bluuebunny/crossref_metadata_2025_split huggingface dataset:
Microarray analysis of the PVN of spontaneously hypertensive wistar rats (SHR)
Authors: O'Brien Fiona , Feetham Claire , Barrett‐Jolley Richard 
Published: June 2021

The paraventricular nucleus (PVN) of the hypothalamus is a key autonomic regulation area in the brain known to influence sympathetic nerve activity. Identified neurons project from the PVN to the spinal cord and can modulate cardiac function (Coote JH 1995). The PVN has been linked to cardiovascular diseases such as hypertension, where an elevation in sympathetic nerve activity is observed (Guyenet PG 2006 and Zhang K et al. 2002). For example, the activity of ‘spinally projecting’ PVN neurons is altered in models of heart failure (Patel KP 2000) and lesioning of the PVN prevents the development of spontaneous hypertension in SHRs (Takeda K et al. 1991).In this study, we performed transcriptome analysis of tissue from the PVN of 4 SHR and 4 normal Wistar Kyoto rats. In brief, tissue was isolated by 2mm diameter tissue punch from the dorsal PVN, homogenised and RNA extracted with Qiagen RNAeasy minikits in accordance with the manufactures instructions. The highest quality scoring 3 normal and 3 SHR were then analysed with an Illumina Beadchip microarray. Data was analysed in MatLab. Log2 normalised expression sets were created and genes with low absolute expression values or variance were filtered out (lowest 10%) (Kohane, I. et al. 2003). Significant differential transcription was then identified with an all permutation (t‐test). FDR and correct “q‐values” were created following the methods of Storey et al. 2003. Significantly increased/decreased genes were subjected to biological function enrichment analysis using PANTHER.Both PCA (PC1 vs PC2) and cluster analysis showed complete separation of the SHR and normotensive populations and 1592 genes in total were differentially expressed by more than 1.3 fold (and q‐value<0.05). Of particular interest to autonomic control of cardiovascular function was the enriched clusters of genes associated with circulatory control (AVP, angiotensin and Substance P, and the α‐1B adrenergic receptor), however, some genes associated with “inflammaging” were also differentially expressed such as superoxide dismutase 1 enzyme (SOD) and Il1b.Our results illustrate that there are distinct PVN‐specific changes in gene expression that occur in spontaneously hypertensive rats. In particular, we show that genes important in circulatory control and fear/stress responses are enriched in the PVN, further supporting a role for the PVN in cardiovascular diseases. Further experiments are needed to verify the functionality of these changes in gene expression, such as brain‐slice electrophysiology and whole‐animal experiments. The results of these experiments will be crucial in understanding the role of PVN in cardiovascular diseases such as hypertension.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cellular Senescence in Primary Muscle Fibroblasts is Associated with Changes in Extracellular Matrix Remodeling
Authors: Judge Jennifer L , Brooks Susan V 
Published: June 2021

BackgroundAdvanced aging leads increased stiffness of the skeletal muscle extracellular matrix (ECM), which is associated with declines in skeletal muscle function and mass. Previous work by our group has shown increased stiffness of the ECM is associated with changes in ECM modification (hydroxyproline content, advanced glycation end products), without changes in total ECM area in aged mouse muscle tissue (Wood et al., 2014). Here we investigated whether changes in ECM modification with age are the result of cellular senescence in muscle fibroblasts, which play an active role in ECM production and modification. We hypothesize that muscle fibroblasts become senescent with advanced age, and that cellular senescence is associated with altered ECM modification that could lead to stiffness.MethodsHindlimb muscles from 3–4 month old C57Bl/6 mice were digested in collagenase and primary fibroblasts were cultured and used between passages 3–5. Cells were confirmed to fibrogenic by expression of Tcf4 and absence of MyoD. Cellular senescence was induced using 200 μM hydrogen peroxide (H2O2) for 2 hours. RNA was collected after 72 hours to assess cellular senescence, and changes in collagen and ECM modification transcripts by quantitative PCR. Cellular senescence was evaluated by senescence associated beta‐galactosidase staining.ResultsH2O2 resulted in positive beta‐galactosidase staining, suggesting cellular senescence. H2O2 also increased mRNA transcripts associated with cellular senescence: p16 and p21. Fibrillar collagen 1α1 and 3α1 transcripts were unchanged, while non‐fibrillar collagen 4α1 was increased with H2O2. Prolyl 4 hydroxylase, an important enzyme in collagen fiber synthesis, was increased, as was tissue inhibitor of metalloproteinases (TIMP) 1 and 2. Matrix metalloproteinase 2 levels were unchanged.ConclusionsInduced cellular senescence with H2O2 in muscle fibroblasts demonstrated changes in mRNA transcripts that promote collagen synthesis (prolyl 4 hydroxylase) and inhibit matrix degradation (TIMP 1 and 2) without changing de novo fibrillar collagen transcripts. These results suggest that muscle fibroblast senescence may be a cellular mechanism that leads to increased stiffness in ECM through changes in matrix modification and processing.Support or Funding InformationThis work was supported by K12GM111725 and AG051442.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Activation of Growth Signaling in Skeletal Muscle by Growth Arrest Specific‐6
Authors: Wyson Brenna M. , Matsumura Marc S. , Deyle Michael R. , Knowlton M. Nekel , Arroyo Juan A. , Hyldahl Robert D. , Thomson David M. 
Published: June 2021

Growth‐arrest specific protein 6 (Gas6) is believed to be involved in activation of cell proliferation and growth of many tissues through its activation of the Akt and MAPK signaling pathways downstream of its primary tyrosine kinase receptor, Axl. Currently the role of Gas6 in skeletal muscle has not been elucidated. The purpose of this study was to determine whether skeletal muscle is responsive to Gas6 stimulation. To do this C2C12 myoblasts were incubated in 100 ng/ml of recombinant mouse Gas6 for 15–180 minutes. Gas6 treatment increased Akt phosphorylation, which peaked at 30–60 minutes. We identified Axl expression via immunofluorescence in rat tibialis anterior muscles. Next, soleus (SOL) and plantaris (PLT) muscle strips from 10–12 week old Lewis rats were incubated with or without 200 ng/ml of Gas6 for 30 minutes. Gas6 failed to increase Akt or Erk phosphorylation in slow‐twitch SOL muscles, but increased Erk phosphorylation (but not Akt) in fast‐twitch PLT muscles. We conclude that Gas6 activates Akt or Erk signaling in myoblasts and mature muscle respectively. This suggests that Gas6 may potentially regulate regeneration and/or growth as well as many other cellular processes in skeletal muscle.Support or Funding InformationBYU Gerontology Program GrantThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Differential Diagnosis of pancreatic ductal adenocarcinoma based on differential expression of select miRNA in pancreatic tissue and plasma
Authors: Figueiredo Rafael Zimak , Vieira Nivaldo Farias , Cirino Mucio Luiz Assis , Neto Fermino Sanches Lizarte , Gaspar Alberto Facury , da Cunha Tirapelli Daniela Pretti , Sankarankutty Ajith Kumar , Kemp Rafael , dos Santos José Sebastião 
Published: June 2021

The differential diagnosis, treatment and prognosis of pancreatic ductal adenocarcinoma (PA) are still a challenge in clinical practice. The aim of the current study was to assess the role of select plasma and tissue microRNAs (miRs‐21, ‐23a, ‐100, ‐107, ‐181c e ‐210) as biomarkers for the diagnosis of PDCA. Samples of plasma (PAp Group, n=13), pancreatic tumors (PAt Group, n=18) and peritumoral regions (PPT Group, n=9) were collected from patients during the surgical procedure. The control group consisted of samples collected from patients submitted to pancreatic surgery for trauma or from cadaveric organ donors (PC Group, n=7). Healthy volunteers donated blood for plasma collection (PCp Group, n=6). The relative abundance of six miRNAs was measured in all groups using real‐time PCR, and serum CA 19‐9 levels were determined in Groups PA and PC, with the difference being considered significant when p <0.05. In tissue samples, there was a difference in the expression of miRNA‐21 (p=0.005) and miRNA‐210 (p=0.008) across the PAt, PC and PPT groups; miRNA‐21 distinguished group PAt from PC and PPT and miRNA‐210 distinguished group PAt from PC. Serum CA 19‐9 showed 88% accuracy (ROC‐AUC 0.95; 0.84 – 1), 83% sensitivity, and 100% specificity at a cut‐off of 37 ng/dl. The PAp group showed overexpression of miR‐181c (95% CI1.67 – 17.68; p<0.00001) and miR‐210 (95% CI0.16 – 5,20; p=0.03) compared to the PCp group. The combination of tissue expression of miR‐21, ‐210 and serum CA 19‐9 showed 100% accuracy for the diagnosis of PA, and the accuracy of plasma miR‐181c (PAp × PCp) was also 100%. These findings suggest that the combination of tissue miRNAs and serum CA 19‐9 can improve accuracy, but plasma miRNA expression can be used as a noninvasive diagnostic test for PA, although further studies are necessary to validate these results as well as to evaluate the utility of miRNA expression as a biomarker related to survival and to the response to therapy in PA.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Quantifying the Genotype to Phenotype Map in Developing Mice
Authors: Richbourg Heather A , Young Nathan , Takenaka Risa , Marcucio Ralph 
Published: June 2021

ObjectiveQuantify phenotypic variance of face shape within mice after ablation of the gene encoding the E3 ubiquitin ligase, nitric oxide synthase‐interacting protein (Nosip).MethodsMouse embryos from a nitric oxide synthase‐interacting protein (NOSIP) knock‐out model were collected at e12.5 via cesarean section (n=39). Two‐dimensional images were taken of the head, and DNA was collected from the limb bud for genotyping via PCR. Additionally, skulls were collected post‐natally (23 – 263 days of age (n=25)), and were scanned via microCT (55kV, 0.2 second integration time, 145uA). Anatomical landmarks were manually placed along the skull, creating three‐dimensional coordinates. Data were corrected for rotation and scaling differences through a Procrustes transformation using MorphoJ. Standard geometric morphometric analyses were subsequently performed, and data were compared using principal components analyses (PCA).ResultsAll wildtype (WT) embryos appeared to have normal phenotypes (n=8), but there was a large phenotypic variation in mutants (Fig. 1), ranging from normal (n=19) to severe forebrain malformations (n=7) or resorption (n=6). Adult HET mice (n=9) also showed phenotypic variation, including lateral malocclusion (n=1), opaque eyes (n=4), or no eyes present (n=1). After correcting for skull size, there was minimal variation within the WT adults, but there was specific separation between HETs with normal phenotypes and abnormal phenotypes. Animals were separated by PC1 (left‐side of snout width asymmetry; 24.5% of total variation) and PC2 (snout length; 16.3% of total variation) (Fig. 2). Along the PC1 axis, three HETs are clustered within the WTs; however, PC2 separates the two HETs with an abnormal phenotype (no eyes and lateral malocclusion), while the remaining HET within the WT cluster has a normal phenotype. Visualizing the same PCA relative to sex does not distinguish unique clusters, suggesting that phenotype is not related to sex.ConclusionsThis model presents a unique opportunity to understand and study phenotypic variation within a HET genotype, as well as when comparing HETs to WTs, producing variation in both embryos and adults. The malformations present in the HET embryos resembles a sonic‐hedgehog loss‐of‐function phenotype, so more work will be performed to determine the affected signaling pathways and cellular processes.Support or Funding InformationTravel support was given by the American Association of Anatomists. This project is supported by NIH funding (R01DE01823, R01DE01963,R01DE021708).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Spa47 Oligomerization Plays a Key Role in ATPase Activation and Function in the Shigella Type III Secretion System
Authors: Burgess Jamie Lee , Burgess Robert Alan , Dickenson Nicholas 
Published: June 2021

Several Gram‐negative pathogens, including Shigella, utilize a conserved type three secretion system (T3SS) for infection. The type three secretion apparatus (T3SA) provides a unidirectional conduit for injection of effector proteins from the bacteria to the host cell cytoplasm. The conserved T3SS ATPase found at the base of the T3SA is believed to provide the energy for apparatus formation and effector secretion, however, the mechanisms of ATP hydrolysis, effector secretion, and apparatus formation by the T3SS ATPase remain unclear. We have recently shown that Spa47, the T3SS ATPase of Shigella, forms a unique activated oligomeric species during the course of purification. Using our recently published crystal structure of Spa47 as a guide, we modeled an activated Spa47 oligomer, finding that ATP hydrolysis may be supported by specific side chain contributions from adjacent protomers within the complex. To explore the dynamics of Spa47 protomer interaction in vitro and in vivo, we performed a series of ATP hydrolysis, fluorescence microscopy and Shigella invasion experiments. We observe a positive Hill coefficient with the monomeric Spa47 species that is not present with the oligomeric Spa47 species in vitro. Interestingly, we continue to see a positive Hill coefficient even with the addition of engineered ATPase inactive Spa47 point mutants to WT Spa47 monomer species. We have utilized live cell fluorescence microscopy to show that these ATPase inactive Spa47 point mutants localize to the T3SA though they do not support secretion. Furthermore, each of the ATPase inactive Spa47 point mutants as well as an oligomerization deficient Spa47 construct exhibits a dominant negative phenotype. Together, these results suggest that while ATP hydrolysis is not the major driving factor for Spa47 localization to the apparatus base, the Spa47 oligomer may undergo a series of concerted ATP hydrolysis events that are critical for apparatus function, similar to ATP synthase. As we better understand the nuances of T3SS ATPase mechanism and function, the results of this work will provide a strong platform for future work to perhaps target the T3SS ATPase for novel therapeutics.Support or Funding InformationNational Science Foundation Graduate Research Fellowship and National Institute of Health (1R15AI124108‐01A1)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Homarus Gemmarus: Seriated CT‐Scan and Re‐Constructions: Horizontal, Sagittal and 3D
Authors: le floch‐prigent patrice pierre , Verdeille stéphane 
Published: June 2021

The european lobster is a large, common, marine, crustacean decapod.MaterialAn entire animal was acquired still alive and deeply frozen in extension of the abdomen. It was 26,5 cm long; 7 cm width at its maximal dimension of the cephalo‐thorax. The CT‐scan apparatus was a Siemens somatom sensitive, whole body apparatus with 128 stellar detectors.MethodsThe entire animal was CT‐scanned all along the rostro‐caudal axis, in axial cross‐sections and reconstructed in the two other planes (horizontal and para‐sagittal) and in 3‐ dimensional ones. The frozen lobster was laid in prone position on the radiological table. It was CT‐scanned with an acquisition (100 kV, 18 mAs) in the axial plane: the axial cross‐sections (every 0,8 mm, 1 mm thick) were joined, and frontal. Two different penetrations were used: B30 for the soft parts and B70 for the hard parts (cartilage and extra‐skeleton).Results500 frontal cross‐sections were obtained from the native axial acquisition. 120 para‐sagittal cross‐sections were reconstructed from the preceding ones, and 80 horizontal ones as well. 11, 3D reconstructions were realized with several incidences: superior, inferior, rostral, caudal, postero‐superior, antero‐inferior, antero‐lateral and superior (right and left), lateral (right and left), median sagittal.ConclusionThe seriated CT‐scan and the reconstructions afforded very numerous morphological elements in Homarus gammarus, and also in the study of their relationships. The muscular masses presented very few differenciation. The rare cartilaginous structures were well vizualized. A voluminous aerial poach at the upper and rostral part of the cephalo‐thorax corresponded to the two parts of the stomach: cardial (rostral) and pyloric (caudal) ones. The reconstructions were very contributive.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Prionace glauca (Pole shark): seriated CT‐scan and reconstructions: horizontal, sagittal and 3D
Authors: Le Floch‐Prigent Patrice Pierre , Verdeille Stéphane 
Published: June 2021

The pole shark (Prionace glauca, Linnaeus 1758) is a common, pelagian condrichtyan.MaterialThe entire, rostral extremity (28,4cm long) of an adult specimen was CT‐scanned all along the rostro‐caudal axis, in axial cross‐sections and reconstructed in the two other planes and in 3‐dimensional ones. The CT‐scan apparatus was a Siemens somatom, whole‐body apparatus with 64 detectors.MethodsThe frozen shark's head was laid in prone position on the radiological table. It was CT‐scanned with an acquisition (100kV, 18mAs) in the axial plane. The frontal cross‐sections (every 0,8 mm, 1 m thick) were joined. Two different penetrations were used: B30 for the soft parts and B70 for the hard parts (cartilage and extra‐skeleton).Results355 frontal cross‐sections were obtained from the native, axial acquisition. 100 para‐sagittal cross‐sections were reconstructed from the preceeding ones and 100 horizontal ones as well. 16, 3D reconstructions were realized with several, different incidences and penetrations.ConclusionThe seriated CT‐scan and the reconstructions gave numerous morphological elements in Prionace glauca, as well as in the study of their relationships. The muscular masses presented very few differenciation. The cartilaginous structures were well vizualized, as the eye and the central nervous system, the teeth were particularly easy to study, on the reconstructions, noticeably the successive raws in the inferior jaw.We thanks the “Le Venec” Company for the delivery of the animal material and Mr Lionel Feuillassier (Marinarium, Muséum National d'Histoire Naturelle), both in the town of Concarneau (Brittany, France).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 Influence of Warm Ischemic Time on the Viability of Deceased Organ Donor Derived Bone Marrow
Authors: Sherry Aubrey M. , Johnstone Brian H. , LaFontaine Michael , Woods Erik J. 
Published: June 2021

It was recognized 60 years ago that deceased donor bone marrow (BM) could be a useful source of viable hematopoietic stem cells (HSCs) for transplantation. Validated procedures have since been published for the recovery of viable deceased donor HSCs and their safe use in immune tolerance clinical trials. It is the goal of Ossium Health to streamline the process of obtaining deceased donor derived BM on an industrial scale. It has been shown by previous studies that BM is tolerant to a wide range of cold and warm ischemic times during collection. CD34+ HSCs prefer a hypoxic environment and remain quiescent in vivo. This observation indicates that BM procured during routine organ and tissue recovery may remain viable for extended periods following cardiac arrest and, thus, will be suitable for transplantation. Ossium is evaluating the range of ischemia times to determine if organ and tissue donor bone marrow recovered using a standardized approach is suitable for transplantation.Vertebral bodies (VBs) and ilia of 34 deceased organ donors were recovered by Ossium's network of organ procurement organizations (OPOs) over warm ischemia times ranging from 1:03–22:03 (hr:min). Recovered VBs were then cooled and shipped overnight on wet ice to Ossium Health, Indianapolis, IN, for BM recovery using grinding and elution methods. The CD34+ HSC population was enumerated using flow cytometric ISHAGE protocols and a mean viability over the entire ischemia range was determined to be 79.3 ± 3.7% (mean ± SEM) as determined by 7‐Aminoactinomycin D (7‐AAD).While gross membrane viability is a useful measure, engraftment potential correlates strongly with functional viability as measured through colony forming unit (CFU) assay, and engraftment of transplanted units requires the presence of Granulocyte/Macrophage (GM) colonies specifically. To confirm that the deceased donor marrow would be suitable for engraftment, the CFU assay was performed and CFU‐GM were scored. Over the entire range of ischemic times, an average of 37.05 ±11.16 CFU‐GM (mean ± SEM) per 105 nucleated cells plated was observed.Rapid neutrophil and platelet engraftment in living donor allogeneic grafts has previously been correlated with a dose containing CFU‐GM of 5.6 per 105 TNC in for an average patient of 70kg. The results of this preliminary study indicate deceased donor marrow can easily meet this requirement. These data demonstrate the potential to recover viable and functional HSCs after extended periods of warm ischemia; thus, establishing the feasibility and utility of banking cells from deceased organ and tissue donors.Support or Funding InformationThis work was funded by a Grant from the NIH‐NIAID (1R43AI129444‐01A1, PI: Woods, EJ), and Ossium Health, Inc.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Tumor suppressor protein PALB2 has strand exchange activity independent of recombinase Rad51
Authors: Deveryshetty Jaigeeth , Kanikkannan Lakshmi , Korolev Sergey , Doisy E. A. 
Published: June 2021

PALB2 (Partner and Localizer of BRCA2) is recombination mediator protein (RMP) involved in homology dependent repair (HDR) of double strand breaks (DSB). PALB2 works synergistically with BRCA2 by promoting Rad51 recombinase filament formation on single‐stranded (ss) DNA to initiate strand exchange essential for HDR. Numerous inherited PLAB2 mutations are reported to predispose to cancers such as breast and pancreatic cancers and rare genetic disease, Fanconi Anemia. However, the molecular mechanism PALB2 function is not well characterized. Here we report the DNA binding properties of PALB2 DNA‐binding domain (DBD), affinity and stoichiometry of interaction with various DNA substrates and identify residues involved in DNA binding. Further, we report an unanticipated activity of PALB2, where PALB2 promoted strand exchange in the absence of recombinase. This activity was enhanced in the presence of recombinase Rad51 and is abolished upon mutations of PALB2 DNA‐binding residues. Several properties also resemble those reported for Rad52. Our study demonstrates that PALB2 has multiple roles in homologous DNA repair.Support or Funding InformationSiteman Cancer centerThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.