bluuebunny/crossref_metadata_2025_split huggingface dataset:
The effect of diet on the metabolic rate and microbiome of Gromphadorhina portentosa
Authors: Contreras Heidy L. , Lorenz Todd 
Published: November 2023

The metabolic phenotype of an animal is shaped by the developmental trajectory of its gut microbiome. Although the core microbiota of an organism is stable throughout adulthood, there are certain parts of the microbiome that can be altered through an array of environmental stressors, such as diet. So far, there is a lot of research focused on the effects that diet has on the metabolism and gut microbiome of vertebrate mammals. However, not a lot of work is focused on how a change in diet can affect the metabolic rates, or the microbiome, of invertebrates. In its natural habitat Gromphadorhina portentosa, the hissing cockroach, feeds opportunistically on fruits and may also consume decaying animal material and small insects. In this study we measured the effects of diet on the metabolic rate, and microbiota, of G. portentosa. Prior to data collection we normalize the microbiota of forty male cockroaches by placing them on a control diet. Preliminary data showed that roaches would move all gut contents through their digestive system within 7 days. Resting metabolic rates for normalized cockroaches was measured after cockroaches had been placed for 2 weeks under the control diet. Fecal samples were also collected at this time. Ten male roaches were placed under one of four diet treatments (control, high protein, high carbohydrate or high lipid) for a total of 8 weeks. During this experimental period, resting metabolic rates were obtained on a weekly basis using flow through respirometry. Fecal samples were once again collected at the end of the 8‐week experimental period. Cultivation independent metagenomic sequencing was accomplished so that changes in the microbiota of roaches could be identified. Previous work has shown a shift in the microbiota of organisms fed varying amounts of fat vs. protein, were animals fed a high protein diet showed an increase in bacteroidetes with was an inverse relationship to the firmicute population. Furthermore, an increase in metabolic rates have also been seen when animals are given a high protein diet. We expect G. portentosma to respond in a similar manner when their diet is altered.Support or Funding InformationULV Faculty Research Grant
α1G T‐type Ca2+ channel plays a dual role in control of pulmonary microvascular endothelial cell P‐selectin surface expression
Authors: Zheng Zhen , Hu Xingang , Chen Hairu , Sellak Hassan , Alexeyev Mikhail F , Wu Songwei 
Published: November 2023

Signaling pathways directing alveolar endothelial cell activation are of pivotal importance in controlling lung inflammation as lung alveolar capillaries constitute the major site for neutrophil trafficking. We had previously established that the expression of an α1G T‐type Ca2+ channel, the only voltage‐gated Ca2+ channel identified in lung endothelium, is restricted to the alveolar capillary network. Notably, the α1G‐mediated Ca2+ entry is pro‐inflammatory as it specifically leads to P‐selectin surface expression, which enables intra‐capillary neutrophil sequestration. More recently we observed that, in pulmonary microvascular endothelial cells (PMVECs), the α1G directly interacts with endothelial‐type nitric oxide (NO) synthase (NOS3) and blockade of the α1G abolishes agonist‐stimulated NO generation. These observations suggest that the NOS3/NO signaling cascade, known to inhibit endothelial exocytosis, is also activated downstream of α1G activation. The present study tested whether the α1G‐mediated Ca2+ entry activates NO generation which in turns leads to inhibition of α1G‐dependent P‐selectin surface expression. In PMVECs over‐expressing rat α1G and NOS3 loaded with an NO‐sensitive fluorophore, DAF‐FM, real‐time fluorescence imaging was utilized to assess the NO production in response to high external [K+] (40 mmol/L) stimulation that is to depolarize the endothelial cell membrane potential from the resting level to a voltage at which the T‐type “window current,” i.e., a constant α1G‐mediated Ca2+ entry, occurs. Depolarization of plasma membrane via high [K+] elicited a sustained NO production that was nearly abolished by pharmacologic NOS blockade as well as T‐type channel blockade with L‐NAME (1 mmol/L) and mibefradil (5 μmol/L), respectively. Next, in PMVECs stably transduced with a GFP tagged P‐selectin, fluorescence microscopy was employed to examine the real‐time dynamics of the P‐selectin in response to plasma membrane depolarization. Indeed, depolarization provoked rigorous P‐selectin translocation from cytosol to the plasma membrane leading to P‐selectin surface expression. While such depolarization‐stimulated P‐selectin surface expression was nearly abolished by T‐type channel blockade (mibefradil, 5 μmol/L), it was significantly potentiated by NOS blockade (L‐NAME, 1 mmol/L). Taken together, these findings revealed that the NO signaling cascade is a specific physiological target of the Ca2+ signal initiated by α1G activation and that the α1G‐dependent NO generation subsequently limits the α1G‐mediated P‐selectin surface expression, as in a negative feedback mechanism. In this regard, α1G serves a dual role in control of endothelial cell pro‐inflammatory phenotype.Support or Funding InformationHL‐066299
Activation of Adenylyl Cyclase in Pulmonary Microvascular Endothelial Cells Increases Microparticle cAMP Content
Authors: Sayner Sarah L , Hargett Leslie , Scruggs April , Ramila KC , Yarbrough Thomas , Bauer Natalie 
Published: November 2023

RationaleMicroparticles (MPs) are submicron intact vesicles released from many cell types constitutively or following activation. MPs play a role in cellular homeostasis, injury and act as biomarkers. These submicron vesicles can encapsulate a variety of functional proteins and RNAs/miRNAs; however, little is known about whether they encapsulate signaling molecules, such as cyclic adenosine monophosphate (cAMP). Thus, we investigated the hypothesis that MPs from endothelial cells contain cAMP and that pulmonary microvascular endothelial cells (PMVEC) can be stimulated to increase MP‐cAMP content.MethodsMPs were collected from the media of unstimulated pulmonary and aortic endothelial cells as well as pulmonary artery smooth muscle cells (PASMCs) by ultracentrifugation. MPs were also collected from (PMVECs) following treatment with agonists to stimulate endogenous cAMP production from the transmembrane adenylyl cyclase (AC) or bicarbonate‐stimulation of the soluble AC (AC10) in the presence or absence of the phosphodiesterase (PDE) 4 inhibitor, rolipram. Cyclic AMP in the MPs as well as cellular lysates was measured by immunoassay or mass spectrometry and normalized to protein concentration.ResultsWhile cAMP was detected in MPs derived from various endothelial cell types as well as PASMCs, the cAMP content was highest in PMVECs compared to the other cell types. Further, stimulation of PMVECs with agents that increase the near membrane cAMP pool led to an increase in cAMP content in MPs.ConclusionMicroparticles released from the vasculature contain cAMP while stimulation of transmembrane ACs in PMVECs increases MP‐cAMP content.Support or Funding InformationThis work was supported by the Parker B. Francis Fellowship Foundation Award (SLS) and HL121513 (SLS), AHA 11SDG7390037 (NB) and T32 HL076125 (LH, AS, TY)
Lipopolysaccharide and Isoproterenol Alter Sodium Bicarbonate Cotransporter Phosphorylation in Pulmonary Microvascular Endothelial Cells
Authors: Sayner Sarah L. 
Published: November 2023

The hallmark of acute respiratory distress syndrome (ARDS) is damage to alveolar capillaries leading to increased endothelial permeability. Bacterial induced sepsis is a primary cause of pulmonary endothelial barrier dysfunction. Our recent studies reveal that the lipopolysaccharide (LPS)‐induced increase in pulmonary endothelial permeability is dependent upon bicarbonate stimulation of soluble adenylyl cyclase isoform 10 (AC10 a.k.a. sAC). Further, bicarbonate stimulation of AC10 is dependent upon sodium, suggesting a role for Na+‐bicarbonate cotransporters (NBCs). Further, it has been reported that forskolin stimulation of transmembrane ACs and generation of subplasma membrane cAMP inactivates NBCn2 via PKA, potentially reducing the availability of bicarbonate for stimulation of AC10. Thus, we investigated the expression of NBCs in the lung and pulmonary endothelium and the phosphorylation status of NBCn2 in response to barrier disruptive (LPS) or barrier protective (isoproterenol) agents. To examine the phosphorylation status of NBCn2, endothelial cells were treated with either LPS or isoproterenol and rolipram, the phosphodiesterase 4 inhibitor. Immunoprecipitation was performed on whole cell lysate with the NBCn2 or control IgG antibodies and western blot analysis with PKA Phospho‐Ser/Thr antibody. Our results revealed the differential expression of NBCs in the pulmonary circulation. Further, while LPS led to a decrease in the phosphorylation of NBCn2 at a PKA specific site, isoproterenol and rolipram increased PKA‐phosphorylation of NBCn2. Thus, while transmembrane AC activity increases NBCn2 phosphorylation, LPS decreases NBCn2 phosphorylation and could act as a mechanism to regulate the bicarbonate source necessary to stimulate AC10.Support or Funding InformationThis work is supported by the Parker B. Francis Fellowship Foundation Award (SLS) and HL121513 (SLS)
Characterization of Ectonucleotidase Expression in the Rat Carotid Body: Potential Regulation by Hypoxia?
Authors: Salman Shaima , Vollmer Cathy , Nurse Colin A 
Published: November 2023

The carotid body (CB) maintains homeostasis in the respiratory and cardiovascular systems of mammals by initiating reflex responses during exposures to blood‐borne stimuli such as low O2 (hypoxia). Thus, stimulation of CB chemoreceptors during hypoxia results in compensatory hyperventilation and activation of the sympathoadrenal system. CB hyperactivity has been linked to several clinical morbidities including sleep apnea, chronic obstructive pulmonary disease, hypertension, obesity, and heart failure. The purine ATP and its derivative, adenosine, contribute significantly to the excitatory CB afferent discharge during hypoxia (Conde et al., 2012, J Appl Physiol 112: 2002–10; Nurse and Piskuric, 2013, Semin Cell Dev Biol 24: 22–30). Given that extracellular nucleotides and products of their hydrolysis by ectonucleotidases are known to play a key role in purinergic signaling in the brain and tissues innervated by the autonomic nervous system, we investigated ectonucleotidase expression in the rat CB. Ectonucleoside triphosphate diphosphohydrolase (E‐NTPDase) is the dominant family of ectonucleotidases and subtypes of this enzyme family catalyze the hydrolysis of purines and regulate their levels in the synaptic cleft. In extracts of whole rat CBs, RT‐PCR and sequence analyses revealed the presence of E‐NTPDase‐1 and ‐2, as well as ecto‐5′‐ectonucleotidase (Nt5e) mRNA. Interestingly, expression of ectonucleotidases, E‐NTPDase‐1 (CD39) and Nt5e (CD73), has been proposed to have a protective role in mouse intestinal epithelia during hypoxia (Synnestvedt et al., 2002, J Clin Invest 110: 993–02). This raised the possibility that ectonucleotidases might be regulated by hypoxia in O2‐sensitive organs such as the CB and its neural crest‐derived counterpart, the adrenal medulla. Indeed, microarray analysis of a rat O2‐sensitive immortalized adrenomedullary chromaffin cell line (MAH cells) suggests that chronic hypoxia exerts a selective hypoxia inducible factor‐2 alpha (HIF2a)‐dependent upregulation of E‐NTPDase‐2 transcript. Future studies will determine the cellular localization of ectonucleotidases in the rat CB and the potential role of HIF2a in the regulation of E‐NTPDase‐2 expression during chronic hypoxia.Support or Funding InformationSupported by grants from Canadian Institutes of Health Research and Natural Sciences and Engineering Research Council of Canada
Early Postnatal Exposure to Intermittent Hypoxia Results in Significant Alterations in White Matter Integrity in a Rat Pup Model of Apnea of Prematurity
Authors: Darnall Robert , Chen Xi , Sirieix Chrystelle , Xia Luxi , Gimi Barjor 
Published: November 2023

Exposure to severe hypoxemia in the human newborn infant results in white matter (WM) injury. However, premature infants are exposed to multiple episodes of less severe intermittent hypoxia (IH) associated with apnea of prematurity (AOP) and immature respiratory control. Often these events are not detected clinically and it is unknown whether IH during this period is associated with WM injury. To answer this question we exposed newborn rat pups to IH from postnatal day (P) 2 to P12 (6 decreases to 5–8% oxygen per hour alternating with an hour of normoxia, 24 hours a day, 7 days a week). A control group was treated identically, except that the periods of hypoxia were replaced with room air. This period is roughly equivalent to a preterm infant from 28 to 40 weeks postmenstrual age. At P20 we performed Diffusion Tensor Imaging (DTI) under isofluorane anesthesia in 11 IH exposed and 9 control pups using a 9.4 T small animal magnetic resonance imaging system, using 6 gradient directions with b=1000 with a slice thickness of 1 mm and an in‐plane resolution of ~160 × 160 μm. Compared to controls, exposure to IH resulted in a decrease in fractional anisotropy (FA) and radial diffusion (RD) and an increase in axial diffusion (AD) in several major WM regions including the corpus callosum (P=0.004) and cingulate (P<0.001) consistent with the loss of white matter integrity. We conclude that in the rodent repeated exposures to even mild hypoxia in the early postnatal period results in white matter injury. Future studies should be done to determine whether similar MRI changes occur in premature infants with apnea of prematurity and if so, whether IH associated with AOP can be treated more effectively.Support or Funding InformationNIH PO1 HD36379, American SIDS Institute
Chronic Hypoxia Downregulates Colocalization of PKG with its Target BK Channel Proteins in Ovine Cerebral Arteries
Authors: Thorpe Richard Burdette , Williams James M , Pearce William J 
Published: November 2023

BACKGROUNDPrevious studies have shown that chronic hypoxia ablates the endothelium‐dependent NO‐mediated vasorelaxation pathway of which protein kinase G (PKG) is a key effector. Direct activation of PKG likewise demonstrates attenuated PKG influence on 5HT‐induced vascular tone in chronic hypoxic cerebral arteries compared with normoxic arteries. Furthermore, these effects have been shown to correlate with reduced BK channel activity (potassium currents). Taken together, this suggests that the direct influence of PKG on BK channel function is diminished in conditions of chronic hypoxia.OBJECTIVEGiven that PKG phosphorylates multiple known targets, this present study explores the hypothesis that the proximal association of PKG with its target BK‐channel proteins is altered by chronic hypoxia, and that this diminished colocalization accounts for a significant loss of function with regards to cerebral vasorelaxation.METHODSPregnant and Non‐Pregnant ewes were held at sea level (normoxic) or at 3820m (hypoxic) for the final 110 days of gestation. Middle cerebral arteries from non‐pregnant adults and term fetal sheep were prepared for organ culture with ±8‐pCPT‐cGMP. Colocalization of PKG with the alpha and beta isoforms of the BK channel proteins, as well as colocalization of BK‐alpha and ‐beta proteins with each other, was determined by confocal microscopy.RESULTSChronic hypoxia reduced the ability of PKG to localize with its target BK channel proteins. Chronic hypoxia also reduced the colocalization of BK‐alpha with BK‐beta channel proteins. Activation of PKG with 8‐p‐CPT‐cGMP in organ culture did not overcome these effects of chronic hypoxia on PKG colocalization.CONCLUSIONSPKG‐mediated activation of BK channels accounts for a significant portion of PKG‐induced vasorelaxation in ovine middle cerebral arteries. Chronic hypoxia attenuates the ability of PKG to associate with BK channel proteins, and thereby attenuates the influence of PKG on BK channel activity. This loss of association explains in part how chronic hypoxia ablates the ability of PKG to relax 5HT‐induced vascular tone.Support or Funding InformationThe work reported here was supported by USPHS Grants HL54120, HD31266, HL64867 and the Loma Linda University School of Medicine
Activation of the hypoxia‐inducible factor pathway induced by PHD2 deficiency enhances the effect of running training in mice
Authors: Nunomiya Aki 
Published: November 2023

BackgroundRecently, altitude training has widely been accepted by elite athletes as an efficient way to improve the sea‐level performance. The adaptation to the deprivation of O2, which is called hypoxic response and mediated by hypoxia‐inducible factors (HIFs), seems to contribute to the benefit of altitude training. However, the veracity of this benefit is still controversial, and the effect of biological processes mediated by the HIF pathway on the improvement in exercise performance after exercise training, if any, needs to be demonstrated. The main regulator of HIFs activity is prolyl hydroxylase domain protein 2 (PHD2). Previous reports showed that Phd2 deficiency induced HIFs activation and subsequent hematopoietic and angiogenic adaptations (Minamishima et al., 2008, Takeda et al., 2007). Overall, we believe that the Phd2‐deficient mouse model enables us to elucidate the influence of the activated HIF pathway on the exercise training effect. We therefore hypothesised that the stabilization of HIFs and the subsequent hypoxic response induced by Phd2 deficiency contribute to the improvement in exercise training. To examine this hypothesis, we investigated the maximal exercise capacity of Phd2‐deficient mice before and after a 4‐week regimen of exercise training.MethodsBecause Phd2 knockout is embryonic lethal, estrogen receptor (ER) agonist‐induced Phd2 cKO mouse using Cre‐loxP system was used. Two weeks after tamoxifen, an estrogen‐agonist, maximal running test using a rodent treadmill was performed. Control mice (n = 16) and Phd2 cKO mice (n = 15) were then divided into two groups: trained and non‐trained. The mice in the training group underwent 4‐week treadmill training on a rodent treadmill. After 4‐week period, all mice of both groups were tested again for their maximal running capacity and were sacrificed for blood and tissue examination.ResultsBecause of Phd2 deficiency, HIF‐α proteins accumulated. In line with increases in plasma erythropoietin levels (P < 0.01), the Phd2 cKO mice exhibited marked increases in haematocrit values (P < 0.01) and haemoglobin concentrations (P < 0.01). Increased expression of plasma vascular endothelial growth factor (P < 0.05) induced angiogenesis in the skeletal muscle of the Phd2 cKO mice (P < 0.05). Importantly, trained Phd2 cKO mice showed a significantly higher gain in running time by a factor of approximately 1.53 compared with the trained control mice (P < 0.05, Figure). On the other hand, we did not find any difference between the non‐trained control and non‐trained Phd2 cKO mice (P = 0.21). Collectively, these data suggest that the combination of training and the activation of the HIF pathway is important for maximizing the effect of running training.ConclusionsWe concluded that the activation of the HIF pathway induced by PHD2 deficiency enhanced the effect of running training.Support or Funding InformationThis work was supported by a JSPS Grant‐in‐Aid for JSPS fellows (26‐3209).
Neonatal maternal separation increases microglial activation in brainstem regions controlling the laryngeal chemoreflex
Authors: Baldy Cécile , Boisjoly‐Villeneuve Samuel , Fournier Stéphanie , Tremblay Marie‐Ève , Kinkead Richard 
Published: November 2023

Laryngeal chemoreflex (LCR) is immature in preterm infants and results in prolonged apneas, O2 desaturations and bradycardias. Previous work from our laboratory has shown that in newborn rat, neonatal maternal separation (NMS) interferes with LCR development and aggravates cardio‐respiratory complications. This effect was more important in males. Microglia are the immune cells of the central nervous system; during early life, they contribute to brain development by playing an important role in synaptic pruning. These cells are activated during stress and their activation is characterized by morphological changes. Indeed, activated microglia are more round (ameboid shaped) with short and few processes like macrophages, whereas inactivated microglia have several ramified processes scanning for abnormal cells and contacting synapses. Because neonatal stress (in the form of NMS) affects LCR development, we hypothesised that NMS increases microglial activation in regions which are involved in the LCR, particularly the nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (NX). Rat pups were separated from their mother 3 hours each day during 10 days from post‐natal day 3 to 12 (P3 to P12). Control animals were left undisturbed. At post‐natal days 14–15, brains were harvested for Iba‐1 immunostaining. Microglia were counted in the NTS and NX by light microscopy. Cell morphology was assessed in each microglia by measuring morphological index (ratio between cell body and arborisation areas), cell density and scattering (nearest neighbour distance) in NTS and NX. Stress increase significantly morphological index in both region of interest and in males and females. NMS increases microglia density in males in NTS and NX. Stress increases also microglial scattering in the N10 but this effect is significant only on P15 males and females. We conclude that NMS elicits microglial activation in the NTS and NX in a sex‐dependent fashion. Based on these morphological changes we propose that NMS may interfere with microglia's ability to perform synaptic pruning during early life. This, in turn may contribute to abnormal LCR function in pups.Support or Funding InformationThis research was supported by CIHR (RK).