bluuebunny/crossref_metadata_2025 huggingface dataset:
Chronic binge alcohol (CBA) dysregulates mesenteric adipose tissue phenotype in Simian Immunodeficiency Virus (SIV) infected macaques
Authors: Simon Liz , Ricks Joshua , Berner Paul , Stouwe Curtis Vande , Molina Patricia E
Published: November 2023

Alterations in lipid metabolism, with or without an associated fat “redistribution” or lipodystrophy lead to a greater risk of insulin resistance or frank diabetes mellitus in people living with HIV/AIDS (PLWHA). PLWHA with alcohol use disorders (AUDs) have higher odds of presenting with lipodystrophy and altered adipokine profiles, which have been linked to metabolic dysregulation in anti‐retroviral therapy (ART)‐treated PLWHA. Our studies have shown significant reduction in circulating adiponectin levels and dysregulation of whole body glucose‐insulin dynamics in CBA/SIV compared to sucrose (SUC)/SIV macaques. To determine whether these changes are associated with adipose tissue dysregulation, male rhesus macaques were administered daily CBA or sucrose (SUC) starting 3 months prior to intrarectal SIVmac251 inoculation and continued throughout the study period. ART or placebo was initiated 2.5 months after SIV infection and continued through the study period. Mesenteric adipose tissue was excised from four treatment groups (SUC/SIV±ART and CBA/SIV±ART) at study endpoint (11 months post SIV inoculation). Mesenteric adipose tissue of CBA/SIV macaques had decreased adipocyte size and a significant increase in the number of mast cells, immune cell infiltration and collagen staining compared to that of SUC/SIV macaques. Moreover, adipose derived‐stem cells of CBA/SIV macaques showed decreased differentiation compared to SUC/SIV macaques. These findings suggest that CBA exacerbates dysregulation of adipose tissue phenotype potentially contributing to metabolic dysregulation in SIV infection.

Support or Funding Information

The work was supported by National Institutes of Health (NIH) grants: P60 AA09803
IFN‐α inhibits Adipogenesis via regulation of JAK/STAT1 signaling
Authors: KYOUNGRAN LEE , Suhkneung Pyo
Published: November 2023

Obesity is characterized by hypertrophy and/or by the differentiation or adipogenesis of pre‐existing adipocytes. Some cytokines have been known to be negative regulators of adipocyte differentiation and obesity is associated with cytokines secreted by adipose tissue. It has been reported that IFN‐α is involved in the development of type 1 diabetes, lipid disorder and insulin resistatance. However, the effect of IFN‐α on adipogenesis remains unknown. In this study, we examined the inhibitory effect of IFN‐α on adipocyte differentiation and its mechanisms of action using colorimetric assay and Western blotting. IFN‐α inhibited the accumulation of lipid droplets and the expression of adipogenesis related genes. The inhibition of adipogenic differentiation by IFN‐α occurred primarily in the early stages of differentiation. In addition, IFN‐α arrested the cell cycle at the G0/G1 phase and caused decreased expression of CDK2 and p21. IFN‐α treatment increased STAT1 phosphorylation and STAT1 knockdown by siRNA prevented the ability of IFN‐α to inhibit the expression of PPARγ and C/EBPα in 3T3‐L1 cells. These data suggest that IFN‐α inhibits adipocyte differentiation during the early stage of adipogenesis by regulating the expression of PPARγ and C/EBPα through JAK/STAT1 signaling pathways in 3T3‐L1 preadipocytes.
Repeated Binge‐Like Alcohol Intoxication‐Induced Adaptive Immunity in Perilymphatic Adipose Tissue
Authors: Benavides Vienna Isabel , Song Kejing , Molina Patricia E , Souza‐Smith Flavia M.
Published: November 2023

Adipose tissue inflammation, characterized by the presence of major histocompatibility complex class II cells (MHC II), macrophages, and T cell activation, has been proposed to lead to the development of insulin resistance. In our previous studies, we showed that acute alcohol intoxication (AAI) induces mesenteric lymphatic hyperpermeability, promotes perilymphatic adipose tissue (PLAT) inflammation, disrupts adipokine profile, and increases systemic circulating lipopolysaccharide (LPS) levels, suggesting a potential role of AAI on metabolic dysregulation. Moreover, repeated binge‐like alcohol intoxication (RBAI) was also shown to increase PLAT inflammatory cytokine expression. Whether RBAI‐induced PLAT inflammation is associated with dysregulation of the adaptive immune system remains unknown. We hypothesized that RBAI promotes PLAT gene expression of T cells, macrophages, dendritic cells, and monocytes, exacerbating PLAT inflammation and increasing the risk for metabolic dysregulation. Male Sprague Dawley rats were surgically instrumented with a gastric catheter. RBAI was produced by daily administration of 2.5 g/kg/day of alcohol (12.5% alcohol w/v) or isocaloric dextrose in Vanilla Ensure (116 kcal/kg/day) for 3 consecutive days. PLAT was isolated at 30 minutes or 24 hours after the last RBAI episode. Gene expression of markers for T cells (CD4), macrophages (CD11c), monocytes (CD14), and dendritic cells (MHCII) was determined by Real Time PCR. RBAI resulted in a significant increase in gene expression of markers for T cells, macrophages, and dendritic cells at 30 min post final alcohol binge when compared with PLAT isolated from controls and from RBAI animals at 24h post final binge. No significant differences in monocyte gene expression were detected between the groups. These results suggest that RBAI promotes adaptive immune cell infiltration to PLAT. We predict that these cells contribute to the increase in PLAT cytokine expression detected in our previous studies and speculate that together; PLAT inflammatory milieu and innate immunity cell infiltration, promote localized insulin resistance.

Support or Funding Information

Supported by LSUHSC‐NO Department of Physiology, STRIDE Undergraduate Research Fellowship (APS and NHLBI 1 R25 HL1155473‐01)
Phrenic and Cervical Afferents Following Spinal Cord Injury
Authors: Nair Jayakrishnan , Jorgensen Marda L , Reier Paul J , Fuller David D
Published: November 2023

The phrenic nerve contains a mixture of motor, sensory, and sympathetic fibers. The diaphragm has few spindles and tendon organs, and a relatively greater population of unmyelinated afferents. Unmyelinated fibers are important in regulating diaphragm blood flow, and myelinated fibers convey information regarding diaphragm muscle contraction. The anatomical projections and distribution of phrenic afferents in the cervical spinal cord have not been described in detail in adult mammals. Using adult Sprague‐Dawley rats, our first purpose was to characterize the neuroanatomical projections of myelinated phrenic afferents in the cervical spinal cord and describe the distribution of non‐myelinated mid‐cervical afferents. Our second purpose was to test the hypothesis that cervical spinal cord injury (SCI) rostral to the phrenic nucleus would alter the distribution of myelinated phrenic afferents and non‐myelinated mid‐cervical afferents. We developed a new method to label myelinated phrenic afferents which consisted of prolonged soaking of the phrenic nerve with 1% cholera toxin‐β (CT‐β). In spinal‐intact rats, CT‐β‐positive afferents were relatively sparse, projected exclusively to the spinal cord ipsilateral to the nerve soak in 9 of 10 rats, and terminated in the dorsal horn (laminae II and III) and intermediate grey region (laminae IV and V). However, following both acute and chronic SCI (lateral cervical hemisection at C2), CT‐β‐positive phrenic afferents could be detected in the spinal cord contralateral to the side of CT‐β application in 11 of 12 rats. In additional studies, CT‐β was delivered directly to the C4 dorsal root ganglion, and this approach confirmed that robust detection of contralateral projecting cervical afferents only occurred in rats with SCI. Further experiments used immunochemistry to detect calcitonin gene related peptide (CGRP) and isolectin‐B4 (IB4) as markers of unmyelinated cervical afferents in spinal cord from rats with C2 hemisection (>12 wks post‐injury) or uninjured control. Quantitative assessment of staining intensity showed significant and bilateral increases in CGRP‐ and IB4‐positive fibers in the C4 dorsal horn after injury (P<0.05 vs. control). We conclude that both unmyelinated cervical afferents and myelinated phrenic afferents undergo considerable remodeling following cervical SCI

Support or Funding Information

1R01NS080180‐01A1 (DDF)
Oxidative Enzyme Activity in Respiratory and Non‐Respiratory Muscle of Hypoxic Mice and Human Respiratory Patients
Authors: O'Connor Karen M. , Lewis Philip , O'Leary Andrew J. , Burns David P. , NiHici Tamara , Mayooran Nithiananthan , Hinchion John , O'Halloran Ken D.
Published: November 2023

Despite the clinical relevance, little is known about the effects of long‐term exposure to hypoxia on respiratory muscle form and function. Recently, we demonstrated that exposure to chronic sustained hypoxia (CSH) causes respiratory muscle weakness due to oxidative stress (1). Redox proteomics with mass spectrometry revealed widespread oxidation of enzymes including those of the mitochondrial respiratory chain (1). We sought to explore citrate synthase activity in muscle samples from normoxic and CSH exposed mice. We have also initiated a study in human respiratory patients to explore the relationship between hypoxaemia and muscle aerobic capacity.

C57BL6/J mice were exposed to normoxia or CSH (FiO2 = 10%) with/without antioxidant supplementation (either 1% N‐acetyl cysteine (NAC) or 1mM Tempol) for a period of six weeks (n=4–8 per group). Animals were anaesthetized with 5% isoflurane and euthanized. The diaphragm, soleus, extensor digitorum longus and tibialis anterior were excised. Following approval, external intercostal (n=10) and latissimus dorsi (n=9) muscle samples were harvested from respiratory patients under‐going thoracic surgery in Cork University Hospital. Patients presented with a wide range of pre‐operative blood oxygenation (PaO2) values breathing room air, including COPD patients. Homogenisation was performed on muscle samples, followed by protein quantification using a bicinchoninic assay (BCA). Citrate synthase activity was measured spectrophotometrically, as an index of oxidative enzyme activity (aerobic capacity), using commercial kits.

Values are reported as mean±SEM. In mouse, diaphragm and limb muscles showed differential responses to hypoxia. Antioxidant supplementation with either NAC or Tempol prevented the hypoxia‐induced reduction in mouse diaphragm citrate synthase activity (P<0.0001). In human muscle, there was a significantly higher citrate synthase activity (P=0.018) in external intercostal compared with latissimus dorsi. Our pilot study examining citrate synthase activity as a function of PaO2 in respiratory patients is suggestive of a positive correlation, revealing low enzyme activity in two hypoxaemic patients (PaO2 < 60mmHg).

The working respiratory muscles are particularly susceptible to hypoxia‐induced reduction in citrate synthase activity. Recent work from our group has revealed considerable redox remodelling and metabolic reprogramming in respiratory muscle following CSH exposure (1). Preliminary data from human external intercostal muscle suggest a similar outcome to mouse studies. Antioxidants may have application as an adjunctive therapy in respiratory diseases characterised by hypoxia.

Support or Funding Information

The Physiological Society
Cholera Toxin‐B Coated Protocells as a Targeting Drug Delivery System to Motoneurons
Authors: Porras Maria Alejandra Gonzalez , Gregory Ashley , Durfee Paul , Brinker Jeffrey , Sieck Gary , Mantilla Carlos
Published: November 2023

At present, there are limited delivery systems to target neurons in the central nervous system. Neurological diseases are commonly hindered by treatments lacking drug selectivity to the neurons of interest. In particular, motoneuron diseases will likely require selective targeting of therapeutic interventions, which can be delivered via the peripheral nervous system. Nanoparticles are novel drug delivery systems with exceptional therapeutic potential. Specifically, mesoporous silica nanoparticles coated with a lipid bilayer (protocells) are a suitable treatment vehicle to target the peripheral and central nervous system because they offer advantageous fusion characteristics including 1) increased surface area with larger loading capacity, 2) stability and 3) improved biocompatibility. A number of natural toxins exist that already target the nervous system and could be used as a targeting strategy. Cholera toxin produced by the bacterium Vibrio cholerae has an atoxic subunit (CTB) that binds to gangliosides present on neuronal membranes and is a reliable motoneuron tracer. We hypothesized that CTB‐coated protocells specifically target motoneurons. In culture, CTB uptake is evident in motoneuron‐like NSC‐34 cells but not in L6 muscle cells, consistent with the suitability of CTB as a motoneuron targeting strategy. Fluorescently labeled protocells (130 nm in diameter) were bound to CTB using biotin‐neutravidin. CTB‐protocells showed significantly greater motoneuron uptake compared to non‐CTB coated protocells when quantified by measurements of intracellular protocell fluorescence intensity. Confocal images were used for co‐localization of fluorescently labeled protocells and lysosomes. Protocells effectively escape endo‐lysosomal degradation delivering their cargo intracellularly. Biocompatibility was evidenced by lack of cytotoxicity in cultured motoneurons and unimpaired cellular proliferation. To evaluate the targeting efficacy of CTB‐protocells in tissue, we utilized isolated diaphragm muscle‐phrenic nerve preparations. Evidence of protocell uptake at diaphragm neuromuscular junctions measured by protocell fluorescence intensity supports uptake into phrenic motoneuron axon terminals. In conclusion, CTB‐protocells are a promising, customizable approach for targeting and delivering drugs to motoneurons in vivo.
Pulmonary Collagen Content in 1‐Year‐Old Rats Exposed to Postnatal Hyperoxia
Authors: Sobotik Atzie , Pegelow David , Goss Kara , Haraldsdottir Kristin , Braun Rudolf , Barton Greg , Tetri Laura , Eldridge Marlowe
Published: November 2023

Introduction

Bronchopulmonary dysplasia (BPD) is a chronic lung disease of preterm infants exposed to oxygen therapy postnatally. Although it is a vital, life‐saving treatment, oxygen therapy is closely associated with long‐term consequences, including decreased alveolarization and reductions in pulmonary vasculature cross‐sectional area. Previous studies using a BPD rodent model have demonstrated an increase in lung collagen content within the first 20 days of birth. This suggests that the aberrant lung development in the BPD animal model is associated with pathological changes in the extracellular matrix (ECM) (i.e. increased collagen content). No study to our knowledge has looked at collagen accumulation later in life as a result of early postnatal hyperoxia treatment. Furthermore, it is currently unknown whether or not there are sex differences in the ECM remodeling. The purpose of this study is to determine if 14 days of postnatal hyperoxia exposure affects collagen accumulation in the lungs of 1‐year‐old male and female rats.

Methods

An established rat model of BPD using Sprague‐Dawley rats was. Within 12 hours of birth, rats were placed in a hyperoxic (HYP, 85% O2) chamber, or normoxic (NORM, 21% O2) chamber. After 14 days in respective conditions, all rats were allowed to age until 1 year in room air. The aged rats were sacrificed, and tissues were harvested and flash‐frozen in liquid nitrogen. Colorimetric assays were performed on lung tissue to assess overall hydroxyproline content as a measurement of collagen. Histology was also analyzed to visually assess collagen levels using Masson's trichrome‐stained lung slides. Four pictures were taken per slide, each picture was divided into 9 sections, and a blinded party scored each segment's collagen content on a scale of 1–3 (0=none, 1=mild, 2=moderate, 3=severe). Two‐way ANOVAs were used to determine the effects of sex, HYP exposure, and their influence on lung collagen.

Results

The colorimetric assay revealed that aged hyperoxic females (n=7) demonstrate a 23% increase in collagen content compared to aged normoxic females (n=5, p=0.02). Aged HYP males (n=6) demonstrated similar pulmonary collagen content compared to aged normoxic males (n=3) based on colorimetric assay. However, histopathologic examination revealed increased collagen deposition in both males and female aged HYP rats (p=0.07, n=2–3).

Conclusions

These results suggest that females exposed to HYP in early‐postnatal life are more susceptible to collagen accumulation in the lungs, which may be indicative of increased lung injury as compared to males.

Support or Funding Information

NIH‐NHLBI R01 – HL115061

NIH‐NHLBI R01 Supplement – HL1150613
Predictors of Mortality Following Systemic LPS Injections During a Critical Period of Neonatal Development
Authors: Mayer Catherine A. , Desai Vilok , MacFarlane Peter M
Published: November 2023

We showed previously the presence of a developmental window (postnatal age 10 days, P10) during which the respiratory neural control system exhibits a heightened vulnerability to a systemic injection of LPS. The vulnerability is associated with an attenuated hypoxic ventilatory response (HVR) and an unexpected high degree of mortality (~50% of rat pups) and is unique to the P10 age group because neither younger (P5) nor older (P20) rats appeared to be affected by LPS. Since only about half of the P10 rats appear to be vulnerable to LPS, we investigated whether the mortality could be predicted by prior examination of: 1) the magnitude of their hypoxic ventilatory response (HVR); and 2) changes in serum cytokine levels. Rat pups received an intraperitoneal injection of LPS (0.1mg/kg) or saline at P10. Plethysmography was performed at 2hrs post injection to determine the magnitude of the HVR; a blood sample was also taken immediately after plethysmography (via facial vein puncture) in unanesthetized rats to determine changes in serum TNFα levels using an ELISA. Rats were returned to the mother and monitored closely throughout the following 8hr time period for signs and symptoms of imminent death (color change, labored breathing, lethargy). 42% of rats treated with LPS died between 4–7 hrs after LPS whereas the remaining 58% survived. However, in the rats that died, the HVR at 2 hrs after LPS was reduced (0.64 ± 0.07 ml/g/min) compared to both saline treated rats (1.14 ± 0.13 ml/g/min) and survivors of LPS (0.97 ± 0.13 ml/g/min). The attenuated HVR of rats that died from LPS was associated with elevated serum TNFα levels (884.1 ± 89.4pg/ml; p<0.05) compared to saline rats (not detectable) and animals that survived LPS. These data suggest a sub‐population of rats exhibit a heightened and lethal vulnerability to LPS exposure that can be predicted by the magnitude of their HVR and serum TNFa levels prior to death. The cause of the vulnerability remains unknown although we speculate it may involve an aberrant “inflammatory” response to an otherwise sub‐lethal exposure to the endotoxin, resulting in a fatal disturbance in the cardio‐respiratory control system.

Support or Funding Information

Department of Pediatrics, Rainbow Babies and Children's Hospital, Case Western Reserve University
Effects of A2A and 5‐HT2A Antagonists on Hypoxic and Hypercapnic Ventilatory Response in Rats Exposed to Chronic Sustained Hypoxia
Authors: Moya Esteban A , Powell Frank L
Published: November 2023

Different patterns and durations of hypoxic exposure cause different forms of plasticity in the neural control of ventilation (Pamenter & Powell, F1000Prime Rep, 5: 23, 2013). For example, acute intermittent hypoxia (AIH) causes long term facilitation (LTF) of ventilation and phrenic nerve activity, while chronic sustained hypoxia (CSH) causes ventilatory acclimatization to hypoxia (VAH). In both LTF and VAH, ventilation in normoxia is increased after the hypoxic stimulus is removed. Previous work has shown that 5HT2a serotonin receptor activation (Gq pathway) is necessary for LTF but not VAH. However, recent work (Nichols et al., J Appl Physiol, 112: 1678–1688, 2012) shows A2A adenosine receptor activation (Gs pathway) is also involved in LTF with more severe AIH (PaO2 = 25–30 Torr vs. 35–45 Torr). We hypothesized that the Gs pathway also contributes to VAH and tested this in adult male rats by blocking the Gs and Gq pathways before and after 7 days of CSH (PO2 = 80 Torr) and measuring ventilation and the hypoxic ventilatory response (HVR) with barometric pressure plethysmography. Consistent with previous work, Gq blockade with a 5HT2a antagonist (Ketanserin, 1 mg/Kg i.p.) caused no significant changes before or after CSH. Gs blockade with an A2A agonist antagonist (MSX‐3, 1mg/Kg i.p.) had no effect before CSH but after CSH it significantly increased ventilation in acute normoxia (1777±352 vs. 2396±268 ml/min*kg) and hypoxia (2063±195 vs. 2403±450 ml/min*kg). This contrasts with the effect of Gs blockade during severe AIH to significantly decrease LTF and does not support our hypothesis. However, further experiments are necessary to determine the effects of blocking Gs and Gq pathways during CSH.

Support or Funding Information

NIH RO1 HL‐081823
Ventilatory and Neurochemical Effects of Microdialysis of the Ionotropic GABAA Receptor Agonist Muscimol into the Ventral Respiratory Column of Awake Goats
Authors: Langer Thomas M , Neumueller Suzanne , Burgraff Nicholas , Crumley Emma , Pan Lawrence , Hodges Matthew R , Forster Hubert V
Published: November 2023

We have previously dialyzed atropine, a nonspecific muscarinic receptor blocker in the ventral respiratory column (VRC) of chronically cannulated awake goats and observed an increase (J. Appl Physiol. 114, 694–647, 2012) or no change (Respir Physiol 205: 7–15, 2014) in ventilation (VI) and breathing frequency (f). The unexpected effects on VI and f with attenuating excitatory muscarinic receptor activity may have been due to the local compensatory increase in the excitatory neuromodulators serotonin (5‐HT) and substance P (SP) measured in the effluent dialysate mock cerebrospinal fluid (mCSF). To determine if a similar mechanism of compensation accompanied increased inhibition in the VRC, we dialyzed DAMGO, a μ‐opioid receptor agonist and found unexpectedly that absolute VI and f did not change (unpublished data). Analyses of effluent mCSF dialysate showed no changes in local 5‐HT or SP, but instead showed a decrease in GABA. Thus, these findings suggested that perturbation of G‐protein coupled receptors to not uniformly elicit local compensatory changes in neurochemicals and/or VI. In the present study, we dialyzed the ionotropic GABAA receptor agonist muscimol (0.5, 1.0, and 10.0 mM) into the VRC of awake goats (n=6), hypothesizing that it would inhibit ventilation as previously shown by others. Dialysis of muscimol at all doses had no effect on VI or f (P<0.05), but during the hour following drug application VI and f were increased (P<0.05) at all doses of muscimol. Associated with these studies, we found no changes in local SP, 5‐HT, GABA or glycine for all doses of muscimol. However, in 3 of 6 goats there was an increase in the concentration of norepinephrine in the effluent dialysate in response to 10 mM muscimol. Norepinephrine remained unchanged during dialysis of the lower doses of muscimol and in all of our previous dialysis studies. The results of these and previous studies support the concept of interdependence among local neurochemicals, and furthermore suggest that the mechanism of compensation differs between attenuation of excitatory and inhibitory G‐protein coupled receptors and between perturbation of G‐protein and ionotropic inhibitory receptors.

Support or Funding Information

National Heart, Lung, and Blood Institute Grants HL‐112996, HL‐007852, and the Department of Veterans Affairs
Combined effects of intermittent hyperoxia and intermittent hypoxia on respiratory control development in rats
Authors: MacDonald Carolyn A , O'Toole Caitlin A , Butler Jesse A , McDonough Amy B , Bavis Ryan W
Published: November 2023

Preterm infants may experience hyperoxia due to early exposure to atmospheric O2 levels (relative hyperoxia) or through administration of supplemental O2. Although intermittent hyperoxia has been found to alter breathing in neonatal rats, hyperoxia alternates with bouts of hypoxia clinically. In this study, we exposed newborn rats for 14 days to four treatments: control (room air), sustained hyperoxia (30% O2; SH), intermittent hypercarbic hypoxia (10% O2/6% CO2, 5 episodes h−1; I10/30), or intermittent hyperoxia + hypercarbic hypoxia (10% O2/6% CO2 to 40% O2/0% CO2, 5 episodes h−1; I10/40); 30% O2 was used as the background O2 level for I10/30 and I10/40 to mimic the relative hyperoxia of preterm birth. SH and I10/40 rats had reduced carotid body responses to hypoxia, but only SH rats exhibited blunted ventilatory responses to hypoxia. CNS plasticity may compensate for abnormal carotid body function in I10/40 rats, or SH could alter breathing through additional mechanisms. In either case, intermittent hypercarbic hypoxia appears to diminish the effects of intermittent hyperoxia on the respiratory phenotype.

Support or Funding Information

Supported by NIH grants HL114001 and GM103423.
Effect of Omega‐3 Polyunsaturated Fatty Acids on Insulin Resistance are Sex‐dependent: A Meta‐analysis of Randomized Controlled Trials
Authors: Garg Manohar Lal , Abbott Kylie , Thota Rohith , Acharya Shamsunder , Burrows Tracy Lal
Published: November 2023

Interventions that improve Insulin Resistance (IR) may be able to prevent or delay the onset of type 2 diabetes in at‐risk populations. Cohort and cross‐sectional studies suggest there is an inverse relationship between omega‐3 polyunsaturated fatty acids (n3PUFAs) and IR, however randomised control trials (RCTs) have had equivocal findings. Recent findings by our research group suggest that effects may be sex‐dependent. The aim of this systematic review and meta‐analysis is to determine whether the results of RCTs differ according to sex.

Five databases were searched (Medline, EMBASE, PubMed, CINAHL and Scopus) to locate RCTs that included a dietary or supplement‐based n3PUFA intervention and measures of IR in adult populations. Studies reporting HOMA‐IR were pooled for meta‐analysis. Sub‐group analysis was conducted to explore sex‐dependent differences.

Thirty‐two trials were included in the systematic review, including a total of 2326 participants (mean age 53.4 years; BMI 29.7). Of those included, five were conducted in males only, seven in females, and the remaining included both sexes (n=20). Sixteen trials were pooled for meta‐analysis (males n=2; females n=4; combined n=10). There was considerable heterogeneity in terms of trial duration (mean 12.8 weeks; range 3–48 weeks), n3PUFA dose and placebos used. When all studies were pooled they showed no significant effect of n3PUFA on HOMA‐IR, with mean difference of intervention vs control (95%CI): −0.23 (−0.382, 0.337) (p=0.902). Subgroup analysis showed n3PUFA intervention significantly improved HOMA‐IR in females: −0.559 (−9.989, −0.130) (p=0.011), but had a non‐significant trend towards deterioration of HOMA‐IR in males: 1.382 (−0.183, 2.947) (p=0.083). No trials conducted in males found a positive effect of n3PUFA supplementation on insulin resistance.

Findings from this systematic review and meta‐analysis support a sex‐dependent effect of n3PUFA intervention on markers of IR, with a protective effect evident in females only. n3PUFA interventions could potentially prevent the development of type 2 diabetes in females. Further research is required to further elucidate potential mechanisms of these sex‐dependent differences.

Support or Funding Information

n/a
Mathematical modeling of the microenvironment beneath a surface pH electrode tip
Authors: Occhipinti Rossana , Calvetti Daniela , Boron Walter F , Somersalo Erkki
Published: November 2023

The time course of extracellular surface pH (pHS), recorded by a blunt pH microelectrode, is the key physiological data to assess membrane permeability to CO2 (PM,CO2) when exposing an oocyte to a solution containing CO2/HCO3−. In an effort to extract PM,CO2 values from pHS measurements, we previously developed a reaction‐diffusion mathematical model of CO2 influx into a spherical cell. The model accounts for the slow interconversion of CO2 and HCO3− as well as the reactions of a multitude of non‐CO2/HCO3− buffers. When compared to physiological experiments, the model can predict the essential features of the measured pHS transients. However, it is clear that further improvements in the quantitative agreement of the model with the physiological data, will require additional refinements of the model, including a detailed modeling of the micro‐environment under the pHS electrode tip, when pushed against the cell membrane.

In the present study, we propose two computational models for studying the effect of the pHS electrode on pHS measurements. In the first approach, we develop a two‐dimensional model of the pHS electrode and the extracellular microenvironment underneath the electrode by assuming that the electrode‐membrane interface is an infinitely thin homogenous disc within which diffusion occurs only in the tangential direction along the disc surface as well as at the interface between the disc and the bulk extracellular fluid. In addition, transmembrane gas exchange occurs between the disc and the cytoplasm, in a direction perpendicular to the disc. Mathematically, this model is described by two‐dimensional reaction‐diffusion equations that are solved numerically by a combination of finite difference (FD) schemes and stiff solvers. Because this model does not reveal the dependence of the pHS transient on the distance of the pHS electrode from the cell membrane, in the second approach, we develop a three‐dimensional reaction‐diffusion model of the pHS electrode tip and of its surrounding environment, where the reaction‐diffusion equations are now solved by a combination of the finite element method (FEM) with stiff solvers.

Our preliminary results corroborate the hypothesis that diffusion is slower in the micro‐environment under the electrode tip, resulting in substantially larger pHS transients than those on a free membrane surface. The work points towards the possibility of defining a reliable electrode gain function that corrects the effect of the electrode.

Support or Funding Information

Supported by grants from NIH (U01‐GM111251, K01‐DK107787)
Three‐dimensional Microenvironment with Uniaxial Topology Induces Myoblast Maturation
Authors: Ko Ung Hyun , Park Suk‐Hee , Choung Jinseung , Lee Junho , Shin Jennifer
Published: November 2023

Introduction

With the recent advancement in micro‐manufacturing technology, there have been remarkable research progresses in utilization of synthetic scaffolds for tissue engineering applications. The synthetic scaffold exhibits high sustainability of sub‐micron structure, enabling researchers to mimic the micro‐environments of the in vivo tissues. However, the manufacturing resolution of the synthetic scaffold still needs improvements to provide truly the 3D environment for the cells by reflecting the dimensions and structural features the cellular extracellular matrix (ECM). On the other hand, hydrogels such as collagen, fibrin, or matrigel can provide 3D environment yet with weak mechanical strength and stability.

Materials and Methods

In this study, we incorporated collagen with the electrospun scaffold to realize the sub‐micron uniaxial pattern with 3D substrate environment. The synthetic scaffold was fabricated by electrospinning method using polycaprolactone (PCL) polymer, and the uniaxial topology was regulated by the tilted gap collectors. Mouse myoblasts (C2C12s) were expanded on electrospun scaffold and collagen gel was added on the scaffold after one day. C2C12 cells were cultured for 6 days in total and subsequently differentiated for 7 days in DMEM supplemented with 5% equine serum.

Results and Discussion

We found that our hybrid scaffold system effectively induced cellular elongation compared to ordinary uniaxial synthetic scaffolds and promoted myogenic differentiation. This elongation effect was initially induced by the uniaxial cue of synthetic backbone, and 3D environment additively increased cellular sustainability leading to higher elongation. These highly elongated cells were then induced to aggregate to form 3D myofibril whose maturation was further evoked by the application of electric field stimulation during the differentiation stage.

Conclusion

Conclusively, we expect that the concept of our hybrid scaffold can be utilized in the field of skeletal muscle engineering and the underlying mechanism behind the role of physical cues in cellular physiology would have potential to be applied broadly in understanding the diseases and therapeutics in general.

Support or Funding Information

Acknowledgements: This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MEST) (2013R1A2A2A01017014).
Cross‐Linked Polypeptide Multilayer Films Facilitates the Cardiac Differentiation of Mouse iPSCs
Authors: Wang Si‐Han
Published: November 2023

Well‐designed biomaterials, which mimic physiological microenvironment and mechanical properties, can guide stem cell differentiation. Polyelectrolyte multilayer films allow for modulation of thickness, stiffness, and adhesiveness and is a useful method for substrate coating to modulate cellular behaviors. In this study, mouse induced pluripotent stem cells (iPSCs) were cultured on polypeptide multilayer films built up by the alternate deposition of cationic poly‐L‐lysine and anionic poly‐L‐glutamic acid, cross‐linked with EDC/sulfo‐NHS, and coated with gelatin. The characteristics of film thickness, surface roughness, and adsorbed film mass were measured by AFM and QCM. Cardiomyocyte differentiation was evaluated by RT‐PCR analysis of cardiac markers, cardiac troponin T (cTnT) and myosin light chain‐2a (MLC‐2a). After embryonic bodies of iPSCs were cultured for 3 days and then induced for cardiac differentiation for 0, 3, and 6 days, compared with iPSCs on culture dish, iPSCs on 6‐layer cross‐linked polypeptide films demonstrates significantly higher expressions of both cTnT and MLC‐2a on day 3 and day 6. Our results indicate a possible way to facilitate iPSCs toward cardiomyocyte differentiation on polypeptide multilayer films.

Support or Funding Information

This work was supported by the Ministry of Science and Technology, Taiwan. (Grant number: NSC 102‐2628‐B‐010‐010‐MY3).
Sphingosine 1‐phosphate Potentiated Endothelial Cell Attachment on De‐cellularized Human Umbilical Vein as a Scaffold for Vascular Tissue
Authors: Chen Wei‐Min , Hsia Kai , Yao Chao‐Ling , Lu Jen‐Her , Lee Hsinyu
Published: November 2023

Cardiovascular disease (CVD) is the leading cause of mortality and morbidity. Choosing a proper material for bypass grafting and avoiding postoperative coagulopathy is critical. In this study, human umbilical veins (HUV) was applied as natural allograft scaffolds and de‐cellularized to remove antigens from HUV donor. De‐cellularized scaffold was then seeded with human umbilical vein endothelial cells (HUVECs) or endothelial progenitor cells (EPCs) in the presence of sphingosine 1‐phosphate (S1P) in rotational cultural conditions. Previous study shows that S1P is a key biolipid signaling molecular that regulates endothelial cell growth and survival. In addition, S1P has been shown to reduce the shedding of syndecan‐1 on endothelial cell surface. Therefore, S1P was hypothesized to enhance re‐attachment of cells onto scaffold and stabilize the surface glycocalyx layer of endothelial cell, which may inhibit blood coagulation. Adhesion assays and H&E staining results showed that S1P indeed enhanced cell adhesion on scaffold. In addition, immunofluorescence staining results showed that S1P prevented syndecan‐1 shedding, but not glypican‐1. Furthermore, S1P treated scaffold reduced platelet adhesion in whole‐blood coagulation tests. Taken together, our results suggested that S1P inhibited coagulation by protecting glycocalyx shedding. Therefore, S1P treated re‐cellularized HUV scaffolds may be potentially suitable for vascular grafts.
A new twist to the primitive mechanical fate identifies an aged‐associated TGFβ1 nexus in vascular smooth muscle cells
Authors: Zhu Wanqu , Isak Abraham , Monticone Robert E , Wang Mingyi , Lakatta Edward G , An Steven S
Published: November 2023

Central arterial stiffening is an independent predictor of cardiovascular morbidity and mortality in the U.S., and is adversely associated with advancing age. The transgression of age‐associated arterial stiffening is classically defined by pro‐inflammatory remodeling of the central arteries, culminating in altered deposition of extracellular matrix (ECM) and hyperplastic changes within the vascular smooth muscle (VSM) layer. Surprisingly, our understanding of age‐associated mechanical changes in the VSM compartment is largely unclear, and genetic/epigenetic nexus remains elusive. Here we adapted Fourier transform traction microscopy and magnetic twisting cytometry for the single‐cell analyses on the identity and state of mechanical properties of individual VSM cells isolated from the central aorta of young (8 months) vs. old (30 months) Fisher 344 cross‐bred Brown Norway rats. Compared with young VSM cells, old VSM cells were bigger in size and spontaneously exercised greater traction stress upon tuned elastic matrix. Old VSM cells also exhibited higher cytoskeletal (CSK) stiffness that is long‐lived in culture and, in the absence of inflammatory mediators, persistent across a wide range of matrix rigidities in a pathophysiologic spectrum. Moreover, Old VSM cells expressed higher levels of the latent TGFβ1 and showed intrinsically active TGFβ1 signaling. On the one hand, exogenous addition of TGFβ1 appreciably increased CSK stiffness in young, but not in old VSM cells. On the other, GW788388 and A8301, pharmacological inhibitors of TGFβ1 receptors, decreased the phosphorylation of Smad2/3 and normalized in turn the stiffness of old VSM cells in a dose‐dependent manner. Taken together, our findings demonstrate an age‐associated VSM contraction mechanophenotype that is independent of immune inflammatory responses, and provide a nodal point for potential therapeutics to rejuvenate and mitigate the central arterial stiffening.

Support or Funding Information

NIH R01 HL107361
Foods Consumed from Non‐School Cafeterias and Vending Machines: NHANES 2006–2012
Authors: Onufrak Stephen J
Published: November 2023

Nutrition guidelines for foods and drinks offered in non‐school cafeterias and vending machines are being used as a strategy to increase availability of healthy food items. However, little is known about who consumes food from these sources and the types and nutritional characteristics of the foods consumed. Using day 1 dietary recall data from NHANES 2006–2012 (N=15,701 adults with valid recall data), we estimated the prevalence among adults ≥ 20 years old of consuming foods from these sources, the most commonly consumed foods, and the nutritional characteristics of foods consumed. Overall, 4.1% of adults consumed items on the recall day from vending machines (mean energy intake from vending sources among consumers=276kcal) and 3.0% consumed items from a cafeteria (mean energy intake from cafeteria sources among consumers=643 kcal). Consuming foods from vending was more commonly reported by males (5.0%) compared to females (3.2%) but consuming foods from cafeterias did not differ by gender. Soft drinks, candies, salty grain snacks, potato chips, and cookies collectively accounted for approximately 67% of all energy consumed from vending machines. Leading sources of energy from cafeterias were grain mixtures (e.g. sandwiches, pizza, pasta dishes, and rice dishes), white breads/rolls, chicken, and mixed dishes of meat/poultry/fish with starch items and vegetables (e.g. stews). Calories from added sugars accounted for nearly half of total energy intake from vending and approximately 12% of total energy intake from cafeteria sources. Among those who consumed foods from cafeterias, mean total vegetable/ legume intake was 0.8 cups and mean total fruit intake was 0.2 cups from cafeteria sources. Vegetable and fruit intake from vending sources was much less (0.04 cups total vegetables/legumes and 0.03 cups total fruit from vending sources among vending consumers). A better understanding of who is consuming foods from cafeterias and vending machines and what is consumed will better inform efforts to design nutrition guidelines for these settings and to estimate the potential publichealth impact of these standards.

Support or Funding Information

Done as part of official work duty. No other funding.
Preparation of an anatomical donor using the Sikon Soft‐Cure method
Authors: Sikon Richard R.
Published: November 2023

The options for preparing the Anatomical Donor can vary from program to program. Traditionally this involves the use of formalin or phenol based solutions. These solutions, while very effective can alter the presentation of the tissues and limit the scope of use. These uses include not only anatomical applications but also surgical and research.

Preparation of donors using the Sikon Soft‐Cure Embalming method does not significantly alter the tissues but rather leaves them in a normal‐like state with regards to appearance, flexibility and function. This process is entirely dependent upon the composition of the solution and a thorough understanding of the changes that occur during the process.

The presentation of the donor following the process of diffusion and perfusion may often be confused with the normal decomposition process. A thorough understanding of preservation processes and the effects of chemical changes within the tissues are key to the success of embalming donors prepared with the Sikon Soft Cure Method.
Plant Sterol Concentration in Plasma and Intake in Plant‐Based Diets
Authors: Jaceldo‐Siegl Karen , Lütjohann Dieter , Sirirat Rawiwan , Haddad Ella
Published: November 2023

Background

Limited information is available on intake and plasma concentration of plant sterols among vegetarians.

Objective

We measured plasma concentration of plant sterols, and estimated phytosterol intake in a representative sample (n=916) of the Adventist Health Study‐2 cohort with a range of plant‐based diets (10% vegans, 28% lacto ovo‐, 11% pesco‐, 5% semi‐vegetarians, and 46% non‐vegetarians).

Design

Plasma concentration of campesterol, sitosterol, cholestanol, lathosterol and cholesterol were measured by gas‐liquid chromatography. Campesterol, β‐stitosterol, and stigmasterol intake was estimated from six 24‐hour dietary recalls using plant sterol data of foods from USDA SR 27 and available literature. Differences by diet were determined using ANCOVA with Tukey‐Kramer's test and 95% CI to adjust for multiple comparisons.

Results

The analytic cohort included 66% women, and 46% Blacks. Mean (SD) age was 60 (14) y, and cholesterol 191 (39) mg/dL. Mean (95% CI) plasma sitosterol was significantly higher in vegans 4.6 (4.3–4.9) μg/ML compared to non‐vegetarians 3.9 (3.8–4.0) μg/ML and semi‐vegetarians 3.8 (3.4–4.2) μg/ML. Plasma campesterol or the non‐plant sterols (cholesterol, cholestenol, and lathosterol) did not differ across diets. Mean β‐stitosterol intake in mg/d among vegans (105.2), lacto ovo‐vegetarians (85.5), and pesco‐vegetarians (85.0) each was higher compared to non‐vegetarians (61.9) (each comparison, p<.0001). Range of intake (from non‐vegetarian to vegan) was 16.7–23.3 mg/d for campesterol, and 10.0–14.8 mg/d for stigmasterol. Compared to non‐vegetarians, intake of each of these phytosterols was significantly higher among pesco‐vegetarians, lacto ovo‐vegetarians, and vegans (each comparison, p<.0001).

Conclusion

We provide evidence that intake and plasma concentration of plant sterols may be higher among a variety of plant‐based diets compared to a non‐vegetarian diet.

Support or Funding Information

Funding for this project was made possible by Unilever. Collection of blood and dietary recalls was funded by the National Cancer Institute.
Soy positively impacts cecal microbiota of ovariectomized rats selectively bred for low aerobic capacity
Authors: Liu Tzu‐Wen , Zidon Terese M. , Welly Rebecca J. , Park Young‐Min , Britton Steven L. , Koch Lauren G. , Padilla Jaume , Vieira‐Potter Victoria J. , Swanson Kelly S.
Published: November 2023

Menopause associates with obesity and insulin resistance (IR) that are likely attributed to loss of ovarian hormone production. Soy is known to ameliorate these menopause‐associated metabolic dysfunctions for reasons that are not clear. The gut microbiome has been shown to mediate gut integrity and impact systemic inflammation associated with obesity. Soy‐mediated changes to gut microbial communities may represent one potential mechanism for its beneficial metabolic impacts in the setting of ovarian hormone insufficiency.

OBJECTIVE

To determine the impact of soy on cecal microbiota in intact vs. ovariectomized (OVX) rats bred for low‐running capacity (LCR), a model previously shown to mimic human menopause.

METHODS

20‐wk old LCR rats were either OVX or sham operated (SHM) and fed either soy‐rich (soy) or soy‐free (control) diets for 28‐wk, resulting in a 2×2 factorial design (n=10/group): 1) OVX/soy; 2) SHM/soy; 3) OVX/control; 4) SHM/control. Diets were isocaloric and comparable in protein, fat, carbohydrate, and fiber contents. DNA from cecal digesta collected at sacrifice were extracted, followed by 16S rRNA amplicon Illumina MiSeq sequencing and analysis using QIIME 1.9.1 and LEfSe.

RESULTS

As previously reported, soy reduced adiposity and improved insulin sensitivity of these OVX rats. Principal coordinates analysis (PCoA) of weighted and unweighted UniFrac distances of cecal microbiota revealed a sharp separation (p<0.05) between soy‐ and control‐fed groups. Alpha diversity measures revealed lower (p<0.05) species richness in rats fed soy vs. control. The soy‐rich diet decreased (p<0.001) the Firmicutes:Bacteroidetes ratio. Linear discriminant analysis (LDA) effect size analysis identified that rats fed soy had decreased relative abundance of Firmicutes (Clostridiaceae, Peptostreptococcaceae), Actinobacteria, and Spirochaetales, but increased relative abundance of Bacteroidetes (Prevotellaceae), Cyanobacteria, and Betaproteobacteria. OVX rats had increased relative abundance of Anaerovibrio compared to SHM rats. A positive correlation (p<0.001) was found between glucose intolerance and an undefined genus in S24‐7 family.

CONCLUSION

Soy significantly decreased the Firmicutes:Bacteroidetes ratio, suggesting a beneficial shift in gut microbial communities in our rat model mimics human menopause.

Support or Funding Information

(CBIS to V. Vieira‐Potter) by Grant Number P50AT006273