bluuebunny/crossref_metadata_2025 huggingface dataset:
The Regulatory Role of microRNA-21 in Coronary Microcirculation in Metabolic Syndrome
Authors: Juguilon Cody , wANG YANG , Enrick Molly , Gadd James , Clark Alyssa , Yanez Katherine , Yin Liya
Published: May 2024

Coronary vasculature is essential to supply oxygen and nutrients to the heart and maintain normal cardiac function. Obstruction of large vessels of coronaries (atherosclerosis) is well studied, but coronary microvascular dysfunction (CMD), which is associated with diabetic cardiomyopathy, Takotsubo cardiomyopathy, Ischemia with the non-obstructive coronary artery (INOCA), and HFpEF, is understudied. CMD is characterized by impaired endothelial-dependent vasodilation, but detailed mechanisms have yet to be elucidated. The anti-fibrotic role of microRNA-21 (miR-21) is reported in the HFrEF induced by ischemia/reperfusion or pressure overload, but how miR-21 regulates CMD is not entirely understood. Our recent study shows miR-21 ablation ameliorated impaired vasodilation in isolated coronaries of mice fed on high fat and high sugar (HFHS) diet. In this study, we examined the coronary flow reserve (CFR), the relationship between myocardial blood flow (MBF) and cardiac work (doppler) under hyperemia, cardiac function (echocardiography and exercise exertion test), fibrosis and gene expression of miR-21 knockout and wild-type (WT) mice fed on chow or HFHS diet. Our preliminary data show that MBF and CFR were decreased, cardiac index, stroke volume, and running distance were reduced, and the E/E’ ratio was increased in WT mice fed on the HFHS diet compared to the WT mice fed on the chow diet. The perivascular fibrosis was increased in the WT mice fed on the HFHS diet compared to WT mice on the chow diet. The ablation of miR-21 reversed all these changes in the mice treated with HFHS, suggesting miR-21 deficiency ameliorated CMD in metabolic syndrome. Moreover, we used endothelial-specific miR-21 knockout to study the endothelial miR-21 function. Interestingly, the endothelial mi-R21 KO phenocopied the global knockouts, suggesting the benefit of miR-21 deficiency to coronary microvascular function in metabolic syndrome is from endothelial cells. The study provides the potential therapeutic strategy to target vascular endothelial cells with miR-21 antagomir for CMD in metabolic syndromes.

AHA 970663,1R56HL165207,1 R01 HL165207-01A1 to LY and 1 F31 HL156726-01A1 to CJ.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Mechanisms of Iron-Mediated Renal Injury in a Mouse Model of Sickle Cell Disease
Authors: Sako Mohammad , Patel Margi , Walker Savannah , Kasztan Malgorzata
Published: May 2024

Background: Sickle cell disease (SCD) is a hematologic disorder characterized by polymerization of the mutated hemoglobin S and sickling of red blood cells, leading to hemolysis, release of free heme/iron, and subsequent tissue iron accumulation. The kidney is one of the most frequently affected organs by iron overload, and chronic hemolysis is associated with chronic kidney disease severity in SCD. The potential role of iron overload in causing the observed renal injury in SCD has never been established. The goal of our study is to identify the impact of iron accumulation on the renal phenotype in SCD. We hypothesized that renal iron accumulation causes injury by oxidative stress dependent ferroptosis and/or mitochondrial dysfunction and subsequent kidney damage, that is worse in males than in females. Methods: We used 20-week-old male and female humanized SCD mice (HbSS) and age-matched genetic controls (HbAA). Mice were treated with either vehicle or the iron chelator Deferiprone (DFP, 100 mg/kg/day) for 8 weeks. Urinary excretion of protein and albumin as markers of renal injury were measured. Glomerular filtration rate (GFR) was measured transdermally after injection of FITC-sinistrin. Thiobarbituric acid reactive substance (TBARS) assay was performed for assessment of lipid peroxidation. Mitochondrial bioenergetics was assessed by Oroboros O2k Fluorometer of permeabilized renal cortices. Results: We previously demonstrated comparable iron accumulation in the kidney of male and female mice. However, HbSS males developed a more severe renal injury than females at 20 weeks of age, evidenced by proteinuria (2.52 ± 0.16 vs. 0.55 ± 0.16 mg/24hrs, p=0.0008) and albuminuria (76.44 ± 6.85 vs. 33.9 ± 8.81 μg/24hrs, p=0.0005). There was higher lipid peroxidation, a hallmark of ferroptosis, in the renal cortex of female HbSS than HbAA mice (1.31 ± 0.07 vs. 1.07 ± μM/mg, p=0.04). Bioenergetics studies revealed significantly lower state 3 oxygen consumption rate (108.2 ± 18.1 vs 164.8 ± 9.3 pmol/(s*mg), p=0.05), signifying lower oxidative phosphorylation capacity in HbSS males. In addition, there was significant reduction in state 4◦ respiration (36.4 ± 3.4 vs. 52.8 ± 3.4 pmol/(s*mg), p=0.03) in male HbSS mice. I also detected a decrease in maximum complex IV activity in male HbSS mice (325.5 ± 63.8 vs. 639.5 ± 51.3 pmol/(s*mg), p=0.02). Treatment with DFP improved proteinuria (2.66 ± 0.09 vs. 1.3 ± 0.14 mg/24hrs, p=0.0011) and GFR (165.3 ± 9.4 vs. 236.6 ± 12.6 μl/min, p=0.0058) in male HbSS mice. Conclusion: Iron accumulation causes renal injury which may be attributed to ferroptosis and mitochondrial dysfunction in SCD mice.

Funding: NHLBI—R00HL144817 and ASN Norman Siegel Research Grant to MK.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Mindfulness-based Stress Reduction and Endothelial Function in Chronic Kidney Disease
Authors: Cruz Julianna , Zanuzzi Matias , Jeong Jinhee , DaCosta Dana , Park Jeanie
Published: May 2024

Introduction: Chronic Kidney Disease (CKD) is associated with increased cardiovascular disease risk in part due to endothelial dysfunction. Studies have suggested that mindfulness-based stress reduction (MBSR) may have beneficial physiologic effects on cardiovascular disease risk; however, the potential beneficial effects of MBSR on endothelial function in CKD are unknown. We tested the hypothesis that MBSR improves endothelial function in patients with CKD. Methods: Participants with CKD (Stages III-IV) were enrolled and randomized to 8 weeks of MBSR versus the Health Enhancement Program (HEP), a structurally parallel control intervention to MBSR. Each intervention consisted of daily homework and 2.5hr weekly classes of mindfulness meditation (for MBSR) and health education (for HEP). Each intervention also included a day-long retreat. Endothelial function was quantified as brachial artery reactive hyperemia index (RHI) using peripheral artery tonometry (EndoPAT). RHI and offce blood pressure were obtained at baseline and at the end of the intervention. Intervention groups were also stratified into subgroups based on normal (RHI ≥2.10) and abnormal (RHI <2.10) baseline values for secondary analyses. Results: Data were obtained from 37 participants with CKD who were enrolled and randomized to MBSR (N=21) versus HEP (N=16). There were no differences in baseline characteristics, hemodynamics or RHI between groups. There were no significant differences in baseline versus end of study RHI within (HEP p=0.76; MBSR p=0.57) and between intervention groups (p=0.36). There was no difference in change in RHI after MBSR between the normal and abnormal RHI subgroups. Conclusions: MBSR did not have significant effects on endothelial function measured as RHI among individuals with CKD.

Supported by NIH Grants R33AT010457 and R01HL135183.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Female mice with menopause exhibit coronary microvascular disease
Authors: Yang Mingya , Li Xinrui , Ramirez Francisco , Makino Ayako
Published: May 2024

Menopause is a natural process as women age, and women with menopause are at higher risk of having cardiovascular diseases than men at the same age. Coronary microvascular disease (CMD) is one of the causes of angina, and the prevalence of CMD is higher in women than in men, and more postmenopausal women have CMD than premenopausal women. However, the causes of CMD in postmenopausal women are not clear. In this study, we investigate whether and how menopause leads to coronary microvascular disease in female mice. Menopausal mice were generated by administering a 20-day injection of 4-Vinylcyclohexene diepoxide (VCD, 160 mg/kg/day in sesame oil, i.p.). VCD gradually destroys primordial and primary follicles in ovaries and leads to ovary failure while keeping the ovary tissues intact; this model is close to the natural progression of human menopause. The vehicle, sesame oil, was administered in control mice. Coronary flow velocity reserve (CFVR) was measured to assess coronary microvascular function. Isometric tension was conducted in third-order coronary arteries (CAs). Capillary density was measured and compared in the left ventricle. Experiments were performed at 6 weeks (perimenopause period) and 8 weeks (post-menopause period) after the injection of VCD or vehicle in female mice. There was no significant difference in CFVR between VCD-injected mice and vehicle-injected mice 6 weeks after the injection. However, at 8 weeks after the injection, VCD-injected mice exhibited a significant decline in CFVR compared to the control mice. Coronary microvascular dysfunction can be driven by attenuated vascular relaxation in small coronary arteries and/or capillary rarefaction. We first conducted isometric tension and found that endothelium-dependent relaxation was significantly attenuated in VCD-injected mice at 8 weeks after the injection, but not at 6 weeks, compared to the controls. There was no significant difference in EC-independent relaxation between the two groups at any age. Interestingly, there was no significant difference in capillary density between VCD-injected mice and vehicle-injected mice at 6 weeks and 8 weeks after the injection. Postmenopausal mice, but not perimenopausal mice, develop CMD due to attenuated endothelium-dependent relaxation. This suggests that improving endothelium-dependent relaxation is the potential mechanism to treat CMD in postmenopausal women.

Supported by NIH R01HL142214 and DOD W81XWH2110472.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Effect of Maternal High Fat Diet on the Hypothalamus-Pituitary-Adrenal Gland (HPA) Axis of the Offspring in Sheep
Authors: Ahir Digishaben , Han Hyungchul , Jellyman Juanita
Published: May 2024

Maternal consumption of a high fat diet (HFD) during pregnancy is associated with metabolic dysfunction and altered HPA axis function in the offspring. These effects are well established in species with short gestation length (e.g., rodents), but less established in species with longer gestation periods more comparable with humans. The objective of the study was to determine the effect of maternal HFD on basal and stimulated cortisol secretion in the offspring at 16 weeks of age in sheep. All experiments were approved by the Institutional Animal Care and Use Committee at Cal Poly Pomona. Pregnant ewes were assigned to control (n=3) and HFD (n=3) groups. The control group was fed farmers best complete ewe and ram pellets. The HFD group was fed the same diet with an additional 30% rumen protected fat (Megalac, Volac Wilmar Feed Ingredients Ltd). After birth lambs were weaned at 12 weeks of age and given free access to control pellet diet with grains. To assess adrenal cortisol secretion at 16 weeks, lambs were infused intravenously with adrenocorticotropic hormone (ACTH; 2 ug/kg cortrosyn in 1 mL saline). Blood samples (1 mL) were taken through the jugular catheter at −30, −15, and 0 min (immediately before) and 5, 15, 30, 60, 90, and 120 min after ACTH administration for measurement of plasma cortisol by ELISA. Data were analyzed by two-way repeated measures ANOVA with a Tukey posthoc test. Basal plasma cortisol tended to be higher in HFD (32±8 ng/mL) than control (13±3 ng/mL) lambs, but this did not reach statistical significance. Plasma cortisol was higher than baseline from 15 to 90 minutes after ACTH administration in all lambs (p<0.05). The increment in plasma cortisol was higher in HFD than control lambs from 15-90 minutes (p<0.05). Accordingly, the maximum increase in plasma cortisol at 60 minutes and was higher in HFD (245±8 ng/mL) than control lambs (121±50 ng/mL p<0.05). These preliminary data suggest that maternal consumption of a high fat diet during pregnancy may increase basal and stimulated activity of the HPA axis in sheep.

The study was funded by a Strategic Interdisciplinary Research Grant (SIRG) at California State Polytechnic University, Pomona.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Neutrophil NADPH oxidase regulation by oxidative phosphorylation and accessory glucose metabolic pathways
Authors: Jobe Tyler , Stephan Jonah , Hill Bradford , Wysoczynski Marcin
Published: May 2024

Neutrophils produce reactive oxygen species (ROS) to extinguish pathogens and regulate intracellular signaling. The major producer of ROS in neutrophils is the NADPH oxidase, which is fueled by NADPH generated via the oxidative pentose phosphate pathway. It remains unclear how other accessory pathways of glucose metabolism and mitochondrial activity influence the oxidative burst in neutrophils. The respiratory burst in bone marrow-derived neutrophils was stimulated with phorbol 12-myristate 13-acetate (PMA) in the presence or absence of an inhibitor of glycogen breakdown (glycogen phosphorylase inhibitor; GPi), inhibitors of mitochondrial respiration (antimycin A and rotenone), or an inhibitor of serine biosynthesis (NCT503). Respiratory burst was also measured under conditions of limited glucose and glutamine. ROS production was measured via extracellular flux analysis or via dihydrorhodamine fluorescence. In both male and female mouse neutrophils, inhibition of glycogen breakdown by GPi delayed initiation of the oxidative burst in response to PMA (n = 6, p<0.05, Student’s T-test and Repeated Measure’s ANOVA). Inhibition of mitochondrial oxidative phosphorylation or inhibition of the serine biosynthetic pathway sustained ROS production in PMA-stimulated neutrophils (n = 6, p<0.05, Student’s T-test). Removal of glucose and glutamine showed a truncated oxidative burst (n = 6, p<0.05, Student’s T-test). Ultimately, these findings suggest that glycogen breakdown fuels the initial activation of the oxidative burst, while mitochondrial activity and the serine biosynthesis pathway control the magnitude and duration of neutrophil ROS production. Furthermore, utilization of exogenous glucose and glutamine are necessary for the entire oxidative burst. Future studies will continue to illustrate the mechanisms of neutrophil metabolic plasticity as well as connect their significance in the context of wound healing, exercise, and chronic inflammatory diseases.

NIH: R01HL141191, NIH: P01 HL078825.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Implications of alcohol use for aging-associated skeletal muscle dysfunction
Authors: Simon Liz , Bourgeois Brianna , Welsh David , Molina Patricia , Rodriguez-Graciani Keishla
Published: May 2024

Background: Alcohol use decreases skeletal muscle (SKM) strength and function. Alcohol use is disproportionately on the rise among older individuals, with 25% engaging in heavy drinking and 11% reporting binge drinking. The significant increase in life expectancy in the United States leads us to predict that alcohol use will be an important factor exacerbating SKM function in chronological aging, increasing the risk of poor physical performance and frailty. The objective of the study was to assess the impact of recent alcohol use on measures of physical performance and on muscle differentiation and bioenergetic capacity among people over the age of 60. Methods: We conducted a prospective cross-sectional study. Sedentary people over the age of 60 years were recruited from the Greater New Orleans Area. Participants were assessed for physical performance and vastus lateralis muscle biopsies taken for biochemical analysis. Recent alcohol misuse was defined as phosphatidyl ethanol (PEth), a biomarker of alcohol use, levels >20 ng/ml. Results: Preliminary analysis of data from 13 participants (PEth> 20 ng/ml, n=5) showed no differences in hand grip strength (Means± SD-31.08±9.8 and 33.17±8.4 kg respectively), 30-sec sit to stand test (10.5± 1.8 and 10.6± 3.9 number respectively), and Timed up and go Test (13.5± 2.6 and 11.4± 3.1 seconds respectively) among participants with PEth < 20 ng/ml compared to those with PEth >20ng/ml. Data collected from myotubes derived from myoblasts isolated from SKM biopsies from participants with PEth >20 ng/ml showed increased proton leak (indicator of poor mitochondrial bioenergetic health) and lower myotube density (p=0.06, Cohen’s d = 1.9) compared to myotubes from participants with PEth <20 ng/ml. Conclusions: Preliminary analysis indicates that recent alcohol use dysregulates muscle bioenergetic and differentiation capacity that are not associated with significant differences in physical performance tests. Ongoing analysis will identify confounding factors, and whether cumulative lifetime alcohol use contributes more significantly to physical performance and frailty measures among older people.

R21AA030869, Pilot funding Department of Physiology LSUHSC-NO, P60AA009803.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
An Acute Bout of Subconcussive Head Impacts Does Not Change Intracranial Pressure
Authors: Ryan Caleb , Gorini Pereira Felipe , Olesh Margaret , Mick Lauryn , Kawata Keisuke , Johnson Blair D.
Published: May 2024

Purpose: Subconcussive head impacts (SHI) are known to cause a variety of negative sequelae including increased plasma concentrations of neuronal structural and glial proteins, attenuated neurovascular coupling, and impaired vestibular function. Traumatic brain injuries are associated with an increase in intracranial pressure (ICP). However, it is unknown whether an acute bout of SHI increases ICP. We tested the hypothesis that estimates of ICP via ultrasound assessments of optic nerve sheath diameter (ONSD) would be greater following a single session of repetitive SHI versus a control condition. Methods: 14 healthy participants (age: 19±2 y; BMI: 21±4 kg/m2, 3 women) with at least 3 y of soccer heading experience completed two experimental conditions in a randomized crossover design. 20 soccer headings were used to induce repetitive SHI and 20 soccer kicks were used as the control condition. A soccer ball was projected from a soccer JUGS machine (40 km/h, 12.2 m away) every 30 s for both conditions. ONSD assessed via ultrasound is associated with direct measures of ICP. Subjects were in the supine position during ONSD (transverse plane, each eye; B-mode linear array ultrasound), heart rate (HR; 3-lead ECG), and mean arterial blood pressure (MAP; photoplethysmography) measurements conducted at baseline prior to heading/kicking and at 2 h, 24 h, and 72 h post. ONSDs were measured 3 mm under the bulb in triplicate for each eye and the mean values were used for analyses. Inter-eye ONSD values were also averaged for analyses. Clinical intervention to alleviate elevated ICP is indicated when ONSD is > 5.0 mm. Data were analyzed using a mixed-effects model with repeated measures with time and condition as within subject factors. Data are presented as mean ± SD if statistically significant. Results: No subjects had an ONSD > 5.0 mm in either eye following heading or kicking. Left ONSD was greater than baseline at 2 h post heading (2.89 ± 0.25 mm vs. 2.99 ± 0.27 mm; p = 0.0157) and greater than baseline at 2 h (2.92 ± 0.37 mm vs. 2.98 ± 0.31 mm; p = 0.0352) and 72 h (3.08 ± 0.26 mm; p = 0.0067) post kicking. Left ONSD at 72 h post kicking was greater than heading (3.08 ± 0.26 mm vs. 2.93 ± 0.25 mm; p = 0.0022). There were no interaction effects for right ONSD (p = 0.4686), average ONSD (p = 0.1178), HR (p = 0.1039) or MAP (p = 0.3889). Conclusion: Neither the heading nor kicking intervention elicited a clinically relevant ONSD that would require medical attention. Contrary to our hypothesis, an acute bout of 20 soccer headers did not evoke greater ONSD in healthy subjects versus kicking. Therefore, our data indicate that a single session of SHI does not increase ICP.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Generation of an Inducible Avp Knockout Mouse Line Using CRISPR/Cas9 and Cre/LoxP
Authors: Khan Shaza , Chen Lihe , Khundmiri Syed , Chou Chung-Lin , Knepper M.A.
Published: May 2024

Background: Arginine vasopressin (AVP) is a peptide hormone coded by the Avp gene, synthesized in the hypothalamus and secreted by the posterior pituitary. Dysregulation of AVP secretion contributes to a variety of human diseases. Previous studies of functional roles of AVP have been largely dependent on the use of Brattleboro rats, which manifest a spontaneous mutation in the Avp gene and lack circulating AVP. Despite their utility, Brattleboro rats are diffcult to breed owing largely to the fixed nature of the Avp mutation, resulting in increased neonatal death and behavioral effects in the adults. Consequently, commercial breeders have ceased production, despite a continued need. To fill this need, we have now created a mouse model with a tamoxifen-inducible deletion of AVP. Methods: We employed CRISPR/Cas9 to target the desired regions (first intron and 3' downstream) within the Avp gene. Successful insertion of the two loxP sites was confirmed by PCR using primers flanking the targeted regions. Restriction enzymes BamHI or EcoRI were used to confirm correct targeting. For additional confirmation, PCR products were cloned into the TA-cloning vector and sequenced. Mice harboring the floxed allele were mated to B6.Cg-Tg(CAG-cre/Esr1*)5Amc/J mice that globally express a tamoxifen-inducible Cre recombinase. Results: The resultant homozygous conditional knockout mice ( Cre+ Avpflx/ flx) are viable, fertile, and show no signs of polydipsia or polyuria prior to induction, indicating that the conditional alleles retain their wild-type function. Induction of Cre-mediated recombination by administration of tamoxifen (2-3 mg q.d., i.p. for 5 days) in 8-week-old mice resulted in a decrease in urine osmolality from 2076±138 to 122±6 mOsm/kgH2O on day 31 after induction (p<0.0001). Sanger sequencing demonstrated the expected 1245 bp deletion at the Avp locus. The kidney parenchyma was structurally normal with no evidence of medullary atrophy 6 weeks after induction. Immunoblotting of AQP2 in the inner medulla showed a significant decrease in AQP2 band density in Cre+ Avpflx/ flx mice to 27±14 % of values in Cre− floxed control mice (p<0.05). Conclusion: We developed an inducible AVP knockout mouse line ( Cre+ Avpflx/ flx) that will be shared with the research community and are likely to be useful for further study of regulation of water balance and polycystic kidney disease, as well as neural, vascular, and metabolic roles of vasopressin.

The work was funded by the Division of Intramural Research, National Heart, Lung and Blood Institute.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Ventilatory long-term facilitation following acute intermittent hypoxia in unanesthetized rats: influence of short hypoxic episodes and changes in metabolic rate
Authors: Jones Aaron , Mitchell Gordon
Published: May 2024

Plasticity is a hallmark feature of the neural system controlling breathing. Acute intermittent hypoxia (AIH) elicits plasticity in multiple respiratory motor pools ( e.g., phrenic, intercostal, XII, etc.) and has emerged as a promising therapeutic approach to restore lost respiratory (and non-respiratory) function in people with spinal cord injury and other clinical disorders that compromise movement. In anesthetized rats during the rest phase, AIH-induced phrenic long-term facilitation is enhanced when the AIH consists of short (1-min) versus longer (5-min) moderate hypoxic episodes (PaO2 ~40-50 mmHg), even with the same cumulative duration of hypoxia (15-min). However, since respiratory plasticity is influenced by a variety of behavioral and physiological factors such as sensory feedback, lung/chest wall mechanics and arousal state, the effectiveness of specific AIH protocols may differ when implemented in unanesthetized rats. Since it is unknown how hypoxic episode duration impacts ventilatory long-term facilitation (vLTF) in unanesthetized rats during the rest phase, we initially tested the hypothesis that short (1-min) hypoxic episodes (FIO2 = 0.09) elicit vLTF in young male Sprague-Dawley rats via whole-body plethysmography; metabolic CO2 production was also assessed. Following 15, 1-min hypoxic episodes, significant vLTF was observed, and this effect was more pronounced when ventilation was normalized to metabolic rate ( i.e. VE/VCO2; p < 0.05). AIH-induced vLTF was accompanied by hypometabolism and a post-hypoxic temperature decrease (p < 0.01), consistent with previous work. Additional studies will be performed to compare these results with “traditional” AIH protocols ( i.e., 5-min hypoxic episodes), and across the rats’ rest versus active phase. In perspective, these results set the stage for future investigations concerning the optimization of AIH protocols in unanesthetized rodent models, including rodent models of injury or disease.

NIH HL148030 & 147554 (Mitchell).

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
C2 Spinal Cord Hemisection Reduces Mitochondrial Volume Density in Phrenic Motor Neurons
Authors: Haggaz Ali , Fogarty Matthew , Mantilla Carlos B. , Sieck Gary C.
Published: May 2024

The diaphragm muscle (DIAm) is the major inspiratory pump for breathing. Neural control of the DIAm during breathing involves the recruitment of fatigue resistant motor units comprising smaller phrenic motor neurons (PhMNs), which innervate type I and IIa fibers. More fatigable DIAm motor units comprise larger PhMNs that innervate type IIx/IIb fibers, which generate greater forces but are more fatigable. The fatigable motor units are only infrequently recruited for expulsive behaviors of the DIAm. Thus, there are marked differences in the activation history of DIAm motor units. Consistent with their higher activity levels, previous studies from our lab have shown that smaller PhMNs and type I and IIa DIAm fibers have greater mitochondrial volume density (MVD). PhMNs located in the cervical spinal cord (C3-C5) receive direct excitatory input from medullary premotor neurons, which is primarily ipsilateral. Thus, C2 spinal cord hemisection (C2SH) disrupts the neural pathway essential for breathing and significantly reduces DIAm activity on the same side. Therefore, we hypothesized that C2SH significantly reduces MVD of smaller PhMNs. In Sprague Dawley rats, PhMNs on both sides were retrogradely labeled by intrapleural injection of cholera toxin B (CTB) 3 days prior to C2SH (right side). In addition, DIAm EMG activity was recorded on both sides by inserting bipolar electrodes. Mitochondria were labeled by intraspinal injection of MitoTracker Red and imaged in identified PhMNs using 3D confocal microscopy. A series of optical slices (0.5 mm) of PhMNs were deconvolved to improve contrast, then thresholded for MitoTracker labeling followed by 3D reconstruction, providing detailed images of mitochondrial morphometry and distribution within PhMNs. In sham rats, the MVD of smaller (lower somal surface area) PhMNs was higher compared to larger PhMNs. C2SH reduced DIAm EMG activity on the ipsilateral side across a 14-day period, while activity on the contralateral side increased. After 14 days, C2SH reduced MVD by ~25% in smaller PhMNs on the ipsilateral side. We conclude that the reduced activity of smaller PhMNs induced by C2SH induced mitochondrial morphological changes, most likely due to reduced mitochondrial biogenesis. These results indicate that reduced DIAm activity associated with mechanical ventilation may impose mitochondrial remodeling that reduces mitochondrial oxidative capacity in PhMNs. These adaptations may disrupt mitochondrial function and impair weaning from mechanical ventilation.

Research support by NIH grants HL166204, HL146114 and AG44615.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Effects of aging and exercise on Mitochondria-ER contacts (MERCs) in skeletal muscle
Authors: Allen Ryan J. , Kronemberger Ana , Pereira Renata O. , Anderson Ethan J. , Shi Qian , Song Long-Sheng , Lira Vitor A.
Published: May 2024

Mitochondrial dysfunction is a prominent factor contributing to age-associated loss of muscle mass and force (a.k.a., sarcopenia). Yet, events preceding age-related mitochondrial dysfunction, which may allow the development of new therapies, remain poorly understood. Physical coupling of the outer mitochondrial membrane (OMM) to the endoplasmic or sarcoplasmic reticulum (ER), termed mitochondria ER contact sites (MERCs), modulate several mitochondrial processes influencing cellular function. In an energetically demanding and primarily post-mitotic tissue, we hypothesized that alterations in MERCs correlate with the development of sarcopenia. We also hypothesized that MERCs respond to exercise training and may contribute to beneficial muscular remodeling in older adults. To address these, we studied three age groups of male C57BL6N mice: young adults (Y, 5 mo. of age), old (O, 20 mo. of age), and very old (VO, 31 mo. of age). An additional group of old mice underwent 8-9 weeks of treadmill training (OET). Muscle mass and contractile function decreased with aging; VO animals displayed ~25% decline in maximum tetanic force vs. Y mice (p<0.0001). Fatigability (assessed via repetitive sub-maximal contractions) increased by ~10% with aging but was attenuated in the OET group (p<0.001). In saponin-permeabilized muscle fibers, aging did not affect mitochondrial oxidative capacity. However, H2O2 emission was nearly doubled in the O group (p<0.01) and this trend was partially alleviated by exercise training. Assessed via TEM, aging decreased the size of MERCs (i.e., length of SR and OMM apposition) by around 20% (p<0.01). Interestingly, OET animals displayed ~50% increase in MERC coverage (i.e., proportion of OMM covered by SR; p<0.0001). Immunoblots of cellular MERC fractions revealed that MERC proteins associated with calcium exchange were altered with age and exercise. The OMM channel, VDAC1, was reduced by 20% in O muscles (p=0.02), while Grp75, a tether protein, was ~25% higher in OET (p=0.02). Quantitative proteomic analyses revealed various MERC proteins being modified by aging and exercise. Our findings indicate that changes in MERC structure and protein composition may underlie early events preceding the development of sarcopenia and that exercise-mediated adaptations at MERCs may contribute to the beneficial adaptations in aging skeletal muscle.

Supported by the University of Iowa Fraternal Order of Eagles Diabetes Research Center (UIowa FOEDRC).

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Effects of aging on vasoconstrictor responses of diaphragm arterioles
Authors: Kenney Nathan , Horn Andrew , Schulze Kiana , Weber Ramona , Poole David , Behnke Bradley , Musch Timothy
Published: May 2024

Introduction: In health, the diaphragm is the most important inspiratory muscle and the only skeletal muscle that is continuously recruited throughout life. Because of the diaphragm’s importance to generating the exercise hyperpnea and wealth of evidence that diaphragm fatigue and exhaustion can impair exercise tolerance, we were interested in whether the exercise intolerance that attends older populations might be associated with altered diaphragm vasomotor control. While it is known that aging compromises peripheral skeletal muscle vasomotor control, it is not known if similar impairments are present in the respiratory muscle vasculature, namely the diaphragm. Therefore, we tested the hypothesis that advanced age decreases diaphragm arteriole contractile responses to alpha-adrenergic agonists. Methods: 4–5 month old Fischer rats and 22–24 month old Fischer rats were divided by age into two groups: young (Y) and old (O). In Y and O, diaphragm first order (1A) arterioles (~200μm diameter, ~2μm in length) were isolated, cannulated, and pressurized to develop spontaneous tone. Following this, contractile responses to cumulative doses of norepinephrine (NE) and phenylephrine (PE) (10−9 to 10−4M) were determined. Results and Conclusion: The physical characteristics of the two groups differed as O were heavier than Y. However, with respect to the diaphragm 1A arterioles, there were no differences in spontaneous tone development (%) or maximal diameter (μm) between Y (38 ± 4%; 132 ± 19μm) and O (37 ± 4%; 157 ± 14 μm; both P >0.05). In diaphragm 1A arterioles, the maximal vasoconstrictor response to NE was also not different between O and Y animals (O: 37 ± 8% vs. Y: 45 ± 1%; p > 0.05). However, the maximal vasoconstrictor response to PE was greater in Y compared to O (O: 29 ± 3% vs. Y: 39 ± 5%; p ≤ 0.05). These data support that aging may reduce a1 adrenergic receptor mediated vascular control, altering the vasoconstrictive responsiveness of diaphragm resistance vessels. We are increasing our group sizes to test the robustness of this conclusion.

This work was supported by the Sustained Momentum for Investigators with Laboratories Established (SMILE) Grants awarded to T.I.M and D.C.P. by Kansas State University College of Veterinary Medicine and the National Institute on Aging 1R15AG078060 awarded to B.J.B and D.C.P. A.G.H and K.M.S. were financially supported by Ruth L. Kirschstein National Research Service Awards 1F31HL167618-01 and 1F31HL170643-01, respectively.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Disruption of ER Stress CHOP in beta cells protects from HFpEF pathogenesis
Authors: Srinivas Balaji , Fortuno Paula , Belmadani Souad , Rhaleb Nour-Eddine , Matrougui Khalid
Published: May 2024

Introduction: Heart Failure with Preserved Ejection Fraction (HFpEF) is a multifactorial cardiovascular disorder characterized by diastolic dysfunction, hypertension, and metabolic disturbances. In this study, we aimed to investigate the effect of endoplasmic reticulum (ER) stress C/EBP homologous protein (CHOP) in beta cells on the development of HFpEF. Methods: Eight-week-old male mice, both CHOPflox/flox and CHOPbeta cells −/− were divided into four groups: control ER stress CHOPflox/flox and ER stress CHOPbeta cells −/− mice subjected to standard diet and water. ER stress CHOPflox/flox and ER stress CHOPbeta cells −/− mice fed a high-fat diet (HFD) and L-NAME (0.5 g/L) for five weeks. A comprehensive cardiovascular, metabolic, and histological evaluation was conducted. Results: Following five weeks of HFD and L-NAME, ER stress CHOPflox/flox mice exhibited both clinical and molecular manifestations of HFpEF, encompassing diastolic dysfunction (elevated E/E' ratio), normal cardiac ejection fraction, hypertension, metabolic aberrations, cardiac hypertrophy paired with fibrosis pulmonary (lung) edema, renal injury with fibrotic changes, and reduced exercise tolerance. Vascular endothelial dysfunction was observed. Western blot analysis unveiled a diminished expression of phosphorylated endothelial nitric oxide synthase (p-eNOS) in mesenteric resistance arteries (MRA), concomitant with qRT-PCR data revealing elevated levels of inflammatory markers (Cox2, iNOS, NLRP3) and ER stress markers (CHOP, ATF6, PERK, XBP1, IRE1) in both MRA, heart, and pancreas. Interestingly, ER stress CHOPbeta-cells−/− mice subjected to an HFD and L-NAME were protected from the development of HFpEF and its associated pathologies. These mice displayed improved cardiac and vascular endothelial function, reduced ER stress, and decreased inflammatory factors compared to their ER stress CHOP flox/flox. Conclusion:Our research findings indicate that ER stress CHOP in beta cells plays a pivotal role in the development of HFpEF. Thus, targeting ER stress CHOP in beta cells could be a promising strategy for preventing HFpEF.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Retinoic acid modulation guides human-induced pluripotent stem cell differentiation towards left or right ventricle-like cardiomyocytes
Authors: Zhang Hengliang , Sen Payel , Hamers Jules , Sittig Theresa , Dendorfer Andreas , Merkus Daphne
Published: May 2024

Background: Cardiomyocytes (CMs) induced from human induced pluripotent stem cells (hiPSCs) by traditional methods are a mix of atrial and ventricular CMs and other non-cardiomyocytes. Retinoic acid (RA) plays an important role in regulation of the spatiotemporal development of the embryonic heart. Aim: Create engineered heart tissue (EHT) with left and right ventricular phenotype from hiPSCs by intervening with specific concentrations of RA during hiPSC differentiation towards CM. Methods: hiPSC were derived by reprogramming skin fibroblasts and erythroid progenitors. Different concentrations of RA (Control without RA, LowRA with 0.05 μM and HighRA with 0.1 μM) were administered during third to sixth days of the differentiation process. EHTs were generated by assembling CMs derived from hiPSC (hiPSC-CM) at high cell density in a low collagen hydrogel. EHTs were further matured and grown in a customized biomimetic tissue culture system, that provides continuous electrical stimulation, medium agitation and stretch. Finally, RNA extraction and tissue fixation were performed on CMs and EHTs for RT-qPCR and immunofluorescence staining analysis, while RNA sequencing was conducted on EHTs to examine how RA affects gene expression associated with function and structure of EHTs. Results: HighRA-hiPSC-CMs exhibited highest expression of maturity genes MYH7 and cTnT. The expression of TBX5, NKX2.5 and CORIN, which are transcription factors associated with left ventricular development, was also the highest in the HighRA-hiPSC-CM. The expression of TBX5 and NKX2.5 was also found to be highest in the HighRA-EHTs, while expression of right ventricular transcrition factors TBX20 and ISL1 was highest in control-EHTs. RNAseq of EHTs showed augmented extracellular matrix gene expression in HighRA-EHTs. The HighRA-EHTs were functionally most similar to left ventricle with the highest contraction force, the lowest beating frequency, and the highest sensitivity to hypoxia and isoprenaline. Conclusion: By interfering with the differentiation process of hiPSC with a specific concentration of RA at a specific time, we were able to successfully induce CMs and EHT with a phenotype similar to that of the left ventricle or right ventricle. Our research paves the way for future studies on in vitro left ventricular or right ventricular function, personalized drug screening, and precision medicine.

German Center for Cardiovascular Research (DZHK81Z0600207 to D.M. and P.S.) China Scholarship Council (CSC202108410141) and Henan Provincial Medical Science and Technology Research Project.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Piezo2 sensory innervation during kidney development
Authors: McLarnon Sarah , N’Guetta Pierre , Tassou Adrien , Geron Matan , Scherrer Gregory , O'Brien Lori L
Published: May 2024

Piezo2 is a mechanically activated ion channel expressed in a subset of sensory neurons, including in the bladder. Our lab has recently mapped innervation of the developing kidney and found that renal nerve development begins at embryonic day 13.5 (E13.5) and is tightly aligned with smooth muscle actin (SMA+) arterial development. However, the presence and spatiotemporal innervation of Piezo2+ sensory nerves in the kidney remains unknown. In the current study, we hypothesized that Piezo2+ sensory nerves would be present early in kidney development adjacent developing nephrons and that these axons innervating the kidney could be traced back to Piezo2+ cell bodies in the dorsal root ganglia (DRGS). To test this hypothesis, we performed immunofluorescent staining to label all nerves (Tubb3+, pan-neuronal marker) and Piezo2 in kidneys from wildtype mice at early (E13.5), mid (E15.5) and late (post-natal day 0, P0) nephrogenesis. To determine the signaling pathways of Piezo2+ nerves in the kidney to the peripheral nervous system (PNS), we utilized retrograde tracing with a Cre-driven fluorescently (pAAV-Flex-tdTomato) labeled virus injected into the kidney of heterozygous Piezo2Cre/+ adult mice (n=4, 10 weeks). The injected kidney, the contralateral control, and cervical to sacral DRGs were collected and fluorescent imaging was performed with confocal microscopy. We found that by E15.5, Piezo2+ sensory nerves were present in the developing kidney which constitute a small subset of all nerves (Tubb3+). These Tubb3+Piezo2+ sensory nerves persist through nephrogenesis (P0) and in the adult mouse kidney. Piezo2+ sensory nerves were found in nerve bundles running along SMA+ vasculature as well as observed adjacent glomeruli (Nphs2+) and proximal tubules (Lrp2+). Importantly, we also identified Piezo2+ sensory nerves in the renal cortex from human kidney samples. Retrograde tracing experiments revealed Piezo2+ sensory nerves in the kidney originate from lower thoracic and lumbar DRGs. Taken together, these data suggest that Piezo2+ sensory nerves are present in the mouse and human kidney and are capable of transmitting signals back to the PNS. As Piezo2 is known to be expressed in juxtaglomerular cells, Piezo2+ sensory nerves observed in close proximity suggests a previously unrecognized kidney cell-nerve crosstalk. Future studies will investigate Piezo2+ sensory activity and feedback in the kidney to understand the role of Piezo2 in renal mechanosenstion.

NIH R01DK121014 to LO.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
CD8 T cells exert a critical role in the transition from left heart failure to lung remodeling and right ventricular hypertrophy
Authors: Wang Dongzhi , Weng Xinyu , Chen Yingjie
Published: May 2024

Chronic left ventricular (LV) failure or heart failure (HF) often progress to pulmonary remodeling and right ventricular (RV) hypertrophy even with optimal medical care. Cytotoxic CD8+ T cells play an important role in modulating virus infection and tissue injury by producing proinflammatory cytokines and cytolysis of the infected or injured cells. However, the role of CD8+ T cells in heart failure (HF)-induced lung remodeling and RV hypertrophy is unknown. To determine the role of CD8 T cells in modulating HF progression under preexisting HF conditions, wild type mice with existing LV failure produced by transverse aortic constriction (TAC) were randomized to depletion of cytotoxic CD8+ T cells, regulatory T cells (Tregs), or both by using specific blocking antibodies. The cardiac function, lung inflammation, fibrosis, vascular remodeling, and right ventricular remodeling were determined. As anticipated, LV failure caused lung inflammation and activation of pulmonary CD4+ T cells and CD8+ T cells. Interestingly, depletion of CD8+ T cells dramatically attenuated the increase of lung weight, lung inflammation, lung vascular remodeling, and RV hypertrophy in mice with existing LV failure. LV failure was associated with an increased ratio of activated T cells to Tregs, and Treg depletion exacerbated lung inflammation and the progression of LV failure. Treg depletion also exacerbated the lung CD4+ and CD8+ T cell infiltration, and the percentage of effector memory T cells (Tem) in HF mice. Depletion of CD8+ T cells was suffcient to rescued HF mice from the exacerbated lung inflammation and the progression of LV failure that was caused by Treg depletion. These findings demonstrate that CD8+ T cells play a critical role in promoting the transition from LV failure to lung remodeling and RV hypertrophy.

NIH funding.

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.