bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti‐Inflammatory Muscle Macrophage Phenotype is Predictive of Resistance Training Gain in Older Individuals
Authors: Lee Jonah Dan , Erfani Rod A. , Campbell Marilyn S. , Mula Jyothi , Peterson Charlotte A.
Published: June 2021

Macrophages are responsible for phagocytic and reparative functions essential for the initiation and resolution of muscle damage‐induced inflammation. Muscle macrophages are phenotypically characterized by their polarization: either pro‐inflammatory M1 (classic) or anti‐inflammatory M2 (alternative) macrophages. We tested the hypothesis that different polarized macrophages would be associated with muscle adaptability in response to a chronic hypertrophic stimulus in the elderly. M1 and M2 macrophages were quantified from vastus lateralis biopsies in older (n=8; 68.3y) individuals before and after a 12‐week progressive resistance training program. An increase (p<0.05) in the M2, but not (p>0.05) in the M1, macrophage population was observed as an influence of training. Surprisingly, the strength gained in the resistance training program tended to correlate (P=0.065) to the pre‐training amount of M2 macrophages present in resting muscle. In vitro experiments are underway to study the muscle cell‐macrophage interaction directly. Our findings suggest anti‐inflammatory macrophages respond to and also promote a beneficial response to resistance training in aged‐muscle. Therapeutic interventions that affect the balance between muscle macrophage subpopulations may influence the effectiveness of utilizing resistance training as sarcopenia countermeasure.

Funding: NIAMS R01AR060701
Wnt3a a potent modulator of myogenic differentiation and muscle cell function
Authors: Romero-Suarez Sandra , Brotto Marco
Published: June 2021

Previously we showed that conditioned media (CM) from MLO‐Y4 osteocytes accelerates myogenesis of C2C12 myoblasts. Here we examined molecular and functional events occurring when C2C12 cells were treated with one of the key factors released by osteocytes, Wnt3a; which induced a significant nuclear translocation of β‐catenin, a hallmark of canonical Wnt signaling. This pathway showed to play a crucial role in the enhancement of myogenic differentiation produced by both 10ng Wnt3a and 10% MLO‐Y4 CM. The potent effects of Wnt3a on stimulating the β‐catenin signaling pathway strongly correlated with changes in calcium oscillations. In addition, fully differentiated myotubes that were treated with Wnt3a displayed an enhanced sensitivity to caffeine. These data are consistent with the upregulation of a key myogenic regulatory factor (MRF), MyoD and some of the essential genes related to intracellular calcium homeostasis (i.e., IP3‐R1,3; Stim‐2; Camk2d; SERCA‐1,2). Our new findings suggest that Wnt3a works as a potent modulator of myogenic differentiation and muscle cell function. We propose that Wnt3a may be an important secreted factor from osteocytes that mediates bone to muscle signaling by modulation of intracellular calcium homeostasis leading to gene regulatory changes that stimulate myogenesis. Support: NIH RC2 AR058962 and Missouri Life Sciences Research Board to MB.
Reversal of Age‐Related Mitochondrial Dysfunction In Vivo by Treatment with the Mitochondrially Targeted Therapeutic SS‐ 31
Authors: Siegel Michael P , Kruse Shane E , Knowels Gary , Szeto Hazel H , Rabinovitch Peter S , Marcinek David J
Published: June 2021

In vivo mitochondrial function and energy homeostasis in skeletal muscle declines with age. The ability to reverse this dysfunction with an acute treatment would represent a new paradigm for improving skeletal muscle function in the elderly. Here we test whether the novel, mitochondrially‐targeted peptide SS‐31 restores mitochondrial energetics in aged mouse skeletal muscle in vivo. We treated young and old mice with an acute 3mg/kg dose of SS‐31 and assessed in vivo mitochondrial function and energetic state after one hour using a unique combined magnetic resonance and optical spectroscopy system. In vivo mitochondrial coupling (P/O), phosphorylation capacity, and energy state (PCr/ATP) significantly declined with age. SS‐31 significantly improved all three measures of mitochondrial energetics in skeletal muscle of aged mice, but had no observed effect in young mice. Reversal of the age‐related decline in capacity and energy state may be due to both SS‐31's antioxidant activity and a more direct facilitation of mitochondrial respiratory activity in aged muscle. These results suggest that short‐term treatment with mitochondrially targeted therapeutics may represent a new approach to reverse the effects of aging on skeletal muscle energetics. This was supported by NIH grants AG001751 and AG028455.
Mitochondrial function, but not glucose uptake, is disrupted in skeletal muscle from Bmal1−/− mice
Authors: Garcia-Cazarin Mary L , Lefta Mellani , Esser Karyn A , Andrade Francisco H
Published: June 2021

Molecular clocks regulate circadian rhythms including muscle function and metabolism. Mice lacking Bmal1, a circadian clock transcription factor, showed a decreased in mitochondrial density in skeletal muscle. We hypothesize that mitochondrial function and glucose uptake are compromised in skeletal muscle of Bmal1−/− mice. O2 consumption was measured by polarography in mitochondria from 18 week‐old gastrocnemius muscles. Content of respiratory complexes, UCP3 and VDAC1 was measured in isolated mitochondria by western blot. Glucose uptake was measured with [3H] glucose upon insulin stimulation in the soleus muscles of 13 and 36 week‐old mice. Blood glucose was measured using a commercial glucose meter in 7, 13 and 36 week‐old mice. Serum insulin was measured with an Elisa kit in 13 and 36 week‐old mice. Respiration states 3 (ADP‐driven) and 5 (uncoupled) were 47±6% and 50±8% lower in mitochondria isolated from the gastrocnemius muscles of Bmal1−/− vs. control mice. There were no differences in the content of respiratory complexes, UCP3 and VDAC1. Blood glucose was 61% higher in 7 week‐old Bmal1−/− mice but there was no difference at other ages. Serum insulin was significantly lower in 36 week‐old Bmal1−/− mice. Basal and insulin‐stimulated glucose uptake rates were not different in soleus muscles at any age. We suggest that the molecular clock regulates mitochondrial function but not glucose uptake in mice.
Gene profiling of muscle specific VEGF deficient mice: Linking angiogenesis to metabolism?
Authors: Olenich Sara Ann , Audet Gerald N , Szeszel-Federowicz Viola , Chen Dongquan Chen , Olfert I Mark
Published: June 2021

Vascular endothelial growth factor (VEGF) is a multifunctional protein critical in vascular growth/development and essential for angiogenesis, particularly in response to exercise. Using microarray analysis, we examined skeletal muscle (gastrocnemius) gene responses in sedentary (sed) and 8‐week treadmill trained (tra; 1 hr, 5 d/wk) muscle‐specific VEGF deficient mice (KOSed and KOTra) compared to littermate control mice (WTSed and WTTra) to gain insight into the potential non‐angiogenic mechanisms involving VEGF. When comparing sedentary groups (KOSed vs WTSed) and trained groups (KOTra vs WTTra) there were 349 and 126 differentially expressed genes, respectively. Ingenuity pathway analysis revealed potential novel functions of VEGF in association with lipid and carbohydrate metabolism between KOSed and WTSed. Between KOTra and WTTra there were associations with gene networks involved in cell signaling, protein synthesis, endocrine system disorders and metabolic disease. Data also confirm the vital association of VEGF in skeletal muscle structure and function, but more importantly identify potential novel relationships between VEGF and metabolism, endocrine and metabolic disorder not previously recognized. Future studies examining the role of VEGF as a metabolic regulator may open new avenues linking angiogenesis and metabolic diseases.
The response of genes implicated in muscle oxidative metabolism and oxidative stress to acute exercise in COPD
Authors: Slot Ilse , van den Borst Bram , Hellwig Valéry , Kelders Marco , Schols Annemie , Gosker Harry
Published: June 2021

Skeletal muscle oxidative capacity is impaired and oxidative stress enhanced in COPD, which contributes to muscle dysfunction. We aimed to test the hypothesis that COPD patients have impaired exercise‐induced response of muscle oxidative metabolism‐ and stress‐related genes, especially during exercise‐induced oxygen desaturation.

Resting and 4h post‐incremental cycling quadriceps biopsies were obtained from 28 COPD patients and 15 controls. 13 COPD patients desaturated (SaO2 fall ≥4%) and 15 did not. Expression of a broad panel of genes related to oxidative/glycolytic metabolism and oxidative stress were determined by RT‐qPCR.

VO2 peak, peak load and produced work were significantly lower in COPD compared to controls, yet most genes responded alike (eg. PGC1α/β, PPRC1, HKII and MnSOD). Notably, HMOX1, implicated in hypoxia and oxidative stress, showed a stronger response in COPD patients, especially in those without desaturation.

Although the overall response of metabolic genes to exercise seemed normal in COPD, these responses may actually be relatively greater, since patients did on average 66% less work during incremental cycling. The greater exercise‐induced increase in HMOX1 expression in COPD and the role of oxygen desaturation herein requires further investigation.

This research was performed within TI Pharma T1–201.
Respiratory Load Compensation Responses of External Intercostal Muscle in Conscious Rats
Authors: Jaiswal Poonam , Davenport Paul
Published: June 2021

The external intercostals are primary respiratory muscles responsible for the maintenance of efficient respiration. Conscious animal load compensation; in particular external intercostal muscle responses to intrinsic, transient tracheal occlusions (ITTO) have not been reported. We hypothesized that ITTO in conscious rats will recruit the external intercostal muscle with increased activation. Adult male rats were instrumented with a tracheal occluder and bilateral EMG electrodes in the external intercostal muscle (EI‐EMG). After 5 days of surgical recovery, the rats received ITTO via cuff inflations for 3–8 sec separated by 10–15 sec of unoccluded breath, for a total trial time of 20 mins. The trial was divided into before, during, between and after ITTO. The EI muscle was recruited 86.3% of time during the ITTO with an increased activation of EI muscles; EMG amplitude during ITTO was 157.8% compared to before ITTO. Two distinct patterns of activation were observed. Large onset amplitude response followed by phasic activity was the predominant pattern. In some cases, gradual increases in EI‐EMG were observed with maximal activation towards the end of ITTO. These results show recruitment of EI muscle activity in response to increased respiratory loads. EI muscles play an important role in mediating chest wall respiratory load compensation responses to ITTO in conscious rats.
Impact of TrkB signaling on recovery of phrenic activity after cervical spinal cord injury in rats
Authors: Sieck Gary C , Zhan Wen-Zhi , Mantilla Carlos B
Published: June 2021

Upper cervical spinal cord injury disrupts descending inspiratory‐related drive to phrenic motoneurons (PMNs) paralyzing the diaphragm muscle (DIAm). There is gradual recovery of rhythmic DIAm activity ipsilateral to injury suggesting strengthened spared synaptic inputs to PMNs. We previously showed that intrathecal treatment with brain‐derived neurotrophic factor (BDNF) enhances recovery of rhythmic DIAm activity after C2 spinal cord hemisection (C2SH). We hypothesized that TrkB signaling in PMN is necessary for functional recovery post‐C2SH. Using a recently validated intrapleural technique, daily injections of TrkB siRNA were administered to adult male Sprague‐Dawley rats. Bilateral DIAm EMG activity was chronically recorded before and after C2SH to assess recovery of rhythmic DIAm activity. In the vehicle‐treated group, functional recovery was evident in ~10% of C2SH rats at 7D and ~40% of rats at 14D post‐injury. In contrast, none of the TrkB siRNA treated rats displayed functional recovery up to 14D post‐C2SH. Transport of intrapleural Cy3‐labeled siRNA was verified in retrogradely‐labeled phrenic motoneurons. Our results indicate that BDNF/TrkB signaling at PMNs is necessary for functional recovery of rhythmic DIAm activity after C2SH.

Supported by NIH grant R01 HL096750 and Mayo Clinic
State dependent modulation of breathing in rats during urethane anesthesia
Authors: Pagliardini Silvia , Greer John J , Dickson Clayton T
Published: June 2021

Breathing is an automatic process that is highly influenced by brain state, with abnormalities occurring more frequently during sleep. Although rodents are commonly used for respiratory neurobiology studies, they show irregular sleep patterns and cycles, which make studies of sleep‐dependent breathing challenging. The objective of this study was to determine if sleep‐like brain state alternations apparent under urethane anesthesia are correlated to changes in breathing parameters and respiratory muscle modulation, and to determine if they mimic changes occurring during natural sleep.

Rats were anesthetized and respiratory parameters (rate, minute ventilation and EMG activity in diaphragm, genioglossus, abdominal and facial muscles) were recorded across different brain states as assessed by neocortical and hippocampal EEG. Results were compared with recordings in natural sleep. Our data show that brain state alternations under urethane anesthesia are correlated with changes in breathing rate, variability and respiratory muscle modulation and closely reproduce changes observed in natural sleep. Of particular interest is the present demonstration of prominent state dependent modulation of both genioglossus and abdominal muscle activity, which is a robust feature of sleep and sleep‐like brain state transitions.

Supported by CIHR (JJG) and NSERC (CTD). SP is a Parker B. Francis fellow.
Electrophysiological properties of RVLM pre‐sympathetic and CVLM neurons involved with the sympatho‐inhibitory component of the baroreflex in rats submitted to chronic intermittent hypoxia
Authors: Moraes Davi JA , Zoccal Daniel B , Machado Benedito H
Published: June 2021

We evaluated the effects of CIH on the baroreflex sympatho‐inhibitory pathways. Male Wistar rats were submitted to CIH for 10 days while control rats were maintained in normoxia. Using the working heart‐brainstem preparations we recorded respiratory and sympathetic nerves [thoracic (tSN) and lumbar (lSN)] simultaneously intracellular recordings of RVLM pre‐sympathetic and CVLM neurons. Baroreflex was activated by stimulation of aortic depressor nerve (ADN) during inspiration and expiration. In the control rats (n=32), the decrease in the tSN and lSN in response to ADN stimulation was more pronounced during the expiratory phase. CIH (n=31) enhanced the inhibitory tSN (−87 ± 6 vs −71 ± 3%; p<0.05) and lSN (−90 ± 3 vs −65 ± 4%; p<0.05) responses to ADN stimulation during the expiratory phase. CIH enhanced the inhibition of pre‐sympathetic (−83 ± 5 vs −59 ± 3%; p<0.05; n=20), as well the stimulation of CVLM neurons (+79 ± 3 vs +55 ± 4%; p<0.05; n=23) produced by ADN stimulation during expiratory phase only in neurons with expiratory modulation. However, in the presence of synaptic blockade no changes were observed in the electrophysiological properties (excitability and input resistance) of RVLM pre‐sympathetic and CVLM neurons after CIH. We suggest that changes in the respiratory network contribute to the observed increase in the baroreflex gain after CIH. Supported by FAPESP and CNPQ.
Quantification of burst‐to‐burst variability of basal phrenic nerve discharge in adult in vivo rat
Authors: Kiridly Adam , Rahim Tania , Reid Inefta M , Solomon Irene C
Published: June 2021

The respiratory control system is highly dynamic; however, typical measures of respiratory variability are often limited to computing the mean±SE and/or the coefficient of variation (CV), which do not distinguish between short‐term and long‐term variability. To overcome this limitation, some studies have employed Poincaré plot (PP), irregularity score (IS), or approximate entropy (ApEn) analyses; however, these analysis methods have not been implemented and compared on a single data set. To address this issue, we used all of the above analysis methods to quantify burst‐to‐burst variability of basal phrenic nerve discharge (n=300 bursts) recorded from anesthetized or decerebrate adult rat. We found that while all methods provide insight into the overall burst‐to‐burst variability, PP, IS, and ApEn analyses provide novel information about the patterning of the data spread. Moreover, these methods provide novel information about the short‐term vs long‐term variability. We suggest that these additional analysis methods be used to more fully capture patterns of long‐term variability and provide complementary insight into the dynamics underlying the respiratory control system. Supported by HL63175
Cardio‐Pulmonary Coupling II: Reduced Stroke Volume Effects on Time Phase Signals′ Combination at Integrative Site in CNS
Authors: Sun Xingguo
Published: June 2021

INTRODUCTION

Since 2011 physiology conference, we introduced a New dynamic theory of breathing control: a complex model integrates multi‐systems. Another abstract described reduced left ventricle ejection fraction (EF) decrease the amplitude of arterial O2, CO2 and [H+] oscillations.

HYPOTHESIS

We assessed the hypothesis that except low EF, reduced Stroke Volume (SV) decrease the combined amplitude of arterial O2, CO2 and [H+] oscillations signals at integrative site due to different time courses blood arrive at chemical sensors.

METHODS

Smilar to another abstract; reduced SV, which results blood arrive at aorta and carotid body from different SV (time phase difference), was used to calculate the combined magnitudes of PO2 and PCO2 oscillation arrive at integrative site at CNS.

RESULTS

The decreased SV (low CO) results the blood arrives at aorta and carotid by different SV, the weak combined signal (magnitude) for next breath at integrative site at CNS. This will results that mean of O2 and CO2 at central chemical sensors is unstable, and then breath control ¡°gain¡± goes up and down. Low SV may result periodic breathing pattern.

CONCLUSIONS

The same O2 and CO2 oscillations, reduced SV will via low EF mixed chamber and decreased in combining signal for next breath at integrative site are influenced by different time phase. It is another explanation of Cardio‐Pulmonary Intra‐Coupling.
Cardio‐respiratory Coupling is Negligible in a Rodent Septic‐Model
Authors: Hsieh Yee Hsee , Jacono Frank J , Dick Thomas E
Published: June 2021

Cardio‐respiratory failure is a frequent complication of sepsis and is associated with the loss of heart rate variability (HRV) but the effects on ventilatory pattern variability (VPV) and cardio‐respiratory coupling are undetermined. We hypothesize that cardio‐respiratory failure may be associated with a loss of cardio‐respiratory coupling in terms of power spectral analysis of heart rate (HR) and cardio‐ventilatory coupling (CVC). To test this hypothesis, adult male Sprague Dawley (Harlan) rats (n=4) were implanted with an indwelling blood pressure transducer (DSI). Both ventilatory pattern (plethysmography) and blood pressure were recorded for 2h in the morning and 2h in the afternoon for 4d. The septic model was induced by injecting animals with a single bolus of lipopolysaccharide (LPS) (24 mg/kg, intraperitoneal injection) every morning for 3d. After 3 injections, both respiratory and heart rate doubled while the coefficient of variation (CV) did not change. Furthermore, the high frequency component of HRV disappeared. Entropy‐based measures of CVC reveal a decrease in coupling strength after LPS treatment. We concluded that although CV, a measure of HRV, did not decrease in the septic model, cardio‐respiratory coupling was markedly diminished. The loss of coupling may result in gas‐exchange inefficiency which may contribute to cardio‐respiratory failure. Supported by T32 HL007913 and HL087377
Ventilatory pattern variability is associated with long‐term recovery following cardiac arrest and resuscitation in rats
Authors: Xu Kui , Kuang Youzhi , Dick Thomas E , Jacono Frank J
Published: June 2021

Predicting outcomes is essential for intervening to prevent mortality following cardiac arrest and resuscitation. We hypothesize that ventilatory pattern variability (VPV) reflects brainstem cardio‐respiratory regulation and propose to use VPV as a predictor for long‐term survival following cardiac arrest and resuscitation. Spontaneous ventilation was recorded (60 min) using whole‐body plethysmography in male Wistar rats before cardiac arrest (12 min) and 1d after resuscitation. VPV was analyzed including quantification of a nonlinear complexity index (NLCI) by comparing the sample entropy of original and surrogate data sets with similar autocorrelation functions. Overall survival (4d) was determined. VPV was similar before cardiac arrest but differed between survivor (n=5) and non‐survivor (died before 4d, n=2) groups after resuscitation. Non‐survivors had lower respiratory rate and higher coefficient of variation (CV) of respiratory cycle length compared to the survivors (RR: 80±13 vs. 104±11 bpm; CV: 0.27±0.09 vs. 0.15±0.02). NLCI was about 8‐fold higher in non‐survivor as compared to survivor rats (0.3±0.1 vs.0.04±0.02). Changes in VPV might distinguish surviving from non‐surviving rats early in the recovery phase after resuscitation, suggesting that VPV can be used as an outcome predictor following cardiac arrest and resuscitation. Supported by NS 38632, HL 087377, VA Research Service.
Galactooligosaccharides (GOS) decrease the probability of upper respiratory tract infection (URTI) in aged adults, but it is dependent on Mini‐Nutritional Assessment (MNA) score and the percentage of natural killer cells
Authors: Girard Stephanie-Anne , Nieves Carmelo , Downes Eric C , Christman Mary C , Mai Volker , Dahl Wendy J , Duriancik David M , Gardner Elizabeth M , Barkley Phillip L , Langkamp-Henken Bobbi
Published: June 2021

Previously, we showed a reduction of URTI in academically stressed students with GOS. This study determined the effect of GOS on risk of URTI in aged adults over cold/flu season. In a randomized, double‐blinded study, adults (66±1 y) received 0 g (n=38) or 5 g (n=43) of GOS for 24 wk. Nutritional status was evaluated at baseline (wk 0) using the MNA (score <24 out of 30 is associated with risk of malnutrition). Blood was drawn at wks 0, 3 (concurrent with seasonal influenza vaccine administration) and 5 (2 wk post‐vaccination). PBMCs were stimulated in vitro with PHA, and IFNγ production was quantified. Subpopulations of CD45+ cells were determined. Wk 0 MNA scores (27±1) and %NK cells at all wks (15±1%, 14±1%, 15±1%) were not different between groups; however, both interacted with the intervention on probability of URTI. As MNA score increased, probability of URTI decreased, and this effect was greater with GOS vs. placebo. As %NK cells increased, the probability of URTI decreased with GOS and slightly increased with placebo. Percent change in IFNγ between wk 0 and wks 3 (−26±5 vs 13±19, P=0.053) and 5 (−17±8 vs 35±27, P=0.07) differed for placebo vs GOS, respectively. GOS had a greater effect than placebo on decreasing the probability of URTI with improving nutritional status, and unlike placebo, GOS decreased the probability with increasing %NK cells. This effect may be via alterations in IFNγ.

Grant Funding Source: Corn Products International
Sympathetic inhibition attenuates hypoxia induced insulin resistance
Authors: Peltonen Garrett Lee , Scalzo Rebecca L , Schweder Melani M , Szallar Steve E , Binns Scott E , Wood Lacey M , Klochak Anna L , Schroeder Thies , Irwin David C , Hamilton Karyn L , Bell Christopher
Published: June 2021

Acute exposure to hypoxia decreases insulin sensitivity in healthy humans. The mechanism is unclear but increased activation of the sympathetic nervous system (SNS) may be involved. We hypothesized that SNS inhibition would attenuate hypoxia induced insulin resistance. Insulin sensitivity (hyperinsulinemic euglycemic clamp) was determined in 10 men (23±1 years, body mass index 24.2±0.8 kg/m2 (mean±SE)) during normoxia (FIO2=0.21), hypoxia (FIO2=0.11), normoxia and SNS inhibition (48‐hour transdermal clonidine), and hypoxia and SNS inhibition. Oxyhemoglobin saturation was decreased (P<0.01) in hypoxia (63±2%) compared with normoxia (96±0%), and was unaffected by SNS inhibition (P>0.25). The area under the norepinephrine curve (relative to the normoxia response) was increased in hypoxia (137±13%; P=0.02); clonidine prevented the hypoxia induced increase under the norepinephrine curve (94±14%; P=0.43). Compared with normoxia (225±23 nmol/kg fat free mass/pmol/L/min) the glucose infusion rate (adjusted for fat free mass and circulating insulin), required to maintain blood glucose at 5 mmol/Lduring administration of insulin, was decreased in hypoxia (128±30; P=0.03), and unchanged during normoxia and SNS inhibition (219±19; P=0.86), and hypoxia and SNS inhibition (169±23; P=0.23). We conclude that short‐term SNS inhibition attenuates hypoxia induced insulin resistance.

Support: DARPA N66001‐10‐c‐2134
Prolyl Hydroxylase Inhibition Attenuates Mast Cell Degranulation During Systemic Hypoxia Via Upregulation of Heme Oxygenase‐1
Authors: Wood John G. , Moncure Michael , Thomas James H. , Holloway Naomi B. , Gonzalez Norberto C.
Published: June 2021

Acute systemic hypoxia (Hx) causes mast cell degranulation and increased leukocyte adherence in post‐capillary venules of rats. When animals are chronically hypoxic, microvascular inflammation resolves in part due to upregulation of heme oxygenase‐1 (HO‐1), which prevents mast cell activation. Inhibition of prolyl hydroxylase (PHD) increases hypoxia‐inducible factor‐1 (HIF‐1) levels, which promotes HO‐1 expression.

Objective

Our goal was to evaluate whether inhibition of PHD attenuated Hx‐induced mast cell degranulation, and if upregulation of HO‐1 was involved.

Methods

Intravital microscopy was used to measure mast cell degranulation and leukocyte adherence in mesenteric venules of anesthetized rats. The PHD inhibitor ethyl dihydroxybenzoate (EDHB) was injected s.c. to rats 24 hrs prior to experiments. Hx was produced in rats breathing 10% O2.

Results

Hx caused mast cell degranulation and increased leukocyte adherence in the mesenteric microcirculation of vehicle‐ but not EDHB‐treated rats. Superfusion of the mesentery with the HO‐1 inhibitor zinc protoporphyrin caused a significant increase in mast cell degranulation and leukocyte adherence during Hx in EDHB‐ but not in vehicle‐treated rats.

Conclusion

Inhibition of PHD reduces Hx‐induced mast cell degranulation and leukocyte adherence in part through HO‐1 upregulation, which is consistent with a HIF‐1‐dependent mechanism.
Molecular interaction of organic cation transporters with the tetraspanin CD63 modulates transporter trafficking and their trafficking‐associated regulation
Authors: Hirsch Beatrice , Brast Sabine , Grabner Alexander , Holle Svenja , Guckel Denise , Schlatter Eberhard , Ciarimboli Giuliano
Published: June 2021

Human organic cation transporters (hOCT1 to 3) are involved in the transport of endogenous and exogenous organic cations in the liver, kidney and brain. Here we analyse the regulation of hOCT1 to hOCT3 by direct protein‐protein interaction using a mating based Split‐Ubiquitin‐Yeast‐Two‐Hybrid System and pull‐down assays. Specifically, we found that the tetraspanin CD63 is an interacting partner of hOCT2 and hOCT3. A strong co‐localisation of CD63 and hOCT2 was shown on the plasma membrane and in vesicular compartments of HEK293 cells by TIRF‐microscopy and in tissue sections of human kidneys by immunofluorescence. Further immunofluorescence and surface biotinylation experiments after CD63 overexpression revealed a loss of cell surface hOCT2 and its relocalisation into intracellular vesicles. In HEK293 cells stably transfected with hOCT2, CD63‐overexpression caused a decrease, CD63‐knockdown an increase of cation‐uptake. As functional consequences of CD63 knockout in mice we observed that OCT2 lost its specific basolateral localization in the kidney, appearing to be widespread distributed in the cell and that basophil‐mediated histamine secretion, which apparently involves OCT3, was increased. Our data strongly suggest that CD63 regulates the function of hOCT2 and −3 by influencing their trafficking to/from the cell membrane and processing via the intracellular sorting machinery. Supported by the Deutsche Forschungsgemeinschaft (DFG CI 107/4‐1 to 2).
Blood‐Brain Barrier Na/HCO3 Cotransporters: Evidence for a Role in Ischemia‐induced Brain Na Uptake
Authors: Yuen Natalie Yuk-yee , Chen Yi-Je , Boron Walter F. , Praetorius Jeppe , Anderson Steven E. , OˈDonnell Martha E.
Published: June 2021

Increased Na transport across the blood‐brain barrier (BBB) contributes to brain edema during the early hours of ischemic stroke. Our previous studies show that BBB Na‐K‐Cl cotransporter and Na/H exchange are stimulated by ischemic factors and inhibition of these Na transporters by bumetanide and HOE642, respectively, greatly attenuates Na uptake and edema in the rat middle cerebral artery occlusion (MCAO) model of stroke. In the present study, we examined the possibility that one or more Na/HCO3 cotransporters (NBCs) are present at the BBB and contribute to ischemia‐induced brain edema. NBC protein expression and activity were evaluated using Western blot and microspectroflurometric assays, respectively. We found the expression of NBCe1C and NBCn1 in bovine cerebral microvascular endothelial cells (CMEC) and freshly isolated rat brain microvessels. We also observed a Na‐ and HCO3‐dependent, HOE642‐insensitive H+ flux in CMEC that is comprised of both DIDS‐sensitive and DIDS‐insensitive components, indicating the presence of both NBCe1 and NBCn1 activity in the cells. Finally, localized 23Na MR spectroscopy was used to assess brain Na uptake in rats subjected to MCAO. We found that MCAO‐induced Na uptake was significantly reduced by DIDS (40 mg/kg, iv). These findings support the hypothesis that BBB NBCe1C and/or NBCn1 participate in ischemia‐induced brain Na uptake and edema formation.

Supported by NINDS
Adenylyl cyclase 6 determines AVP‐induced membrane abundance and phosphorylation of NKCC2 and NCC
Authors: Rieg Timo , Fenton Robert A , Tang Tong , Uchida Shinichi , Sharik Meaghen , Hammond H Kirk , Vallon Volker
Published: June 2021

The adenylyl cyclase isoform 6 (AC6) is expressed in all tubule segments and the collecting duct. Previously we showed that AC6 is important for AQP2 phosphorylation and urinary concentration. To define the role of this enzyme in membrane abundance and arginine‐vasopressin (AVP)‐induced phosphorylation of NKCC2 (residue S126) and NCC (residues T53, T58 and S71) we studied AC6 wild‐type (WT, n=3–4) and knockout (AC6−/−, n=3–4) mice. Endogenous AVP was suppressed overnight by water loading (5% dextrose/1% ethanol in drinking water). dDAVP (0.1 μg/kg i.p.) or vehicle were applied, kidneys removed 20 min later and responses compared between WT vs AC6−/−. The dDAVP‐induced increase in total membrane NKCC2 expression was blunted in AC6−/− vs WT (1.9±0.3 vs 13±3 fold, P<0.05) and S126‐NKCC2 phosphorylation attenuated (AC6−/−: 0.9±0.1 vs WT: 1.4±0.1 fold, P<0.05). The dDAVP‐induced increase in total membrane NCC expression (0.6±0.1 vs 1.6±0.1 fold, P<0.05) and NCC phosphorylation at T53, T58 and S71 were attenuated in AC6−/− vs WT (0.6±0.1 vs 2.0±0.3, 0.8±0.1 vs 1.8±0.1, and 0.4±0.1 vs 2.9±0.1 fold, respectively, P<0.05). Immunohistochemistry showed reduced membrane abundance for S126‐NKCC2, T53‐ and T58‐NCC in response to dDAVP in AC6−/− vs WT. Our results implicate AC6 in AVP‐induced upregulation of NKCC2 and NCC membrane expression and phosphorylation.
Differential stimulation of duodenal HCO3− secretion by luminal fatty acids
Authors: Akiba Yasutada , Inoue Takuya , Higashiyama Masaaki , Rudenkyy Sergiv , Iwamoto Ken-ichi , Kuwahara Atsukazu , Kaunitz Jonathan D
Published: June 2021

Luminal nutrients enhance mucosal defenses via gut hormone release. We examined the effects of long‐chain (LCFA) or short‐chain fatty acid (SCFA) receptor agonists on duodenal HCO3− secretion (DBS). We measured DBS with pH and CO2 electrodes in a perfused duodenal loop in anesthetized rats. Acetate or propionate, GPR43 agonist phenylacetamide 1 (PA1) or GPR40 agonist GW9508 was luminally perfused +/− iv dipeptidyl peptidase 4 inhibitor NVP728 or hormone receptor antagonist. LCFA receptor GPR40 and SCFA receptors GPR41/43 were expressed in the duodenal enteroendocrine cells. Luminal perfusion of acetate or propionate similarly increased DBS, enhanced by NVP728, while increasing GLP‐2 release in portal blood. Acetate‐induced DBS was reduced by the monocarboxylate transporter inhibitor, suggesting the acetate absorption‐mediated and GLP‐2‐mediated DBS. PA1 dose‐dependently increased DBS, unaffected by NVP728 or CCK‐A receptor antagonist, but inhibited by atropine or 5‐HT4 antagonist. The GPR40 agonist GW9508 increased DBS with GLP‐2 release, enhanced by NVP728. Activation of GPR40 by LCFA and possibly GPR41 by SCFA increases DBS via GLP‐2 release, whereas GPR43 activation stimulates DBS via muscarinic and 5‐HT4 receptor activation. The presence of fatty acid sensing in the duodenal mucosal defenses may be implicated in the pathogenesis of functional dyspepsia syndromes.

VA Merit Review, R01 DK54221
Chitin‐binding protein GbpA of Vibrio cholerae induces interleukin‐8 gene expression in intestinal cells through a TLR2/TLR1/CD14 complex
Authors: Chatterjee Nabendu Sekhar , Ghosh Avishek , Sabui Subrata , Acharya Sagar , Banerjee Kalyan Kumar
Published: June 2021

Vibrio cholerae N‐acetylglucosamine binding protein (GbpA) is a secretory protein which is essential for bacterial survival in environment as well as in human gut. Here we show that GbpA binds to toll‐like receptor 2 (TLR2) and initiates downstream cellular inflammatory signaling pathway by IL8 secretion. GbpA triggered formation of lipid raft microdomain on intestinal cell surface by interacting with TLR2 with a Kd of 76 nM. The microdomain contains TLR1, CD14 and flotilin in addition to TLR2. As a downstream effect, the lipid raft complex triggered IL8 secretion and the effect was specific in HT29 intestinal cells. The critical involvement of the GbpA‐induced lipid raft microdomain complex in IL8 secretion was indicated by the decrease in secretion caused by disruption of the raft by extraction of cholesterol by methyl‐β‐D‐cyclodextrin and by its restoration by extraneous addition of cholesterol. The GbpA‐mediated elevation in IL8 secretion involved activation of NFκB and formation of AP1 complex which eventually increased IL8 promoter activity. Taken together, the results suggest that GbpA is a key player in pathogenesis of cholera by triggering pro‐inflammatory response in infection by inducing downstream activation through clustering of GbpA‐TLR1/2‐CD14 complex in lipid raft microdomain. [Supported in part by grant from CSIR, Govt. of India No. 37(1410)/10/EMR‐II]
High fat diet differentially regulates olfactory receptors in the duodenum of obesity‐prone and obesity‐resistant rats
Authors: Primeaux Stefany , Braymer H. Douglas , Bray George
Published: June 2021

The gastrointestinal tract is an important factor in the regulation of food intake, nutrient sensing and nutrient absorption. Obesity‐prone Osborne‐Mendel (OM) rats are less sensitive to the satiating effects of a duodenal infusion of fatty acids than obesity‐resistant S5B/Pl (S5B) rats, suggesting that the gastrointestinal tract differentially senses the presence of fat in these two strains. Based on microarray analysis of the duodenal enterocytes, it was hypothesized that olfactory receptors present in the duodenum of OM and S5B rats were differentially regulated by the intake of a diet high in fat. OM and S5B rats were fed either a high fat diet or a low fat diet for 2 weeks, in which OM rats consumed more high fat diet than S5B rats. The mRNA levels of four olfactory receptors (OLR1744, OLR50, OLR124, OLR1507) were assessed from the duodenal enterocytes of these rats. The duodenal mRNA levels of OLR1744, OLR124 and OLR1507 were significantly elevated in OM rats fed the high fat diet, but not S5B rats. No differences in the expression of OLR50 receptor mRNA were detected. These data suggest that several olfactory receptors present in the duodenum are selectively regulated in obesity‐prone OM rats fed a high fat diet. Therefore, these receptors may play a role in the sensing and regulation of dietary fat, and may be important for the individual susceptibility to obesity in these two strains. Support: NIDDK 32089 to GAB
Biotinylation of K16 in histone H4 causes chromatin condensation
Authors: Singh Mahendra P , Zempleni Janos
Published: June 2021

Histone modifications play crucial roles in the control of gene activity, nuclear architecture, and genomic stability. Nucleosomes are the basic units of chromatin in eukaryotes, consisting of an octamer of histone core proteins wrapped by ~147 bp of DNA. Biotinylation of lysines (K) in histones H3 and H4 by holocarboxylase synthetase (HLCS) is a rare epigenetic repression mark, e.g., H4K16bio. Here, we tested the hypothesis that H4K16bio contributes toward chromatin condensation and, therefore, gene repression. We generated mutant recombinant histone H4 in which K16 has been replaced with cysteine (H4K16C) for subsequent chemical biotinylation with a thiol reactive reagent (H4K16Cbio). Nucleosomes were assembled by salt dialysis, using recombinant histones and the Widom 601 DNA sequence. DNA binding was analyzed by atomic force microscopy. Core octamers containing H4K16Cbio bound ~29% more DNA compared with octamers containing H4K16 or H4K16C (controls), suggesting biotin‐dependent condensation of chromatin. We conclude that biotinylation of H4 increases the nucleosomal affinity for DNA. We propose that histone biotinylation contributes to gene repression, in addition to the emerging role of HLCS as a member of a multiprotein gene repression complex in human chromatin.

Grant Funding Source: ARD, and NIH DK063945, DK077816, and DK082476.
Notch signaling regulates proliferation and differentiation of the intestinal crypt base columnar (CBC) stem cell
Authors: Samuelson Linda C , VanDussen Kelli L , Carulli Alexis J , Keeley Theresa M , Smith Julie
Published: June 2021

This study examined the importance of the Notch signaling pathway for homeostasis of the CBC stem cell in the intestine. The Lgr5‐GFP reporter mouse strain was used to identify and track CBC stem cells after Notch inhibition with DBZ. Global Notch disruption in adult mice induced rapid CBC cell loss, with reduced proliferation and apoptotic cell death. Furthermore, expression of the CBC stem cell‐specific marker Olfactomedin 4 (Olfm4) was observed to be directly dependent on Notch signaling, with transcription activated through RBP‐Jκ binding sites in the promoter. Notch inhibition also led to precocious differentiation of epithelial progenitor cells into secretory cell types, including goblet, enteroendocrine and tuft cells, as well as an unusual secretory cell type that expressed both Paneth and goblet cell markers. Analysis of Notch function in Atoh1‐deficient mouse intestine demonstrated that the generalized secretory cell hyperplasia induced by Notch inhibition was dependent on this transcription factor, whereas Notch regulation of Olfm4 gene expression was Atoh1‐independent. Our findings suggest that Notch targets distinct progenitor cell populations to maintain adult intestinal stem cells and to regulate cell fate choice in order to control epithelial cell proliferation and differentiation.
Establishment and characterization of novel in vitro models of gastric chief cell and metaplastic lineages
Authors: Weis Victoria G. , Petersen Christine P. , Nam Ki Taek , Whitehead Robert H , Goldenring James R
Published: June 2021

Investigations of preneoplastic metaplasias in the stomach are limited by the lack of cell models for mature mucosal cells and metaplasias. We developed in vitro models of chief cells and spasmolytic polypeptide‐expressing metaplasia (SPEM) cells to study the transdifferentiation of chief cells into SPEM. Utilizing the Immortomouse, which is completely transgenic for tempertature‐dependent TAg, chief cells were isolated and cultured from untreated mice. SPEM cells were isolated from mice treated with L635, an acute SPEM induction model. mRNA and protein expression profiles were used to characterize the cultures. ImChief cultures express most chief cell markers with immunostaining for pepsinogen and Rab3D‐containing granules. After 2 weeks at 39°C, ImChief cells cease proliferating and resemble a more differentiated state similar to in vivo chief cells. ImSPEM cells express the SPEM markers TFF2 and HE4 and produce HE4‐containing granules. They lack expression of most other lineage specific markers. ImSPEM cells retain their proliferative ability for at least 1 month at 39°C, resembling the in vivo properties of SPEM cells. Our results indicate that the expression and proliferation profiles of the novel ImChief and ImSPEM cultures resemble in vivo chief and SPEM cell lineages. These cultures provide the first in vitro system for studying the molecular mechanisms of the metaplastic transition in the stomach.

Department of Veterans Affairs Merit Review Award and NIH RO1 DK071590
Hyperhomocysteinemia decreases intestinal motility leading to constipation
Authors: Givvimani Srikanth , Munjal Charu , Narayanan Nithya , Tyagi Geetansh , Metreveli Naira , Tyagi Suresh
Published: June 2021

Elevated levels of plasma homocysteine (Hcy) known as hyperhomocysteinemia (HHcy), has been implicated in inflammation and remodeling of intestinal vasculature and it also aggravates the pathogenesis of inflammatory bowel disease (IBD). Interestingly, colon is the pivotal site that regulates Hcy levels in the plasma. We hypothesize that hyperhomocysteinemia decreases intestinal motility through MMP‐9 induced intestinal remodeling leading to constipation.

Method

To verify this hypothesis we used C57BL/6J or wild type (WT), CBS+/−, MMP‐9 −/− and MMP‐9−/− + Hcy mice. Intestinal motility was assessed by barium meal studies and daily feces output. Expression of ICAM‐1, iNOS, MMP‐9 and TIMPs were studied by western blot and immunohistochemistry.

Results

Barium meal studies suggest that intestinal motility is significantly decreased in CBS−/+ mice compared to other groups. Fecal output to body weight ratio was significantly reduced in CBS−/+ mice compared to other groups. There was significant up‐regulation of MMP‐9, iNOS, and ICAM‐1 expression in the colon from CBS−/+ mice compared to WT mice.

Conclusions

Results suggest that HHcy decreases intestinal motility due to MMP‐9 induced intestinal remodeling leading to constipation.
Dietary behaviors and acculturation in South Asian college students
Authors: Jasti Sunitha , Sachmechi Amanda
Published: June 2021

Objective

To determine whether the degree of acculturation is associated with dietary behaviors and body mass index (BMI) in first‐generation South Asian college students.

Method

A cross‐sectional online survey assessed acculturation level, dietary behaviors, and BMI of 204 South Asian students at Queens College.

Results

Mean age was 21 y (2.9) and mean BMI was 22.7 (4.1). Mean (SD) acculturation was 3.57 (0.62) on a scale of 1–5, 61% of the sample preferred South Asian food at home, while 30% preferred it at restaurants. The degree of acculturation was not significantly associated with BMI or dietary behaviors assessed in this sample. There was, however, a high prevalence of obesogenic dietary behaviors. Fast food was consumed at least 3 times a week by 40% of the students. Sweetened beverages were consumed by 53% and fried snacks by 50% at least 4–6 times a week; fats (butter, ghee, cream) were added to food at the table at least sometimes by 56%; and only a third (34%) of the participants always chose low fat type for dairy products. Furthermore, a majority of the students (64%) indicated that there are too many nutrition recommendations making it hard to know what to believe and 33% did not know which menu items in a restaurant have more/less fat.

Conclusion

All South Asian college students may benefit from nutrition education regardless of the acculturation level.
Healthy Mediterranean‐like dietary pattern predicts reduced risk of acute myocardial infarction (MI) in women: The Epic‐France (E3N) prospective cohort study
Authors: Ahluwalia Namanjeet , Tondeur Laura , Boutron Marie-Christine , Clavel-Chaperon Francoise
Published: June 2021

Limited data are available on the impact of overall dietary pattern on prospective risk of MI in women particularly in Europe. We conducted a prospective analysis among 71,886 women in the French E3N‐EPIC study (mean age of 52.9 y at dietary assessment using a 208‐item validated FFQ). During a median 11.8 y follow‐up, 171 cases of incident MI were confirmed. Factor analysis identified two dietary patterns: “Western‐Alcohol” (high in processed foods including meats, canned fish, butter, cream, and alcoholic beverages) and “Healthy‐Mediterranean (Med)‐like” (high in fruit, vegetables, non‐processed fish, olive oil, and sunflower oil). Western‐Alcohol pattern was not associated with MI. The absolute risk of MI was 24.8 and 12.8 per 100,000 women in the 4th and 1st quartiles of the Healthy‐Med‐like pattern, respectively. The hazard ratio for 4th vs 1st quartile was 0.58 (95% CI: 0.37–0.92) after adjusting for CVD risk factors, hormone‐replacement therapy and energy intake. Greater adherence to a Med diet pattern may contribute to MI reduction in women. The lack of an association between Western pattern and MI may suggest that alcohol intake counterbalances the atherogenic effect of Western diet.
Association between dietary protein and energy intake, handgrip strength (HGS), and lean body mass (LBM) in hypoalbuminemic maintenance hemodialysis (MHD) patients
Authors: Parsons Tara Rae , Bross Rachelle , Molnar Miklos Z , Kalantar-Zadeh Kamyar
Published: June 2021

BACKGOUND

Protein energy wasting/malnutrition is a significant problem in MHD patients. More LBM and greater HGS are associated with better survival rates in MHD patients. These associations are not well described in hypoalbuminemic MHD patients who have increased morbidity and mortality.

METHODS

Hypoalbuminemic (albumin <4.0 g/dL) MHD patients from DaVita Dialysis Centers in Los Angeles were enrolled in a randomized, controlled study (FREDI) associating meals during dialysis, phosphorus control and nutritional status. A dietitian measured HGS, assessed LBM by dual‐energy x‐ray absorptiometry, and protein and energy intake by interview‐assisted three‐day food record.

RESULTS

The baseline association between protein and energy intake, HGS, and LBM was assessed in 65 (28 male, 37 female) MHD patients, including 33 Hispanics and 32 Non‐Hispanics with a mean (± SD) age of 56 (±13) years.

Body Mass (kg)
LBM (%)
Seum Albumin (g/dL)
HGS‐right hand (kg)
Protein Intake (g/day)
Energy Intake (kcal/day)

75.4 (±16.6)
68.9 (±8.5)
3.71 (±.27)
24.0 (± 8.7)
73.5 (± 31.3)
1573 (± 705)

Protein intake (r=0.38, p=.002), energy intake (r=0.45, p<.001) and HGS (r=0.43, p<.001) were positively correlated with percent LBM.

CONCLUSIONS

These findings suggest that even in hypoalbuminemic MHD patients, protein and energy intake is associated with better nutritional status assessed by HGS and LBM.

Shire US Inc., Wayne, PA 19087.