bluuebunny/crossref_metadata_2025 huggingface dataset:
Vascular Modulation of Adipose function: Role of Endothelial Argonaute 1
Authors: Chen Zhen , Tang Xiaofang , Miao Yifei , Qi Zhijie , Luo Yingjun , Sriram Kiran , Van Keuren‐Jensen Kendall , Fueger Patrick , Wang Qiong , Yeo Gene , Zhong Sheng
Published: June 2021

Among all individuals with Type 2 diabetes mellitus (T2DM), 80% are obese. The dramatic expansion of adipocytes can give rise to chronic hypoxia, poor blood perfusion, accumulation of pro‐inflammatory cytokines, all of which contribute to insulin resistance, key feature of T2DM. Lining the inner surface of microvessels, endothelial cells (ECs) respond to alterations in oxygen, metabolites, and growth factors to further modulate adipose function. However, the molecular and cellular mechanisms underlying the EC‐adipose crosstalk remain largely unknown and have recently attracted great research interests. In this study, we investigated the role of microvascular ECs in adipose function in the context of obesity and insulin resistance. Specifically, we examined the role of microRNA (miR)‐mediated gene regulation in the endothelial modulation of adipose function.

We first used cross‐linking and immunoprecipitation sequencing (CLIP‐seq) to profile the miR targetome in ECs subjected to hypoxia, to which the EC response is often compromised in obesity. CLIP‐seq revealed a panel of angiogenic inhibitors, including thrombospondin‐1 (THBS1; which is positively associated with obesity and insulin resistance) were enriched in miR‐induced silencing complex in ECs under hypoxia. However, their expression was drastically suppressed in ECs by hypoxia. Ina murine T2DM model, mice with EC‐specific AGO1‐knockout (EC‐AGO1‐KO) exhibited a lean phenotype, with lower body weight and fasting glucose level and higher insulin sensitivity, as compared to their wild‐type littermates. The EC‐AGO1‐KO animals also had less body fat content, particularly in the subcutaneous and brown adipose tissues. Histological analysis and RNA‐seq demonstrated that EC‐AGO1‐KO possess higher angiogenic and browning activity and lower inflammation in the adipose depots. Overexpression of THBS1 abolished the protective effect of AGO1 in the subcutaneous tissues, suggesting that AGO1‐KO‐mediated metabolic benefit is in part through suppression of THBS1.

Collectively, these results support a mechanism by which ECs, through AGO1‐mediated miR‐targetome modulate adipose function and metabolic states. Our study provides novel insights linking endothelial function and adipose tissue remodeling. Further, given that endothelial function is often compromised in T2DM, the molecular mechanisms underlying the endothelium‐adipocyte crosstalk may offer new approach to manage metabolic disruption associated with obesity and diabetes.

Support or Funding Information

K99/R00HL122368

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cholesterol Inhibits H2S‐Mediated Vasodilation
Authors: Mendiola Perenkita J , Gonzalez Bosc Laura V , Naik Jay S , Kanagy Nancy L
Published: June 2021

Hydrogen sulfide (H2S), the most recently described endothelium‐derived vasodilator, is predominantly produced in the endothelium by cystathionine γ‐lyase (CSE). We previously demonstrated that within endothelial cells (ECs), H2S‐mediated vasodilation involves activation of transient receptor potential cation channel subfamily V member 4 (TRPV4)‐dependent Ca2+ influx and large conductance Ca2+‐activated potassium (BK) channel. In addition, a single concentration of H2S donor NaHS (1 μM) dilated small mesenteric arteries (<100 μm) but had no effect in large mesenteric arteries (>300 μm). Furthermore, inhibiting CSE generation of H2S with β‐cyanoalanine (BCA, 100 μM) impaired acetylcholine (ACh)‐induced dilation in small mesenteric arteries, but had no effect in large mesenteric arteries. These data suggest that H2S‐induced dilation is prominent and important in small resistance arteries but not in larger arteries. It is possible that decreased availability of TRPV4 and/or BK in larger arteries is responsible for the diminished sensitivity to H2S‐induced vasodilation. Previous reports suggest that membrane cholesterol negatively regulates TRPV4 mobility and BK activity. Our current study tested the hypothesis that H2S‐mediated vasodilation is inhibited by membrane cholesterol. Mesenteric arteries induced vasodilation was assessed in pressurized large rat mesenteric arteries (>300 μm). Isolated pressurized mesenteric arteries (320 μm – 360 μm) were pretreated luminally with vehicle (HEPES) or the cholesterol‐depleting agent, methyl β cyclodextran (MBCD, 100 μM) and exposed to NaHS (100 μM). NaHS‐induced dilation in U46619 pre‐constricted arteries was significantly enhanced by MBCD pretreatment (+vehicle: 6.78% ± 1.6, +MBCD: 17.10% ± 6.8, p < 0.05, n=3–4 animals/group). These studies suggest that membrane cholesterol disrupts H2S‐mediated vasodilation. Ongoing studies in isolated mesenteric arteries of different sizes are investigating endothelial cell membrane cholesterol content and the effect of cholesterol depletion on TRPV4‐dependent Ca2+ influx in large and small mesenteric arteries.

Support or Funding Information

Funding provided by the NIH HL12330 & HL7736‐23

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Role of Tissue Iron in Development of Cardiovascular Delayed Effects of Acute Radiation Exposure
Authors: Russell Abigail , Chittajallu Supriya , Siddiqui Sameen , Unthank Joseph L , Sampson Carol H , Fisher Alexa , Orschell Christie M , Miller Steven J
Published: June 2021

The threat of radiation exposure from terrorist activity, warfare, or accidents creates an urgent need to develop agents to mitigate the acute and chronic effects of high dose rate total body irradiation (TBI). High level short‐term radiation exposure results in rapid organ injury leading to development of the acute radiation syndrome (ARS), which if untreated will result in death within weeks. Survivors of ARS will experience the delayed effects of acute radiation exposure (DEARE), which include development of chronic pathology and dysfunction of multiple organ systems months to years following exposure. However, the factors and mechanisms that regulate the onset and progression of cardiovascular DEARE are unclear. The goal of the current study was to determine the possible role for abnormal iron metabolism in the development of cardiac DEARE, especially vascular pathology. Studies utilized the ARS mouse model with gamma radiation exposure at 853 cGy (LD50) and time points from 1 day to 4 months post‐TBI.

Serum iron was elevated at 1 day post‐TBI, peaked at 2 weeks, and returned to non‐irradiated control values by 4 weeks post‐TBI. A similar trend was seen for transferrin saturation, and both results correlated inversely with red blood cell number, hematocrit, and hemoglobin concentrations. Perls' Prussian Blue staining was used to detect iron (hemosiderin) deposition in heart paraffin‐embedded tissue sections, and myocardial iron was present as early as 2 weeks post‐TBI. Pretreatment of mice with the iron chelator deferiprone in drinking water decreased myocardial iron deposition (P=0.055), but not serum iron, at 2 weeks. A histological assessment of coronary artery endothelial cell (EC) density showed intimal nuclei per unit area significantly decreased as early as two weeks post‐TBI vs. non‐irradiated controls (P<0.05), and the decrease persisted to 4 months post‐TBI. EC density in deferiprone‐treated mice was significantly increased at 2 and 4 weeks post‐TBI compared to irradiated non‐treated mice (P<0.03). Administration of the ferroptosis inhibitor ferrostatin‐1 via daily i.p. injection for 2 weeks post‐TBI resulted in no change in coronary artery EC density compared to vehicle controls. Taken together, the results suggest the presence of tissue iron contributes to endothelial cell loss occurring early following TBI and is a significant event impacting the development of DEARE. This information may facilitate development of novel medical countermeasures to prevent or mitigate cardiovascular DEARE‐related pathophysiology.

Support or Funding Information

NIH 1U01AI107340‐01 (C. Orschell), IUPUI Research Support Funds Grant (S. Miller)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Short‐term Ibuprofen Treatment cause Gender‐Specific Differences in Proteasomal and Mitochondrial functions in Mice Heart
Authors: Tiwari Shuchita , Gomes Aldrin
Published: June 2021

Ibuprofen is the most common pain‐killer widely used for the management of dental pain, headache, migraine and rheumatoid arthritis. Concern has been raised due to adverse cardiovascular side effects associated with the chronic use of ibuprofen. Thus, understanding the short‐term effects of ibuprofen on cardiac function is crucial to the safe use of the drug among individuals with established cardiovascular disorders. The objective of this study was to evaluate whether ibuprofen caused gender‐specific differences in mitochondrial and proteasomal functions in mice cardiac tissue and to gain insights into the molecular mechanisms underlying these differences. To determine whether there is a sex difference on protein homeostasis with respect to ibuprofen treatment 8‐week‐old aged matched male and female C57BL/6J mice were treated with ibuprofen (100 mg/kg/day) dissolved in drinking water for seven days and were randomly assigned to control group and group treated with ibuprofen. Ibuprofen significantly reduced caspase‐like (β1), and trypsin‐like (β2) 26S proteasome activities in both male and female heart. However, ibuprofen reduced chymotrypsin‐like (β5 proteasome activity) in the female heart but not male hearts. Notably, immunoproteasome β1i (expression) and β5i (expression and activity) significantly increased in females relative to males. In contrast, a trend towards the decrease in expression of β2i and β5i immunoproteasome was observed in ibuprofen‐treated male mice. To determine mitochondrial function, measurement of mitochondrial respiratory chain complexes revealed that ibuprofen treatment significantly decreased complex I activity in female heart compared with males. Short‐term ibuprofen treatment also decreased heart weight in male mice but not in female mice. Western blots and oxyblots analysis showed no differences in the expression levels of polyubiquitinated proteins, free ubiquitins and oxidized proteins in both males and females. Our in vitro study with rat cardiac H9c2 cells showed that ibuprofen‐induced reactive oxygen species (ROS) generation, decreased mitochondrial complex II activity, reduced mitochondrial membrane potential and had variable effects on cell viability. In conclusion, our findings suggest that short‐term ibuprofen treatment results in gender‐specific adverse changes in cardiac mitochondrial function and protein homeostasis.

Support or Funding Information

American Heart Association (AHA) 16GRNT31350040 grant (A.V.G.)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Vascular Function in Heart Failure Patients Implanted with a Continuous‐Flow Left Ventricular Assist Device: Impact of Increasing Peripheral Vascular Pulsatility
Authors: Hydren Jay R. , Gifford Jayson R. , Jarrett Catherine L. , Park Soung Hun , Shields Katherine L. , Ratchford Stephen M. , Broxterman Ryan M. , Kithas Andrew C. , Thurston Taylor S. , Selzman Craig H. , Drakos Stavros G. , Richardson Russell S.
Published: June 2021

Vascular function is thought to be attenuated in heart failure patients implanted with a continuous‐flow left ventricular assist device (LVAD), likely due to a decrease peripheral vascular pulsatility, which may contribute to the observed serious cardiovascular complications. However, the impact of increasing pulsatility on vascular function in this population is unknown.

Methods

Fifteen LVAD recipients and fifteen controls, matched for age and physical activity, underwent 45‐minutes of unilateral arm pulsatility treatment, evoked by cuff inflation, distal to the elbow, with a two second duty cycle. Vascular function was assessed by brachial artery flow mediated dilation (FMD, Doppler ultrasound).

Results

Baseline FMD, expressed as percent dilation, was not different between the LVAD recipients and controls (LVAD: 4.1±1.8; Controls: 4.1±1.5%), however, FMD/shear rate was significantly lower (LVAD: 0.10±0.04; Controls: 0.16±0.07) and time to peak dilation significantly longer in the LVAD group (LVAD: 77±21; Controls: 41±16s). The LVAD recipients exhibited a significantly attenuated pulsatility index (PI) compared to controls at baseline (LVAD: 3±2; Controls: 14±7), but experienced a similar PI during the pulsatility treatment (LVAD: 29±21; Controls: 33±15). Both % FMD (LVAD: 6.9±1.9; Controls 6.9±2.7%) and FMD/Shear rate (LVAD: 0.19±0.08; Controls 0.30±0.17) increased similarly in both groups with the pulsatility treatment, while time to peak was unaltered.

Conclusions

LVAD recipients exhibit vascular dysfunction. Pulsatility treatment improved vascular function in both groups, but, most importantly, this intervention restored vascular function in the LVAD recipients to the baseline level of the controls, and may, through this improved vascular health, reduce cardiovascular complications in this population.

Support or Funding Information

This research was supported by the National Institutes of Health under Ruth L. Kirschstein National Research Service Award NIH 1T32HL139451 from the National Heart, Lung, and Blood Institute.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chronic Reductive Stress Impairs Endoplasmic Reticulum Function and Cause Proteotoxic Cardiac Disease
Authors: Soorappan Rajasekaran Namakkal , Shanmugam Gobinath , Wang Ding , Fernandez Jolyn , Jones Dean P , Abel Dale E , Ping Peipei
Published: June 2021

Background

Heart failure (HF) is a growing cause of global human morbidity and mortality and elevated ROS are generalized as drivers of HF pathology. However, antioxidant treatments have failed to offer sufficient therapeutic benefits; the underlying mechanisms remain largely unknown. Here, we hypothesized that chronic reductive stress (cRS) induces endoplasmic reticulum (ER) dysfunction, which leads to proteotoxicity and pathological cardiac remodeling.

Methods

Transgenic mice expressing cardiac specific, constitutively active Nrf2 (caNrf2‐TG) and their non‐transgenic (NTG) littermates at the age of 3 and ~6 months (n=6–8/group) were used to determine ER redox states and protein aggregation levels. Diastolic function was measured by echocardiography.

Results

cRS substantially increased ER glutathione levels and suppressed protein oxidation (DMPO‐adducts) in TG vs. NTG mouse hearts. The co‐localization of KDEL (ER‐marker) with GSH or DMPO, showed by immunofluorescent imaging, indicates a hyper‐reductive redox milieu in the ER. Immunoblotting revealed a dose‐dependent induction of ER‐proteins (CHOP, GRP78, GRP94 and PDI) in TG hearts at 3‐months‐old. However, at 6‐months of age, this response was significantly repressed along with impaired protein folding response in TG mice, evidencing that progressive cRS is associated with a transition from adaptive ER stress‐response to ER dysfunction. Notably, electron micrograph and Proteostat® staining showed a dose‐ and age‐dependent augmentation of misfolded/aggregated proteins in TG hearts; these aggregates were highly ubiquitinated as demonstrated. Patterns of Mitral valve movement showed that ≥6 month‐old TG mice developed overt LV diastolic dysfunction as evidenced by increased mitral inflow velocity (MV E/A ratio).

Conclusion

ER dysfunction and proteotoxic insults triggered pathological cardiac remodeling and diastolic dysfunction in the TG hearts experiencing cRS. The heterogeneity of redox profiles in HF subjects demands further investigation into the potential impact of RS in cardiovascular pathology. Research focusing on correlations between redox stress intensity, redox imbalance and disease progression may offer insights into redox‐based therapies.

Support or Funding Information

This study was supported by funding from NHLBI (HL118067), NIA (AG042860), the AHA (BGIA 0865015F), University of Utah Center for Aging Pilot grant (2009), the Division of Cardiovascular Medicine/Department of Medicine, University of Utah and the start‐up funds (for NSR) by Department of Pathology, the University of Alabama at Birmingham, AL and UABAMC21 grant by the University of Alabama at Birmingham, AL.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Short‐term changes in left ventricular dyssynchrony may lead to congestive heart failure
Authors: Castaneda Diana , Yamazaki Katrina
Published: June 2021

Ventricular dyssynchrony is characterized by a difference in timing or lack of synchrony in ventricular contraction, leading to an inefficiency of pumping blood. Ventricular dyssynchrony (VD) consists of a disruption in the normal electrical conduction system, which leads to congestive heart failure (CHF). Long‐term effects of VD that lead to CHF is well studied but little is known about the short‐term effects associated with VD. Short‐term physiological changes due to VD are crucial because electrical and mechanical abnormalities or structural changes cause a decrease in overall cardiac functionresulting in long‐term effects that may lead to CHF. Such physiological abnormalities can range from metabolic alterations, metabolic remodeling such as changes in substrate utilization, mitochondrial dysfunction, ATP deficiency, impaired contractility, and the progression of cell regeneration and cell growth, etc. Thus, theobjective of this study is to investigate short‐term cellular effects associated of induced VD in vivo. Our hypothesis is that induced left ventricular dyssynchrony (via pacing) in a non‐CHF model may lead to short‐term changes such as alterations in cardiac metabolism and mitochondrial dysfunctionresulting in structural rearrangements. Frogs will undergo ventricular pacing to induce VD. Strain patterns associated with contraction and relaxation will be analyzed to verify dyssynchrony. Paced and non‐paced regions of the heart will be analyzed to determine if reactive oxygen species (ROS) generation and mitochondrial abnormalities lead to cellular changes that may induce apoptosis leading to structural rearrangement, known as ventricular remodeling.

Support or Funding Information

Supported by Cal State LA Minority Opportunities in Research MBRS‐RISE Program fund through NIH NIGMS under Grant No. R25 GM61331.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evidence that Food Intake Controls Diurnal Blood Pressure Rhythm in Mice
Authors: Zhang Dingguo , Jin Chunhua , Speed Joshua , Pollock David M.
Published: June 2021

Blood pressure exhibits a 24‐hour rhythm and peaks in the beginning of the active period in both human and rodents. Previous studies suggested that timing of food intake plays pivotal roles in regulating many physiological processes, such as lipid metabolism, sleep‐wake cycle, immune responsiveness, and even blood pressure. However, whether blood pressure and kidney function is altered by changing feeding time remains largely unknown. The current study was designed to test the hypothesis that feeding during the inactive period (reverse feeding or RF) disrupts blood pressure and kidney function in mice. Male C57Bl/6J mice (12–16 weeks old, n=6) were placed in metabolic cages and had free access to food and water for 3 days, followed by a 6‐day RF with free access to water. Food and water intake, as well as urine output were monitored twice per day at the beginning of active and inactive periods (AP and IP, respectively). Blood pressure was recorded via telemetry transmitters. As expected, we found that mean arterial pressure (MAP) was significantly higher in AP compared to IP when mice were fed ad libitum (114 ± 1 vs. 96 ± 1 mmHg, respectively, p<0.01). However, RF led to an inversion of diurnal MAP (98 ± 5 vs. 112 ± 2 mmHg, p=0.02). Similar effects were observed in heart rate (ad libitum, 659 ± 12 vs. 548 ± 16 bpm, p=0.03; RF, 484 ± 48 vs. 613 ± 22 bpm, p=0.02). In contrast, RF did not alter diurnal rhythm in urine output (ad libitum, 1.0 ± 0.2 vs. 0.2 ± 0.1 ml/12hr, p<0.01; RF, 1.4 ± 0.1 vs. 0.7 ± 0.1 ml/12hr, p<0.01). However, diurnal rhythm in urinary sodium excretion was dampened after RF (ad libitum, 7.3 ± 0.8 vs. 1.0 ± 0.2 mEq/12hr, p<0.01; RF, 6.5 ± 0.8 vs. 4.5 ± 0.7 mEq/12hr, p=0.08). Similarly, diurnal rhythm in urinary potassium excretion was abolished (ad libitum, 7.0 ± 0.9 vs. 0.8 ± 0.2 mEq/12hr, p<0.01; RF, 4.8 ± 0.7 vs. 5.3 ± 0.7 mEq/12hr, p=0.97). We also measured urinary excretion of aldosterone by ELISA and found that its diurnal rhythm remained intact after RF (ad libitum, 990 ± 142 vs. 370 ± 38 pg/12hr, p=0.01; RF, 856 ± 181 vs. 204 ± 42 pg/12hr, p=0.01). These data suggest that RF inverts diurnal blood pressure rhythm and differentially modifies diurnal excretion of water and electrolytes in mice.

Support or Funding Information

Funded by an AHA pre‐doctoral fellowship (18PRE33990345) to DZ and NHLBI P01 HL136267 to DMP. We appreciate Drs. Celso and Elise Gomez‐Sanchez's help with the aldosterone assay.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Impaired pulmonary function and right ventricular morphology in well‐healed burn survivors is related to aerobic capacity and not severity of burn injury
Authors: Samuel T. Jake , Nelson Michael D. , Nasirian Aida , Jaffery Manall , Moralez Gilbert , Romero Steven A. , Cramer Matthew N. , Huang Mu , Kouda Ken , Babb Tony G. , Sarma Satyam , Crandall Craig
Published: June 2021

Acute pulmonary dysfunction—second to inhalation injury—is prevalent in burn survivors, and is associated with a 24‐fold increase in morbidity and mortality. Systemic inflammation is known to persist many years post‐injury and likely compromises the pulmonary vasculature and respiratory gas exchange in well‐healed burn survivors. Whether pulmonary dysfunction in this population is related to right ventricular (RV) morphology and function remains unknown. Twenty‐six well‐healed burn survivors (age: 42 ± 14; body mass index: 28.1 ± 5.8; body surface area [BSA] burn: 49 ± 22 [range: 20–88]) and 11 age, sex and activity matched controls (age: 34 ± 9; body mass index: 28.2 ± 5.3) were recruited for assessment of maximal aerobic capacity (VO2max; via a graded cycle ergometer test), pulmonary function (diffusing capacity of the lungs for carbon monoxide [DLCO]) and RV morphology and function (via magnetic resonance imaging). Burn survivors were grouped post‐hoc by DLCO and classified as having either low (≤80% predicted; n = 6; %BSA burn: 42 ± 11), moderate (80–100% predicted; n = 11; %BSA burn: 42 ± 23), or high (>100% predicted; n = 9; %BSA burn: 61 ± 22) diffusing capacity. Group differences in RV morphology and function were assessed by one‐way ANOVA with Bonferroni correction where appropriate, and relationships between variables were assessed by partial correlation after controlling for body mass. Group differences were observed between those with low and high diffusing capacity in RV morphology (RV end‐diastolic volume [EDV] index: 73 ± 10mL/m2 vs. 81 ± 11mL/m2 vs. 95 ± 15mL/m2 vs. 84 ± 13mL/m2, low vs. moderate vs. high vs. control, respectively; ANOVA P = 0.012), but not RV function (RV ejection fraction: 54 ± 7% vs. 51 ± 9% vs. 53 ± 6% vs. 51 ± 10%, low vs. moderate vs. high vs. control, respectively; ANOVA P = 0.94). However, correlational analyses revealed that these group differences were primarily related to maximal aerobic capacity, after controlling for body mass (RV EDV vs. VO2max, r = 0.48, P = 0.003; DLCO vs. VO2max, r = 0.77, P < 0.001; RV EDV vs. DLCO, r = 0.57, P < 0.001). Taken together, we interpret these data to reflect RV remodeling due to lower maximal aerobic capacity, as opposed to frank impairments related to burn‐induced pulmonary dysfunction, and thus support a role for aerobic exercise training to improve cardiac function in well‐healed burn survivors.

Support or Funding Information

Project funded by NIH GM068865

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Circulating miR‐122‐5p: A Novel Exerkine in Exercise‐induced Angiogenesis
Authors: Wu Jie , Hou Zuoxu , Li Guohua , Zhang Xing , Gao Feng
Published: June 2021

Physical exercise effectively boosts angiogenesis in skeletal muscles, but the mechanisms remain poorly understood. Our previous study showed that 4‐week swimming training (1.5 h×2/d, 7 d/wk×4 weeks) significantly promoted angiogenesis in skeletal muscle. And we had found the expression of several circulating miRNAs profoundly changed. Among which, miR‐122‐5p, a liver specific microRNA, promoted endothelial proliferation significantly. Here we further determined the role of circulating miR‐122‐5p in exercise‐induced angiogenesis. Exercised mice (running wheel training, 1.5 h/d ×9 d) exhibited elevated miR‐122‐5p in serum and aorta (2.132±0.294, 3.275±0.819 fold of Sedentary, p<0.05), concomitant with increased capillary density in the quadriceps marked with fluorescent CD31 (3.562±0.354 fold of Sedentary, p<0.01). Whereas, tail vein injection of miR‐122‐5p antagomir partially decreased the exercise‐induced VEGF expression and CD31 staining in quadriceps, suggesting that elevated circulating mir‐122‐5p partially contributes to the exercise‐induced angiogenesis. In vitro study on primary aortic endothelial cells (ECs) transfected with miR‐122‐5p mimic showed increased tube numbers, migration distance and relative sprouting distance, accompanied with increased expression of p‐ERK/ERK and VEGF. Mechanistically, miR‐122‐5p mimic increased the expression of CD36, CPT1a and CPT1b together with the uptake of fatty acid assessed by BODIPY fluorescence intensity in ECs, suggesting an increased free fatty acid (FFA) metabolism. Transfection of CD36 siRNA blunted the pro‐angiogenic effect of miR‐122‐5p. In conclusion, our findings demonstrate that exercise‐induced proangiogenic effects are partly attributed to elevated circulating miR‐122‐5p via increasing endothelial FFA metabolism.

Support or Funding Information

This study was supported by the grants from the National Natural Science Foundation of China (Grant No. 816770253)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Exosomal Signaling by Skeletal Muscle: Role in Neuromuscular Ageing
Authors: Hemmings Kay , Girithananda Aarthi , Robertson Rebecca , Pugh Jamie , Scalabrin Mattia , Mora Silvia , McArdle Anne
Published: June 2021

Muscle mass and function are reduced with age in mammals and this is associated with disrupted neuromuscular interactions. Skeletal muscle mounts a robust stress response to contractions by the increased expression of Heat Shock Proteins (HSPs), which facilitate muscle remodelling, promote protein folding and clearance of damaged proteins. In contrast, other tissues, including neuronal cells are unable to mount a stress response. It has been proposed that HSPs may be transferred from one cell type to another in exosomes to maintain proteostasis in the recipient cells. Further, HSP generation by muscle following contractions is attenuated in old mice and humans. We hypothesise that this inability of muscles of old mice to produce HSPs in response to contractions results in altered exosomal HSP transfer, and therefore a failure to maintain proteostasis in neighbouring cells including motor neurons, with subsequent neuronal and muscle degeneration (1).

Single muscle fibres were isolated from flexor digitorum bravis (FDB) muscles of adult (6–12 month) mice, and initiated to contract using a non‐damaging electrical stimulation protocol. Media was collected at time points up to 24hrs, and exosomes were purified from the media using ultracentrifugation approaches. NanoSight analysis was used to characterize and quantify the exosomes, and HSP content analysed by western blotting.

Data demonstrated that there was a significant (2–3 fold) increase in exosome release by muscle fibres of adult mice following contraction compared with non‐stimulated controls, and that the exosomal HSP content was also increased. For example, at 4hrs post contraction, there was no increase in HSP25 content of exosomes but a 2 fold increase in HSP60 was detected at this time point compared with non‐stimulated controls. Preliminary data also showed that there was a 3.5 fold increase in HSP25 exosomal content at 24hrs post stimulation.

Additional data demonstrated the potential uptake of exosomes labelled with lipophilic carbocyanine fluorescent dye by immortalized neuronal cells. Additional studies will also examine the effect of age on exosome production by muscle fibres, including HSP content and neuronal uptake, by comparison with muscle from CuZn superoxide dismutase (CuZnSOD) null mice, a model of accelerated muscle ageing.

Support or Funding Information

With thanks to National Institute of Ageing (AG051442), the Physiological Society (summer studentship awarded to RR) and the University of Liverpool for their generous support.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A model of skeletal muscle showing the process of cramp and the mechanism of its relief
Authors: Tasaki Kazuyo Maria , Noble Penelope Jean , Garny Alan , Noble Denis
Published: June 2021

The research reported here is the first time that skeletal muscle cramp has been modelled using biophysically detailed processes, and the first time that the mechanism of its relief has been identified. We extended the Shorten et al (2007) skeletal muscle model to include sarcolemmal Ca2+ transporters, including ICaL, Na+‐Ca2+ exchange, and Ca2+ pump. When stimulated at 100 Hz the model successfully reproduces cramp, defined as a maintained contraction that does not immediately reverse when stimulation is stopped. The incorporation of ICaL is necessary for the model to reproduce the cramp process since block of ICaL, or using the original model without ICaL, removes the cramp mechanism. A further important process in maintaining cramp is accumulation of interstitial [K+] which produces large membrane depolarization. Relief of cramp is shown to require rapid washout of interstitial [K+]. The resulting hyperpolarization enables Na+‐Ca2+ exchange to promote Ca2+ efflux and so relieve the cramped state (Figure 1).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects of the Anticancer Drug Doxorubicin and Dietary Creatine Supplementation on Myogenic Regulatory Factor Expression
Authors: Busekrus Raquel B , Torok Zoltan A , Hydock David S
Published: June 2021

The purpose of this study was to explore the acute effects of doxorubicin (Dox) treatment on myogenic regulatory factor (MRF) expression and determine the impact that creatine monohydrate (Cr) feeding prior to Dox treatment has on the MRF response. Male Sprague‐Dawley rats were randomly assigned to one of three feeding groups: rodent chow supplemented with 2% Cr for four weeks (Cr1), rodent chow supplemented with 4% Cr for one week followed rodent chow supplemented with 2% Cr for three weeks (Cr2), or standard rodent chow for four weeks (Con). After four weeks, animals in each feeding intervention received a 15 mg/kg bolus Dox i.p. injection or a 0.9% bolus saline i.p. injection as a placebo (Sal). One day after injections, animals were anesthetized, and the extensor digitorum longus (EDL) muscles were excised. Expression of the primary MRFs Myf5 and MyoD and the secondary MRFs myogenin and Mrf4 were then quantified with Western blotting. Creatine feeding promoted an upregulation in Myf5 and MyoD (significant diet effect p=0.022 and p=0.017, respectively), and this upregulation was more profound with MyoD expression in Cr2. Significant diet effects were also observed with myogenin (p=0.0011) and Mrf4 (p<0.001) with Cr1 and Cr2 expressing higher levels of myogenin than Con+Sal (p<0.05) and Cr2 expressing higher levels of Mrf4 than Con+Sal and Con+Dox (p<0.05). In conclusion, Cr feeding prior to Dox treatment increased MRF expression suggesting that Cr may enhance signaling for skeletal muscle repair and regeneration following Dox treatment thereby attenuating muscle damage.

Support or Funding Information

David S. Hydock, PhD was supported by a Pilot and Exploratory Projects in Palliative Care of Cancer Patients and Their Families, PEP‐17‐193‐01 ‐ PCSM, from the American Cancer Society.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Skeletal Muscle Stretch‐Induced PGE2 Formation does not Impact the Muscle Mechanoreflex in Decerebrate Rats
Authors: Rollins Korynne , Butenas Alec L , Hopkins Tyler D , Felice Kennedy P , Copp Steven W
Published: June 2021

Mechanical and metabolic signals associated with skeletal muscle contraction stimulate thin fiber muscle afferents and produce reflex increases in blood pressure (i.e., the exercise pressor reflex). There is an interplay between those signals as metabolites may sensitize afferent responsiveness to mechanical signals. A common strategy used to study the mechanical component of the exercise pressor reflex (i.e., the mechanoreflex) in isolation from the metabolic component is to measure the reflex increase in blood pressure during passive hindlimb muscle stretch in an experimental animal preparation. Passive muscle stretch has been presumed to impart a purely mechanical stimulus. However, intermittent but not static mechanical stimulation of cultured skeletal muscle cells was found to increase PGE2 production and PGE2 has been shown to sensitize the muscle mechanoreflex. Based on those findings, the purpose of this investigation was to test the hypothesis that in decerebrate, unanesthetized rats, dynamic but not static hindlimb skeletal muscle stretch would increase interstitial PGE2 concentration. Moreover, we tested the hypothesis that COX inhibition with indomethacin would attenuate the stretch‐induced increase in PGE2 concentration and the increase in blood pressure during mechanoreflex activation. Experiments were performed on 28 young adult male Sprague‐Dawley rats. The microdialysis technique was used to collect triceps surae muscle interstitial fluid at rest and during 4 minutes of 1 Hz dynamic muscle stretch and 4 minutes of static muscle stretch (performed in random order and separated by 10 minutes). Muscle stretch was performed by manually turning the rack and pinion. We found that triceps surae muscle interstitial PGE2 concentration was significantly increased during both static (↑38%, p=0.01) and dynamic (↑56%, p<0.01) stretch above their respective baseline concentrations (n=9). Indomethacin (1μg/kg) significantly reduced baseline PGE2 concentration and prevented the increase in PGE2 during static (n=4) and dynamic (n=5) stretch. Importantly, indomethacin had no effect on the peak pressor response during static (control: 15±4, indomethacin: 16±4 mmHg; p=0.82) and dynamic (control: 17±2, indomethacin: 18±4 mmHg; p=0.68) stretch (n=7). Collectively, these data indicate that static and dynamic hindlimb muscle stretch in decerebrate, unanaesthetized rats increased muscle interstitial PGE2 concentration. The magnitude of increase in PGE2, however, was not sufficient to impact the muscle mechanoreflex.

Support or Funding Information

1R01HL142877 to SWC

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Attenuated Skeletal Muscle Contraction‐Induced Rapid Onset Vasodilation in African Americans
Authors: Kaur Jasdeep , Barbosa Thales C. , Young Benjamin E. , Stephens Brandi Y. , Nandadeva Damsara P. , Brothers R. Matthew , Fadel Paul J.
Published: June 2021

A number of studies have demonstrated that African Americans (AA) have a blunted ability to vasodilate at rest compared to Caucasian Americans (CA). Recently, our laboratory has demonstrated that AA have an attenuated forearm vascular conductance (FVC) response to rhythmic handgrip exercise at moderate and high intensities. Whether these differences in vasodilation also occur at the immediate onset of exercise remains unknown. Herein, we tested the hypothesis that AA subjects have an attenuated increase in FVC following a single muscle contraction, compared to CA subjects. A single muscle contraction model was chosen since it allows for isolation of rapid and robust local vasodilatory responses that are independent of large changes in systemic hemodynamics or sympathetic activation. Young, healthy AA (n = 4) and CA (n = 4) men performed single forearm contractions at 20%, 40%, and 60% of their maximum voluntary contraction (MVC). Heart rate (ECG), arterial blood pressure (Finometer), and brachial artery blood flow (duplex Doppler ultrasound) were continuously measured. FVC was calculated as forearm blood flow (FBF)/mean arterial pressure (MAP). Baseline FVC (CA: 1.04 ± 0.18 and AA: 1.31 ± 0.11 ml/min/mmHg; p = 0.26), FBF (CA: 99.5 ± 16.1 and AA: 105.9 ± 8.7 ml/min; p = 0.54), MAP (CA: 82 ± 2 and AA: 84 ± 2 mmHg; p = 0.46) and MVCs (CA: 56 ± 1 and AA: 58 ± 3 kg; p = 0.45) were similar between the groups. Following single forearm contraction, both groups exhibited intensity‐dependent increases in FBF and FVC; however, these responses were significantly attenuated in AA at all intensities compared to CA. For instance, in response to 20% MVC single contractions, AA had a 34 ± 10% increase in FVC from baseline compared to a 86 ± 31% increase in CA (mixed‐model ANOVA; p < 0.001), and in response to 60% MVC single contractions, AA had a 132 ± 9% increase in FVC from baseline compared to a 198 ± 30% in CA (mixed‐model ANOVA; p < 0.001). These preliminary data suggest that AA have blunted vasodilation in response to single forearm contractions at mild, moderate and high‐intensity workloads.

Support or Funding Information

Supported by UTA College of Nursing and Healthy Innovation

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Aged neutrophils fuel tumor progression: Good cells gone bad
Authors: Mittmann Laura Alice , Schaubaecher Johanna B. , Hennel Roman , Lauber Kirsten , Uhl Bernd , Reichel Christoph A.
Published: June 2021

Neutrophilic granulocytes (neutrophils) represent the first immune cells recruited to the site of injury or infection. Beyond their well‐established role in inflammatory pathologies, these leukocytes are increasingly recognized to modulate initiation, progression, and metastasis of cancer. Recently, ageing of mature neutrophils in the circulation has been identified to be critical for proper immune responses in inflammatory scenarios. The fate and function of aged neutrophils in malignant disorders, however, remain elusive. Here, we demonstrate for the first time that particularly aged neutrophils are recruited to malignant tumors as evidenced in heterotopic (multi‐channel in vivo microscopy) and orthotopic mouse models (multi‐channel flow cytometry) of mammary or head and neck carcinoma. Mechanistically, tumor‐released damage associated molecular patterns (DAMPs) including S100A8/9 and uric acid, but not high mobility group box 1 (HMGB1), initiate responses of these ‘experienced’ immune cells by activating the multimeric intracellular NLRP3 inflammasome (but not NLRP1, AIM2, or NLRC4 inflammasomes) in peritumoral macrophages. This leads to the release of cytokines that, in turn, activate microvascular endothelial cells, ultimately promoting the trafficking of aged neutrophils to neoplastic lesions. Most interestingly, aged neutrophils were found to exhibit a pro‐tumorigenic ‘N2’ phenotype. Accordingly, blockade of NLRP3 activation almost completely abolished responses of aged neutrophils in the peritumoral microvasculature as well as the subsequent infiltration of these immune cells into malignant lesions. Concomitantly, NLRP3 inhibition severely compromised tumor growth to the level of neutrophil‐depleted animals in these experiments, but did not directly affect the proliferation of cancer cells in vitro. Thus, our experimental data uncover a previously unappreciated role of aged neutrophils as potent regulators of tumor progression. Interfering with the recruitment of these immune cells to malignant lesions might emerge as a promising immunotherapeutic strategy to prevent advanced stages of cancer.

Support or Funding Information

This study was supported by Deutsche Forschungsgemeinschaft (DFG), Sonderforschungsbereich (SFB) 914.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Anti‐inflammatory effects of punicalagin on LPS‐activated microglia
Authors: Lee Yueh‐Shan , Lai Yi‐Xin , Li Chia‐Yang
Published: June 2021

Pomegranate (Punica granatum L.) is a popular fruit originated in the Middle East and has been widely planted around the world. It contains various phytochemicals such as polyphenolic compounds and has a wide range of biological activities, including anti‐inflammatory, anti‐tumor, antibacterial, and anti‐oxidant activities. Among many polyphenols of pomegranate, punicalagin is the main component of pomegranate polyphenols. However, the effect of punicalagin on immunoregulation has not been well examined. Since inflammation is associated with several neurodegenerative diseases, the inhibition of inflammatory response has benefit to delay the progression of the neurodegenerative diseases. Hence, the aim of this study is to examine whether punicalagin affects LPS‐induced inflammatory responses in BV2 murine microglial cells. Cell viability was analyzed by MTT assay. The production of nitric oxide (NO) was determined by Griess reagent assay. The expression of cyclooxygenase‐2 (COX‐2), inducible NO synthase (iNOS) and mitogen‐activated protein kinases (MAPKs)‐related proteins was examined by Western blot. The production of pro‐inflammatory cytokines, interleukin (IL)‐1β, IL‐6, and tumor necrosis factor‐α (TNF‐α), was detected using ELISA. Our experimental showed that punicalagin inhibits NO production, attenuates COX‐2 and iNOS expression, decreases IL‐1β, IL‐6 and TNF‐α production in LPS‐activated BV2 microglial cells. In addition, we also found that punicalagin inhibits the phosphorylation of ERK, JNK, and P38 in LPS‐activated BV2 microglial cells. Collectively, these results pointed out that punicalagin has anti‐inflammatory activities in LPS‐activated BV2 microglial cells and suggest that punicalagin might have benefit in attenuating brain inflammation and delaying the progression of the neurodegenerative diseases.

Support or Funding Information

This study was supported by the Ministry of Science and Technology, Taiwan, R.O.C. (Grant No. MOST 107‐2320‐B‐037‐019)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Myeloid Cell Recruitment and Activation Following Ozone Exposure in a Murine Model of Mutant Surfactant Protein‐C Pulmonary Dysfunction
Authors: Venosa Alessandro , Tomer Yaniv , Kopp Meghan , Gow Andrew J , Laskin Debra L , Beers Micheal F
Published: June 2021

The missense isoleucine to threonine substitution at position 73 [I73T] in the Surfactant Protein‐C [SP‐C] gene [SFTPC], an alveolar type‐2 cell [AT2] restricted protein, has been linked to sporadic and familial cases of idiopathic pulmonary fibrosis (IPF). We have developed a hypomorphic SP‐CI73T mouse model which expresses 20% of wild‐type SP‐C output. In this setting, AT2 cells mistraffick pro‐SP‐C on the plasma membrane and temporally acquire a late block in AT2 cell macroautophagy. This is associated with age‐dependent enlargement of alveolar septae, AT2 cell hyperplasia and minor inflammatory cell accumulation, beginning at 16 wk of age. Ozone is a ubiquitous air pollutant that causes parenchymal injury and monocyte/macrophage dominant inflammation. Clinically, acute exposure to ozone is known to exacerbate respiratory injury in susceptible population (i.e. asthma). Therefore, we tested whether SP‐CI73T dysfunction is associated with susceptibility to ozone induced injury (0.8 ppm, 3 h) in young (8–12wk) and aging (52 wk) mouse cohorts. We found that young WT mice exposed to ozone did not cause significant alterations in bronchoalveolar lavage (BAL) fluid immune cell composition or cell numbers at 24 h. Conversely, differential analysis of cytospins and flow cytometry analysis of tissue digest showed increased cell counts with significant neutrophilia and lymphocyte accumulation in SP‐CI73T mice. Analysis of aging cohorts confirmed significant deterioration of lung architecture in air‐exposed 52 wk SP‐CI73T mice. Ozone exposure was associated with histological evidence of perivascular edema and immune cells influx. BAL total protein analysis also showed increased epithelial‐vascular barrier disruption at 24 h and 48 h. Taken together, our findings support the concept that epithelial dysfunction driven by mutant SP‐CI73T expression is associated with increased susceptibility to environmental toxicant and that these effects are heightened with aging.

Support or Funding Information

Grant support: P30ES013508 (MFB), VA 1I01BX001176 (MFB), ES004738 (DL), ES005022 (DL).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.