bluuebunny/crossref_metadata_2025 huggingface dataset:
KCC and NKCC activity affect spike frequency adaptation in CO2‐sensitive avian intrapulmonary chemoreceptors (1092.3)
Authors: Meehan Shane , Bassett Peter , Lonjaret Jean‐Guillaume , Hempleman Steve , Sheridan Krystal
Published: June 2021

IPC are respiratory chemoreceptors that exhibit SFA, have phasic and tonic CO2 sensitivity, and help control breathing in birds. In IPC, large changes in PCO2, pHi, and [HCO3]i occur with each breath. Here we test the hypothesis that Cl‐ transmembrane flux may be involved in IPC CO2 transduction to balance intracellular [HCO3‐] changes. Furosemide (FU), an equipotent inhibitor of both K+ ‐Cl‐ cotransport (KCC), which allows for Cl‐ efflux driven by the K+ gradient, and Na+ ‐K+ ‐2Cl‐ cotransport (NKCC), which allows for Cl‐ influx driven by the Na+ gradient, was given to anesthetized Anas platyrhynchos (N=6) while recording single‐unit IPC. FU progressively reduced SFA at 10‐40 mg/kg (p蠄0.0001). In contrast, tonic IPC discharge was decreased at low FU doses (0.5‐5 mg/kg, p蠄0.0001) but increased at higher dosages (10‐40 mg/kg, p蠄0.0001). Results suggest that IPC SFA and tonic CO2 sensitivity are influenced by KCC and/or NKCC, presumably by reducing ability of the cell to produce Cl‐ counter movements required by phasic [HCO3‐] changes.

Grant Funding Source: Supported by NIH R15 HL087269‐02
CORM‐3 derived CO suppresses NLRP3 inflammasome in cardiac fibroblasts and protects cardiomyocytes from apoptosis in mice with sepsis (1096.6)
Authors: Zhang Wenbo , Xu Xuemei , Mele Tina , Kao Raymond , Cepinskas Gediminas , Martin Claudio , Rui Tao
Published: June 2021

The study is to examine the role of CO derived from CORM‐3 on NLRP3 inflammasome activation in cardiac fibroblast (CF) and its effect on cardiomyocyte apoptosis and myocardial dysfunction in mice with sepsis. Methods: CFs were primed with LPS and followed by ATP treatment with or without CORM‐3. CF NLRP3 expression and NLRP3 inflammasome activation assessed. Cardiomyocyte apoptosis and myocardial function were measured in mice with LPS. Results: LPS priming and ATP treatment to CFs resulted in NLRP3 inflammasome activation as indicated by increase in cleaved caspase‐1 p20 and matured IL‐1β. CFs with NLRP3 siRNA abolished caspase‐1 cleavage and IL‐1β maturation. CORM‐3 attenuated the increase in NLRP3 and pro‐IL‐1β expression in LPS primed CFs and prevented NLRP3 inflammasome activation. CORM‐3 inhibited NLRP3‐mediated ASC pyroptosome formation. Myocardial NLRP3, pro‐capase‐1, p20, pro‐IL‐1β and IL‐1β were increased, cardiomyocyte apoptosis was induced and myocardial contractility was decreased in mice with LPS. CORM3 prevented the myocardial NLPR3 inflammasome activation, attenuated cardiomyocytes apoptosis, and improved cardiac function in mice with LPS. Conclusions: CORM‐3‐derived CO suppresses NLRP3 inflammasome priming and activation in CFs and protects myocardium from apoptosis and dysfunction in mice with endotoxemia. (HSFO 2012‐000212)

Grant Funding Source: Heart and Stroke Foundation of Ontario
The effects of hypersalinity on the physiology and morphology of a livebearing fish (1100.12)
Authors: Castellanos Krystal , Cavazos Corrina , Weaver Kathleen , Garcia Jerome , Weaver Pablo
Published: June 2021

The Poeciliidae family of livebearing fish exhibit broad environmental tolerances and occupy some of the most extreme aquatic environments on earth. One group in particular, the Limia genus, is widespread throughout freshwater habitats in the Dominican Republic and has also colonized hypersaline lagoons at salinity levels more than twice that of sea water. Populations found in hypersaline conditions appear to be smaller and less colorful than their freshwater counterparts. We hypothesize that these changes to morphology may be related to the energetic cost of osmoregulation in an extremely hypertonic environment. Here we use geometric morphometrics to quantify morphological differences, including size and shape, of freshwater and hypersaline populations of L. perugiae. We correlate changes to size and shape with an increase in energy usage by the active ion transport mechanisms of the mitochondria rich cells in the gill tissues.

Grant Funding Source: Supported by Title V STEM, Rose Hills, University of La Verne Faculty Research Grant
Influence of myostatin on skeletal muscle plasticity (1102.22)
Authors: Reisz‐Porszasz Suzanne , Abraham Andrea , Baker Michael , Caiozzo Vincent
Published: June 2021

Goal: to investigate the role of myostatin (Mst) in adult skeletal muscle structure, function and physical performance. We generated the conditional Mst overexpressing transgenic (CMOT) mice in which Mst expression can be switched ON or OFF. Methods: We used mechanical overloading (MO) (bilateral synergist ablation of plantaris muscle) to determine the role of Mst in muscle structure and function in Mst KO (knockout), TG (transgenic), CMOT and WT (wild type) animals. Results: In CMOT mice, Mst level was 5 ‐10 times higher compared to WT followed by doxycyclin treatment. After 6 weeks of MO surgery, plantaris muscle specific tension was the highest, but not significant in CMOT; muscle mass increased in each groups, the biggest increase (200%) was in Mst TG; total cross sectional area and fiber number had increased with the highest rate (2.3x and 1,4x, respectively) in CMOT. Succinate dehydrogenase (SDH) was determined, SDH positive fiber number significantly increased in Mst KO and CMOT; the oxidative fiber ratio was the highest (45.3%), while the SHD negative fiber ration was the lowest (21.6%) in CMOT mice. In CMOT animals, TGF‐beta, insulin, Wnt, LDL, PI3K/Akt and p53 signaling pathways were up‐regulated. Conclusion: We have established an animal model, the CMOT with which we can investigate the correlation between Mst levels and muscle wasting in adulthood and the possible therapeutic intervention of muscle loss.

Grant Funding Source: NIH/NIAMSD R21AR054101
Motor unit recruitment patterns in genioglossus and first dorsal interosseous (1102.4)
Authors: Shumway Karlie , Porfirio David , Bailey Fiona
Published: June 2021

The nervous system regulates force via two means; rate coding and recruitment. In a previous study (Bailey et al. 2007) we reported on rate coding of genioglossus (GG) motor units (MUs) in the context of voluntary tongue movement. Here we focus on MU recruitment and compare GG with first dorsal interosseous (FDI). We recorded single MU action potentials via intramuscular tungsten microelectrodes inserted into the GG and FDI in 9 healthy adults (N=3 men; N=6 women). Recruitment was elicited in FDI via force increments associated with finger abduction (force range 4‐7% of max), while recruitment in the GG was elicited via increments in sound pressure level (loudness range 5‐34% of max) associated with articulation of the vowel /i/. We noted significantly higher firing rates in GG MUs at recruitment relative to FDI (18.9 4.8 vs. 11.0 2.8). Once recruited, GG MUs discharged at significantly higher rates than FDI MUs (20.9 4.7 vs. 12.1 2.7). Moreover, the firing rates of subsequently recruited units was also notably higher for GG than for FDI (21.3 4.8 vs. 12.2 2.7). These findings provide useful insights into motoneuron pools that regulate movements of the tongue in speech and of the hand in grasping. Higher firing rates at recruitment for GG compared to FDI, given comparable pressure/force ranges, suggest hypoglossal motoneurons have a shorter duration after hyperpolarization than spinal motoneurons (Bailey et al. 2007).

Grant Funding Source: Supported by the National Institute of Health Grant (DC 009587)
Short‐ and long‐term effects of ActRIIB receptor ligand trapping agents on muscle mass and downstream signaling in dystrophic (mdx) limb and respiratory musculature (1102.44)
Authors: Potter Ross , Desai Pooja , Nielsen Cory , Corbell Kathryn , Mellon Christopher , Danishyar Amina , Gatti Francesca , Grinberg Asya , Kumar Ravi , Carroll Chad , Carlson C. George
Published: June 2021

These studies examined the translational efficacy and the signaling consequences of in vivo treatment with two soluble ActRIIB receptor ligand trapping agents (Acceleron Pharma) in the mdx mouse model for Duchenne muscular dystrophy. A one month treatment of 30 day old mdx mice with RAP‐031 (10 mg/kg; subcutaneously, twice weekly) produced significant (p<0.05) increases in the body mass of both male (14% increase) and female (22% increase) mice. These increases were associated with proportionately larger increases in limb musculature (gastrocnemius; 30%) than in respiratory musculature (costal diaphragm; 12%). Longer periods of treatment (3 months) with RAP‐435 induced 30% increases in body mass and again produced larger proportional increases in the mass of the gastrocnemius (about 90%) than the costal diaphragm (about 43%). The 1 month RAP‐031 treatment produced relatively moderate increases in fiber density and diameter in the severely dystrophic mdx triangularis sterni (TS) muscle. However, the 3 month treatment with RAP‐435 substantially increased TS muscle mass by producing large increases in both fiber density and fiber diameter. Collagen expression and crosslinking in the mdx costal diaphragm was not influenced by treatment with RAP‐031 for periods of 1 or 3 months. The total expression of smads 2 and 3 were unaffected by RAP‐031 treatment. RAP‐031 treatment increased the expression of Smad 4 in the gastrocnemius but not in the costal diaphragm. The expression of Akt was unaffected by RAP‐031 treatment in either muscle preparation, while the proportion of phosphorylated Akt was slightly increased by RAP‐031 treatment in the gastrocnemius. These results further demonstrate the efficacy of the ActRIIB receptor ligand trapping agents in increasing dystrophic muscle mass, and suggest that muscle‐specific differences in TGF‐beta expression or smad and Akt signaling may contribute to differential effects in the limb and respiratory musculature.

Grant Funding Source: Supported by NIHR15AR055360 to CGC
Thermoregulatory dysfunction in multiple sclerosis patients during moderate exercise in a thermoneutral environment (1104.17)
Authors: Huang Mu , Morris Nathan , Jay Ollie , Davis Scott
Published: June 2021

Impairments in sudomotor function during passive heat stress have been reported in multiple sclerosis (MS), a demyelinating disease of the CNS that disrupts autonomic function. However, little is known regarding exercise induced increases in core body temperature on thermoregulatory mechanisms in MS. Thus, the aim of this study was to test the hypothesis that thermoregulatory function is impaired in MS patients compared to healthy controls (CN) during moderate exercise. Thermoregulatory function in five patients diagnosed with relapsing‐remitting MS and five mass‐matched healthy controls were compared during a single bout of cycling exercise (fixed workload of 70 Watts) for 30‐60 minutes in a climate‐controlled room (25°C, 30% RH). Sweating thermosensitivity (MS: 0.56±0.15 vs CN: 0.81±0.13, p=0.04) was significantly lower while a delay in sweating onset time (MS: 14.8±10.0 min vs CN: 5.6±1.6 min, p=0.07) approached significance in MS patients compared to controls. These altered mechanisms of body temperature regulation likely contributed to a greater observed change in core body temperature measured rectally (MS: 0.84±0.34 °C vs CN: 0.37±0.27 °C, p=0.04) in patients with MS. This observed thermoregulatory dysfunction in MS patients may intensify disease symptoms limiting exercise tolerance.

Grant Funding Source: Kuzell Institute and National MS Society Grant RG4043A1/1
Exercise‐induced hyperthermia exacerbates the rightward displacement in the force‐frequency relationship compared with passive hyperthermia (1104.27)
Authors: Periard Julien , Racinais Sebastien , Thompson Martin
Published: June 2021

This study examined the extent to which exercise‐induced hyperthermia (ExH) influences contractile function and modulates the force‐frequency relationship, compared with passive hyperthermia (PaH). ExH was induced in ten males by cycling at 60% VO2max in 38°C conditions and PaH by sitting in a 48°C chamber. Prior to and following ExH and PaH, an 8 s train of stimulation (10, 20, 50 and 100 Hz; 2 s per frequency) and a potentiated twitch were evoked on the relaxed knee extensors via percutaneous stimulation. Core (~37.0 to ~38.5ºC) and muscle (~34.7 to ~38.5ºC) temperature were increased similarly during both trials (P<0.01). The development of ExH and PaH resulted in a decrease in the 20:50 Hz force ratio, indicative of low‐frequency‐fatigue (P<0.01). This alteration was more pronounced after ExH than PaH (P<0.01). A rightward displacement in the force‐frequency relationship occurred after ExH and PaH (P<0.05), but was exacerbated by ExH (P<0.05). Peak twitch force decreased following ExH (P<0.05) and was maintained after PaH. The exacerbated rightward shift in the force‐frequency relationship during ExH appears to stem from adjustments influencing force summation at low frequencies of stimulation. This is likely due to the development of peripheral fatigue, evidenced by a reduction in peak twitch force production.
Local 5‐methyltetrahydrofolate administration augments NO‐dependent vasodilation in aged human skin (1106.11)
Authors: Stanhewicz Anna , Alexander Lacy , Kenney W.
Published: June 2021

Even in the absence of overt cardiovascular disease, older men and women have reduced vascular endothelial function, evidenced by attenuated nitric oxide (NO)‐dependent cutaneous vasodilation (VD) during local heating. Folic acid and its metabolite 5‐methyltetrahydrofolate (5‐MTHF) improve conduit vessel function, possibly through NO‐dependent mechanisms. We hypothesized that local 5‐MTHF administration would augment NO‐dependent VD in aged skin. Two intradermal microdialysis fibers were placed in the forearm skin of 7 young (Y, 23±1 years) and 7 older (O, 77±3 years) subjects for local delivery of Ringer’s solution (control) or 5mM 5‐MTHF. Red cell flux was measured over each site by laser‐Doppler flowmetry (LDF) during standardized local heating to 42°C. After cutaneous vascular conductance (CVC = LDF/MAP) plateaued, 20mM L‐NAME was perfused at each site to quantify NO‐dependent VD. Data were normalized as %CVCmax (28mM SNP and local heat, 43°C). Local heating plateau (O: 82±3 vs. Y: 96±1%CVCmax; p=0.002) and NO‐dependent VD (O: 26±6 vs. Y: 49±5%CVCmax; p=0.03) were attenuated at the control site in O. 5‐MTHF administration augmented overall (91±2%CVCmax; p=0.03) and NO‐dependent (43±9%CVCmax; p=0.04) VD in O but not Y. Local 5‐MTHF administration restores cutaneous VD through augmented NO‐dependent VD, suggesting that exogenous 5‐MTHF increases NO synthesis in aged cutaneous microvasculature.

Grant Funding Source: Supported by NIH RO1‐AG07004‐25
Is sustained post‐exercise vasodilation associated with an exercise‐induced mast cell activation? (1106.19)
Authors: Romero Steven , Ely Mathew , Sieck Dylan , Buck Tahisha , MacLean David , Halliwill John
Published: June 2021

An acute bout of aerobic exercise elicits a sustained post‐exercise vasodilation which is histamine‐receptor mediated. However, the source of the histamine which activates histamine receptors during recovery from exercise remains unknown. One possible source is degranulation of mast cells. We tested the hypothesis that tryptase, a marker of mast cell degranulation, is elevated during or after exercise. In 11 healthy subjects (6 men; 5 women), we measured interstitial tryptase via microdialysis of the vastus lateralis before, during, and through 90 min after exercise (1 hr of unilateral dynamic knee extension at 60% of peak power; 6 subjects) or rest (1 hr seated upright; 5 subjects). Compared with baseline, interstitial tryptase was increased more during exercise (Δ 60 ± 7%) than during rest (Δ 10 ± 9%, P < 0.05) and tended to remain elevated during recovery (exercise Δ 44 ± 14% vs rest Δ 16 ± 7%, P = 0.11). Thus, localized mast cell degranulation occurs in exercising muscle, and appears to contribute in part to sustained post‐exercise histaminergic vasodilation. This suggests that exercise produces an anaphylactoid signal which may influence skeletal muscle blood flow during exercise.

Grant Funding Source: NIH grant HL115027
Neurovascular responses to whole‐body cold stress in aged humans (1106.6)
Authors: Greaney Jody , Kenney W. , Alexander Lacy
Published: June 2021

Human aging is associated with an augmented pressor response to cold stress; however, whether this is due to alterations in sympathetic nervous system activation remains unclear. We hypothesized that whole‐body cold stress elicits greater sympathetic nervous system responses in older (O) compared to young (Y) adults. Reflex vasoconstriction was induced in 7 Y (24±1 yr) and 6 O adults (66±2 yr) by lowering mean skin temperature (Tsk) from 34 to 30.5℃ using a water‐perfused suit. BP, muscle sympathetic nerve activity (MSNA; peroneal microneurography), and red blood cell flux (laser Doppler flowmetry; dorsum of foot) were measured continuously. Cutaneous vascular conductance (CVC=flux/mean BP) was expressed as a percent change from baseline. The increase in systolic BP during whole‐body cold stress was greater in O adults (∆2±1 Y v. ∆10±2 O mmHg; P<0.05); however, the increase in mean BP was not different (∆5±1 Y v. ∆6±1 O mmHg; P>0.05). Reflex cutaneous vasoconstriction was attenuated in O adults (‐31±6 Y v. ‐14±3 O %∆CVC; P<0.05). MSNA was greater in O adults at baseline (Tsk=34℃; 14±3 Y v. 25±2 O burst/min) and during cooling (Tsk=30.5℃; 14±4 Y v. 37±3 O burst/min; both P<0.05). MSNA increased during cooling in O, but not Y, adults (∆‐1±1 Y v. ∆12±3 O burst/min; P<0.05). These data suggest the MSNA response to cold stress is augmented, but peripheral vascular response is attenuated, in healthy aging.

Grant Funding Source: Supported by HL120471‐01 (JLG), AG007004‐23 (WLK), HL093238‐04 (LMA).
NanoSOD reduces adipose tissue inflammation in obesity (1107.10)
Authors: Saraswathi Viswanathan , Murali Ganesan , Desouza Cyrus , Manickam Devika , Bronich Tatiana , Kabanov Alexander
Published: June 2021

Obesity‐linked hypertension is resistant to antihypertensive drugs and, adipose tissue (AT) inflammation is considered to mediate the pathological consequences of obesity. The objective of this study is to determine the effectiveness of nanoformulated copper/zinc superoxide dismutase (nanoSOD) in reducing AT inflammation in obesity. Wild type mice were fed a high fat (HF) diet (45% fat) for 8 or 13 wk followed by 2 wk treatment with native or nanoSOD. The drug uptake studies showed that nanoSOD was accumulated in the visceral AT to a greater extent than native SOD in obese mice. The mice fed a HF diet for 8 wk followed by 2 wk treatment with nanoSOD showed an increase in SOD content in stromal vascular cells (SVCs) collected from AT and in the plasma. These mice also showed a reduction in nitrotyrosine level, a marker of superoxide induced oxidative stress, in SVCs. The AT inflammatory markers showed a trend towards a decrease. The serum toxicity markers did not alter in nanoSOD compared to other groups suggesting that the nanoSOD is safe to use in vivo. After 13 wk on a HF diet followed by 2 wk treatment, the SVCs derived from AT of nanoSOD treated mice showed a decrease in the mRNA levels of MCP‐1 and MIP‐1α. Also, the MCP‐1 mRNA and protein levels in total AT was reduced to a greater extent in nanoSOD compared to native SOD treated mice. Our data suggest that nanoSOD may be a potential therapeutic agent to ameliorate obesity‐linked cardiovascular risk factors, in particular, systemic hypertention.

Grant Funding Source: National Institutes of Health (NIH‐NIGMS)
Expression of the glycine transporter type 1 (GlyT‐1A) is upregulated by ATF‐4 following physiological stress in human intestinal epithelial cells (1109.14)
Authors: Fultang Livingstone , Howard Alison , Hirst Barry
Published: June 2021

Downstream of activating transcription factor 4 (ATF‐4) in Caco‐2 cells in response to stress. In the present study we show that increased expression of the GlyT‐1a gene following stress is dependent on elements contained within the 5’untranslated region (UTR). Caco‐2 cells transfected with plasmid constructs containing sequences representative of the GlyT‐1a proximal promoter and 5’UTR upstream of a beta‐galactosidase reporter showed increased reporter activity following treatment with thapsigargin (Tg), tunicamycin (Tu), amino acid (AA) starvation, tertbutylhydroquinone(tBHQ) or Diethyl maleate (DEM). Short labeled DNA probes containing a potential amino acid response element (AARE) located in exon 1 of the GLYT‐1a gene demonstrated binding by ATF‐4 in gel shift and super shift assays. qPCR assays performed on short genomic DNA fragments isolated from Caco‐2 cells by chromatin immunoprecipitation (ChIP) with antibodies against ATF4 demonstrated 9, 5 and 2‐fold enrichment of the ATF‐4/AARE interaction at the 5’‐UTR region of GlyT1a following Tu, AA starvation and DEM treatment respectively. Our data suggests that response elements within the 5’UTR including exon 1 and the first 85 bases of exon 2 are required for transcriptional induction of GlyT1a. We propose the direct interaction of ATF‐4 at the identified AARE drives transcriptional up‐regulation of GlyT‐1a following nutrient, oxidative and endoplasmic reticulum stress.

Grant Funding Source: University of Newcastle
Epigenetic impact of curcumin‐primed exosomes in amelioration of blood‐brain barrier dysfunction (1120.9)
Authors: Kalani Anuradha , Kamat Pardip , Familtseva Anastasia , Tyagi Suresh , Tyagi Neetu
Published: June 2021

Exosomes are nano‐units (<200nm), released from diverse cell types in extracellular body fluids. Since exosomes carry biological information from their cells of origin, the idea is to prepare improved molecular conservatory for therapeutics by priming the cells to release improved cellular contents through exosomes. Curcumin provides a promising treatment for cerebral diseases and therefore, the aim of the study is to establish that mouse brain endothelial cells when primed with curcumin (7.5µM), release alleviated exosomal population to recover blood‐brain barrier (BBB) permeability. Homocysteine‐treated endothelial cells were used as a model of BBB disruption and curcumin‐primed (CU‐primed) exosomes were utilized to check BBB disruption along with associated epigenetic measurements. We determined that CU‐primed exosomes reprogrammed endothelial cells, in part, by alleviating epigenetic alterations and improving junction proteins that ameliorate vascular permeability. These results pave the way for the novel translational therapies for cerebral diseases by maintaining and establishing therapeutic conservatories via primed exosomes. Additionally, the shortcomings of the gene therapy can be overcome by using these potentially primed exosomes containing the effector molecules.

Grant Funding Source: HL107640
Analysis of cerebrospinal fluid flow in hydrocephalic mouse models of primary ciliary dyskinesia (1125.1)
Authors: Johnson Katherine , Finn Rozzy , Lee Lance
Published: June 2021

Patients with primary ciliary dyskinesia (PCD) suffer from a variety of disorders that include chronic sinusitis, otitis media, infertility, situs inversus, and hydrocephalus. We have shown that two mouse models of PCD, nm1054 and bgh, have hydrocephalus characterized by dilatation of the lateral ventricles. These models also have male infertility and sinus abnormalities that result from a decrease in respiratory ciliary beat frequency. We therefore hypothesize that the hydrocephalus results from dysfunction of ependymal cilia that prevents proper cerebrospinal fluid (CSF) flow. To investigate CSF flow in vivo, we injected India ink into the lateral ventricles of live mice and histologically demonstrated that ink migrated through the ventricular system of wild type mice but failed to progress past the cerebral aqueduct of either mutant. To validate that this defect was directly due to ciliary dysfunction, we used an ex vivo method to measure ink flow rate over the ependymal cells on dissected ventricular walls. This method demonstrated a decreased flow rate and altered directional flow in the mutant brains, confirming that ependymal ciliary function is required for proper CSF flow.

Grant Funding Source: Supported by the NIH NIGMS (1P20GM103620‐01A1) and an NSF REU (1262744)
The role of TNF‐α in femoral artery occlusion‐augmented autonomic responsiveness (1132.6)
Authors: Lu Jian , Xing Jihong , Li Jianhua
Published: June 2021

Prior studies demonstrated that metabolically sensitive receptors in thin‐fiber muscle afferents contribute to the amplified responses of sympathetic nerve activity and arterial blood pressure observed in rats with femoral artery occlusion (24‐72 hrs). The purpose of this study was to examine the role played by TNF‐α in augmented sympathetic responsiveness induced by stimulation of muscle metabolic receptors following 72 hrs of femoral artery occlusion. First, the results of this study show that the levels of TNF‐α are increased in dorsal root ganglion (DRG) neurons of limbs with 72 hrs of femoral artery occlusion. Second, protein expression of TNF‐α is dominantly observed within DRG neuron supply C‐fiber muscle afferent nerves and TNF‐α receptor type 1 (TNFR1) is largely identified in DRG compared with TNFR2. Third, TNF‐α synthesis suppressor pentoxifylline (PTX) administered previously into the hindlimb muscles significantly attenuates sympathetic and blood pressure responses induced by arterial injection of capsaicin into the hindlimb muscles. Capsaicin was applied to stimulate metabolically sensitive receptors in thin‐fiber muscle afferents nerves. Overall, our data suggest that TNF‐α plays a role in modulating augmented sympathetic responsiveness via metabolic receptors of thin fiber muscle afferent nerves when arterial blood supply to the hindlimb is deficient under ischemic conditions.

Grant Funding Source: NIH P01 HL096570 and AHA EIA 0840130N
PDGF receptor antagonism prevents the increase in kidney ANG II levels in Cyp1a1‐Ren2 transgenic rats with ANG II‐dependent malignant hypertension (1136.1)
Authors: Thompson David , Seth Dale , Davis Porcha , Mitchell Kenneth
Published: June 2021

This study was performed to determine if chronic PDGF receptor blockade prevents the augmentation of intrarenal ANG II levels in Cyp1a1‐Ren2 transgenic rats with ANG II‐dependent malignant hypertension. Male Cyp1a1‐Ren2 rats (n=5/group) were induced to develop malignant hypertension by dietary administration of indole‐3‐carbinol (I3C, 0.3% wt/wt) for 10 days. Systolic blood pressures (SBP) were measured daily by tail‐cuff plethysmography. On day 10 of I3C administration, rats were subjected to conscious decapitation and trunk blood was collected for measurement of plasma renin activity (PRA) and plasma ANG II levels. The kidneys were harvested for determination of total kidney ANG II levels. Dietary I3C resulted in increases in SBP (130±5 to 171±9 mmHg, P<0.05), PRA (64±6 vs. 5±1 ng ANG I/ml/hr, P<0.001), plasma ANG II levels (186±26 vs. 139±10 fmol/ml, P<0.05), and total kidney ANG II content (550±31 vs. 253±24 fmol/g, P<0.001). Chronic imatinib administration (60 mg/kg/day, oral gavage) did not prevent the development of hypertension (187±10 vs. 141±2 mmHg, P<0.001) or the increase in PRA (72±6 vs. 5±1 ng ANG I/ml/hr, P<0.001), but prevented the I3C‐induced augmentation of intrarenal ANG II levels (285±128 vs. 253±24 fmol/g). The present findings indicate that PDGF receptor activation contributes to the increase in the elevations of intrarenal ANG II levels in ANG II‐dependent malignant hypertension.

Grant Funding Source: 5P30GM103337
AT2 receptor protects high sodium‐induced decrease in ACE2/Mas receptor expression and increase in blood pressure in obese rats (1136.8)
Authors: Ali Quaisar , Hussain Tahir
Published: June 2021

High sodium (Na) intake is a risk factor for the pathogenesis of hypertension, especially in obesity. Whereas high Na diet increases blood pressure (BP) associated with a decrease in renal MasR/ACE2 expression, chronic AT2R stimulation lowers BP concomitant with an increase in MasR/ACE2 expression in obese Zucker rats. However, role of AT2R in high Na‐induced hypertension and regulation of ACE2/MasR expression in obesity is not known. Male obese rats were treated with AT2R agonist C21 (1mg/kg/day oral) while maintained on either normal (NSD, 0.4%) or high Na diet (HSD, 4%) for 2‐weeks. Telemetry monitoring of BP showed an increase in systolic BP (SBP) by 20 mm Hg in obese HSD compared to NSD group; which was prevented by C21 treatment. Compared to control, HSD caused diuresis which was further increased in HSD+C21 group. The rats on HSD had increase in body and kidney weight; which was lowered by C21 treatment. HSD reduced the cortical MasR and ACE2 activity which was increased by C21 treatment. AT2R, ACE expression and renin activity were same in all the groups. However AT1R, although unchanged with HSD or C21 treatment, was decreased in HSD+C21 group. This study clearly demonstrate that chronic AT2R agonist treatment prevents high Na‐induced increase in BP which was associated with an increase in the beneficial ACE2/MasR axis in obese rats indicating the potential role of AT2R against Na‐sensitive hypertension in obesity.

Grant Funding Source: NIH R01‐DK61578
Intronic mutations in GLA causes Fabry disease (1138.5)
Authors: Ortolano Saida , Vieitez Irene , Souto Olga , Garcia‐Jimenez Concepcion , Pintos Guillem , Navarro Carmen
Published: June 2021

INTRODUCTION: Fabry disease (FD) is a rare X‐linked disorder, due to a defect of lysosomal α‐Galactosidase A (GalA), which determines cellular storage of globotryalosil ceramide (Gb3), eventually causing multysistemic dysfunctions. Male patients are diagnosed by enzyme activity assay and female patients by genetic study. AIMS: To characterize molecular defects in a family of patients with unconfirmed diagnosis of FD. PATIENTS & METHODS: Male with impaired GalA activity and his 7 years old daughter, presenting FD symptoms. Patients carry genetic defects in GLA up to date validated as polymorphisms (rs207122, rs2071397). RT‐PCR was performed to asses GLA transcripts levels in patient's derived leukocytes. GalA and Gb3 expression were evaluated by immunofluorescence in daughter's skin biopsy. RESULTS: GLA transcript levels are significantly reduced in both patients. Immunofluorescence of skin biopsy revealed lysosomal inclusions of Gb3 mainly in dermal fibroblasts, while GalA present lower expression, compared to a control biopsy. CONCLUSIONS: The association of the intronic mutations rs207122, rs2071397 affects transcription of GLA and causes Gb3 accumulation, determining FD phenotype.

Grant Funding Source: Instituto Carlos III Spain
Cytotoxicity of peracetic acid in L929 fibroblasts (1139.2)
Authors: Faria Gisele , Viola Kennia , Carlos Iracilda , Kuga Milton , Guerreiro‐Tanomau Juliane , Tanomaru‐Filho Mario
Published: June 2021

Recently peracetic acid (PA) has been reported in the dental literature as a possible root canal irrigating solution. It has the advantage of bringing together in one product antibacterial activity and ability to remove smear layer. For selection of root canal irrigating solution it should take into account not only its therapeutic benefits, but also the possible cytotoxic effects, since it can contact the periapical tissues. The aim of this study was to evaluate the cytotoxicity of PA at different concentrations in comparison with 2.5% sodium hypochlorite (NaOCl) through the study of the L929 fibroblasts metabolism and the cytoskeleton. L929 fibroblasts in culture were exposed to different concentrations of PA and NaOCl and to the culture medium for 24 hours (control). After this period, it was evaluated the cell metabolism by methyl tetrazolium (MTT) assay and the cytoskeleton by staining actin and α‐tubulin. Data were analyzed by two‐way ANOVA and Bonferroni post test (α = 0.05). The solutions had an effect on cell viability/metabolism in a dose‐dependent manner. The cell viability decreased progressively as the concentration of irrigating solutions was increased. 0.5% AP showed similar cytotoxicity to 2.5% NaOCl (p>0.5); the AP at concentrations of 1%, 2% and 2.5% showed higher cytotoxicity than 2.5% NaOCl (p<0.5). The cytoskeleton suffered progressive structural changes as the concentrations of the solutions were increased. It was concluded that the AP is more cytotoxic than NaOCl. However, further studies evaluating other cell parameters and employing other PA‐based solutions need to be performed.

Grant Funding Source: Supported by São Paulo Research Foundation (FAPESP) #2012/18203‐2
The potential of N‐methylpyrrolidone to prevent osteoporosis and enhance bone regeneration in vivo (1139.5)
Authors: Gjoksi Bebeka , Ghayor Chafik , Weber Franz
Published: June 2021

Osteoporosis is a chronic, skeletal disease highly prevalent in post‐menopausal women influenced by hormonal factors causing a huge burden on health care in an aging society. We previously demonstrated that NMP is a bioactive drug which enhances bone regeneration in vivo and acts as an enhancer of bone morphogenetic protein (BMP) in vitro. NMP also inhibits osteoclast differentiation and attenuates bone resorption. The present study used animal models established for the evaluation of enhanced osteoporosis through double ovariectomy (OVX) and treated systemically with different doses of (NMP). With this treatment we want to describe the preventive effect of NMP on bone mass loss.

Female Sprague‐Dawley rats with an approximate weight of 220‐250g were randomly divided into sham‐operated group (Sham) either treated or not with NMP and four ovariectomized subgroups as OVX (control), OVX treated with three different graded doses of NMP. Bilateral ovariectomy or Sham operations were performed as previously designated and the weight of the animals was measured weekly. We assessed the pharmacological effects of NMP against osteoporosis by evaluating the body weight, serum biochemical parameters, bone mineral density (BMD), dynamic histomorphometry of bone and bone histomorphology.

The results showed that over a 15 week period, in OVX rats the increase in body weight, serum osteocalcin levels and decreases of BMD were significantly reversed by NMP treatment. Additionally, the effect of the three different treatment dosages of NMP correlated dose‐dependently with their effect on bone resorption. Mineral apposition rate (MAR) showed the protective activity of NMP through promotion of bone formation and suppression of bone resorption.

These results suggest that NMP has a remarkable antiosteoporotic activity, and may be a promising candidate for treatment of postmenopausal osteoporosis induced by estrogen deficiency.

Grant Funding Source: Swiss National Science Foundation
Expression of vitamin D receptor and 1α‐hydroxylase in epicardial fat: association to plasma 25‐hydroxyvitamin D level in coronary artery disease patients (1139.8)
Authors: Corsi Romanelli Massimiliano , Vianello Elena , Briganti Silvia , Malavazos Alexis , Dozio Elena
Published: June 2021

25‐hydroxyvitamin D (25OHD) is the main circulating vitamin D form and 1α‐hydroxylase (CYP27B1) is the key enzyme which activates 25OHD in tissues. In adipose tissue, reduced vitamin D and increased vitamin D receptor (VDR) levels have been associated to fat accumulation and inflammation.

Alterations in epicardial fat (EAT) biology (i.e. increased fat thickness and inflammation) have been described in patients with coronary artery disease (CAD) but it is unknown whether low plasma level of 25OHD and/or inadequate levels of VDR and CYP27B1 at EAT level may contribute to these EAT functional alterations.

Our aim was to evaluate in CAD patients the relationship between plasma 25OHD level and gene expression of VDR and CYP27B1 at EAT level.

Plasma 25OHD was quantified by an immunoluminometric assay. Gene expression was performed by microarray.

CAD patients had low 25OHD level. After patient stratification according to 25OHD level, we observed increased expression of VDR and reduced expression of CYP27B1 at EAT level only in patients with 25OHD deficiency (< 20 ng/mL) and insufficiency (20‐30 ng/mL).

We may conclude that in CAD patients, EAT tries to compensate reduced plasma 25OHD level by increasing VDR expression. Anyway, due to the impairment in 25OHD activation, the local vitamin availability seems to be reduced. How this may contribute to EAT functional alterations observed in CAD needs further investigation.

Grant Funding Source: Supported by DiaSorin S.P.A., Vercelli, Italy
Stress‐induced depression‐ and anxiety‐like behaviors are associated with the imbalance of redox state: the protective effect of PDE2 inhibition (1144.3)
Authors: Xu Ying , Ruan Lina , Zhang Hanting , O'Donnell James
Published: June 2021

The correspondence between the prevalence of exposure to oxidative stress and the development of depression/anxiety is lacking. Chronic stress alters the expression of genes affecting redox state, such as excess levels of reactive oxygen species (ROS) and reduced superoxide dismutase (SOD) expression, which induce functional abnormalities in the brain. Phosphodiesterase 2 (PDE2) is highly expressed in the limbic system, which may play an important role in both the development and the treatment of stress‐related depression. The present study investigated the relationship between PDE2 activity and the ROS expression after stress and how PDE2 inhibitor and PDE2 silence regulate stress‐induced depression/anxiety‐like behaviors through affecting redox state. The results suggested that corticosterone‐induced ROS expression was positively related to PDE2 levels, which was consistent with in vivo data. Pretreatment with protein kinase G inhibitor KT5823, but not protein kinase A inhibitor H89, reversed the effect of PDE2 inhibition, suggesting the involvement of cGMP‐dependent signaling. This study provides the evidence that PDE2 inhibition may represent a novel therapeutic target for stress‐related psychiatric disorders.
The mechanism of amphetamine‐mediated trafficking of the dopamine transporter (1145.1)
Authors: Underhill Suzanne , Thiels Edda , Amara Susan
Published: June 2021

Acute amphetamine (AMPH) exposure elevates extracellular dopamine by a variety of mechanisms including increasing the rate of internalization of the plasma membrane dopamine transporter (DAT). We report here that AMPH also activates the small GTPases, Rho and Rac. In recombinant expression systems, primary cultures and midbrain slices, activation of Rho and the downstream Rho‐kinase (ROCK) triggers endocytosis of DAT. Inhibition of RhoA activity blocks AMPH‐induced DAT internalization, while Rac1 inhibition has no significant effect on DAT trafficking. ROCK inhibitors also prevent AMPH‐induced DAT internalization. Intriguingly, we found that AMPH must enter the cell to initiate this cascade. Further, intracellular AMPH also increases cAMP, leading to the phosphorylation of Rho by protein kinase A (PKA) that inactivates Rho and serves as a break on DAT internalization, thus demonstrating an important interaction between PKA‐ and RhoA‐dependent signaling in mediating the actions of AMPH. In agreement with our observations in cell lines and brain slices, we have also shown that the activation of receptors that couple to PKA in dopamine neurons within the brain can antagonize the behavioral effects of AMPH in mice. These observations imply the existence of a novel intracellular target that mediates the effects of AMPH on Rho and cAMP signaling and suggest new pathways to target in order to disrupt AMPH action.

Grant Funding Source: Supported by the National Institute of Mental Health Intramural Program.
Microarray approach in experimental hyperthyroidism reveals two novel responsive genes in the rat heart (1148.8)
Authors: Moriscot Anselmo , Rosanski Andrei , Takano Ana Paula , Garcia Antonio , Campos Juliane , Ferreira Julio , Barreto‐Chaves Maria Luiza
Published: June 2021

In order to get further insight upon the effects of Thyroid hormone (T3) in the heart, we have used a microarray approach to identify responsive genes in rats submitted to T3 injections for 12, 24h and 7 days. Firstly, we have compared 5 different algorithms to analyze microarray data and found MAS5 to be the best fit in our model. We have detected novel responsive genes at the intercalated disk and sarcomere M‐band levels. qRT‐PCR analysis and western blot confirmed a transient increase in cadherin‐13 expression (2.7 fold for qRT‐PCR and 2.4 for Western Blot, both p<0.05) peaking at 24 hours of T3 treatment and a progressive decrease of M‐protein gene expression (0.2 fold, p <0.05, at 7 days of T3 treatment). Because M‐protein is a structural sarcomeric protein and has a predicted role in sarcomere stability we decided to further investigate cellular/molecular mechanisms. M‐protein is up regulated in hypothyroidism (3.4 fold) and repressed by T3 in a dose dependent manner in vivo and also in cultured neonatal cardiomyocytes. Reporter assays revealed that the M‐protein promoter is responsive to T3. siRNA M‐protein knock down showed reduced contractility, highlighting the importance of this protein for sarcomere stability. We conclude that T3 quickly and strongly repress M‐protein at the transcriptional level, which might help to explain the long‐term deleterious effects of T3 in the heart.

Grant Funding Source: FAPESP and CNPq
Characterization of cardiovascular performance during ischemia‐reperfusion injury in obese vs. lean Ossabaw swine (1150.4)
Authors: Sassoon Daniel , Goodwill Adam , Conteh Abass , Noblet Jillian , Mather Kieren , Tune Johnathan
Published: June 2021

This study examined the effects of obesity/metabolic syndrome (MetS) on left ventricular pressure‐volume relations at baseline and in response to ischemia‐reperfusion injury. Measurements were obtained in lean and obese Ossabaw swine at rest, following 30 min occlusion of the left circumflex artery, and 2 hours after the release of the occlusion. Baseline end‐diastolic volume (83±5 ml vs. 74±5 ml) and cardiac output (2215±389 ml/min vs. 2511±389 ml/min) were not significantly different in lean vs. MetS swine. However, mean arterial pressure was ~40% higher in MetS swine at rest and during reperfusion, while heart rate increased ~15‐20 beats/min with ischemia‐reperfusion in both groups. Assessment of inotropic status revealed that both cardiac output and blood pressure were augmented relative to end‐diastolic filling volumes in MetS vs. lean swine under all conditions (p<0.001). These apparent differences in cardiac function were associated with a marked reduction in the time to ischemia‐induced ventricular fibrillation in lean (17.5±7 min; n = 3 of 6) vs. MetS (3.3±1 min; n = 4 of 6) swine. These findings suggest that increases in blood pressure and cardiac function in MetS, at rest and following ischemia‐reperfusion injury, are independent of changes in filling volume and could contribute to increased susceptibility to fatal ventricular arrhythmias.

Grant Funding Source: Supported by HL117620; This publication was made possible in part by Grant Number TL1 TR000162 (A. S
Syndecan‐4 promotes myocardial stiffness by regulating collagen expression and cross‐linking in response to pressure overload (1152.2)
Authors: Herum Kate , Lunde Ida , Skrbic Biljana , Tønnessen Theis , Louch William , Hasic Almira , Sjaastad Ivar , Linke Wolfgang , Gomez Maria , Christensen Geir
Published: June 2021

Pressure overload of the heart leads to myocardial stiffening due to excessive production and cross‐linking of collagen, thereby compromising diastolic function. We recently identified the focal adhesion proteoglycan syndecan‐4 (syn4) as important for collagen expression in response to mechanical stress. Here we investigate the effect of syn4 deletion on the mechanical properties of the left ventricle following pressure overload. Passive tension increased in skinned muscle fiber bundles from wild‐type (WT) mice following aortic banding and this response was significantly blunted in syn4‐/‐ mice. Total collagen content determined by hydroxyproline quantification was increased in WT but not in syn4‐/‐ mice. Furthermore, mRNA levels of the collagen cross‐linking enzyme lysyl oxidase (LOX) were dramatically increased in WT left ventricles and mechanically stressed cardiac fibroblasts, whereas this response was blunted in syn4‐/‐ mice and cardiac fibroblasts, suggesting impaired collagen cross‐linking following mechanical stress in mice lacking syn4. Finally, recombinant extracellular syn4 and collagen I formed a precipitate when mixed in vitro, suggesting a direct role for syndecan‐4 in collagen fiber organization. In conclusion, we demonstrate reduced passive tension in left ventricular tissue of syn4‐/‐ mice possibly due to reduced ECM production and attenuated collagen cross‐linking.

Grant Funding Source: Supported by the Norwegian Health Association and the Norwegian Research Council
Inhibition of lysyl oxidase activity reverses fibrosis and improves cardiac function (1152.8)
Authors: El Hajj Milad , El Hajj Elia , Bradley Jessica , Gardner Jason
Published: June 2021

The collagen crosslinking enzyme, lysyl oxidase (LOX), is elevated in human dilated cardiomyopathy and heart failure, but it is not known if LOX plays a causative role in disease. However, clinical evidence shows that diuretics with LOX inhibitory properties are more effective at slowing the progression of heart failure. Using the aortocaval fistula (ACF) rat model of volume overload, we assessed the efficacy of LOX inhibition to reverse cardiac fibrosis and dysfunction in rodents with established cardiac disease. Serial echocardiography was used to evaluate cardiac structural and functional changes. LOX inhibition (100 mg/kg/d BAPN; osmotic minipumps) was initiated 8 wks post‐surgery in ACF and SHAM‐operated rats. Treatment continued for 6 wks and hearts were collected for analysis. Left ventricular (LV) sections were stained with PSR for collagen volume fraction (CVF). Protein expression was assessed in LV homogenates by WB. Untreated ACF had significantly increased LV LOX expression (65% increase), concomitant with LV dilatation (43% increase), depressed fractional shortening (26% decrease), increased collagen cross‐linking (52% increase), and interstitial fibrosis (CVF; 135% increase) relative to SHAM (p<0.05). LOX inhibition reversed fibrosis (CVF; Treated ACF vs SHAM; p=NS) and partially restored cardiac function (20% increase vs Untreated ACF). However, there was no significant reversal of LV hypertrophy or dilatation. LV NADPH oxidase‐4 was increased in untreated ACF (52% increase vs SHAM), but was not different in treated ACF, indicating a possible role for oxidative stress in the observed beneficial effects of LOX inhibition. These data identify inhibition of LOX activity as a potential therapeutic approach for the regression of cardiac fibrosis and improvement of function in heart failure patients.

Grant Funding Source: Supported by LSUSOM REF grant (jdg) and the American Heart Association GSA 11GRNT7700002 (jdg)