bluuebunny/crossref_metadata_2025 huggingface dataset:
Autonomic and metabolic changes induced by fructose overload: role of baroreflex (1086.4)
Authors: Santos Fernando , Moreira Edson , Sartori Michelle , Irigoyen Maria
Published: June 2021

The aim of this experiment was to identify the changes in autonomic and metabolic parameters in a model of metabolic syndrome induced by fructose overload, with focus on the role of baroreflex, as evaluated by sinoaortic denervation. Four experimental groups (rats) were used, followed by 90 days: control(C), denervated control (D), treated with fructose(F) and denervated treated with fructose(DF). The treated groups were submitted to a fructose overload(10%) in drinking water. The caloric intake was higher in F(32%) and DF(18%) groups compared to C, inducing higher body weight gain in F(41%) and DF(27%) when compared to C group. The consumption of D was lower (12%) than the C group as well as body weight gain(8%). The overload of fructose was able to change the fasting plasma glucose baselines values, promoting increasing of glycaemia in both F and DF groups and leading to insulin resistance in the F group. Blood pressure values were also increased in F(17%) and DF(18%) compared to C group. The variability parameters were altered in the three experimental groups (D, F and DF), evidenced by the reduction of the pulse interval variance. Furthermore, the vagal modulation indicators are decreased and blood pressure variability was increased, as expressed by the total variance, in both denervated groups (D and DF). Treatment with fructose was able to induce characteristic changes of metabolic syndrome, such as hypertension, obesity, increased blood sugar and dysautonomia. The absence of the baroreflex was able to mitigate metabolic changes, but when associated with fructose overload caused a greater autonomic dysfunction. The results of the present study indicate a marked influence of baroreflex in the development of metabolic syndrome.

Grant Funding Source: Supported by FAPESP 2011/04111‐6
Salt intake during pregnancy influences offspring organ growth (1087.2)
Authors: Peres Maria , de Oliveira Ivone , Furukawa Luzia , Guimarães Carina , Heimann Joel
Published: June 2021

Introduction: Industrialized food consumption is responsible for high salt ingestion, mainly in western countries. Circulating and tissue renin‐angiotensin system (RAS) activity is influenced by the degree of salt intake. RAS is responsible for blood pressure regulation and hydroelectrolitic balance. High salt intake during gestation may influence the health of adult offspring. As a working hypothesis, the alterations observed in generation F1 may be transmitted to generations F2 and F3. Therefore, in this study some effects of high salt diet during gestation were evaluated in adult male rats of generation F1.

Material and Methods: Fifteen adult male Wistar rats, born from mothers fed normal (N group, n=9) or high salt diet (H group, n=6) during the whole gestation, were followed from birth until 15 weeks of age. Body and organs weight (heart, adrenals, kidneys, testicles, epididimes, and retroperitoneal fat), urinary sodium and potassium and creatinine excretion were measured. Daily sperm production and sperm transit time in the epididymal cauda were evaluated by counting homogenization‐resistant testicular spermatids or epididymal spermatozoa, respectively.

Results: (mean±SEM)

Parameter
N
H
p value

Body weight (g)
458.78±25.93
438.08±20.91
0.128

Retroperitoneal fat (mg/g)
5.03±2.92
8.24±1.52
0.029

Left testis/body mass (mg/g)
3.50±0.31
3.92±0.30
0.022

Right kidney/body mass (mg/g)
3.21±0.12
3.59±0.17
<0.001

Left kidney/body mass(mg/g)
3.34±0.21
3.60±0.18
0.026

Heart/body mass (mg/g)
3.38±0.23
3.79±0.40
0.024

Sperm production (x106/g/day)
13.05±1.92
12.51±2.65
0.648

Sperm transit time (days)
7.93±0.39
6.99±0.99
0.331

Discussion: Damages on cardiovascular and renal systems caused by high salt intake are well documented and molecular mechanisms are still been studied. Reproductive alterations have been reported, regarding both male and female performance. Our results disclosed higher kidney, heart, testicles and adipose tissue mass in the first generation born from mothers fed high salt diet uring gestation indicating a possible disturbance in organ growth. The difference in testicular mass between the experimental groups was not accompanied by functional alterations.

Grant Funding Source: Support by FAPESP (2012/22415‐5, 2012/22801‐2)
Acute hypoxia‐induced endothelial‐dependent suppression of Ca2+ waves in pulmonary arterial myocytes of sheep (1089.14)
Authors: Sandy Scott , Romero Monica , Paez Ricardo , Francis Michael , Taylor Mark , Longo Lawerence , Wilson Sean
Published: June 2021

Ca2+ responses are important to acute hypoxia‐induced pulmonary arterial contraction. To better understand these signals and the role of functional endothelium we performed confocal imaging of Fluo‐4 loaded pulmonary arterial myocytes of adult sheep using the en‐face preparation. The present studies tested the hypothesis that functional endothelium suppresses resting as well as acute hypoxia (~4%) dependent Ca2+ waves in pulmonary arterial myocytes by examining endothelium intact and denuded arteries. Automated analyses were performed to determine the magnitude and kinetics of the oscillatory Ca2+ responses. Endothelium removal decreased the amplitude and area under the curve (AUC) of Ca2+ waves but did not significantly impact the duration, or the rate of rise and fall. Acute hypoxia in endothelium intact arteries suppressed the Ca2+ amplitude, AUC, as well as the duration, and rate of rise and fall. However, endothelium removal prevented these acute hypoxia‐induced reductions in the magnitude and kinetics of the Ca2+ responses. These data illustrate that functional endothelium suppresses pulmonary arterial myocyte Ca2+ responses in resting conditions and in response to acute hypoxia, an effect that could influence either excitation‐contraction or excitation‐transcription coupling.

Grant Funding Source: Supported by MRI923559, NIH R03HD69746, R01HD31226, R01HD3807, P20MD6988 and LLUSOM
ASIC1‐dependent Ca2+ influx is required for NFATc3 activation in chronic hypoxia‐induced pulmonary hypertension (1090.12)
Authors: Jernigan Nikki , Plomaritas Danielle , Giermakowska Wieslawa , Herbert Lindsay , Nitta Carlos , Gonzalez Bosc Laura
Published: June 2021

The development of chronic hypoxia (CH)‐induced pulmonary hypertension is associated with 1) the activation of the Ca2+‐dependent transcription factor, nuclear factor of activated T‐cells isoform c3 (NFATc3), and 2) enhanced acid sensing ion channel 1 (ASIC1)‐dependent Ca2+ influx in pulmonary arterial smooth muscle cells (PASMC). Therefore, we hypothesize that ASIC1‐mediated Ca2+ influx increases NFATc3 nuclear translocation in pulmonary vascular smooth muscle. To test this hypothesis we first examined the effect of ASIC1 inhibition on NFATc3 nuclear translocation in PASMC in response to endothelin‐1 (ET‐1; 10‐7 M). Inhibition of ASIC1 with psalmotoxin‐1 (20 nM) diminished ET‐1 induced Ca2+ influx and NFATc3 nuclear import. In addition, ET‐1 mediated pulmonary arterial vasoconstriction and Ca2+ influx following CH (1 wk @ 380 mmHg) were blunted in ASIC1 knockout mice (ASIC1‐/‐) compared to the ASIC1 wildtype mice (ASIC1+/+). Similarly, CH increased pulmonary vascular smooth muscle NFATc3 nuclear translocation in ASIC1+/+ mice, but not in ASIC1‐/‐ mice. These CH‐induced responses were additionally associated with decreased right ventricular systolic pressure and right ventricular hypertrophy in ASIC1‐/‐ mice. Taken together, these data suggest that ASIC1‐dependent Ca2+ influx is required for the activation and nuclear translocation of NFATc3 during the development of pulmonary hypertension.

Grant Funding Source: Supported by R01 HL92598, R01 HL111084, T32 HL07736, R01HL088151, R01 HL088192
Microparticles stimulated by hypoxic pulmonary hypertension have differing effects on vascular cell growth than microparticles from severe occlusive pulmonary arterial hypertension (1090.7)
Authors: Yarbrough Thomas , Bauer Natalie
Published: June 2021

The release of microparticles (MPs), intact, sub‐micron vesicles containing signaling molecules such as proteins, lipids, mRNAs, and miRNAs, is significantly increased in pulmonary arterial hypertension (PAH). MPs contribute to vascular cell proliferation, and the mechanism of increased proliferation in the arteriopathy of PAH is still unclear. Thus, we propose that circulating MPs from pulmonary hypertensive rats will stimulate pulmonary microvascular endothelial (PMVEC) and/or pulmonary artery smooth muscle cell (PASMC) proliferation. PMVECs and PASMCs were treated with MPs from controls, 3 wk hypoxia only (Hx) rats, or 3 wk Su/Hx/Nx rats. Interestingly, although the Su/Hx/Nx model shows severe vascular remodeling, MPs from the hypoxic rat had the most significant effects on PASMC proliferation by shifting the log phase of growth. Western blot analysis shows corresponding changes in smooth muscle cell markers. PMVECs were treated similarly with 5 wk MPs from the Su/Hx/Nx progressive model, resulting in a shift of log growth phase. Our data suggest that circulating MPs isolated from 3 wk Hx‐treated rats may signal to PASMCs to increase proliferation, whereas MPs from a severe occlusive model of PAH stimulate changes in proliferation rates of PMVECs. Combined these data suggest that hypoxic pulmonary hypertension and severe occlusive PAH result in different signaling MPs with unique vascular cell effects.

Grant Funding Source: Supported by NHLBI T32 HL076125 to TMY and AHA 11SDG7390037 to NNB.
Ventilatory chemoreflex sensitivity in sleep apnea is decreased with high dose ascorbic acid (1092.13)
Authors: Leuenberger Urs , Aswegen Caleb , Blaha Cheryl , Mast Jessica
Published: June 2021

Obstructive sleep apnea (OSA) is associated with increased sympathetic activity, hypertension and augmented activity of the arterial chemoreflex. To test the hypothesis that oxidative stress, presumably due to intermittent hypoxia, contributes to these abnormalities, we determined blood pressure (BP), heart rate (HR), muscle sympathetic nerve activity (MSNA, peroneal microneurography), minute ventilation (MV) and the responses to 2‐8 breaths of nitrogen (N2, 10 trials aiming for O2 saturation nadirs of 70‐90% [SpO2, oximetry]) before and after infusion of the antioxidant ascorbic acid (AA, 45 mg/kg over 20 min, followed by 15 mg/kg over 1 hr) in 8 patients with OSA (age 49±4 yrs, 7 male/1 female, apnea‐hypopnea‐index 42±10 events/hr). While AA did not affect BP (pre vs. post: 89±2 vs. 90±3 mmHg; P=NS), HR (pre vs. post: 67±5 vs. 66±4 beats/min; P=NS), MSNA (pre vs. post: 48±4 vs. 45±3 bursts/min; P=NS) and MV (pre vs. post: 11.1±0.8 vs. 11.6±0.9 L/min; P=NS), the ventilatory responses to N2 induced transient hypoxia (expressed as the slope of peak MV vs. SpO2 nadir) decreased from ‐0.372±0.059 pre‐AA to ‐0.288±0.058 L/min/%SpO2 post‐AA (P<0.05). Thus, ventilatory chemoreflex sensitivity which is thought to be enhanced in OSA decreases after AA. This suggests that the enhanced chemoreflex activity in patients with OSA is at least in part due to oxidative stress.

Grant Funding Source: Supported by P01 HL096570 and UL1 TR000127
The role of miR‐21 in propofol‐induced neurotoxicity in developing human neurons (1093.2)
Authors: Twaroski Danielle , Bai Xiaowen , Olson Jessica , Yan Yasheng , Liu Yanan , Bosnjak Zeljko
Published: June 2021

Recent studies in animal models have suggested that anesthetics such as propofol, when administered early in life, can lead to neurotoxicity. These studies have raised safety concerns regarding the use of anesthetics in the pediatric population. Human embryonic stem cells (hESCs) are capable of differentiating into any cell type and represent a promising model by which to study mechanisms governing anesthetic‐induced neurotoxicity in humans. In this study, we assessed the effects of propofol on hESC‐derived neurons and the role of microRNAs (miRs) in the toxicity observed.

hESCs were differentiated into neurons following a four‐step differentiation protocol. Cell death in the hESC‐derived neurons was assessed using TUNEL staining and microRNA expression was assessed using qRT‐PCR. miR‐21 was overexpressed using a miR‐21 mimic and the expression of the miR‐21 targets of interest was assessed by Western blot.

Exposure to 20 µg/mL propofol for 6 hours induced significant cell death in the hESC‐derived neurons and downregulated several microRNAs, including miR‐21. Overexpression of miR‐21 significantly attenuated the increase in cell death following propofol administration. In addition, several targets within the miR‐21 pathway (eg. Sprouty 2 and pSTAT3) were significantly altered by propofol exposure, suggesting that miR‐21 is contributing to the propofol‐induced neurotoxicity.

Grant Funding Source: Supported by P01GM066730 and R01HL034708 (NIH) and FP00003109 (Advancing Healthier Wisconsin) to ZJB
Bone marrow cells apoptosis in experimental anemia (1093.4)
Authors: Zaporozhets Tetiana , Sanyk Oleksandr
Published: June 2021

Environmental pollution consisting of toxic metals and ionizing radiation is a serious problem throughout the world. Our objective was to compare the level of markers (BCL‐ 2 and p53) of the bone marrow cells apoptosis in experimental gamma‐ irradiation and chronic lead intoxication. Methods: guinea pigs (n = 24) were divided into two groups. The first group of animals received a single dose of 4.5 Gr of gamma irradiation extracorporeal (aplastic anemia model). The second group received a 4% lead acetate solution for 60 mg/kg body weight per day for 27 days (anemia associated with violation of the synthesis of porphyrins). The level of Bcl‐ 2 and p53 was determined immunohistochemically. We observed:1) In the irradiated animals the number of cells with a marker protein Bcl‐ 2 was reduced to 45.23 % and the number of cells with p53 marker was increased to 112.8 % compared to control animals. 2) In animals with chronic lead intoxication the number of cells with a marker protein Bcl‐ 2 was decreased to 16.42 % and the number of cells with p53 marker was increased to 61.37 % compared to control animals. We conclude that apoptosis of bone marrow cells increases with aplastic anemia and anemia associated with violation of the synthesis of porfirins, which is a major pathogenetic mechanism of anemia. It may be one of the factors capable of causing deficiency of stem cells in this pathology.
TRPC channels are involved in endothelin 1 mediated inhibition of renin release from juxtaglomerular cells (1095.12)
Authors: Ortiz‐Capisano M. Cecilia
Published: June 2021

Renin is produced by, stored in and released by juxtaglomerular (JG) cells. Increases in intracellular calcium (Ca) decrease renin release. I’ve found that JG cells contain multiple isoforms of canonical transient receptor potential (TRPC) Ca‐permeable channels. Among those, TRPC1, 4 and 5, are known to function as store‐operated Ca channels. Endothelin (ET)1 binds to G‐coupled protein receptors whose activation leads to a phospholipase C (PLC)/ IP3 signaling pathway, potentially stimulating store‐operated Ca channels. I hypothesized that endothelin inhibits renin release by activating TRPC store‐operated Ca channels. RT‐ PCR revealed expression of both ETA and ETB receptors in freshly isolated mouse JG cells. Incubation of primary cultures of JG cells with ET1 (10nM) decreased renin release by 28% from 0.84±0.07 to 0.60±0.07 µg ANGI/ml/hr/mg prot. (p<0.001). An IP3 blocker (2APB, 0.1mM) returned renin release back to control. Addition of the TRPC blocker (SKF 96365, 50µM) completely reversed the Ca‐mediated inhibition of renin release by ET1 to 0.99±0.07 µg ANGI/ml/hr/mg prot. These results suggest that 1) JG cells express ETA and ETB receptors, 2) ET1 mediates inhibition of renin release in JG cells by activation of ETA or ETB receptors and subsequent PLC/IP3 signaling cascade activation, and 3) ET receptors link ET1 and TRPC store‐operated Ca channels to mediate ET1‐mediated Ca inhibition of renin release.

Grant Funding Source: NIH 5PO1HL090550‐03
Lipopolysaccharide‐induced inflammation is associated with receptor for advanced glycation end products in human endothelial cells (1096.5)
Authors: Wang Liqun , Chen Tangting , Wu Jianbo , Huang Qiaobing
Published: June 2021

The vascular endothelium is the center of events leading to the development of inflammatory lesions. The activation of endothelial cells is of significance in acute as well as chronic inflammatory states such as septic shock and atherosclerosis, respectively. Receptor for advanced glycation end products (RAGE) plays a role in immune reaction to recognize specific molecular patterns as pathogen recognition receptors. However, the function of RAGE in LPS‐induced inflammation is unclear. In this study, we investigated whether RAGE is enrolled in lipopolysaccharide (LPS)‐induced cell activation in human umbilical vein endothelial cells (HUVECs) using a combination of western blotting, confocal microscopy, and functional assays. We found that LPS can directly increase the expression of RAGE in a time‐ and concentration‐ dependent manner. And anti‐RAGE antibody inhibited LPS‐mediated barrier high‐permeability and apoptosis of HUVECs. Furthermore, LPS‐induced NF‐κB activation and IκBα degradation was also significantly attenuated by anti‐RAGE antibody. Given these results, RAGE plays an important role in LPS‐induced inflammation and could be a candidate as a therapeutic target for various systemic inflammatory diseases.

Grant Funding Source: AHA
Signal‐transduction pathways in hemolysis‐associated right‐ventricular failure (1097.12)
Authors: Olonisakin Tolani
Published: June 2021

The pulmonary arterial pressure at which the right ventricle fails in patients with sickle‐cell disease occurs at markedly lower pressures than in pulmonary hypertension not associated with hemolysis. The mechanism by which the right ventricle fails and the reason that dysfunction occurs at relatively low pulmonary arterial pressures in sickle‐cell disease remains poorly understood. Preliminary data from our group suggests that hemolysis, through activation of Ras homolog gene family member A (RhoA), causes activation of NADPH oxidase 2 (Nox2) and calcium/calmodulin‐dependent protein kinase II in the right ventricle leading to the production of reactive oxygen species, downstream mitochondrial destruction and ultimately, maladaptive remodeling of the right ventricle. We proceeded to examine whether RhoA activation indeed activates CaMKII via Nox2 and to see if this signaling axis drives myocyte dysfunction as evidenced by eNOS uncoupling and mitochondrial dysfunction. Six hours following exposure of rat neonatal cardiac myocytes (RNCM) to lysed red cell plasma resulted in a marked increase in superoxide production (P<0.01, n=15) that was significantly reduced by co‐incubation with the Nox2ds peptide (P<0.001). Bioenergetic analyses of RNCM showed a significant decrease in oxygen consumption rate after exposure to hemolyzed blood. Transfection studies showed an increase in RhoA expression after silencing Nox2 and CaMKII genes, suggesting that if RhoA is linked to these proteins, its activation may be upstream of Nox2 and CaMKII. Ongoing studies using peptide inhibitors are being employed to further elucidate the roles of the aforementioned proteins in hemolysis‐associated myocyte dysfunction.
The European eel NCCβ encodes for an electroneutral NaCl cotransporter that is not sensitive to thiazide diuretics (1099.1)
Authors: Moreno Erika , Cutler Christopher , Chávez‐Canales María , Vazquez Norma , Gamba Gerardo
Published: June 2021

The renal thiazide‐sensitive NaCl cotransporter (NCC) in mammalian nephron plays a key role in salt transport of the distal convoluted tubule and thus in modulation of arterial blood pressure and potassium secretion. Only one gene encoding NCC has been described in mammals. However, in the teleost European eel, two genes encoding putative isoforms of NCC were identified: NCCα, from kidney, and NCCβ from intestine. For the NCCα the identify degree with flounder, rat and human NCC is 70, 65, and 45%, respectively and for the NCCβ is 55, 55, and 43, respectively. No functional characterization of these variants has been reported. By RT‐PCR we cloned the full‐length cDNA of eel NCCβ isoform and inserted into the Xenopus expression vector pgh19. The insert contains 3437 bp, with an open reading frame of 3132, encoding a 1043 amino acid protein. Functional expression was assessed in Xenopus oocytes, microinjected with 10 ng of cRNA in vitro transcribed from NCCβ cDNA, by measuring the tracer 22Na+ uptake. Rat NCC was used as a control signal. Our results revealed that injection of oocytes with NCCβ cRNA induced the appearance of Na+ uptake mechanism that is fully dependent on extracellular Cl‐, constituting a NaCl cotransporter. However, in strict contrast to rat NCC, eel NCCβ is not inhibited by thiazide type diuretics. In addition, it is also not stimulated by WNK3 kinase that is a powerful activator of mammalian NCC. We conclude that NCCβ encodes a non‐thiazide sensitive NaCl cotransporter.
Glucose infusion increases mRNA expression of deiodinase 1 & 2, and thyroid hormone receptor β‐1 in a fasting‐adapted mammal (1101.12)
Authors: Martinez Bridget , Soñanez‐Organis José , MacKenzie Duncan , Crocker Daniel , Ortiz Rudy
Published: June 2021

Food deprivation in mammals is typically associated with reduced glucose, thyroid hormone (TH) concentrations and deiodinase (DI) content and activity to suppress metabolism. However, in prolong‐fasted, metabolically active elephant seal pups, the hypothalamic‐pituitary‐thyroid axis (HPT axis), which is responsible for the production of TH, is up‐regulated. The functional relevance of this apparent paradox is unknown and demonstrates variability in the regulation of TH levels, metabolism and function in food‐deprived mammals. To address our hypothesis that increased glucose increases peripheral TH receptor beta‐1 (THrβ‐1) mRNA expression and content in prolong‐fasted pups, we assessed the cellular TH‐mediated response to a glucose challenge by measuring TH profiles during a glucose tolerance test (GTT) and comparing the changes in profiles in early and late fasted pups. THrβ‐1 mRNA expression increased 59% ± 24 and 336% ± 95 by the 60 and 120 minute mark, respectively. Protein expression of THrβ‐1 also increased during the GTT, 40% ± 4 and 80% ± 23 at 60 and 120 minutes, respectively. mRNA expression of DI1 and DI2 increased by the 120 minute time point, 100% ± 34 and 400% ± 63, respectively. The data suggests that cellular TH pathways are responsive to changes in glucose, which in turn maybe regulated by TH. This suggests that the HPT axis is active and functional during prolonged food‐deprivation in metabolically and physically active mammals.

Grant Funding Source: Supported by NIH/NHLBI 442606‐RO‐29013
Role of body composition and circulating hormones on resting energy expenditure in obese pregnant women (1101.9)
Authors: Perez‐Rodriguez Marcela , de Haene Jessica , Castañeda Diana , Stotland Naomi , Murphy Elizabeth , Talavera Juan , Havel Peter , King Janet
Published: June 2021

The extra energy required for synthesis and maintenance of new tissue increases the REE during pregnancy. The net change may vary 8‐fold with higher increases seen in obese than non‐obese women. Our objective was to identify body composition and hormonal factors influencing the REE of obese women during the last half of pregnancy. A total of 64 women with BMI >25 kg/m2 , 18‐42 years of age, and no history of chronic disease were recruited at 20 weeks gestation. Three were eliminated due to extreme obesity (BMI=53 kg/m2) and 2 for missing values giving a total of 59 participants. REE, body composition, and insulin resistance (HOMA‐IR), prolactin, progesterone, estradiol, leptin, adiponectin, and IGF‐1 were measured at 20, 28, and 34 weeks gestation. During the 14‐wk study, the REE increase averaged 200 kcal/d. The change varied from a 400 kcal/d decrease to a 590 kcal/d increase. Multivariant models predicted 62% of the variability in REE at 20 weeks, 41% at 28 and 34 weeks, and 24% of the change in REE from 20 to 34 weeks. Significant determinants varied with time of pregnancy: FM, FFM, progesterone and prolactin at 20 wks; FM and FFM at 28 wks; FM, FFM, and IGF‐1 at 34 wks; and FFM and IGF‐1 change predicted the REE change from 20 to 34 wks. Body composition and metabolic hormones only explained 24 to 60% of the REE variability in obese pregnant women suggesting that other factors play a role.

Grant Funding Source: NIH Grant R01HD46741.
Neutrophil accumulation after contraction‐induced muscle injury is dependent upon P‐ and E‐selectin (1102.13)
Authors: Sloboda Darcee , Brooks Susan
Published: June 2021

Neutrophils accumulate in skeletal muscle after contraction‐induced injury and are thought to exacerbate damage to muscle fibers. The mechanism by which neutrophils enter the muscle is not known, but adhesion proteins on vascular endothelial cells may play a role. We tested the hypothesis that the adhesion proteins P‐ and E‐selectin are critical for neutrophil accumulation after contraction‐induced injury and that blunting neutrophil accumulation would reduce damage. To test our hypothesis, we injured mouse muscles in situ with a series of lengthening contractions. Mice were injected with either antibodies that block P‐ and E‐selectin or control antibodies. In control muscles, injury caused a 50% force deficit, overt damage to muscle fibers and a 15‐fold increase in neutrophil content after two days. Mice treated with blocking antibodies to P‐ and E‐selectin showed a decrease in neutrophil content but no change in force deficit or damage to muscle fibers. Our data suggests that neutrophil accumulation after contraction‐induced injury is dependent upon P‐ and E‐selectin but that blunting neutrophil infiltration after contraction‐induced injury does not always reduce damage.

Grant Funding Source: AG000114
The effects of doxorubicin administration on intramuscular nitric oxide concentration in rat skeletal muscle (1102.28)
Authors: Lalanne Josee , Fabris Sergio , MacLean David
Published: June 2021

Doxorubicin (DOX) is a frontline anticancer therapeutic which has proven effective in the treatment of breast cancer. However, very little is known regarding the effects that DOX has on skeletal muscle metabolism, specifically its influence on nitric oxide (NO) production. Therefore, the purpose of the present study was to determine NO concentrations in the plantaris (P, n =6) and gastrocnemius (G, n =6) muscles of Sprague‐Dawley rats after DOX administration at a single dose of 1.5 mg/kg. Muscle samples were collected at 24, 48, 72, 96, 120, 144, 168 and 192 hours post injection. . Following administration, intramuscular NO increased (P<0.05) in the G after 120 (0.99±0.04 mmol/kg dw), 144 (0.91±0.03 mmol/kg dw) and 168 hours (0.79±0.03 mmol/kg dw) compared to control (0.62±0.07 mmol/kg dw). Similarly, NO increased (P<0.05) in the P after 120 (0.89±0.02 mmol/kg dw) and 144 hours (0.89±0.03 mmol/kg dw) compared to control (0.66±0.08 mmol/kg dw). Other than the observation that NO remained higher in the G but not in the P group at 168 hr, there were no other differences between fiber types. These data clearly suggest that DOX administration stimulates intramuscular NO production in a time dependent manner. In addition, these findings may represent the first step in better understanding NO deregulation in skeletal muscle upon DOX administration.

Grant Funding Source: Supported by NSERC
Temporal variation in skeletal muscle atrophic pathways following transection of the sciatic nerve (1102.31)
Authors: Callahan Zachary , Menke Chelsea , Schaeffer Paul
Published: June 2021

The purpose of this study was to investigate the temporal changes in gene expression and morphology in muscles of the hindlimb following transection of the sciatic nerve. A sham surgery was performed on the contralateral limb and was used as a control. Mice were sacrificed 3, 10, 28, or 56 days after surgery. There was no significant difference in muscle mass between sham and transected limbs at 3 days post‐surgery, but tissue mass was significantly lower following transection at the other three time points. Transected muscle mass stabilized by 28 days post‐surgery with no further loss. Myocellular cross sectional area was significantly smaller at 28 and 56 days post‐transection surgery. Additionally, tissue fibrosis area was significantly greater at 56 days post‐transection. In transected muscle, gene expression analysis revealed higher expression of MAFbx and MuRF‐1 (genes involved in atrophic pathways) at 3 days post‐surgery while expression was significantly lower at 28 days post‐surgery. We saw no difference in gene expression of Fox01 or Fox03 (two upstream regulators of MAFbx and MuRF‐1) for any of the conditions. Western blot analysis revealed that MuRF‐1 protein expression was significantly higher in transected muscle at all times except 56 days post‐surgery. Fox01 protein expression in transected tissues was significantly lower in 10, 28, and 56‐day groups. These data suggest that the transection of the sciatic nerve results in the activation of pathways that initially increase atrophy, but that long‐term effects are less clear.

*This work was supported by NIH grant number 1R15DK085 497‐01A1.

Grant Funding Source: Supported by NIH grant number 1R15DK085 497‐01A1
Roles of FOXO signaling and mitochondrial dynamics in muscle disuse atrophy (1102.46)
Authors: Yeo Dongwook , Kang Chounghun , Song Chao , Zhang Tianou , Ji Li Li
Published: June 2021

Prolonged inactivity results in skeletal muscle atrophy during which oxidative stress and inflammation play an important role. Only sparse data are available regarding mitochondrial dynamic in muscle disuse atrophy. PURPOSE: To investigate whether hindlimb immobilization (IM) and remobilization (RM) alter mitochondrial dynamics in mice. METHOD: FVB/N mice (age 2 mo, N=35) were divided into: 1) control (CON); 2‐4) IM with banding of one hindlimb for 1, 2, and 3 wks (1,2,3 w‐IM) and 5) 2‐wk IM followed by 1‐wk RM with band removed. RESULT: FOXO1a content was increased by 4.0 & 6.3‐fold, respectively, in 2w‐ & 3w‐IM, (P<0.01) and by 50% (P<0.01) in RM vs. 3w‐IM. Phospho (P)‐FOXO1a levels were elevated in all IMs and RM groups vs. CON (P<0.01). In contrast, FOXO3a levels showed 51 & 42% (P<0.01) decrease, respectively, at 1w & 2w‐IM, whereas P‐FOXO3a was increased only in 2w‐IM (P<0.05). Mitofusion‐2 content was reduced to ~ 50 & 25% (P<0.01), respectively, in 2w‐ & 3w‐IM but increased by 3.7‐fold in RM (P<0.01). Fission protein‐1 was upregulated by 2.2 & 2.4‐fold (P<0.01) at 1w‐ & 2w‐IM, but was lowered at 3w‐IM (P<0.01). CONCLUSION: FOXO1 upregulation and phosphorylation play an important role in muscle disuse atrophy which may explain the activation of proteases shown in our previous study. Decreased fusion and increased fission protein expression may make the mitochondria more fragmented and catabolic.
Anti‐inflammatory effect of PEGylated dexamethasone liposome for tourniquet‐induced acute ischemia‐reperfusion injury in mouse skeletal muscle (1102.5)
Authors: Zhang Dongze , Yuan Fang , Tu Huiyin , Liu Jinxu , Wang Dong , Muelleman Robert , Li Yu‐Long
Published: June 2021

Ischemia‐reperfusion (IR) triggers a cascade of cellular and systemic inflammation and leads to cellular apoptosis and tissue necrosis in the skeletal muscle. In present study we investigated if anti‐inflammation improves skeletal muscle function. Hindlimbs of C57/BL6 mice were subjected to 3h ischemia and 4 wks reperfusion via placement of release of a tourniquet at a greater trochanter. Treatment of Dexamethasone (Dex, 1 mg/kg/day) for 4 weeks had a strong anti‐inflammatory effect but did not improve muscle contraction. However, treatment of Dex at the first 2 weeks induced a mild anti‐inflammatory effect and had the recovery of muscle contraction. Additionally liposome‐Dex (14 mg/kg) was bolus‐intravenously injected before IR due to severe side effects of Dex. Live imaging showed that this drug had a high target in IR muscles and stayed in the muscles at the first 2 wks of IR. Liposome‐Dex not only decreased expression of IL‐1β and NFkB‐p50, but also improved muscle contraction, compared to IR group. These data suggest that anti‐inflammatory effect of liposome‐Dex in early stage of IR is beneficial to muscle function recovery from IR without Dex‐associated side effects.
Heat stroke decreases cytokine and chemokine gene expression in the hypothalamus of rats (1104.15)
Authors: Audet Gerald , Ramon Jose , Dineen Shauna , Leon Lisa
Published: June 2021

We showed previously that rats display ~1°C core temperature (Tc) increase during ~4 days of heat stroke (HS) recovery; however, the mechanisms mediating this response remain unidentified. It was suggested that hypothalamic damage mediates this Tc response, but there is a lack of histological evidence in support of this. We hypothesized that local hypothalamic cytokines and/or chemokines, which are known regulators of Tc, may be mediating the elevation in Tc during HS recovery. The hypothalamus of Fischer 344 rats was examined for 83 cytokine/chemokine genes (real‐time PCR) at Tc,Max (41.8°C), 1, 3, and 10 days of HS recovery (3‐8 rats/group). IL‐1β was elevated ~14‐fold at Tc,Max and returned to baseline by 24 hours; IFNγ and IL‐10 were decreased ~3‐fold at 24 hours. All cytokines returned to baseline by 3 days. Most chemokines were downregulated ~2‐ to 4‐fold through 24 hours, but shifted towards increased expression by 3 days. Despite normal Tc at 10 days of recovery, some chemokine changes persisted. Contrary to our hypothesis, most cytokines/chemokines showed significant down regulation during HS recovery, which may indicate increased translation in our model. Importantly, resumption of normal Tc at 10 days with persistent alteration in the hypothalamic cytokine/chemokine profile may indicate greater susceptibility to a subsequent HS event. Author views not official US Army or DoD policy.
An excitatory projection from median preoptic area to the dorsomedial hypothalamus contributes to the activation BAT thermogenesis (1104.28)
Authors: Tupone Domenico , Morrison Shaun
Published: June 2021

Disinhibition of brown adipose tissue (BAT) sympathoexcitatory neurons in the dorsomedial hypothalamus (DMH), by reduced discharge of their input from GABAergic, in the preoptic area (POA), is thought to be a key step in the cold‐defense and febrile activations of BAT sympathetic nerve activity (SNA) and BAT thermogenesis. Here we present evidence supporting the hypothesis that a population of median preoptic area (MnPO) neurons projecting to the DMH provides the excitatory drive to DMH neurons that is essential for cold‐defense and febrile BAT thermogenesis.

To determine if the MnPO contains putative BAT thermogenesis‐promoting neurons that are activated during cold exposure and project to DMH, we injected the retrograde tracer, CTb into DMH and exposed the rats to a cold (10°C) ambient temperature to elicit BAT thermogenesis. We observed a larger number of c‐fos and CTb immunoreactive neurons in the MnPO of cold‐exposed rats than in those maintained at 25°C, confirming the existence of cold‐activated neurons in MnPO that project to DMH. In urethane/chloralose‐anesthetized, Wistar rats, bilateral nanoinjection of the GABAA agonist, muscimol (120nl, 1mM), into the medial preoptic area (MPA), a region containing warm‐sensitive neurons, did not decrease cold‐evoked BAT SNA, consistent with the hypothesis that the MPA is not the source of DMH excitation required for BAT thermogenesis. However, subsequent nanoinjection of muscimol (120nl, 1mM) into the MnPO completely reversed cold‐evoked BAT SNA. Moreover, in other experiments, the activation of BAT SNA following PGE2 nanoinjection into MPA was also inhibited by muscimol nanoinjection into MnPO. These data demonstrate that the MnPO contains essential thermogenesis‐promoting, BAT sympathoexcitatory neurons.

Grant Funding Source: NIH NS40987
The effect of the head or neck cooling on body core temperature and thermal sensation in humans (1104.6)
Authors: Matsuda‐Nakamura Mayumi , Wada Satoshi , Marui Shuri , Sato Nobuo , Nagashima Kei
Published: June 2021

Some animals have mechanisms that selectively cool the brain during hyperthermia. However, it remains controversial if humans have the mechanism. In the present study, we investigated the effect of the head or neck cooling on tympanic temperature as an indicator of brain temperature in normothermic and hyperthermic conditions. Eight healthy male subjects performed four local cooling trials (facial fanning, cooling of the forehead and neck, and the control). Subjects sat on a chair, putting on a water‐perfusion suit to maintain skin temperature at 33°C (normothermic condition) or 38°C (hyperthermic condition). Tympanic, esophageal, and skin temperatures were continuously monitored. Temperature sensation and thermal pleasantness were reported by the subjects. After 10‐min baseline period, local cooling was conducted for 15 min. During the head or neck cooling, the local skin temperature significantly decreased, however, there was no significant difference between tympanic and esophageal temperatures in normothemic condition. Only in hyperthermic condition, a significant difference between tympanic and esophageal temperatures was observed. After facial fanning, tympanic temperature was lower than esophageal temperature. While facial fanning produced a slight change in thermal pleasantness in normothermic condition, facial fanning produced a clear “pleasant” feeling in hyperthermic condition. These results may suggest that facial fanning cools the brain in hyperthermic condition. However, local skin cooling of the forehead and neck may not effect on the selective brain cooling.

Grant Funding Source: This study was supported by JSPS KAKENHI Grant Number 25870831
Regional difference in blood flow response to hypercapnia (1106.12)
Authors: Miyaji Akane , Ikemura Tsukawa , Hamada Yuka , Hayashi Naoyuki
Published: June 2021

To investigate whether vascular responses to hypercapnia differ among various blood vessels, we recorded relative changes of mean arterial pressure (MAP) and blood flow values in the middle cerebral artery (MCA), retinal artery (RA), and choroidal vessels (CV) associated with an increase in end‐tidal CO2 pressure (PETCO2) induced by breathing CO2 rich air (5%) for 3 min in 30 healthy males (23.6 ± 0.3 years). PETCO2 and MAP significantly increased from baseline values by 16.5 ± 1.2% and 4.9 ± 1.0%, respectively (P<0.05). Blood flow values in MCA, RA, and CV significantly increased by 11.2 ± 4.9%, 7.6 ± 2.0%, and 18.4 ± 4.5%, respectively (P<0.05). There was a significant inter‐individual correlation in relative changes solely between RA and CV (r=0.47). It was suggested that vascular responses to hypercapnia differ among vessels, even among vessels similarly having vasodilatory response to CO2.
Acute intermittent hypoxia alters leptin signaling pathways in arcuate nucleus (1107.1)
Authors: Moreau Jason , McCoy Aaron , Jones Douglas , Ciriello John
Published: June 2021

The effect of acute intermittent hypoxia (AIH) on central pathways involved in metabolic homeostasis was investigated in male wild type Sprague‐Dawley (WT) or leptin deficient (Kilo rat; a homozygous knockout of the leptin gene) rats. Rats were exposed to AIH (8 h) or normoxic control conditions and their hypothalamic arcuate nuclei (ARC) were assessed for changes in protein signaling associated with satiety. AIH reduced body weight, food intake and active cycle locomotion without altering adipose tissue mass compared to WT normoxic controls. These differences were not present in Kilo rats. Plasma concentration of leptin was significantly increased after AIH in WT rats, but was not detectable in the Kilo rat. In ARC, phosphorylation of STAT3 and pro‐opiomelanocortin (POMC) expression were higher in the AIH WT rats, but these differences were absent in Kilo rats. Additionally, POMC‐expressing neurons were activated as determined by immediate early gene FRA‐1/2 expression in the AIH WT rats. Finally, ERK1/2 and its phosphorylation were lower in response to AIH in the WT rat, but these differences were eliminated in the Kilo rat. These data suggest that AIH induces significant alterations to body energy balance through changes in the secretion of leptin which exerts effects on satiety‐inducing pathways within the hypothalamus. Supported by HSF of Ontario.

Grant Funding Source: Heart and Stroke Foundation of Ontario
Molecular support for high paracellular absorption in the intestine of flying mammals (1109.18)
Authors: Caviedes‐Vidal Enrique , Brun Antonio , Fernández‐Marinone Guido , Gatica‐Sosa Claudia , Price Edwin , Amweg Ayelen , Ortega Hugo , Karasov William
Published: June 2021

Flying mammals absorb water‐soluble nutrients (e.g. glucose) at the intestine mainly by the paracellular route [across tight junctions (TJ) between adjacent enterocytes], while non‐flying mammals mainly rely on the transcellular route (across cell membrane using transporter proteins). We predicted that insectivorous bats Tadarida brasiliensis, compared to Mus musculus, would show enhanced expression of CLDN‐2 (CLDN=claudin) and OCLN (occludin) TJ proteins, which have been described as responsible for the permeability of polar water‐soluble solutes. We assessed the level and spatial distribution of CLDN‐2, ‐3, ‐5, ZO‐1 and OCLN by immunohistochemistry along the villus axis in the proximal intestine. Immuno‐specificity of the antibodies was checked by western blot. No differences were found for CLDN‐3 and ZO‐1 between species. We confirmed low expression of CLDN‐2 in mice (Holmes et al. 2006, Gene Expr Patterns 6: 581‐8; no signal detected in our preparations), but in bats the villus tips exhibited CLDN‐2. OCLN was expressed along the entire villus axis but with a stronger signal for the bat than for mice, giving additional support to our prediction. A third difference was that CLDN‐5 was strongly expressed along the villus in bats, whereas in mice its presence was not detected. Findings suggest potential molecular mechanism(s) underlying the observed higher paracellular absorption in bats compared to mice.

Grant Funding Source: NSF IOS 1025886
Measurements of the stomachs and duodenums of fasted and sated rats (1110.2)
Authors: Lorber Mortimer
Published: June 2021

Rats feed until sated. Thus, gastric and duodenal dimensions likely increase. Under Na pentobarbital the stomach and proximal duodenum were exposed via a midline incision. Measurements were made by a calipers and digital micrometer. Eight fasted adult females and eight which ate chow until sated, as judged by pausing or preparation for sleep, were studied. They ate (mean + s.d.) 4.74 + 1.17 g of chow. Body weights and visceral dimensions were compared by unpaired t‐tests. A P value < 0.05 was deemed significant. The fasted:sated mean + 1 s.d. values were: Body Weight (g) 186 + 9:195 + 10 (ns). Gastric values (cm): Width 1.94 + 0.13: 3.86 + 0.53; Rostrocaudal Height 1.78 + 0.15: 2.84 + 0.30; Dorsoventral Thickness 0.73 + 0.16: 1.70 + 0.25. (P < 0.001 for all). Proximal duodenal values (cm) were: Rostrocaudal Height 0.53 + 0.09: 0.55 + 0.17 (ns); Dorsoventral Thickness could not be obtained because the proximal duodenum is bound to the peritoneum. Separation would be unphysiological. All fasted rats but one had dorsoventrally concave duodenums. All sated rats but one had flat duodenums, suggesting some gastric emptying had occurred. The ratios of mean sated:fasted dimensions were 1.99 for Width, 1.60 for Height, and 2.33 for Thickness. Thus, the stomach markedly thickens. Its height increases least and its width, which doubles, is intermediate. In achieving satiety the rat stomach enlarges both significantly and anisotropically. ,

Grant Funding Source: Personal
Upregulation of RhoA/Rho kinase pathway and downregulation of cGMP/PKG pathway in smooth muscle impair peristaltic reflex and pellet propulsion in the colon of diabetic mice (1110.5)
Authors: Kendig Derek , Nalli Ancy , Mahavadi Sunila , Grider John , Murthy Karnam
Published: June 2021

Abnormalities in extrinsic and intrinsic innervation and in the distribution of interstitial cells of Cajal are known to contribute to altered gut motility in diabetes. Smooth muscle dysfunction in relation to diabetes has not been fully explored. Aim. To test the hypothesis that upregulation of RhoA/Rho kinase pathway and downregulation of cGMP/PKG pathway contribute to increased muscle tone in diabetes. Methods. Colonic smooth muscle cells from type 1 (NOD) or type 2 diabetes (db/db) mice were used to measure i) RhoA and PDE5 expression; 2) ACh‐induced Rho kinase activity and muscle contraction; 3) NO‐induced PDE5 activity, cGMP levels and muscle relaxation. Colonic segments were used to measure peristaltic reflex and pellet propulsion. Results. Smooth muscle cells isolated from the colon of NOD and db/db mice showed an increase in RhoA and PDE5 expression, Rho kinase and PDE5 activity, and muscle contraction, and a decrease in cGMP levels and muscle relaxation relative to control mice. Consistent with an increase in RhoA and PDE5 expression, orad contraction was increased by 20%, caudad relaxation decreased by 60%, and pellet propulsion decreased by 35% in db/db mice relative to control. Discussion. In diabetic colon upregulation of the RhoA/Rho kinase pathway and downregulation of the cGMP/PKG pathway causes an increase in muscle contraction and decrease in relaxation leading to a decrease in pellet propulsion.
Butyrate increases and propionate decreases the frequency of propulsive contractions in guinea pig proximal colon (1110.7)
Authors: Hurst Norm , Kendig Derek , Murthy Karnam , Grider John
Published: June 2021

INTRODUCTION: Colonic bacteria produce short‐chain fatty acids (SCFAs) by fermentation of dietary carbohydrates and fiber. The production of SCFAs is greatest in proximal colon where propulsion is likely to be highly dependent on chemical/nutrient stimuli. AIM: To determine the effect of individual SCFAs on propulsive contractions in guinea pig proximal colon. METHODS: Proximal colon was excised, cannulated and placed in Kreb’s buffer in an organ bath. After equilibration, the colon was distended with 1 ml of Krebs’s buffer alone or containing sodium salts of acetate, butyrate, or propionate at 10‐100 mM. Motility was videorecorded, spatiotemporal maps generated, and the number of full‐length propulsive contractions during a 5 min period determined. RESULTS: The basal rate of propulsive contractions in Kreb’s buffer was 2.9±0.7 per 5 min. Butyrate and propionate had concentration‐dependent and opposing effects on propulsive contractions; acetate had no effect. Butyrate significantly increased propulsive contractions (maximal increase of 207% (p<0.05) at 30 mM) whereas propionate abolished propulsive contractions (91‐100% inhibition at 10‐30 mM). Control Kreb’s buffer containing 10‐100 mM NaCl had no effect. CONCLUSION: SCFAs have differential effects on propulsive contractions in guinea pig proximal colon with butyrate being excitatory and propionate being inhibitory.

Grant Funding Source: Supported by DK34153