factors are identified for a given liver cancer patient in the precision medicine era.Support or Funding InformationThis project was supported by NIHR01CA176012 and R01CA188506 (to Dr. Gen‐Sheng Feng).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Upregulation of Stromal‐derived Factor‐1 by Hypoxia Requires Hypoxia‐Inducible Factor‐1α and Transforming Growth Factor‐1
Authors: Strickland Jenna D. , Garrison Dominique , Copple Bryan 
Published: June 2021

Following both acute and chronic liver injury, expression of the chemokine stromal‐derived factor‐1 (SDF‐1), or CxCl12, is upregulated and plays a vital role in the activation of down‐stream signaling pathways implicated in either driving hepatic injury progression or leading to its repair. For example, while SDF‐1 has been implicated in promoting tumor growth and metastasis in patients with hepatocellular carcinoma, in acute injuries it may play an important role in liver regeneration. This suggests SDF‐1 may represent a novel target for therapeutic intervention in both acute and chronic liver diseases. However, the mechanism by which SDF‐1 is upregulated following hepatic injury is not fully known. The present study determined whether the transcription factor, hypoxia‐inducible factor‐1α (HIF‐1α), regulates SDF‐1 in primary mouse hepatocytes. To activate HIF‐1α in vitro, primary hepatocytes were incubated for 72 hrs in either room air or hypoxic (1% O2) conditions. Under hypoxic conditions, SDF‐1 mRNA levels were upregulated after 72 hr in WT mouse hepatocytes, an effect that was prevented in hepatocytes isolated from either HIF‐1α or HIF‐1β knockout mice. We demonstrated previously that upregulation of several genes in hepatocytes during hypoxia requires autocrine release and activation of transforming growth factor‐1 (TGF‐β1). To determine if TGF‐β1 is required for the upregulation of SDF‐1 during hypoxia, primary mouse hepatocytes were pretreated with a TGF‐β1 receptor antagonist (SB‐431542) and placed in hypoxic conditions for 72 hrs. Pretreatment with SB‐431542 completely inhibited upregulation of SDF‐1 by hypoxia, suggesting TGF‐β is required for upregulation of SDF‐1 in hypoxic hepatocytes. Additionally, while treatment of hepatocytes with TGF‐β1 upregulated SDF‐1, this effect did not require HIF‐1α, suggesting that TGF‐β1 is downstream of HIF‐1α activation. We previously demonstrated that hypoxia upregulates thrombospondin‐1 in hepatocytes, which could be responsible for activation of latent TGF‐β1. Upregulation of SDF‐1 by hypoxia, however was not different between hepatocytes isolated from wild‐type or thrombospondin‐1 knockout mice. The results indicate that hypoxia activates HIF‐1α in hepatocytes, which leads to activation of latent‐TGF‐β1. TGF‐β1 then acts in an autocrine fashion to upregulate SDF‐1. However, the mediator responsible for converting latent TGF‐β to its active form remains unknown.Support or Funding InformationNIH grant 2 R01 DK073566NIH Training Grant T32 ES007255This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.