bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of aging on oxygen permeability of wild type (WT) and AQP1‐RhAG double knockout (dKO) mouse red blood cells
Authors: Zhao Pan , Boron Walter F 
Published: June 2021

The dogma that all gases move through red blood cell (RBC) membranes by dissolving in and diffusing through the membrane lipid is no longer correct. Our preliminary data on mouse RBCs genetically deficient in both aquaporin‐1 (AQP1) and Rh complex (mainly RhAG) suggest that these two proteins account for ~60% of membrane O2 permeability. Using stopped‐flow (SF) analysis of hemoglobin (Hb) absorbance spectra, we now examine the effect of aging on the rate constant of Hb deoxygenation (kHbO2) of RBCs from WT mice and mice genetically deficient in both AQP1 and RhAG (double knockout, dKO). Studying mice at five ages (2~3 months, 6 months, 12 months, 18 months, and 24 months), we find that RBCs from WT mice at ages from 2~3 months through 12 months all have similar kHbO2 values (N = 6 mice/group). RBCs from WT mice at 18 (N = 6) and 24 months (N = 2) have mean kHbO2 values that—compared to RBCs from WT mice 2~3 months old—decrease by 8% at 18 months and by 10% at 24 months. These decreases in mean kHbO2 do not yet reach statistical significance, using a Holm‐Bonferroni correction. Regarding RBCs from dKO mice, we observe similar kHbO2 values for all ages from 2~3 months to 18 months (N=5 for 18 months), inclusive. RBCs from dKO mice at 24 months are currently under investigation. We plan to extend this study to RBCs from mice genetically deficient in just AQP1 or RhAG. If the trends continue (i.e., for RBCs from aging WT but not dKO to exhibit a fall in kHbO2), it would be consistent with the hypothesis that the non‐AQP1/RhAG component of RBC membrane O2 permeability is independent of aging, whereas the AQP1 and/or RhAG component declines with aging.Support or Funding InformationSupported by ONR N00014‐15‐1‐2060, ONR N00014‐16‐1‐2535 to WFBThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The right ventricle has more resident immune cells than the left ventricle
Authors: Gorr Matthew W , Sriram Krishna , Muthusamy Abinaya , Chinn Amy M , Insel Paul A 
Published: June 2021

The right ventricle (RV) pumps deoxygenated blood from the systemic circulation to the lungs for reoxygenation. Diseases such as pulmonary hypertension, pulmonary fibrosis, and chronic obstructive pulmonary disease are associated with hypoxia that leads to an increase in pulmonary vascular resistance (PVR), resulting in RV hypertrophy and dysfunction. The RV may differ from the left ventricle (LV) in the pathobiology in response to increases in afterload and no specific therapy exists for RV dysfunction. In order to gain insight into the biology of the RV compared to the LV, and perhaps uncover potentially novel therapeutic approaches for RV dysfunction, we performed RNA‐sequencing on RV and LV tissue from rats in normal ambient conditions or subjected to hypoxia (10% O2) for two weeks. Gene ontology and pathway analysis of the RV compared to the LV revealed multiple transcriptomic differences under ambient conditions and in particular, increased expression in the RV of genes related to immune function. Immune cell profiling by flow cytometry of rat cardiac digests, analyzed as the percentage of CD45+/CD11b/c+ cells in the cell population, revealed that in both ambient conditions and hypoxia, the RV had a significantly greater percentage of double‐positive cells compared to the LV (normoxia: average % ± SEM, 8.55 ± 1.71 RV vs. 2.04 ± 0.62 LV and hypoxia: 6.59 ± 2.71 vs. 1.55 ± 0.4; P <0.005, two‐way ANOVA, n=4 rats/group). Hypoxia had no significant effect on these values (P>0.05). These data reveal that the RV has a greater number of resident immune cells compared to the LV, possibly contributing to differences in pathobiology. Further investigation may reveal immune‐related therapeutic strategies for RV disease.Support or Funding InformationSupported by Department of Defense (W81XWH‐14‐1‐0372)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Modeling the Evolution of Crocodyliform Cranial Shape Through Time
Authors: Wilberg Eric , Turner Alan , Smaers Jeroen 
Published: June 2021

Crocodyliforms have a rich fossil record, attained global distribution, and filled a wide variety of ecological niches throughout their history. The group has explored a broad range of cranial morphologies through time, particularly in relation to rostral and supratemporal fossa (STF) form. These character complexes are functionally related, with the snout acting as the prey capture apparatus and the STF as the origin for the pseudotemporalis and/or adductor mandibulae externus profundus musculature. A correlation between STF size and snout shape has been previously suggested (e.g. taxa with long slender snouts tend to have larger STF, possibly related to the importance of the aforementioned muscles in their prey capture strategy). To investigate evolutionary patterns of these cranial modules we quantified snout and supratemporal fossa shape using sliding semilandmark‐based geometric morphometric methods. We then modeled the evolution of these modules across a time‐calibrated phylogeny of Crocodyliformes, testing for differences in allometric patterning (using new functions in the R package bayou and phylogenetic ANCOVA as implemented in evomap). Our results indicate a significant shift in the covariation of STF relative to snout length at the origin of Eusuchia. In comparison with non‐eusuchian crocodyliforms, eusuchians exhibit less change in STF size relative to snout length. This shift in phenotypic covariation is potentially related to a change in pattern (but not strength) of developmental integration between these modules.As these modules are related to feeding, this shift in phenotypic covariation may indicate an important transition in the evolution of the crocodyliform feeding apparatus at the origin of Eusuchia. That non‐eusuchian crocodyliforms tend to have a larger STF suggests a reliance on muscles occupying the dorsal temporal fenestra, while the relatively smaller STF of eusuchians may mark a shift to an increased role of the pterydoideus musculature responsible for the extreme bite forces of extant crocodylians. If such a model holds true, it may serve to best explain the observed shift in covariation between these modules. The timing of this transition in our analysis matches closely with a previous study modeling the evolution of bite force in the group based on a different osteological proxy. Collectively this suggests that the modern crocodylian feeding apparatus developed around the origin of the crown group, coincident with the establishment of the eusuchian secondary palate.Support or Funding InformationNational Science Foundation grant NSF‐DEB 1754596This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Quantifying Ubiquitination Signaling with a Chemical Proteomics Strategy
Authors: Li Yunan , Luo Ang , Erber Luke , Chen Yue 
Published: June 2021

Stoichiometric analysis of posttranslational modifications is an emerging strategy for absolute quantification of the modification's fractional abundance. In this study, we present a quantitative chemical proteomic workflow for stoichiometric analysis of ubiquitination, named Isotope‐BAlanced Quantification of Ubiquitination (IBAQ‐Ub). The strategy utilizes a new amine‐reactive chemical tag (AcGG‐NHS) for highly efficient labeling of amine residues. The new chemical tag is structurally homologue to the GG remnant of ubiquitin on modified lysine following trypsin cleavage and therefore enables the generation of structurally identical peptides from ubiquitinated and unmodified lysine residues following trypsin digestion and the secondary stable isotopic labeling. Quantitative analysis of labeled peptides with HPLC and high resolution mass spectrometry allows unbiased measurements of absolute site‐specific ubiquitination stoichiometry. Using recombinant protein standards and isotopic serial dilution, we showed that the strategy is highly robust, sensitive and accurate. We further designed a SILAC‐based experiment to demonstrate the efficient labeling of lysine with the new AcGG tag in complex cell lysates. Application of the IBAQ‐Ub workflow for the analysis of enriched histone fractions allowed us to measure the stoichiometry of histone H2B C‐terminal ubiquitination. Comparative analysis of stoichiometric dynamics upon proteasome inhibitor treatments showed a dramatic decrease of histone H2B ubiquitination levels and a significant increase of polyubiquitination K48 linkage. Thus, this work provides an efficient chemical proteomics tool for quantitative stoichiometric analysis of ubiquitination signaling pathways in cells and diseases.Support or Funding InformationUniversity of Minnesota start‐up fund and National Science Foundation (CHE‐1753154)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Role of WWOX and Zfra in limiting neurodegeneration
Authors: Chang Nan‐Shan , Lin Yu‐Hao 
Published: June 2021

Human and animal newborns with WWOX/Wwox gene deficiency suffer severe neural diseases, seizure, microcephaly, metabolic disorders, and early death. Neuronal heterotopia, which accounts for seizure, occurs in the newborns due to enhanced migration of cortical neurons to the epicortex. When tumor suppressor WWOX is Tyr33 phosphorylation, it strongly inhibits cancer growth in vitro and in vivo. When Tyr33 is de‐phosphorylated and Ser14 is phosphorylated, endogenous pS14‐WWOX is associated with enhanced growth of cancer stem cell spheres and solid tumors, and progression of neurodegeneration in patients with Alzheimer's disease (AD). We recently reported that a small zinc‐finger like Zfra peptide, e.g. Zfra1‐31 or 4–10, dramatically suppresses Ser14 phosphorylation in WWOX (>90% pS14‐WWOX), which leads to significant suppression of cancer growth and restoration of memory loss by blocking neurodegeneration in triple transgenic mice (3xTg) for AD. Also, when 3xTg mice received Zfra via tail vein injections at month 3, Zfra prevented age‐dependent loss of short‐term and long‐term memory in these mice. Zfra accelerates the degradation of aggregated proteins (e.g. amyloid beta) and activates a novel spleen non‐T/non‐B Hyal‐2+ CD3‐CD19‐lymphocyte, designated Z cells. Intringuingly, activated Z cells have a memory anticancer function, even though these cells have never gotten a chance encountering cancer cells or cancer cell antigens. Whether Z cells block neurodegeneration is unknown. Wwox heterozygous mice have an enhanced memory loss with age, compared to 3xTg mice. This is due, in part, to aggregation of a cascade of proteins, including initiators TRAPPC6AΔ, TIAF1, and SH2GLB2 (designated TTS) for causing amyloid beta and tau aggregation in the brain. In Wwox knockout mice, protein aggregates can be found in the cortex and hippocampus in 3 weeks after birth. In parallel, downregulation of WWOX protein in middle‐aged humans may lead to the TTS cascade of protein aggregation that allows formation of amyloid plaques and tau aggregation in old ages. Our study reveals an inhibitory role of WWOX and Zfra in suppressing neurodegeneration.Support or Funding InformationResearch was supported by the Ministry of Science and Technology, Taiwan (MOST 105‐2320‐B‐006‐046, 105‐2320‐B‐006‐036,106‐2320‐B‐006‐017, and 106‐2320‐B‐006‐061), the Department of Defense, USA (W81XWH‐08–1‐0682), the National Health Research Institute, Taiwan (NHRI‐EX107‐10734NI) to NS Chang.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Impact of the lipid bilayer on energy transfer kinetics in the photosynthetic protein LH2
Authors: Tong Ashley L , Schlau‐Cohen Gabriela , Ogren John , Cao Jianshu , Chenu Aurelia , Gordon Sam , Blankenship Robert , Lu Yue 
Published: June 2021

Photosynthetic purple bacteria convert solar energy to chemical energy with near unity quantum efficiency. The light‐harvesting process begins with absorption of solar energy by an antenna protein called Light‐Harvesting Complex 2 (LH2). Energy is subsequently transferred within LH2 and then through a network of additional light‐harvesting proteins to a central location, termed the reaction center, where charge separation occurs. The energy transfer dynamics of LH2 are highly sensitive to intermolecular distances and relative organizations. As a result, minor structural perturbations can cause significant changes in these dynamics. Previous experiments have primarily been performed in two ways. One uses non‐native samples where LH2 is solubilized in detergent, which can alter protein structure. The other uses complex membranes that contain multiple proteins within a large lipid area, which make it difficult to identify and distinguish perturbations caused by protein‐protein interactions and lipid‐protein interactions. Here, we introduce the use of the biochemical platform of model membrane discs to study the energy transfer dynamics of photosynthetic light‐harvesting complexes in a near‐native environment. We incorporate a single LH2 from Rhodobacter sphaeroides into membrane discs that provide a spectroscopically amenable sample in an environment more physiological than detergent but less complex than traditional membranes. This provides a simplified system to understand an individual protein and how the lipid‐protein interaction affects energy transfer dynamics. We compare the energy transfer rates of detergent‐solubilized LH2 with those of LH2 in membrane discs using transient absorption spectroscopy and transient absorption anisotropy. For one key energy transfer step in LH2, we observe a 30% enhancement of the rate for LH2 in membrane discs compared to that in detergent. Based on experimental results and theoretical modeling, we attribute this difference to tilting of the peripheral bacteriochlorophyll in the B800 band. These results highlight the importance of well‐defined systems with near‐native membrane conditions for physiologically‐relevant measurements.Support or Funding InformationNational Science Foundation Beckman Young Investigator Department of EnergyThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Isolation and Identification of C‐Mannosylated Hsc70 Conjugates in RAW 264.7 Macrophage‐Like Cells
Authors: Parsons Michael Arland 
Published: June 2021

Post‐translational addition of carbohydrate molecules is often necessary for proteins to be fully active. This process, known as glycosylation, usually involves a carbon‐nitrogen or carbon‐oxygen bond. There is, however, a form of glycosylation that involves formation of a carbon‐carbon bond. C‐linked glycosylation in higher organisms consists of the addition of α‐mannopyranoside from dolichyl‐phosphate mannose onto the C‐2 carbon of tryptophan's indole functional group, generating a residue known as C‐mannosyltryptophan. Though the function of this modification is not well characterized, the addition of α‐mannopyranoside to tryptophan residues is thought to play an important role in the innate immune response. (Muroi et al.; Hartmann and Hofsteenge; Hofsteenge et al.)It is known that the consensus sequence for C‐mannosylation is Trp‐x‐x‐Trp/Cys (x could be any amino acid), in which the first tryptophan is modified. (Furmanek and Hosteenge) This canonical sequence has been found in several signaling molecules such as cytokines and cytokine receptors.It has been shown that Hsc70 interacts with proteins containing the C‐mannosyltryptophan residue. With this in mind, Hsc70 pulldowns were done by the Rakus Lab in an attempt to capture some of these conjugates. Several proteins were identified using this procedure, and mass spectrometry showed a mass shift of 162 Da on tryptophan containing peptides – indicative of a C‐mannosyltryptophan residue.One of these modified peptides was identified as belonging to a Type I cytokine receptor – Interleukin 12 Receptor β2 (IL12Rβ2). Notably, one of its target molecules, interleukin‐12 (IL12), also contains the canonical sequence and has been shown to contain a C‐mannosyltryptophan residue. (Doucey et al.)Anti‐ IL12Rβ2 antibodies were used in an attempt to isolate IL12Rβ2 directly. The same was done for a similar protein IL12Rβ1. While IL12Rβ2 appeared to be present in western blots, mass spec data failed to confirm this. IL12Rβ1 was not seen in western blots or mass spec. Upon successful isolation, a more thorough mass spec analysis can confirm or deny the existence of C‐mannosyltryptophan residues in these receptors.Support or Funding InformationMarshall University Chemistry Department; NSF (OIA‐1738707)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Direct Current Electric Fields Induce Collective Migration of Human Corneal Epithelial Monolayers
Authors: Contreras Adam , Zhu Kan , Guo Liang , Lee Rachel , Campanello Leonard , Losert Wolfgang , Zhao Min 
Published: June 2021

Bioelectricity is an endogenous biological electric field (EF) that can be measured in vivo. A biological direct current (DC) EF is produced by ion fluctuations, and is a common response to wounds in various tissues. Our lab previously illustrated DC EF responses in corneal wounds in rodent models in vivo and in human corneal epithelial cell (hCEC) culture. We have observed that both endogenous DC EF and applied exogenous DC EF are capable of guiding cell migration in the process of electrotaxis. However, we have yet to examine exogenous DC EF responses in epithelial monolayers, which are more representative of a wound edge. Our aim was to characterize collective migration of epithelial monolayers to model electrotaxis of a wound edge. We hypothesized that exogenous DC EF can improve collective migration in hCEC monolayers, and therefore DC EF might be applicable for medical treatments of corneal wounds. To test the first part of this hypothesis, we applied exogenous DC EF to hCEC monolayers. We used a standard method of cell tracking combined with an advanced method of particle image velocimetry (PIV) analysis to extrapolate collective migration data. We observed that applications of DC EF to hCEC monolayers induced collective migration and directionality ‐ the directional precision of migration parallel to the EF. Increasing DC EF voltages of 0, 50, and 200 mV/mm2 positively correlated with increasing migration rates. In controls (0 mV/mm2), we observed no directionality, but at DC EF voltages of 50 and 200 mV/mm2 we observed a comparable directionality response, guiding cell migration parallel (0 degrees) to the DC EF. Further, directionality was promptly reversed following DC EF reversals (180 degrees). We concluded that DC EF provides a precise guidance signal for hCEC monolayers that might be applicable for corneal wound healing therapies. Future investigations will aim to dissect cellular mechanisms of the collective migration response to DC EF and to examine the feasibility of DC EF applications to corneal wounds in vivo.Support or Funding InformationNIH, NEI ‐ 2R01EY019101‐05A1 to MZ; DOD, MURI ‐ FA9550‐16‐1‐0052 to WL and MZThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Neural Circuits and Olfactory Responses to Neurotropic Viruses in Trout
Authors: Mancha Fabiola , Salinas Irene , Huertas Mar 
Published: June 2021

Vertebrate olfactory receptors (OR) are directly exposed to microorganisms, such as viruses, due to their direct contact with the external environment. A previous study showed that nasal delivery of rhabdovirus IHNV (Infectious hematopoietic necrosis virus) in fish activate nasal immune responses marked by an increase of chemokine CCL19 and prostaglandin synthase expression in olfactory epithelia (OE), and infiltration of CD8+ cells in the OE.We hypothesize that nasal immune responses are activated by action potential signals generated by activated olfactory receptor (OR) neurons in the OE and olfactory bulb (OB). Moreover, this neural circuit can be traced from a specific OR cell type (crypt cell) in the nose to a specific site in the olfactory bulb. We tested our hypothesis by measuring olfactory responses to live attenuated IHNV virus by electro‐olfactogram (EOG). We also visualized the IHNV neural circuit after activation of specific OR, and consequent internalization of molecular receptor and IHNV mixed with Alexa dextran 488 3000 MW.Our results showed different EOG olfactory responses to live attenuated IHNV and to the medium where the virus was grown (negative control) in rainbow trout. Olfactory responses followed a dose‐response pattern typical of OR. Cross adaptation studies also showed that live attenuated IHNV activates a set of receptors different from those activated by virus‐free supernatants. Recordings of the OB responses by electroencephalogram are under development. Preliminary tracings show fluorescent oval shaped OR in the apical border of the olfactory lamella (putative crypt cells) that extended to the ventral side of the olfactory bulb. This neural circuit differs from those visualized after exposure of trout OE to the food odorant serine.Combined, our results adds evidence for a new olfactory function in trout, which serves as a first layer of pathogen detection in vertebrates.Support or Funding InformationThis material is based upon work supported by the National Science Foundation under Grant No. 1755348This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Human Skeletal Muscle‐on‐a‐Chip
Authors: Naik Akshata R , Pernal Sebastian , Lewis Kenneth T , Wu Yaobin , Wu Hongkai , Carruthers Nicholas J , Stemmer Paul , Jena Bhanu P 
Published: June 2021

A comprehensive understanding of the emergent properties of in vitro skeletal muscle growth and development is still in its infancy. To address this gap in our current knowledge, we have developed a stretchable micropatterned 3D human skeletal muscle platform that recapitulates organized and parallel growth of muscle cells and fibers as opposed to the randomly oriented cells growth on a 2D glass surface. Mass spectrometry of the muscle cells growing on the 3D platform express key myogenic proteins for myoblast fusion required in the formation of muscle tissue, and proteins involved in mitochondrial health and biogenesis, in contrast to cells growing on 2D glass surface. These results demonstrate that the engineered human muscle cells grown on the 3D platform holds great promise to further establish the emergent properties of in vitro skeletal muscle growth and development for a wide range of biomedical applications such as transplants, drug screening and gene therapy.Support or Funding Information

Interdisciplinary Biological Sceinces (IBS) fellowship
Graduate Research Assistantship, Department of Physiology
Richard Barber Fellowship
National Science Foundation grants EB00303, CBET1066661
This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.