bluuebunny/crossref_metadata_2025 huggingface dataset:
JGA‐Derived CCN1 is a Systemically Acting Angiogenic Hormone
Authors: Desposito Dorinne , Moon Ju‐Young , Riquier‐Brison Anne , Peti‐Peterdi Janos
Published: November 2023

The kidneys are major regulatory organs of whole body homeostasis including body fluid and electrolyte balance, blood pressure maintenance, and blood cell production. Renal control mechanisms of these key systemic physiological functions include the local production and secretion of peptide hormones, and their actions at distant organ sites. Classic examples are erythropoietin, and renin, which is released from the highly perfused juxtaglomerular apparatus (JGA). The present studies aimed to test the hypothesis that the newly identified JGA‐derived cysteine‐rich protein CCN1 is released into the blood and serves as a novel systemic angiogenic hormone promoting general cardiovascular health. Blood was collected before and at different times after bilateral nephrectomy in both mice and rats. Plasma CCN1 levels measured by CCN1 ELISA and immunoblotting continuously decayed after nephrectomy, suggesting that the kidney/JGA is a significant source of systemic CCN1. Immunostaining of mouse and human kidneys showed a robust and constitutive expression of CCN1 in the JGA. The tube formation assay of glomerular endothelial cells (GEnCs) was performed to test the effect of exogenous CCN1 on in vitro angiogenesis. CCN1 appeared to be more effective on in vitro angiogenesis than the gold‐standard angiogenic factor VEGF. Other classic in vitro assays of GEnC cell proliferation and cell migration (scratch opening assay) showed similar robust effects of CCN1. In vivo, CD34 immunostaining of kidney tissues collected from C57BL/6 mice treated with subcapsular injection of CCN1 peptide showed a significant increase of angiogenesis compared to untreated mice. Kidney, aorta, heart, and brain tissues were collected from mice with JGA‐specific CCN1 KO before and three weeks after tamoxifen induction, and sections stained for TUNEL assay of apoptosis. Compared to nearly undetectable levels in normal tissues before, a high number of vascular endothelial cells in all tissues stained TUNEL+ after JGA CCN1 deletion, indicating the presence of widespread systemic endothelial injury when renal CCN1 synthesis is compromised. These findings provide strong support for our new concept that JGA‐derived CCN1 is released into the blood, acts as a new systemic angiogenic hormone, and is essential for maintaining vascular integrity throughout the body.

Support or Funding Information

NIH R01 64324, AHA, ADA
Adenosine A1 receptor‐operated calcium entry in renal afferent arteriolar smooth muscle cells is dependent on postnatal maturation of canonical transient receptor potential isoform 3 channels
Authors: Peixoto‐Neves Dieniffer , Soni Hitesh , Buddington Randal K , Adebiyi Adebowale
Published: November 2023

Adenosine, a regulator of cardiovascular development and renal function constricts renal afferent arterioles by activating its cognate A1 receptors (AA1Rs). Activation of AA1Rs elevates intracellular Ca2+ ([Ca2+]i) concentration in renal vascular smooth muscle cells, mechanisms of which are not fully resolved. Whether AA1R expression and function in preglomerular microvessels are dependent on postnatal kidney maturation are also unclear. Here, we show that selective activation of AA1R by 2‐chloro‐N6‐cyclopentyladenosine (CCPA) does not stimulate store‐operated Ca2+ entry in afferent arterioles isolated from infant pigs. However, CCPA‐induced [Ca2+]i elevation is dependent on phospholipase C, protein kinase C, and canonical transient receptor potential isoform 3 (TRPC3) channels. The basal [Ca2+]i concentration was unchanged in afferent arterioles isolated from newly farrowed (0 day) pigs when compared with their 20‐day old counterparts. Whereas, CCPA‐induced [Ca2+]i elevation was ~ 43% larger in afferent arterioles from 20‐day old pigs. AA1R protein expression levels in the kidneys and afferent arterioles were unaltered in 0‐day versus 20‐day old pigs. By contrast, TRPC3 channel protein expression was ~ 92 and 78% higher in 20‐day old pig kidneys and afferent arterioles, respectively. These data suggest that activation of AA1Rs elicits receptor‐operated Ca2+ entry in porcine afferent arterioles, an effect dependent on postnatal maturation of TRPC3 channels.

Support or Funding Information

NIH R01 DK101668
Role of gap junctions in renal vascular conducted response
Authors: Moller Sophie
Published: November 2023

The juxtaglomerular apparatus (JGA) is an essential structure in the regulation of renal function and embodies two major functions: tubuloglomerular feedback (TGF) and renin secretion. Both mechanisms function to regulate renal microcirculation, in addition to a plethora of systemic effects. The JGA is a unique structure to study intercellular communication. Intercellular connections, gap junctions (GJ), allow exchange of ions, nutrients, and small signaling molecules between neighboring cells. GJs consist of two connexons, one from each cell, with connexins (Cx) as their building blocks. Several Cx isoforms are expressed in the JGA [1], making the JGA a functional syncytium. Disruption of the signaling pathways in the JGA is associated with reduced TGF response, dysregulation of renin secretion, and hypertension [2,3]. The aim of the study was to determine if reduced intercellular communication affects the conduction of TGF‐induced vasoconstriction in the afferent arteriole.

Experiments were performed using the isolated perfused juxtamedullary nephron preparation. Kidneys were isolated from wild type (WT) and knockout (KO) of both Cx40 and Cx45 mice. Kidneys were perfused with a Tyrodes buffer containing 5% BSA and an amino acid mixture, at pH 7.4. The inner renal surface contains a unique nephron population and allows access to the afferent arteriole. Local afferent vasoconstriction was induced by electrical pulse stimulation (300ms, 90V), administered via 2M NaCl filled microelectrode (0.5–0.8 MΩ) at the glomerular entrance. These electrical stimulations emulate the initiation of vascular TGF responses. Renal perfusion pressure was kept constant at 95 mmHg. Inner afferent arteriolar diameter was measured at the stimulation site (0 μm) and every 50 μm, at distances upstream (up to 450 μm), following a protocol consisting of a 5 min acclimation period, 30 sec baseline, 30 sec electrical pulse stimulation, and 30 sec recovery measurements. Responses between WT and KO of both Cx40 and Cx45 mice were compared.

Preliminary data shows that electrical stimulation at the glomerular pole of the afferent arteriole reduces the diameter in all four groups at the local stimulation sites, with the vasoconstriction effect attenuated at upstream sites. A loss of vasoconstriction in response to stimulation is observed at 200 μm in Cx40 KO mice, and at 250 μm in Cx45 KO mice. In the WT mice, attenuation in the vasoconstriction is observed further upstream, at 450 μm in Cx40 WT mice. In the Cx45 WT mice no attenuation is observed at the measured distance of 300 μm.

These results suggest that both Cx40 and Cx45 may play a significant role in the propagation of the afferent vasoconstriction elicited by electrical stimulation, and thus in the TGF signaling in the JGA.
A New View of Macula Densa Cell Biology
Authors: Shroff Urvi Nikhil , Perin Laura , Peti‐Peterdi Janos
Published: November 2023

Macula densa (MD) cells of the juxtaglomerular apparatus (JGA) perform important, well‐characterized physiological regulatory functions including tubuloglomerular feedback‐mediated control of glomerular hemodynamics and renin release. Established chemical mediators synthesized and released from MD cells include ATP, adenosine, PGE2, and NO that are of non‐protein nature. However, the unique features of MD cell organelles, the apically located giant nuclei and the robust size of protein synthesis organelles at the cell base suggest that the MD is a massive protein factory. The present study aimed to characterize the basic MD cell biological processes and their regulatory mechanisms. Immunofluorescence localization of the key molecular players in DNA synthesis, cell cycle regulation, protein synthesis, processing, and vesicular release was performed in fixed mouse and human kidney sections. The high level and MD‐specific expression of the key cell cycle regulators Geminin and Wee1 suggested that MD cells are in cell cycle arrest in the G2/M phase. Staining for γ–tubulin revealed multiple centrosomes in some MD cells suggesting DNA endoreduplication (endocycling) and multiploidy. The robust and MD‐specific labeling of secreted angiogenic peptides (Ccn1, Pappa‐2, Gdf15) was localized in numerous small intracellular vesicles, suggesting the high rate of synthesis of these proteins and their release. The high level of MD‐specific expression of unique elements of the actomyosin motor complex (Myh10, Ablim1, Dctn2, Gsn) was also consistent with the presence of a well‐controlled vesicular release (exocytosis) machinery. In summary, MD cells have a unique cell biology and genetic program to power a robust synthetic and secretion machinery of various novel MD‐specific proteins that likely perform important functions in renal tubulo‐interstitial, vascular, and glomerular tissue remodeling.

Support or Funding Information

NIH R01 64324, AHA, ADA
Altered expression of aquaporin water channels in a rat model of chronic diarrhea due to bile acid malabsorption
Authors: Yde Jonathan , Borg Johan , Fenton Robert A. , Moeller Hanne B.
Published: November 2023

Background

Bile acid malabsorption (BAM) is a condition where excess amounts of bile acids (BA) in the colon cause chronic watery diarrhea. BAM occur following resection or inflammation of the terminal ileum, cholecystectomy or as a primary condition, where the feedback regulation of bile acid (BA) synthesis is defective. The mechanisms by which BAs in the large intestine cause diarrhea are largely unknown. My aim is to determine if aquaporin water channels (AQPs) in the colonic epithelium are involved in the development of diarrhea during BAM.

Methods

Six rats kept in metabolic cages were fed a diet containing 1% BA for 10 days as a model for BAM. Six other rats fed standard chow served as controls. Colonic epithelial cells were isolated by Ca2+‐chelation. Cells were processed for RNA and protein analyses. Colonic tissue was also prepared for histology.

Results

BA‐fed rats had greater feces dry weight and feces water content compared to rats fed standard chow. BA rats also had a higher water intake than controls. There were no differences in urine output or urine osmolality between groups. In contrast to controls, BA rats did not gain weight over time. The difference in weight was significant at the end of the experiment. Semi‐quantitative real‐time PCR of epithelial cells showed that organic solute transporter β (Slc51b) mRNA was increased in the colonic epithelium from the BA‐treated group compared to the control group indicating activation of the nuclear bile acid receptor (FXR). Aqp3 mRNA was increased in the distal colonic epithelium in the BA‐treated group, while mRNA for Aqp8 was increased in both the proximal and the distal colonic epithelium. mRNA for Aqp1, ‐4 and ‐7 was not different between groups. AQP3 was detected as a non‐glycosylated and a glycosylated form by western blotting. The glycosylated form was significantly reduced in the distal colonic epithelium of BA rats compared to controls. AQP8 was significantly increased in the distal colonic epithelium of BA rats compared to controls.

PAS‐staining showed no clear differences in the histology of the colons from BA rats compared to colons from control rats. The expression pattern of AQP3 and ‐8 as shown with immunohistochemistry was not different between BA rats and controls.

Conclusion

In a rat model of BAM we find increased expression of Aqp3 and Aqp8 mRNA and alterations of protein expression of AQP3 and ‐8 in the distal colon indicating a role for AQPs in the development of diarrhea during BAM.

Support or Funding Information

Support for my PhD‐study comes from the Faculty of Health's Research training supplement as well as from my main supervisor, Hanne Moeller, who received a Starting grant from Aarhus University Research Foundation, when she was hired as associate professor.
A Study on Mechanism and Potential Application of Novel Chalconoid Derivatives as Anti‐Secretory Therapy for Diarrheal Diseases
Authors: Yibcharoenporn Chamnan , Chavasiri Warinthorn , Chatsudthipong Varanuj , Muanprasat Chatchai
Published: November 2023

Secretory diarrhea is one of major health problems that mainly results from Cl− secretion through apical Cl− channels including cystic fibrosis transmembrane regulator (CFTR) and calcium‐activated chloride channel (CaCC). Naturally derived chalcone (isoliquiritigenin or ISLQ) has previously been shown to inhibit CFTR with IC50 of about 20 μM without affecting CaCC. The aims of this study were to optimize chalconoid derivatives for their inhibitory effect against both CFTR and CaCC, and to explore their underlying mechanisms. The screening of chalconoid derivatives using short‐circuit measurements in T84 cells, a human intestinal epithelial cell line, identified an chalcone derivative CHAL‐025 that inhibited both CFTR and CaCC with higher potency than ISLQ (IC50 for CFTR 1–5 μM and IC50 for CaCC 3–10 μM). CHAL‐025 at concentrations up to 20 μM showed no cytotoxic effect on T84 cells. The inhibition of CFTR‐mediated chloride secretion was reversible and only observed when CHAL‐025 was added at the apical side. In addition, IC50 of CHAL‐025 in inhibiting CFTR did not change when basolateral membrane was permeabilized, suggesting that CHAL‐025 is apically active and the inhibitory effect of CHAL‐025 is insensitive to membrane potential. CFTR inhibitory potency of CHAL‐025 was not affected by pretreatment with inhibitors of protein kinase A (PKA), protein phosphatase (PP) and phosphodiesterase (PDE), indicating that mechanisms of CFTR inhibition by CHAL‐025 did not involve PKA, PP and PDE. Mechanism of CaCC inhibition by CHAL‐025 was also investigated and will be presented. Importantly, CHAL‐025 inhibited chloride secretion induced by bile acid across T84 monolayers, which is an experimental model mimicking bile acid‐induced diarrhea. In conclusion, the present study identified a chalcone derivative that is of potential utility in the treatment of secretory diarrheas resulted from both CFTR and CaCC overstimulation.

Support or Funding Information

This work was supported by grants BRG5980008 and DBG5980001 from the Thailand Research Fund and Mahidol University and by the Faculty of Science, Mahidol University and the Medical Scholars Program, Mahidol University.
A Screen for Potassium‐sensitivity in Yeast Reveals Endosomal Sorting Factors that Regulate the Renal Outer Medullary Potassium (ROMK) Channel
Authors: Mackie Timothy D. , Welling Paul A. , Subramanya Arohan R. , Brodsky Jeffrey L.
Published: November 2023

The renal outer medullary potassium (ROMK) channel is a major route for potassium secretion from distal nephron principal cells in response to changes in dietary potassium intake. In addition, ROMK acts in tandem with the sodium‐potassium‐chloride cotransporter‐2 (NKCC2) to facilitate sodium reabsorption in the thick ascending limb. Genetic perturbations in ROMK cause an autosomal recessive salt‐wasting disorder in newborns known as Bartter Syndrome Type II, while these same mutations provide heterozygote individuals with resistance to developing hypertension. To date, ROMK anterograde trafficking through the secretory pathway and endocytosis from the plasma membrane in renal epithelia are well studied, however little is known about the factors that regulate either the post‐endocytic fate of ROMK or the fate of ROMK in the early secretory pathway. To rectify this deficiency in our understanding of how ROMK is regulated, we developed an assay utilizing a genetic screening technique in the budding yeast Saccharomyces cerevisiae. In brief, yeast were rendered sensitive to low potassium by deletion of the two native potassium transporters, Trk1 and Trk2. ROMK was then transformed into these cells, and crossed with a collection of 5,000 unique strains, each of which have a single ORF deleted. Strains that grow better on low potassium media are hypothesized to be deficient in negative regulators of ROMK. Several factors related to endosomal sorting were identified through this screen, including members of the class‐C core vacuole‐endosome tethering (CORVET) complex, which brings endocytic vesicles together to form early endosomes, as well as the endosomal complexes required for transport (ESCRT), which sort ubiquitinated cargo into multi‐vesicular bodies prior to degradation in the lysosome/vacuole. These findings were validated by serial dilution growth assays in independently constructed yeast strains. Next, these findings were validated in a more relevant model. Cell surface biotinylation of HEK293 cells treated with siRNA to knockdown human orthologues of screen hits indicated that CORVET and ESCRT also deplete ROMK from the plasma membrane in human cells. These genetic data lead us to propose that the plasma membrane population of ROMK is endocytosed and sorted to the lysosome for degradation in a CORVET and ESCRT‐dependent manner. In addition, hypothesis‐driven studies in yeast showed that ROMK is also partially regulated at the level of the endoplasmic reticulum (ER) via ER‐associated degradation (ERAD). Together, these findings enhance our understanding of the cellular itinerary of this critical ion channel and may lead to the development of novel therapies for patients with Bartter syndrome and other disorders of potassium balance.

Support or Funding Information

This work was supported by grants GM75061 and DK79307 to J.L.B from the National Institutes of Health
Effect of Diuretics and Gender on Kcnj16‐mediated Effects in Salt Sensitive Hypertension
Authors: Palygin Oleg , Levchenko Vladislav , Prudnikov Oleh , Nguyen Christine , Staruschenko Alexander
Published: November 2023

Hypokalemia is a potentially life‐threatening imbalance characterized by a serum potassium concentration of less than 3.5mmol/L (severe hypokalemia < 2.5mmol/L) caused by a deficit in total body potassium stores induced by dietary, genetic or iatrogenic factors. Inwardly‐rectifying K+ channels Kir4.1 (Kcnj10) and Kir5.1 (Kcnj16) are highly expressed on the basolateral membrane of distal renal tubules and contribute to Na+ reabsorption and K+ secretion through the direct control of trans‐epithelial voltage. To define the importance of Kir5.1 in electrolyte balance and potassium homeostasis in the body, we generated a Kcnj16 knockout in Dahl salt‐sensitive (SS) rats (SSKcnj16−/−). SSKcnj16−/− rats fed a normal 0.4% NaCl diet (Dyets) had severe hypokalemia (3.8±0.03 vs 2.00±0.06, mmol/L; SS vs SSKcnj16−/− rats correspondingly, p<0.05, N≥9) and reduced blood pressure. Female rats had an even more profound change in blood potassium (3.5±0.04 vs 1.55±0.04 mmol/L; SS vs SSKcnj16−/− rats correspondingly, p<0.05, N≥11). Overall fractional excretion analyses further revealed that SSKcnj16−/− rats had a salt‐wasting phenotype with increased FENa+, FEK+, and FEMg2+. When fed a high salt diet (4% NaCl; Dyets) SSKcnj16−/− rats experienced 100% mortality triggered by diuresis‐natriuresis and a further drop of blood potassium level below 1.4mmol/L. To reduce mortality and preserve potassium wasting under high salt conditions, we supplemented the drinking water of the SSKcnj16−/− rats with inhibitors of sodium channels and transporters critical for maintenance of electrolyte homeostasis in the Thick Ascending Limb (TAL) (furosemide; 150mg/L), Distal Convoluted Tubule (DCT) (Hydrochlorothiazide (HCTZ) 75mg/L) and Cortical Collecting Duct (CCD) (benzamil 15mg/L) (N≥6 for males and females in each treatment group). Epithelial sodium channel (ENaC) inhibitor benzamil, but not other diuretics, rescued SSKcnj16−/− rats from mortality on the high salt diet, but removal of benzamil from the drinking water after two weeks of high salt treatment still resulted in mortality. Interestingly, treatment with HCTZ delayed mortality only in males and had no effect in females, emphasizing the role of the DCT segment specifically in male SSKcnj16−/− rats. Together, our data with diuretics targeting NKCC, NCC and ENaC revealed that only benzamil treatment rescued SSKcnj16−/− rats when challenged with a high salt diet, and furthermore we have demonstrated gender differences important in regulation of Kcnj16‐mediated effects in the kidney.

Support or Funding Information

Medical College of Wisconsin Neuroscience Research Center/Advancing a Healthier Wisconsin pilot grant #9520217 (to OPa) and National Institutes of Health grants HL108880, HL122662 (to AS)
The Divergent Redox Sensitivities of d‐ and a‐ epithelial sodium channel (ENaC) subunits
Authors: Helms My N , Wu Wei
Published: November 2023

In mice, ENaC is a heterotrimer with three well characterized subunits (αβγ). In humans, an additional ENaC subunit (δ1) and its slice variant (δ2) has recently been identified. The physiological roles of δ‐ENaC subunits are not clear. Using the Xenopus laevis oocyte expression system, full‐length cDNAs for human ENaC subunits, and two electrode voltage clamp measurements, we modeled oxidative stress and compared the effects of oxidized glutathione (GSSG) and H2O2 on αβγ‐ENaC and δβγ‐ENaC whole cell current. High extracellular GSSG treatments (400μM) increases the redox potential, and models antioxidant depletion. 400μM GSSG significantly decreased ENaC activity in mouse lung cells and in oocytes expressing αβγ‐ENaC. GSSG, however, increased whole cell current in oocytes expressing δ1βγ‐ENaC by 38% (n=12). Likewise, whole cell current increased 27% in δ2βγ‐ENaC expressing oocytes (n=13) following GSSG perfusion. High extracellular H2O2 perfusion (10μM) did not change αβγ‐ENaC whole cell current in oocytes (n=6). However, 10μM H2O2 perfusion increased δ1βγ‐ENaC whole cell current 13% (n=8), and increased δ2βγ‐ENaC whole cell current 19% (n=8) within 1 min in oocytes. The H2O2‐induced increase in δ2βγ current was not reversible following washout. The divergent effects following GSSG and H2O2 exposures observed in αβγ‐ and δβγ‐ENaC expressing oocytes may have significant relevance in health and disease. Additional studies are needed to clarify the role of δ1 and δ2 ENaC subunits in vivo.
Cerebrovascular Responses to Trigeminal Nerve Stimulation in Humans
Authors: Ahmed Amar , Edwards Louisa , Braz Igor Dutra , Fisher James P
Published: November 2023

The mammalian diving reflex, evoked by cold water face immersion, is characterized by peripheral vasoconstriction, an increase in blood pressure (BP) and a concomitant reduction in heart rate (HR). Trigeminal nerve stimulation (TGS) partly mediates these reflex induced cardiovascular adjustments, which are thought to conserve blood flow and oxygen for delivery to the brain and heart. We aimed to characterise the cerebrovascular responses to TGS under isocapnic conditions, and those evoked by generalized sympathetic activation during thermoreceptor stimulation with ice water hand immersion (cold pressor test; CPT).

In eight healthy men (23±6 years) continuous recordings of middle cerebral artery velocity (MCA Vmean; transcranial Doppler), HR (ECG), mean BP (Finometer) and the partial pressure of end‐tidal carbon dioxide (PETCO2) were obtained. Cerebral vascular conductance index (CVCi) was calculated as; MCA Vmean/MAP. Participants underwent 3 minutes of CPT, with ice water (4°C) right hand immersion, and 3 minutes of TGS evoked by ice packs placed on the forehead and cheeks. Trials were randomized and separated by a 20‐minute resting period. Supplemental CO2 was added to the inspired air in order to maintain isocapnia throughout the TGS and CPT trials.

During TGS a decline in heart rate was observed, while mean BP rose progressively (p<0.05). MCA Vmean did not change significantly with TGS (baseline; 56.9 ± 3.4 cm/s, minute 3; 56.2 ± 3.3 cm/s, p>0.05), while CVCi was reduced (baseline; 0.79 ± 0.50 cm/s, minute 3; 0.66 ± 0.47 cm/s, p<0.05). In contrast, CPT was associated with a transient increase in HR during the first minute, and a constant elevation in mean BP (p<0.05). MCA Vmean was significantly increased during CPT (baseline; 57.5 ± 2.9 cm/s, minute 3; 64.6 ± 3.6 cm/s; p<0.05) and CVCi decreased (baseline; 0.73 ± 0.41 cm/s, minute 2; 0.64 ± 0.38 cm/s, p<0.05). PETCO2 remained unchanged from baseline in both TGS and CPT trials.

These preliminary findings suggest that cerebral perfusion is elevated during generalized sympathetic activation evoked by the CPT, whereas MCA Vmean remains unchanged during TGS under isocapnic conditions.

Support or Funding Information

JPF is funded by the British Heart Foundation.
Fatigue and Motor Unit Activation Are Determined by Neither Load Nor Time Under Tension During Resistance Exercise
Authors: Morton Robert , Sonne Michael , Zuniga Amanda Farias , Jones Amanda , Keir Peter , McGlory Chris , Phillips Stuart
Published: November 2023

Recent evidence has established that repetition load determines neither acute rates of muscle protein synthesis or resistance training‐induced hypertrophy. Our objective was to extend on these findings by examining if skeletal muscle hypertrophy following resistance exercise training is instead determined by the degree of motor unit activation. Ten recreationally active young men (mean ± SD; 21 ± 3 y, 177 ± 6 cm, 82 ± 9 kg) completed three sets of unilateral knee extension exercise to volitional failure under four conditions that varied in the speed of contraction (time under tension [TUT]) and load (% one repetition‐maximum [%RM]): 80%RM Regular (80R; 1 eccentric: 1 pause: 1 concentric), 80%RM Slow (80S; 3:1:3), 30%RM Regular (30R; 1:1:1) and 30%RM Slow (30S; 3:1:3). Each participant completed two training sessions separated by one week, performing two of the four conditions in each session (one on each leg). Electromyography (EMG) and maximum voluntary forces (MVF) were used to assess motor unit activity and fatigue, respectively. Load was higher in the heavy‐load (80R and 80S) conditions (145 ± 18 vs. 55 ± 7 kg), the TUT per repetition was higher in the slow (80S and 30S) conditions (5.4 ± 0.6 vs. 2.8 ± 0.5 s), repetitions per set were different in each group such that 80S < 80R < 30S < 30R (80S: 6 ± 1, 80R: 9 ± 2, 30S: 14 ± 4, 30R: 20 ± 4), volume was lower in the slow conditions (2556 ± 776 vs. 3668 ± 1076 kg) and the total TUT including each repetition was different in all conditions such that 80R < 80S < 30R < 30S (80R: 76 ± 20, 80S: 99 ± 17, 30R: 158 ± 19, 30S: 225 ± 52, p<0.05). The EMG amplitude was higher in the heavy‐load conditions during the first (65 ± 15 vs. 46 ± 18 %MVE) and last (89 ± 26 vs. 72 ± 32 %MVE, p<0.05) repetitions of each set. The EMG amplitude increased during each condition (23 ± 18 %MVE) but the integrated EMG (iEMG) signal was highest in the light‐load (30R and 30S) conditions (27 ± 13 vs. 16 ± 6 %MVE·s, p<0.01). MVF decreased between each set in every condition (pre: 240 ± 32, set 1: 218 ± 33, set 2: 204 ± 29, set 3: 194 ± 34 N, p<0.01). Total TUT was significantly correlated with iEMG (r=0.41, p=0.009). These data demonstrate that fatigue during resistance exercise training is unrelated to load or TUT when resistance exercise is performed until volitional failure. In addition, significant motor unit activation occurs when resistance exercise is performed until volitional failure even at lower loads and, although amplitude may be higher with heavier loads, the total (integrated) EMG response is greater during resistance exercise using lighter loads.
Does maximal isokinetic power at exercise intolerance approximate critical power?
Authors: Yong Stanley , Swisher Austin , Ferguson Carrie , Cannon Daniel T
Published: November 2023

Tolerance to high‐intensity exercise is described by a hyperbolic power‐tolerable duration (P‐tLIM) relationship. The asymptote of the P‐tLIM relationship, termed critical power, demarcates sustainable from non‐sustainable exercise. However, characterizing this relationship is time consuming – typically 4 constant power exercise tests to intolerance are required. Alternatively, at the limit of tolerance to supra‐critical power exercise, a 3 min all‐out sprint approximates critical power (Murgatroyd et al., Eur J Appl Physiol 114:1863–74, 2014). We aimed to determine whether maximal isokinetic power (30 s in duration) measured immediately at the limit of tolerance approximates critical power.

Ten participants (3 females, 7 males, 22±2 yr, 73±11 kg, VO2peak 3.62±0.87 L.min−1) completed constant supra‐critical power exercise tests to intolerance, immediately followed by a maximal isokinetic effort at 80 rpm for 30 s. Mean isokinetic power over the final 20 s was compared to the critical power asymptote determined from four variable‐duration constant power tests to intolerance. Mean bias and limits of agreement were calculated using the method of Bland & Altman.

Critical power estimated from the P‐tLIM relationship was 165±54 W with 95% confidence interval of 157, 172 W. Of the four constant power tests used to define P‐tLIM, isokinetic power was not different to critical power only following the longest duration trial (p>0.05, 186±65 vs 165±54 W). However, mean bias and limits of agreement were 21 W and ±88 W, respectively. In the remaining trials, isokinetic power immediately following the limit of tolerance was greater than critical power (p<0.05).

Isokinetic power measured immediately following the limit of tolerance consistently overestimated critical power. While not different to critical power, the closest estimation showed more than 20 W bias with limits of agreement nearly 12 times the span of the 95% confidence interval. Thus, brief maximal isokinetic power (30 s) immediately following the limit of tolerance does not approximate critical power.
Aortic Pulsatility and Arterial Stiffness After Acute Combined Resistance and Endurance Exercise
Authors: Wakeham Travis R , Greenlund Ian M , Durocher John J
Published: November 2023

Endurance exercise reportedly decreases arterial stiffness, while resistance exercise can increase arterial stiffness. Previous training studies suggest that increases in arterial stiffness might be prevented by combining the two modes of exercise within a workout, especially when resistance exercise is completed before endurance exercise. However, the post‐exercise aortic hemodynamic responses to acute combined exercise, and the effect of order, have not been studied. Therefore, we hypothesized that post‐exercise aortic pulsatile load (aortic pulse pressure × heart rate) and carotid‐femoral pulse wave velocity (cfPWV) would be lower after a resistance then endurance (RE) exercise session compared to an endurance then resistance (ER) exercise session. We examined 15 healthy adults (24±2 years; 12 males and 3 females) before and after RE and ER in a randomized order, with approximately one week between testing sessions. Training sessions incorporated 30 minutes of running at 70% of age‐predicted maximal heart rate and approximately 30 minutes of resistance training, which included six common exercises (three sets of 10 repetitions for each). Aortic pulsatile load was 39±10% above baseline at 30‐min post ER, whereas it was only 28±10% above baseline at 30‐min post RE (P<0.02 between conditions). The changes in cfPWV from pre‐ to 30‐min post‐RE (5.8±0.3 vs. 5.5±0.2 m/s) were similar to those from pre‐ to 30‐min post‐ER (5.7±0.2 vs. 5.4±0.2 m/s). Thus, transient increases in cfPWV following exercise might be prevented by combining resistance and endurance exercise within the same workout. Additionally, the lower pulsatile load after RE could result in less pulsatile stress to end organs. These findings may be beneficial when prescribing exercise to healthy or subclinical populations.
IL‐6 Contributes to Exercise Induced Recovery of Hepatic Insulin Action in Obese Mice
Authors: Peppler Willem T. , MacPherson Rebecca E.K. , Panasevich Matthew R. , Meers Grace M. , Rector R. Scott , Wright David C.
Published: November 2023

Obesity is associated with both systemic and hepatic insulin resistance. After only a single bout of exercise, the obesity‐induced impairment in hepatic insulin action is recovered; however, the biological mechanisms that underlie this phenomenon are not known. As circulating levels of interleukin‐6 (IL‐6) are increased after a single bout of exercise, we hypothesized that IL‐6 may contribute to the exercise‐induced recovery of hepatic insulin action in obese mice. To evaluate this hypothesis, we used three experimental approaches. First, C57BL/6 male mice were fed a low fat diet (LFD, 10% kcals from lard) or high fat diet (HFD, 60% kcals from lard) for 7 weeks, which led to increases in body mass, adiposity, and impaired glucose tolerance. Mice from the HFD group were then subjected to a single bout of exercise (2 hours of treadmill exercise at 15 m·min−1) or cage control, allowed to recover for 2 hours, and the liver was collected 15 minutes after IP insulin (1 U·kg1). Acute exercise recovered HFD‐induced impairments in insulin‐induced phosphorylation of protein kinase B (AKT) at serine 473 (pAKTSer473), a key insulin signaling protein. This occurred alongside increases in circulating IL‐6 and led to increases in indices of hepatic IL‐6 signaling, including phosphorylation of signal transducer and activator of transcription 3 (pSTAT3) and mRNA expression of suppressor of cytokine signaling 3 (SOCS3). Second, to determine whether IL‐6 was required for the recovery of induced‐insulin pAKT Ser473 with exercise, mice were injected with a neutralizing antibody against IL‐6 (IL‐6 NA) or an IgG control 1 hour prior to exercise. The IL‐6 NA completely ablated exercised‐induced increases in hepatic pSTAT3, which occurred in parallel with an attenuated recovery of pAKTSer473, suggesting that IL‐6 contributes to exercise‐induced recovery of hepatic insulin action. Finally, to determine if IL‐6 directly regulates hepatocellular pAKTSer473, primary hepatocytes were isolated from lean mice and treated with free fatty acid (FFA: 250 μM oleate, 250 μM palmitate) for 24 hours as a means to increase lipid accumulation and insulin resistance. Cells were then treated with IL‐6 for 90 minutes, including insulin stimulation for the final 15 minutes. IL‐6 induced increases in pSTAT3 protein content were attenuated by FFA treatment. In addition, the FFA‐induced reduction in insulin‐stimulated pAKTSer473 was not rescued by IL‐6 treatment. Together, this data suggests that the recovery of hepatic insulin action after exercise in obese mice may involve an IL‐6 dependent mechanism; however, the effect of IL‐6 may not be directly in the hepatocytes.

Support or Funding Information

This work was partially supported by a NSERC grant (341158 to DCW) and a Veterans Affairs VHA‐CDA2 grant (IK2BX001299 to RSR).
Blunted Autonomic Response To Head Up Tilt In Individuals With Intellectual Disabilities: Preliminary Results
Authors: Hilgenkamp Thessa , Schroeder Elizabeth C. , White Daniel W. , Baynard Tracy , Fernhall Bo
Published: November 2023

Individuals with intellectual disabilities (ID) have limited cardiorespiratory capacity, not explained by lack of motivation or lack of understanding the testing procedures. In individuals with ID who have Down syndrome, low cardiorespiratory capacity has been linked to autonomic dysfunction. Previous research suggests autonomic dysfunction may also be present in individuals with ID without Down syndrome but this has not been tested.

PURPOSE

To compare the autonomic response to head up tilt of individuals with ID to a control group without ID.

METHODS

Thirteen individuals with ID and 12 individuals without ID were instrumented with an ECG‐lead and finger‐plethysmography for continuous heart rate and blood pressure recordings. After resting supine for 10 min, participants underwent 5 min head‐up tilt at 70 degrees with an electronic tilt table, followed by a 10 min supine recovery. The last five minutes of every position was used to calculate cardiovascular measures and time‐domain heart rate variability measures as non‐invasive indices of autonomic function.

RESULTS

Individuals with ID showed different responses compared to individuals without ID for stroke volume, R‐R‐interval (RRI), root mean square of successive differences (RMSSD) and the proportion of times the change in consecutive intervals exceeds 50 milliseconds (pNN50) (p<0.05; Table).

CONCLUSION

These preliminary results suggest a blunted response to head‐up tilt in individuals with ID, but our findings need to be confirmed with a larger sample.

Support or Funding Information

The research leading to these results has received funding from the People Programme (Marie Curie Actions) of the European Union's Seventh Framework Programme (FP7/2007–2013) under REA grant agreement n°625455.

Results in means (standard deviation). RRI=beat‐to‐beat interval, RMSSD= root mean square of successive differences, pNN50= the proportion of times the change in consecutive intervals exceeds 50 milliseconds

Baseline Supine
Head up tilt
Recovery Supine
Position × Group interaction effect

ID
Control
ID
Control
ID
Control
p‐value

Stroke volume
108 (28)
108 (23)
94 (35)
73 (14)
110 (30)
107 (23)
0.009**

Cardiac output
6.9 (1.5)
5.9 (1.1)
7.1 (2.1)
5.6 (0.8)
7.0 (1.5)
5.9 (1.2)
0.308

RRI mean
962 (111)
1098 (128)
808 (104)
772 (104)
972 (129)
1088 (124)
<0.001**

RMSSD
63 (24)
101 (46)
33 (16)
31 (16)
66 (30)
101 (49)
0.008**

pNN50
36 (17)
53 (16)
12 (12)
10 (11)
37 (19)
53 (19)
0.015*

p<0.05,

p<0.01
Pedometer‐determined physical activities of daily living can affect blood pressure regulation in healthy young adults after postural change
Authors: Yang Hongxia , Kotaki Yumika , Hoshi Junko , Bao Sarina , Tanno Hiromasa , Kanno Emi , Maruyama Ryoko
Published: November 2023

Background

Numerous studies have examined the effects of physical activity (PA) on blood pressure (BP) or autonomic function through exercise interventions. However, few studies have used non‐intervention methods to observe the association between pedometer‐determined physical activities of daily living and BP and autonomic function in healthy young adults.

Objective

The purpose of this study was to examine whether the differences in pedometer‐determined physical activities of daily living cause the variation in BP and autonomic nerve activity in healthy young adults.

Methods

Fourteen Japanese healthy young adults aged 20–24 years participated in this study. A pedometer was used to measure the step count and to calculate other indexes of physical activity for 1 week. It was determined that 5 individuals comprised the high PA group (≥23 METs ■ hr/wk) and 9 individuals comprised the low PA group (<23 METs ■ hr/wk). Weight, height, BMI, muscle mass, body fat percentage, and basal metabolic rates were measured using a body composition analyzer. Then, we measured BP and heart rate variability at rest and during the postural change from the supine position to the sitting position. The ratio of low frequency (LF) components (0.04–0.15 Hz) to high frequency (HF) components (0.15–0.40 Hz) was used as an index of sympathetic nerve activity. HF was used as an index of parasympathetic nerve activity.

Results

The high PA group exhibited significantly higher calorie consumption (P < 0.01) and fat combustion (P < 0.01). The mean diastolic blood pressure value in the sitting position (P < 0.02) for the high PA group was significantly higher than that of the low PA group. After a postural change from the supine position to the sitting position, although there were no significant changes in LF/HF and HF in both groups, systolic blood pressure (SBP) in the high PA group increased significantly (P < 0.05); however, the low PA group had a significant subsequent decrease in SBP after changing from the supine position to the sitting position (P < 0.01).

Conclusions

Through the measurement of pedometer‐determined physical activities of daily living in healthy adults, we concluded that individuals who performed more pedometer‐determined physical activities of daily living can maintain normal responses in blood pressure after a postural change, compared to individuals who performed fewer pedometer‐determined physical activities of daily living.

Support or Funding Information

This work was supported, in part, by a Grant‐in‐aid for Scientific Research (B) (25305018) from the Japan Society for the Promotion of Science.
Altered Hypoxia‐Induced Vascular Reactivity and Endothelial Repair Capacity in Hypertensive Men
Authors: Rocha Helena Naly Miguens , Garcia Vinicius Pacheco , Mattos João Dario , Rocha Marcos Paulo , Campos Monique Opuszcka , de Jesus Mansur Daniel Elias , da Nóbrega Antonio Claudio Lucas , Fernandes Igor Alexandre , Rocha Natália Galito
Published: November 2023

Introduction

Hypoxia induces peripheral vasodilation in healthy subjects, despite the occurrence of sympathoexcitation. Endothelial progenitor cells (EPC) mobilization from bone marrow and nitric oxide released by intact endothelium seem to exert important functions in the vasodilatory response to hypoxia. Even considering that both sympathetic overstimulation and endothelial dysfunction are major contributing factors to the pathophysiology of hypertension, it is still unknown whether those functions are preserved in patients with hypertension.

Aim

To determine the effects of hypoxia on sympathetic activity, peripheral blood flow, and endothelial biomarkers in patients with hypertension.

Methods

Six normotensive men (NT; 44 ± 11 yrs, mean arterial pressure (MAP) 87 ± 6 mmHg) and seven non‐treated patients with hypertension (HT; 44 ± 10 yrs, MAP 110 ± 7 mmHg) were exposed to 5‐min bouts of: (1) Normoxia (21% O2) and (2) isocapnic‐Hypoxia (10% O2). Muscle sympathetic nerve activity [MSNA (microneurography; peroneal nerve)] recordings were obtained throughout the protocol, while femoral artery blood flow measurements (Doppler ultrasound) and blood samples were acquired during the last 30s of each condition. EPC (CD45dim/CD34+/VEGFR2+) and endothelial microparticles (EMP; CD42b‐/CD31+/AnnexinV+) were measured by flow cytometry, whereas nitrite concentration was determined by ozone‐chemiluminescence method.

Results

At baseline, femoral arterial blood flow (HT 361.8 ± 262.6 ml.min−1 vs. NT 344.6 ± 75.5 ml.min−1), EMP (HT 12 ± 8 EMP/μL vs. NT 4 ± 1 EMP/μL) and nitrite (HT 0.35 ± 0.10 μM vs. NT 0.42 ± 0.12 μM) were similar between groups (p>0.05), while EPC were reduced in HT (35 ± 8 EPC/μL) compared to NT (86 ± 24 EPC/μL, p=0.02). During hypoxia, HT presented decreased femoral arterial blood flow (−110.2 ± 43.0 ml.min−1) and EPC (−7 ± 4 EPC/μL), while NT showed increased femoral arterial blood flow (+73.9 ± 37.4 ml.min−1, p=0.01) and EPC (98 ± 35 EPC/μL, p=0.01). MSNA increased only in HT (+12.7 ± 6.7 vs. NT +3.6 ± 1.6 bursts/min, p=0.01). There were no differences in nitrite (HT −0.01 ± 0.07 μM vs. NT 0.06 ± 0.09 μM) and EMP (HT 6.4 ± 16.1 EMP/μL vs. NT 3.5 ± 3.9 EMP/μL) between groups (p>0.05).

Conclusion

These results suggest that patients with hypertension present diminished peripheral vascular reactivity in response to hypoxia, which could be associated with increased sympathetic activity observed in these individuals. Furthermore, EPC only increased in healthy subjects, suggesting an impaired endothelial repair capacity in patients with hypertension.

Support or Funding Information

CNPq, FAPERJ, CAPES and FINEP.
Molecular Basis of Hypoxia Induced Excessive Erythrocytosis
Authors: Azad Priti , Haddad Gabriel
Published: November 2023

Objective

Since polycythemia is a predominant trait in some high altitude dwellers (Chronic Mountain Sickness, CMS, or Monge's disease) but not others living at the same altitude in the Andes, we took advantage of this human “experiment in nature” and studied both populations (with CMS and without, non‐CMS). Although polycythemia could be advantageous at high altitude because it increases O2‐carrying capacity, this adaptive pattern to high altitude has deleterious effects since blood increases its viscosity and induces serious morbidities, such as myocardial infarction and stroke in young adults.

Methods

In order to understand the molecular basis for polycythemia of high altitude, we generated a disease in‐the dish‐model by re‐programming fibroblasts from CMS and non‐CMS subjects. In the past, we have done whole genome sequencing of subjects from this region (CMS and non‐CMS) to study selective sweeps and genetic basis of CMS. In this study, by manipulating expression of some of these genes in using our in‐vitro model system, we are delineating the functional basis and the molecular mechanism(s) linked to the pathology.

Results

As compared to sea level controls and non‐CMS subjects who responded to hypoxia by increasing their RBCs modestly or not at all, CMS cells increased theirs remarkably (up to 60 fold) with a dose‐dependent response to graded hypoxia (1.5, 5, 10% O2). We knocked down SENP1 (a desumoylase) in CMS iPS cells using lentiviral constructs and observed a striking reduction (>90%) of the CMS excessive erythropoietic response to low O2. And, by over‐expressing SENP1 in non‐CMS iPS cells, the hypoxic response in these subjects increased enormously. By further analyzing RBC differentiation in hypoxia, we found that VEGF, GATA1 and Bcl‐xL increased their gene expression in the CMS erythroid cells, in contrast to their minimal expression in the other two populations. We demonstrate also that, by utilizing a SUMO‐GATA1 fused construct, GATA1 desumoylation, a target of SENP1, is critical for the CMS phenotype. Unlike GATA1, the over‐expression of the anti‐apoptotic gene Bcl‐xL (a GATA1 target), only partially rescued the blunted erythroid response to hypoxia in non‐CMS cells.

Conclusion

We conclude that the increased erythropoietic sensitivity to hypoxia in CMS subjects is genetic in nature and depends on an increase in SENP1 expression and its desumoylation mediated activation of GATA1 under hypoxia. By combining the iPS technology with this unique Andean population that has adapted (or mal‐adaptated) to chronic hypoxia over thousands of years, we have built an important in‐vitro model. We believe that utility of this model lies not only for studying hypoxia‐induced polycythemia but also other hypoxia‐driven diseases experienced at sea level.

Support or Funding Information

This study is funded by National Institutes of Health (NIH) grants (1P01HL098053 and 5P01HD32573) to G.G. Haddad