bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sex differences in alcohol‐induced HPA activation following systemic vs icv administration in rats: Neuronal activation studies
Authors: Davis Seth , Wakley Alexa A. , Yap Parry , Pham Charleen , Kiester Christopher , Selvage Dan John 
Published: November 2023

Understanding how alcohol differentially affects the hypothalamic‐pituitary‐adrenal (HPA) axis in males vs. females is important for the development of sex‐specific pharmacological treatments for alcohol abuse. Using intracerebroventricular (icv) administration, our lab previously reported that alcohol activated the hypothalamic‐pituitary‐adrenal (HPA) axis of rats in a sex‐ and estrous cycle‐ dependent manner. This effect was greatest in females in stages of the estrous cycle when the effects of estradiol (E2) are high (proestrus and estrus), as these animals showed greater levels of HPA‐ linked hormonal and neuronal activation by alcohol compared to males and to females in stages of the cycle when E2 levels are low (diestrus I and II). For the current study, we used an intraperitoneal (i.p.) alcohol injection to determine if systemic alcohol activates neuronal HPA activity in a similar manner as centrally delivered alcohol, or whether the many non‐CNS specific effects of the drug in the periphery affect its ability to cause sexually dimorphic activation of the HPA axis. Relative neuronal stimulation levels were determined using single‐ and dual‐ labeling immunofluouresence (IF) staining of the transcription factor and marker of neuronal activity c‐fos (Fos) in the medial parvocellular and magnocellular areas of the paraventricular nucleus of the hypothalamus (PVN), which is known to regulate HPA function. As with icv administration, systemic alcohol induced Fos expression in the medial parvocellular PVN, where cell bodies for corticotropin releasing factor (CRF), the main driver of HPA activation reside. Significant Fos expression was also stimulated in the magnocellular PVN, where cell bodies for arginine vasopressin (AVP), which contributes to HPA activation by CRF, are located. Unlike our results using icv administration, females receiving systemic alcohol exhibited less alcohol‐ induced medial parvocellular PVN Fos than males, with this difference reaching significance in diestrous females. Alcohol also induced less AVP‐ associated magnocellular PVN Fos in females than males following systemic alcohol injection, again with the greatest difference being between males and diestrous females. These results indicate that non‐ CNS specific effects of alcohol in the periphery contribute to HPA responsiveness to the drug in a sexually dimorphic manner, with the male HPA axis exhibiting a significantly greater reactivity to systemic injection of the drug than diestrous females.Support or Funding InformationNIH R01 AA016947‐01A1
Anti‐diabetic effects of class 1 histone deacetylase inhibition in a rodent model of type 2 diabetes mellitus
Authors: Johnson Emily , Marsh Susan 
Published: November 2023

Histone deacetylases (HDACs) have diverse roles in non‐cancerous cells, including an integral role in glucose and fatty acid metabolism. In the heart, class 1 HDACs (HDAC1, 2, 3, and 8) regulate cell growth, and class 1‐specific inhibitors prevent cardiomyocyte hypertrophy. In the pancreas, HDAC inhibition protects β‐cells from apoptosis due to inflammatory insults. Because dysregulated cardiac metabolism, cardiac hypertrophy, and β‐cell apoptosis are complications of type 2 diabetes, we investigated the effects of a class 1 HDAC inhibitor on these outcomes. Type 2 diabetic db/db mice and C57BL/6J controls were treated with romidepsin (FK228), a chemotherapeutic class 1 HDAC inhibitor approved for T‐cell lymphoma, at a low dose (0.56 mg/kg twice a week) for eight weeks. Treatment with romidepsin did not affect heart mass in either genotype. However, in db/db mice romidepsin treatment increased body mass and reduced blood glucose to a level not significantly different from controls. The in vivo blood glucose reduction occurred independent of plasma insulin levels, suggesting that romidepsin is not an incretin; therefore, we investigated the role of class 1 HDACs in intracellular insulin signaling. Results showed that loss of HDAC2 by in vitro siRNA knockdown in AC16 human ventricular cardiomyocytes significantly amplified AKT phosphorylation in response to IGF‐1 treatment compared to IGF‐1 treatment alone. Cumulatively these results show that class 1 HDAC inhibition has anti‐diabetic effects in vivo and suggests that these effects are due to potentiation of intracellular insulin signalling.Support or Funding InformationS.M. is supported by the Washington State University College of Pharmacy. E.J. is supported by the National Science Foundation Graduate Research Fellowship Program.
Augmented Expression of Piezo1 in the Uterine Vasculature During Pregnancy
Authors: Ko Nga Ling , Heal Maci , John Liam , Villalba Nuria , Henrion Daniel , Mandala Maurizio , Osol George J 
Published: November 2023

The maternal uterine circulation undergoes expansive remodeling during gestation to facilitate the many‐fold increase in uteroplacental blood flow required for normal pregnancy outcome. Although we and others have shown that endothelial nitric oxide (NO) is a key mediator of this process, the primary molecular sensor responsible for detecting shear stress and transducing it to eNOS activation has yet to be identified. The recent discovery of Piezo1 ‐ a large, mechanically‐activated cation channel that has been implicated in mechanosensing of shear stress in various cell types, including the endothelium, inspired this study. Notably, transfection of Piezo1 into HEK293 cells, which normally cannot sense shear stress, conferred a shear stress responsiveness evidenced by increased cytosolic calcium and eNOS activation. We hypothesized that Piezo1 may be present in uterine arteries, and that its expression would be augmented during pregnancy and linked to increased eNOS.Main uterine arteries (MUA) were collected from adult (12–14 week old) late pregnant (LP: gestational day 20, n=4) Sprague‐Dawley rats, and from age‐matched nonpregnant (NP, n=4) controls. Following euthanization, MUAs were dissected free of surrounding connective and adipose tissues, and cut open to remove any blood present in the lumen. Segments of MUA were either fixed and permeabilized for immunohistochemistry (IHC), or homogenized for Western Blotting (WB). Tissues were incubated with rabbit polyclonal Piezo1 antibody (Ab), eNOS Ab, β‐tubulin Ab; goat anti‐rabbit IgG‐HRP Ab (and/or Alexa Fluor® 488 Ab). Immunolabeled vessels were imaged using a Zeiss LSM 510 META confocal microscope @ 100x; for WB, proteins were detected with chemiluminescent substrate and analyzed using ImageJ software.Results(1) Piezo1 was detected in the MUA of both NP and LP rats (using either method), and localized to the arterial endothelium (IHC). (2) Both Piezo1 and eNOS expression were augmented in MUAs from LP vs. NP rats. Based on this preliminary information, we suggest that Piezo1 may play a role in transducing shear stress into the activation of eNOS in the uterine circulation during pregnancy, and thereby contributes to the expansive arterial remodeling required to maintain sufficient uteroplacental perfusion. This is the first study to identify this channel in uterine vessels, and to show a gestational effect on its expression. As both shear stress and uterine arterial remodeling are attenuated in preeclampsia (a serious gestational disease that afflicts ten million pregnant women worldwide each year), understanding the underlying mechanosensing/transducing mechanisms may lead to new therapeutic strategies for augmenting uteroplacental blood flow.Support or Funding InformationNIH R21 HD080156
Endothelial dysfunction and outward remodeling in coronary and mesenteric arteries in response to high fat diet in mice
Authors: Dunn Shannon M. , Hilgers Rob H. P. , Das Kumuda C. 
Published: November 2023

It is well established that vascular endothelial dysfunction is an underlying mechanism in the pathophysiology of hypertension, atherosclerosis, diabetes and other cardiovascular diseases (CVD). Additionally, this cardiovascular pathophysiology is accentuated in aged population. We hypothesize that intake of high fat diet would exacerbate age‐related vascular endothelial dysfunction, promoting the onset and propagation of CVD. To test this hypothesis twelve‐week‐old male mice with C57BL6/J background were randomly divided into four groups and fed normal chow (NC) or HFS (42% fat) for a period of 3 or 12 months. We isolated the left coronary artery (LCA) and 2nd‐order mesenteric artery (MA) and mounted them in the wire‐ and pressure myograph. Both young and aged HFS mice had impaired glucose tolerance as well as an increase in cholesterol levels and weight gain compared to both groups of mice on NC. Aged HFS did have a significant increase in body weight (BW) when compared to young HFS. There was no difference in BW between young and aged mice on NC. Endothelium‐dependent ACh‐induced relaxing responses were blunted in MA and LCA from young HFS compared to young NC mice. Strikingly, this endothelial dysfunction was similar compared to aged NC mice. In addition, endothelial dysfunction in LCA and MA was worsened in aged HFS mice. MA from aged HFS mice developed outward hypertrophic remodeling compared to their younger counterparts and aged‐matched NC mice. These findings suggest that high fat diet in aged individuals can accelerate endothelial dysfunction and increase the risk of CVD.Support or Funding InformationThe work presented here is supported by NIH grant R01HL107885 and R01HL107885‐05S1
Lipopolysaccharide‐Induced Inflammation does not Alter Spinal Cord Excitability in Female or Male mice
Authors: Putnam Shea , Criddle Connor , Lopez Alejandro , Wilkinson Katherine A. 
Published: November 2023

Peripheral inflammation can alter the excitability of dorsal horn interneurons and increase flexor reflex strength (Dubner & Ruda, 1992), but the effect of peripheral inflammation on the stretch reflex is not as well understood. Our lab has preliminary evidence that 18 hrs of systemic inflammation induced with lipopolysaccharide (LPS; a bacterial coat protein) causes a blunted response to muscle vibration in mouse muscle spindle afferents, the sensory neurons in the stretch reflex. This would suggest that muscle stretch reflex strength following LPS would be decreased if there were no changes in spinal cord excitability. To test the hypothesis that LPS induced systemic inflammation alters spinal cord excitability, we measured the H reflex, or electrical analog of the stretch reflex, in mice receiving an injection of LPS (0.5 mg/kg) or saline (200 μl) 18 hrs before experimentation. Mice were anesthetized with an intraperitoneal injection of ketamine (100mg/kg) and xylazine (10mg/kg). The sciatic nerve was exposed and stimulated with bipolar tungsten electrodes at current strengths from H wave threshold (T) to 8T (20 × 0.1 ms pulses at 0.1Hz). Bipolar recording electrodes were placed in the 4th dorsal interossei muscle and the results compared between adult male (n=9) and female (n=10) mice given LPS or saline (n=10 for both). We measured the maximum M wave amplitude (Mmax), H wave amplitude (Hmax), and latency of both waves. The percentage of motor neurons activated by electrical stimulation of the muscle spindle afferents (Group Ia) was measured by the ratio of Hmax/Mmax. Increased spinal cord excitability would be reflected in a larger Hmax/Mmax, regardless of any concurrent changes in muscle spindle afferent excitability. No difference in Hmax/Mmax was observed with LPS treatment in either sex (M control = 0.27 +/− 0.08; M LPS = 0.26 +/− 0.08; F control = 0.31 +/− 0.19; F LPS = 0.33 +/− 0.13). Similarly, no change in latency of the H wave was observed with LPS treatment in either sex (M control = 6.9 +/−1.1 ms; M LPS = 6.2 +/− 0.8 ms; F control = 6.1 +/− 1.2 ms; F LPS = 5.9 +/− 0.6 ms). Our results indicate that spinal cord excitability of the stretch reflex is unchanged following LPS treatment. Future studies will determine whether stretch reflex strength is decreased following LPS as would be expected since sensory input is decreased but spinal cord excitability is unchanged.Support or Funding InformationThis work was supported by a CSUPERB New Investigator grant (KAW).
Genome‐wide Assessment of the Pompe (Gaa−/−) Mouse Cervical Spinal Cord Confirms Widespread Neuropathology
Authors: Turner Sara , Hoyt Aaron , Falk Darin , Byrne Barry , Fuller David 
Published: November 2023

Pompe disease, caused by deficiency of acid alpha‐glucosidase (GAA), leads to glycogen accumulation, disruption of cellular architecture and neuromuscular impairments. Respiratory and lingual motor systems often decline first, and respiratory motoneurons typically show profound histopathology and glycogen accumulation at an early age. Although neuropathology is now firmly established in Pompe tissues from both humans and animal models, the associated mechanisms have not been evaluated. Therefore, we studied the cervical spinal cord of adult Pompe (Gaa−/−) and wild‐type (129SVE) mice using histological methods and mRNA analyses with the Affymetrix Mouse Gene Array 2.0ST. Pathway analyses of the mRNA data were done using the Broad Institute's Molecular Signature Database. In separate animals, cervical tissues were stained for terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and ionized calcium binding adaptor molecule 1 (IBA‐1) to assess DNA fragmentation and microglial morphology, respectively. From these analyses, several themes emerged (FDR q values <0.01). First, neuronal loss is an important feature of Pompe disease, and occurs in motoneurons before the onset of overt respiratory symptoms. Putative phrenic motoneurons were TUNEL‐positive, and pathway analyses showed significant upregulation of apoptotic processes in the Gaa−/− cervical spinal cord. Second, pro‐inflammatory mRNA signaling is significantly upregulated in the Gaa−/− cervical cord, and this is associated with greater neuronal co‐localization of microglia. Third, multiple signal transduction pathways are altered in the Gaa−/− cervical spinal cord. Glutamatergic signaling pathways were downregulated, as were well‐known “synaptic plasticity pathways” including genes related to long‐term potentiation. Finally, there was a robust change in cellular metabolism pathways in the Gaa−/− spinal cord that included upregulation of macromolecule, lipid, carbohydrate, and protein metabolic pathways. These data confirm that absence of functional GAA protein triggers a complex neurodegenerative process, and suggest that alterations in synaptic transmission and neuroplasticity processes may contribute to motor phenotypes in Pompe disease. Most importantly, the results underscore the importance of therapeutically targeting neuropathology in Pompe disease.Support or Funding InformationCTSA U54‐TR001012; Craig H. Neilsen Foundation 313369 (SMFT); 2R01HD052682‐06A1 (DDF, BJB)
Single Leg Plantar Flexion Elicits Greater Calf Muscle Oxygen Extraction in Peripheral Arterial Disease
Authors: Luck J. Carter , Muller Matthew D. , Ross Amanda J. , Leuenberger Urs A. , Sinoway Lawrence I 
Published: November 2023

Peripheral arterial disease (PAD) is characterized by atherosclerosis in the large arteries of the lower extremity resulting in impaired oxygen delivery to the gastrocnemius muscle and exercise intolerance. However, large artery hemodynamics do not fully explain symptomology or the responses to therapy in PAD. Recent studies suggest that microvascular and/or mitochondrial dysfunction also contribute to the development of leg pain and poor walking performance. Near‐infrared spectroscopy (NIRS) is increasingly being used to noninvasively measure the oxygen hemoglobin saturation (SmO2) changes that occur within the skeletal muscle during exercise. A reduction in SmO2 during exercise is indicative of greater oxygen extraction. In this study we tested the hypothesis that single leg dynamic plantar flexion elicits enhanced oxygen extraction in PAD vs. healthy subjects. Eight patients with PAD (ankle‐brachial index: 0.64 ± 0.03) and eight age and sex‐matched healthy subjects underwent single leg dynamic plantar flexion exercise in the supine posture. Subjects exercised until fatigue or a maximum of 14 minutes at 20 contractions per minute (0.5 kg to 7.0 kg) with the most symptomatic leg. The primary outcome variable was the amount of oxygen extracted from the medial gastrocnemius during exercise. Heart rate (HR) and mean arterial pressure (MAP) were also quantified on a beat‐by‐beat basis. Changes from baseline to the last 20 seconds of exercise were calculated. Because of symptomatic leg pain, PAD patients terminated exercise at lower workloads. Gastrocnemius muscle oxygen extraction at the end of exercise was significantly greater in PAD patients (78% ± 11%; M ± SEM) than in healthy subjects (23% ± 5%, P < 0.001). Similarly, the minimum SmO2 at the end of exercise was lower in PAD compared to healthy subjects (15% ± 6% versus 60% ± 6%; p < 0.001). HR increased similarly in PAD compared to healthy subjects (15 ± 2 bpm versus 11 ± 2 bpm) whereas the increase in MAP was augmented in PAD (19 ± 2 versus 12 ± 3 mmHg; P = 0.03). The recovery phase SmO2 was blunted in PAD and full recovery was not attained within three minutes post exercise. In conclusion, a threefold increase in oxygen extraction (vs. healthy subjects) is observed in the medial gastrocnemius of PAD patients in response to plantar flexion exercise.Support or Funding InformationExperiments supported by P01 HL096570 (Dr. Sinoway), UL1 TR00127 (Dr. Sinoway), KL2 TR00126 (Dr. Muller), R25 HL115473 (J. Carter Luck). The authors appreciate the technical support of their CRC research collaborators.
The effect of fan use and folic acid supplementation on thermal and cardiovascular strain in healthy aged humans during heat exposure
Authors: Gagnon Daniel , Romero Steven A , Ngo Hai , Poh Paula YS , Kouda Ken , Cramer Matthew N , Jay Ollie , Crandall Craig 
Published: November 2023

Attenuated heat loss and altered cardiovascular adjustments during heat stress are considered important contributing factors to the increased prevalence of death in aged individuals during heat waves. Interventions aimed at reducing thermal and cardiovascular strain during heat stress in the aged are lacking. We therefore examined if fan use (by improving sweat evaporation) or folic acid supplementation (by improving cutaneous vasodilation) can mitigate thermal and cardiovascular strain during heat exposure, as well as modify the limit of thermal compensability in healthy aged humans. Six healthy older individuals (69 ± 3 y, 166 ± 12 cm, 72 ± 16 kg, 2 males/4 females) rested in a thermal chamber at a temperature of 42°C and a relative humidity initially maintained at 30%. After a 30 min baseline period, relative humidity was increased in 2% steps every 5 min until a relative humidity of 70% was reached (100 min). The protocol was performed with no intervention (CON), with a fan that provided ~4 m/s of air velocity (FAN), and following 6 weeks of folic acid supplementation (FOLIC). Esophageal (Tes) and mean skin (Tsk) temperatures, local forearm cutaneous vascular conductance (laser‐Doppler, CVC), forearm vascular conductance (Doppler ultrasound, FVC), local sweat rate (ventilated capsule), heart rate and blood pressure were measured throughout. At 70% relative humidity, fan use and folic acid supplementation both increased CVC relative to control (CON: 1.34 ± 0.24 vs. FAN: 1.95 ± 0.39 vs. FOLIC: 1.81 ± 0.35 units/mmHg, P≤0.05). However, this did not translate into different FVC responses (CON: 4.86 ± 1.95 vs. FAN: 5.08 ± 1.91 vs. FOLIC: 4.41 ± 1.78 mL/min/mmHg, P>0.05). Fan use also increased local sweat rate (0.84 ± 0.27 mg/min/cm2) relative to CON (0.60 ± 0.26 mg/min/cm2, P≤0.05), whereas FOLIC had no effect (0.62 ± 0.21 mg/min/cm2, P=0.64). Relative to CON (Tes: 37.62 ± 0.29°C, Tsk: 38.41 ± 0.25°C), esophageal and mean skin temperatures were greater during FAN (Tes: 37.94 ± 0.63°C, Tsk: 38.97 ± 0.55°C, P≤0.05) but not FOLIC (Tes: 37.58 ± 0.21°C, Tsk: 38.28 ± 0.25°C, P>0.05). In contrast, heart rate and blood pressure were similar during all conditions (both P>0.05). The critical humidity at which an inflection in esophageal temperature was observed did not differ between conditions (CON: 63 ± 2 vs. FAN: 64 ± 5 vs. FOLIC: 63 ± 4%, P=0.64). Under the conditions employed, fan use results in greater esophageal and mean skin temperatures during extreme heat exposure in healthy aged individuals while folic acid supplementation has no effect. Furthermore, neither intervention reduced cardiovascular strain nor did they modify the limit of thermal compensability.Support or Funding InformationFunded by the National Institutes of Health (GM‐068865).
Aging of the antioxidant/inflammatory axis in human lung epithelial cells in vitro mimics aging in animal studies
Authors: Zhou Lulu , Forman Henry Jay , Zhang Hongqiao 
Published: November 2023

The balance of cellular antioxidant/inflammation shifts with aging, which results from increased pro‐inflammatory cytokines and inhibition of the Nrf2 pathway. Human small airway epithelial (SAE) cells were stimulated with lipopolysaccharide at 5 μg/ml and sulforaphane at 2 μM for 12 hours in vitro. The mRNA of cell inflammatory (NF‐κB‐dependent) and Nrf2 mediated production of antioxidant (phase II) genes were analyzed at early and late passage level of SAE. It was discovered that with the treatment of sulforaphane, phase II gene expression decreases as cell passage increases. GCLC and GCLM genes were down regulated, which may result from decreased capacity of cells to respond to electrophiles. However, upon treatment with lipopolysaccharide, NF‐kB dependent genes IL‐1 and TNF‐a were up regulated in company with inhibited phase II gene expression. The up‐regulation of pro‐inflammatory gene expression together with inhibition of Nrf2 mediated genes suggests that aging human epithelial cells gradually lose the ability to induce antioxidant defense, permitting a greater NF‐kB‐regulated increase in inflammatory cytokines. This mimics what has been observed in several previously reported studies of animal organs.Support or Funding InformationSupported by NIH grant ES023864