bluuebunny/crossref_metadata_2025_split huggingface dataset:
Role of Transient Receptor Potential Ankyrin‐1 (TRPA1) in Endothelial‐Specific Effects of Diverse Aldehydes
Authors: Conklin Daniel , Jin Lexiao 
Published: April 2020

IntroductionStructurally diverse aldehydes are found in abundance in the environment including in foods, beverages and tobacco‐derived aerosols. Some aldehydes are associated with cardiovascular disease risk especially in cigarettes smokers, however, the detailed mechanisms by which aldehydes influence cardiovascular health and disease are incompletely known.ObjectiveBecause reactive aldehydes, in general, are considered instigators of endothelium dysfunction, we explored the direct effects of a diverse set of aldehydes in isolated murine blood vessels, i.e., aorta and superior mesenteric artery (SMA).MethodsAldehydes: acetaldehyde (AA); formaldehyde (FA); cinnamaldehyde (CA); crotonaldehyde (CR); and a TRPA1 agonist, allyl isothiocyanate (AITC, mustard oil), were tested for vascular effects in isolated murine aorta and SMA using isometric myography.ResultsAll aldehydes induced endothelium‐dependent relaxation in SMA. AA (1–100 mM), FA (30–1,500 μM), CA (30–300 μM) and CR (1–100 μM) strongly and reversibly relaxed (>75% relaxation) agonist‐induced contractions. High levels of FA (1.2–1.6 mM) and CR (>300 μM) irreversibly altered vascular function in aorta and SMA. To identify the mechanism of aldehyde‐induced relaxations, isolated blood vessels were pre‐contracted with phenylephrine (PE), U46,619 (thromboxane A2 analog), or 60 mM K+ (High K) and then exposed to AA, FA, CA, CR or AITC with intact or mechanically‐impaired endothelium (air‐perfused) as well as in the absence or presence of: 1) Nω‐Nitro‐L‐arginine methyl ester hydrochloride (L‐NAME); 2) an inhibitor of guanylyl cyclase (ODQ); and, 3) a TRPA1 antagonist (A967079; 1 μM). The sensitivity (EC50) but not the efficacy (% decreased tension) of FA‐ and AA‐induced relaxations was dependent on the contractile agonist (i.e., PE < U46,619 ≤ High K) indicating multiple mechanisms. In PE pre‐contracted SMA, vasorelaxations at the lowest levels of AA, FA, CA, CR and AITC were inhibited significantly by mechanically‐disrupted endothelium, L‐NAME, ODQ, and by TRPA1 antagonist except for AA, which was TRPA1 independent. TRPA1 was co‐localized with isolectin immunofluorescent staining in SMA endothelium but not in aorta.ConclusionBecause low levels (physiological) of several aldehydes (FA, CA, CR) induce reversible and efficacious vasorelaxations via TRPA1 channels, we propose that endogenous and exogenous sources of these aldehydes may significantly impact gut (mesenteric) blood flow to augment postprandial hyperemia in aid of digestion and nutrient uptake.Support or Funding InformationResearch funding from the National Institutes of Health: GM127607, HL122676, U54HL120163.
The Role of Glutamate in Maintaining Muscle Spindle Afferent Excitability in Adult Mice
Authors: Ortiz Serena , Kim Enoch , Than Kimberly , Bustos Cebrina , Chu Sarah , Klier Nikola , Occiano Alyssa , Salazar Alexandra , Villegas Natanya , Wilkinson Katherine A. 
Published: April 2020

Group Ia and II muscle spindle afferents are sensory neurons critical to detecting muscle movement and length, which is important in proprioception and motor control. Glutamate is released by synaptic‐like vesicles (SLVs) at muscle spindle afferent nerve endings and exogenous glutamate increases sensory neuron firing rate during stretch (Bewick, et al., 2005). However, the activity of individual muscle spindle afferents was not measured, allowing for the possibility that this increased firing also included other sensory neurons known to have glutamate receptors, like nociceptors. Here we test the hypothesis that glutamate increases the firing rate of individual muscle spindle afferents. We isolated the extensor digitorum longus muscle and the sciatic nerve from adult mice (8–12 wks old) and recorded the activity of individual muscle spindle afferents during exogenous glutamate treatment. The muscle‐nerve preparation was perfused in an oxygenated synthetic interstitial fluid tissue bath, an extracellular recording electrode was placed on the nerve to measure the activity of individual muscle spindle afferents, and a battery of 4s ramp‐and‐hold stretches were performed on the muscle. Glutamate addition led to a significant increase in muscle spindle afferent firing rate during stretch (1mM; n=12, 18.8%). We also decreased glutamate release by blocking the vesicular glutamate transporter 1 (VGLUT1) with the inhibitor xanthurenic acid (XA). We found a significant decrease in muscle spindle afferent firing rates (3mM; n=17, −39%), with 5 afferents exhibiting a complete absence of firing. However, we noticed heterogeneity in muscle spindle afferent responses to glutamate and XA treatment, with some afferents not responding at all to the drug treatments. To confirm our pharmacological findings, we obtained a transgenic mouse line lacking a single copy of vesicular glutamate transporter 1 (VGLUT1; B6.129X1‐Sic17a7tm1Edw/MmcD; Fremeau, 2004). We observed two muscle spindle afferents responding normally to stretch, while another two failed to sustain firing at the end of stretch, something that is only rarely seen in wild type animals. This heterogeneity may be due to differences in VGLUT1 protein levels in individual afferents. We are currently increasing our sample size, but our preliminary results support our hypothesis that glutamate is critical to maintaining muscle spindle afferent firing during maintained stretch. These studies will lead to a better understanding of how muscle spindle afferent stretch sensitivity is regulated.Support or Funding InformationThis work was supported by NIH SC3 GM127195 (KAW) and NIH R25 GM071381 (SO and NV).
The Role of Metadherin in Inflammatory Breast Cancer Progression
Authors: Ortiz-Soto Gabriela , Suárez-Arroyo Ivette J. , Martínez-Montemayor Michelle M. 
Published: April 2020

Inflammatory breast cancer (IBC) is the most lethal and aggressive form of breast cancer where patients have a 43% increased risk of death compared to women with non‐IBC. The lethality of IBC originates from its nature of invading the vascular and lymphatic systems, absence of a typical tumor mass, and the generation of tumor emboli. The arising challenges in IBC include the accurate and early diagnosis and the development of effective targeted therapies. A previous study investigated potential biomarkers involved in tumor spheroid integrity by characterizing IBC’s cell surface proteome. Plasma membrane proteins were identified via “stable isotope labeling with amino acids in cell culture” (SILAC) with subsequent mass spectrometry (MS)‐based quantitative proteomics analysis using the IBC cell line SUM‐149 vs. non‐ cancerous mammary epithelial cells, MCF10A. SILAC results showed overexpression of Metadherin (MTDH) in SUM‐149 cells (>2.0 fold change). Subsequently, we validated the overexpression of MTDH in SUM‐149 IBC cells, IBC tissues and in the emboli. MTDH is a cell adhesion molecule overexpressed in many cancer types, including breast cancer. MTDH promotes cancer progression by modulating various signaling pathways including NF‐kB, which is related to cell survival, proliferation, invasion and metastasis. Therefore, the aim of this study is to assess the functional role of MTDH in IBC progression. To elucidate the mechanism of action of MTDH in colony formation and tumor spheroid we silenced MTDH (CRISPR or shRNA) SUM‐149 and SUM‐190 IBC cells. We assessed the expression of MTDH by immunoblotting and immunofluorescence where we observed reduced MDTH levels. The number of colonies formed in SUM‐149 MTDH silenced cells was ~40% lower when compared to non‐silenced cells. Tumor spheroids of SUM‐149 and SUM‐190 MTDH silenced cells were smaller than non‐silenced cells. Aditionally, MTDH silencing results in a decrease in the phosphorylation of the p65 subunit of NF‐kB while no changes are observed in total protein abundance in SUM‐149 cells. We can conclude from our preliminary results that MTDH serves as a potential IBC target for IBC progression.Support or Funding InformationThis work was supported by NIH NIGMS #GM111171 (MMM), SGRP 2017‐00143 (MMM and GOS), NIMHD #MD007583 (MMM), GM103475 (UPR MMM), Title‐V‐PPOHA #P031M105050, Title‐V‐Cooperative #P031S130068 from the U.S. Dept. of Education and Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health #P20GM103475 (GOS).
Eicosapentaenoic Acid Increases Browning Markers in Subcutaneous Adipose Tissue and Primary Adipocytes from Wild Type and UCP‐1 Deficient Mice
Authors: Ramalingam Latha , Jayarathne Shasika , Pahlavani Mandana , Scoggin Shane , Moustaid-Moussa Naima 
Published: April 2020

ObjectiveUncoupling protein ‐1 (UCP‐1) plays a critical role in brown adipose tissue (BAT) thermogenesis as well as in white adipose tissue (WAT) browning, which primarily occurs in subcutaneous adipose tissue (SAT). We have previously reported that fish oil‐enriched with eicosapentaenoic acid (EPA) reduced obesity, and increased UCP‐1 in BAT of B6 mice fed high fat diets (HF). Furthermore, we and others demonstrated that these protective metabolic effects of fish oil or EPA in BAT were independent of UCP‐1. Our goal here is to determine if EPA exerts anti‐obesity effects in SAT, by increasing browning and thermogenic markers and whether these effects are mediated by UCP‐1.MethodsWild type (WT) and UCP‐1 knock‐out (KO) male mice starting at 6 weeks of age were maintained at thermoneutral environment and fed HF with or without 36g/kg of EPA‐enriched fish oil (800 mg/g) for 14 weeks. The thermoneutral environment was used to induce obesity in UCP‐1 KO mice. SAT was collected for histological, gene and protein analyses. Moreover, SAT was collected from chow diet‐fed WT and KO mice at ambient environment to prepare stroma vascular cells, which were differentiated into adipocytes, then treated with 100uM EPA for 48 hours and harvested for mRNA and protein analyses. Lastly, extracellular flux analyzer was used to measure the mitochondrial capacity and oxygen consumption in differentiated SAT adipocytes. Differences among groups were analyzed using one way ANOVA with significance at p<0.05..ResultsWT mice supplemented with EPA had similar body weight to WT mice fed HF. However, UCP1 KO mice fed HF had higher body weight than all other groups (p<0.05). Interestingly, WT and KO mice fed EPA had smaller SAT adipocytes compared to their respective HF control groups. mRNA levels of fibroblast growth factor‐21 (FGF‐21) and Peroxisome proliferator‐activated receptor gamma coactivator 1‐alpha (PGC1α) which are oxidative markers were not altered in WT groups. However, UCP‐1 KO mice fed had HF or EPA had lower levels of FGF21 and PGC1α compared to l HF WT mice p<0.05, with no differences between the KO genotype. EPA significantly increased gene expression of UCP‐2, UCP‐3 and FGF21 in differentiated SAT adipocytes from both WT and KO compared to control. KO mice fed HF had lower levels of browning markers such as transmembrane protein 26 (TMEM26) and T BOX ‐1 (TBX) in SAT compared to WT mice fed HF, while EPA rescued these only in KO mice, but not in WT mice. Lastly, only the spare respiratory capacity was improved with EPA in both WT and KO primary adipocytes. Additional mechanistic studies are currently underway to further dissect EPA effects on different fat depots in WT vs KO mice.Support or Funding InformationFunding SourcesNIH/NCCIH grant #R15AT008879‐01A1 (N.M.M.)
Expression of Aquaporins AaAQP2 and AaAQP6 in the Osmoregulatory Organs of Mosquito Larvae (Aedes aegypti)
Authors: Guardian Elia Edith Grieco , Donini Andrew 
Published: April 2020

The juvenile stages of the yellow fever mosquito, Aedes aegypti, are found in semi‐permanent water storage containers, septic tanks, and natural coastal rock pools. Changing climates and anthropogenic pollutants affect the ionic (salt) composition of these mosquito breeding sites which can influence the spread and distribution of vector populations. These habitats impose different challenges to the salt and water balance of the mosquitoes which is accomplished by regulation of salt and water transport across epithelial barriers. Aquaporins (AQPs) are transmembrane proteins first discovered as selective water channels and can be classified by their transport capabilities to other small solutes. Six aquaporin isoforms have been identified in A. aegypti mosquitoes, AaAQP1‐6, and here we investigated the protein localization and abundance levels of AaAQP2 and AaAQP6 in the osmoregulatory organs of the aquatic larvae reared in either ion‐poor water (IPW; reverse‐osmosis water), artificial freshwater (FW), or brackish water (BW; 30% seawater). Using custom antibodies with immunohistochemistry and western blotting it was found that AaAQP2‐like and AaAQP6‐like immunofluorescence was primarily intracellular in the Malpighian tubules (MTs) and gastric caeca (GC). Rearing salinity did not affect protein abundance of AaAQP2 in MTs, GC and hindgut which appeared as a putative dimer of ~ 45 to 50 kDa; however, in the hindgut of BW reared larvae an additional band of ~ 30 kDa was apparent which may represent the monomer. Western blots for AaAQP6 are currently being optimized. Results suggest that AaAQP2 and AaAQP6 are required in these osmoregulatory organs regardless of the salinity of the external environment during development. Furthermore, AaAQP2 may have additional functions in the hindgut of larvae that develop in higher salt environments.Support or Funding InformationNSERC ‐ DISCOVERY RGPIN‐2018‐05841
Expression and purification of N‐terminally acetylated Ssa1 chaperone from Saccharomyces cerevisiae
Authors: Griffith Alijah A. , Holmes William 
Published: April 2020

N‐terminal acetylation leads to proteomic diversity by constituting a cellular acetylome wherein proteins exhibit an alternate function and dynamic, allowing cells to react specifically to environmental stressors or cellular perturbation. N‐terminal acetylation is catalyzed by a collection of N‐terminal acetyltransferases (NATs), which transfer acetyl moieties from acetyl‐coenzyme A to α‐amino groups at specific N‐terminal residues. One theoretical target of N‐terminal acetylation is Ssa1, an Hsp70 class chaperone endogenous to Saccharomyces cerevisiae. Ssa1 works in tandem with its co‐chaperone Sis1 to restore the native conformations of aberrantly‐folded proteins through a transient ATP‐driven mechanism. The consequences of N‐terminal acetylation on Hsp70 function remain to be fully characterized. To further probe the functional and mechanistic changes associated with the N‐terminal acetylation of Hsp70, we developed an efficient, user‐friendly, one‐step purification of Ssa1 using previously constructed Protein‐A fusion yeast strains. Our results indicate that our method isolates high yields of pure acetylated and unacetylated Ssa1 without compromising enzymatic activity, allowing for future evaluation of the effects of N‐terminal acetylation on Hsp70 chaperone function and dynamic.Support or Funding InformationResearch supported by RI‐INBRE (#P20GM103430) from NIGMS, NIH.
Vitamin D deficiency contribute to the development of preeclampsia through increased sFlt‐1 production and impaired vascular function
Authors: Yang Weiwei , Li Qinghua , Yin Lingxuan , Si Yanru , Gao Zhiqin , Meng Yuhan , Cao Bin , Wang Yanling 
Published: April 2020

Preeclampsia (PE) is a pregnancy‐specific syndrome that affects 5~8% pregnancies and is a leading cause of maternal and neonatal morbidity and mortality. Although the underlying mechanism remains unclear, PE is widely accepted to be induced, in part, by uteroplacental ischemia, which leads to increased antiangiogenic factors, such as sFlt‐1, which antagonizes the angiogenic factors, causing widespread vascular dysfunction. Vitamin D deficiency (VDD) is a widespread public health issue, and increasing studies suggested that VDD is a possible risk factor for the development of PE; however, the underlying mechanism that contributes to the increased risk of preeclampsia in vitamin D deficient patients is not clear. We tested the hypothesis that vitamin D deficiency contributes to the development of PE in a placental ischemic rat model by increasing sFlt‐1 production and impairing vascular function. We first induced VDD in Sprague‐Dawley rats (DYET 119273 vs 110842, Dyets Inc., USA) prior to RUPP (Reduced Uterine Perfusion Pressure) surgery on gestational day (GD) 14. Blood pressure was monitored via carotid catheter. On GD19, uterine arteries were harvested to determine vascular reactivity on a wire myograph. We found that RUPP surgery significantly increased mean arterial blood pressure as expected(Ctrl+Sham, n=12, 102±10 mmHg; Ctrl+RUPP, n=12, 117±11 mmHg; P<0.01), and this was significantly enhanced by vitamin D deficiency (VDD+RUPP, n=12, 136±11 mmHg; P<0.01). Plasma sFlt‐1 concentrations were significantly increased in VDD rats (n=12, 117±15 pg/mL) compared to the control group(n=12, 49±17 pg/mL)(P<0.05), and VEGF concentrations were significantly decreased(Ctrl, n=12, 1899±371 pg/mL; VDD, n=6, 1071±249 pg/mL, P<0.05). VDD induced endothelial dysfunction in uterine arteries (% relaxation, control, n=6, 95±7%; VDD, n=6, 37±11%, P<0.05). The circulating biomarkers for endothelial dysfunction also significantly increased. The E‐Selectin increased from 5166±1217 pg/mL to 5756±1053 pg/mL (P<0.05). The ICAM‐1 levels increased from 3.15±1.03 ng/mL to 3.50±0.94 ng/mL (P<0.05). The VCAM‐1 level increased from 44±13 ng/mL to 53±11 ng/mL (p<0.05). These results suggest that vitamin D deficiency contributes to the onset of preeclampsia by increased sFlt‐1 production and impaired vascular function.Support or Funding InformationThis research was supported by the National Natural Science Foundation of China (81601318) and Shandong Province Health and Medical Science and Technology Program (2016WS0668), Weifang Medical University (No. 2017BSQD11).
Repeatability of Cerebral Vasomotor Reactivity to Hypercapnia in Humans
Authors: Nandadeva Damsara , Patik Jordan C. , Martin Zachary T. , Fadel Paul J. , Brothers R. Matthew 
Published: April 2020

Impaired cerebrovascular function is a risk factor for the development of various neurocognitive and cerebrovascular diseases including cognitive dysfunction, dementia, Alzheimer’s disease, and stroke. Since hypercapnia elicits a robust cerebral vasodilator response, cerebrovascular function is commonly assessed as an increase in cerebral blood flow to elevations in arterial carbon dioxide (i.e., cerebral vasomotor reactivity ‐ CVMR). Hypercapnia induced by a rebreathing protocol is one such technique used to assess cerebrovascular function. Although this technique is used to evaluate CVMR within and between subjects before and after interventions, to our knowledge, the repeatability of CVMR assessed by rebreathing has not been comprehensively investigated. Moreover, the National Institute of Health’s recent guidelines emphasize the need for rigor and reproducibility of data collection techniques. Accordingly, this study was designed to determine the within‐day repeatability of CVMR response to rebreathing‐induced hypercapnia. Eleven young healthy males (age = 24 ± 3 years, BMI = 24.3 ± 3.8 kgm−2, mean ± SEM) reported to the laboratory following an overnight fast and performed 3 trials of rebreathing‐induced hypercapnia each separated by approximately 2 hours. Heart rate (ECG), respiration (Pneumotrace), beat‐to‐beat blood pressure (Finometer), middle cerebral artery mean blood velocity (MCAv ‐ transcranial Doppler) and breath‐by‐breath end‐tidal carbon dioxide concentration (PETCO2 ‐ capnograph) were continuously measured. Cerebral vascular conductance index (CVCi) was calculated as MCAv divided by mean arterial pressure. CVMR was assessed as the slope of the linear regression between the increase in %MCAv and %CVCi across the range of change in PETCO2 (ΔPETCO2). The increase in %MCAv and %CVCi was also assessed at a ΔPETCO2 of 15 mmHg which was the highest common change in magnitude of hypercapnia achieved across all rebreathing trials among all participants. Repeatability between the 3 trials was assessed using intra class correlation coefficient (ICC). The slopes of %CVCi vs. ΔPETCO2 (3.9 ± 0.4, 3.8 ± 0.3 and 3.6 ± 0.3 %mmHg−1 for 1st, 2nd and 3rd trial respectively, p = 0.82) was not different and showed very good repeatability (ICC = 0.86, p < 0.001) between the 3 rebreathing‐induced hypercapnia trials. Likewise, the % increase in CVCi at ΔPETCO2 of 15 mmHg was not different (40 ± 4, 38 ± 4 and 36 ± 4% for 1st, 2nd and 3rd trial respectively, p = 0.72) and also demonstrated very good repeatability between trials (ICC = 0.86, p < 0.001). Similar results were obtained when MCAv was used as the dependent variable. These results suggest that within‐day CVMR assessed by rebreathing‐induced hypercapnia is highly repeatable and therefore can be used as a reliable index of cerebrovascular function.Support or Funding InformationSupported by UTA College of Nursing and Health Innovation
Group A Streptococcus May Suppress V‐ATPase Activity To Inhibit THP‐1 Macrophage Phagolysosomal Acidification
Authors: Lee Jimin , Nishioka Scott , Snipper Joshua , Okumura Cheryl Y.M. 
Published: April 2020

Infection with Group A Streptococcus (GAS), a human‐specific bacterial pathogen, results in a multitude of diseases ranging from strep throat to toxic shock, and can lead to serious post‐infection sequelae such as rheumatic heart disease. During infection, macrophages engulf bacteria within the phagosome, which then fuses with the lysosome. Subsequent acidification of the phagolysosome allows for degradation of bacteria. Our lab and others have shown that during GAS infection, phagolysosomes are not acidified, allowing GAS to persist. The overall objective of this study was to determine how GAS inhibits phagolysosomal acidification. Our lab has previously shown that secretion of the pore‐forming toxin Streptolysin O (SLO) causes phagolysosomal leakage of relatively large proteins. In our current work, we demonstrate that during infection using a mutant strain lacking SLO or using heat‐killed bacteria that GAS‐infected phagosomes are still not acidified, indicating that lack of acidification is not due to mere leakage. Phagolysosomes undergo acidification using the enzyme complex vacuolar ATPase (VATPase), which needs to properly assemble cytosolic and membrane components to function. To assess whether V‐ATPase assembly was affected, infected cells were fractionated and recruitment of cytosolic V‐ATPase subunits to the phagosome membrane was monitored by Western blot. Our preliminary results indicate that V‐ATPase assembly is not inhibited, suggesting that GAS may instead prevent acidification by inhibiting V‐ATPase activity. We are currently conducting further research to investigate this potential inhibitory mechanism of GAS. Restoring the ability of lysosomes to become acidified and activate proteolytic enzymes may serve as a means to enhance macrophage function, both bactericidal capacity and antigen presentation capability, to elicit effective long‐lasting immunity.Support or Funding InformationAmerican Heart Association (17GRNT3410851), Occidental College Undergraduate Research Center