bluuebunny/crossref_metadata_2025_split huggingface dataset:
Impact of PCSK9 on endothelial efferocytosis in vascular aging
Authors: Ding Zufeng , Liu Shijie 
Published: May 2023

Background and Aims: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a serine protease that binds to low-density lipoprotein receptors. Efferocytosis is the process by which phagocytes remove apoptotic cells. Both PCSK9 and efferocytosis play important roles in regulating redox biology and inflammation, the key factors contributing to vascular aging. This study was designed to investigate the impact of PCSK9 on efferocytosis in endothelial cells (ECs) and its implications in vascular aging. Methods and Results: Studies were performed in primary human aortic ECs (HAECs) and primary mouse aortic ECs (MAECs) isolated from male wild-type (WT) and PCSK9-/- mice, and in young and aged mice treated with saline or the PCSK9 inhibitor Pep2-8. Our findings include that recombinant PCSK9 protein treatment induces defective efferocytosis and aging marker senescence-associated-β-galactosidase (SA-β-gal) expression in ECs, while PCSK9-/- restores efferocytosis and inhibits SA-β-gal activity. Further studies in aged mice showed that endothelial deficiency of MerTK, a critical receptor for efferocytosis that allows phagocytes to detect the presence of apoptotic cells, may be an indicator for vascular dysfunction in the aortic arch. Pep2-8 treatment markedly restored efferocytosis in endothelium from the aged mice. A proteomics study in the aortic arch from aged mice showed that PCSK9-/- significantly downregulates expression of NOX4, MAPK subunits, NF-κB, and secretion of pro-inflammatory cytokines, all known to promote vascular aging. Conclusions: These findings provide first evidence for the ability of aortic ECs to accomplish efferocytosis, and a role of PCSK9 in attenuating EC efferocytosis, leading to vascular dysfunction and acceleration in vascular aging. This work was supported by the National Institutes of Health, including National Institute on Aging (R21AG075548) and National Heart, Lung, and Blood Institute (R01HL162958). The work was also partially supported by the UAMS Bronson Endowment Foundation Award, VCRI Equipment Grant and Shared Resources Voucher Award. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Spinal NMDA receptor activation is necessary to initiate but not maintain ampakine-hypoxia induced phrenic motor facilitation
Authors: Thakre Prajwal , Perim Raphael , Mitchell Gordon , Fuller David 
Published: May 2023

Ampakines are a class of compounds which act as positive allosteric modulators of AMPA receptors. Intravenous (i.v.) delivery of a “low impact” ampakine prior to a single brief hypoxic episode produces a persistent (≥60 min) increase in phrenic nerve output (i.e., phrenic motor facilitation, pMF) (Thakre et al., J. Neurophysiol., 2021). Many forms of synaptic plasticity require an interplay of AMPA and NMDA receptors, including hippocampal long-term potentiation and acute intermittent hypoxia (AIH) induced long-term facilitation (LTF) in the phrenic motor system. Here, we tested the hypothesis that spinal NMDA receptor activation is necessary for pMF induced by a combination of ampakine and a single brief hypoxic episode (A+H). Phrenic nerve recordings were made from an experimental preparation that enables rigorous arterial blood gas control: urethane-anesthetized, ventilated, paralyzed and vagotomized adult male Sprague-Dawley rats. Ampakine CX717 (15 mg/kg, i.v.) was delivered 2-min prior to a 5-min hypoxic episode (partial pressure of arterial O2 ~40 mmHg). In experiment 1, the NMDA receptor antagonist MK-801 (10 μM) or vehicle (physiological saline) was intrathecally delivered at spinal segment C4 prior to A+H. Vehicle treated rats demonstrated greater phrenic burst amplitude at 60-min post-hypoxia (burst amplitude = 47±1 %BL) versus MK-801 treated rats (11±1 %BL; 2-way repeated measures analysis of variance, RM ANOVA, treatment effect, P=0.02, n=5 per group). In experiment 2, intrathecal MK-801 or vehicle (n=5 per group) were delivered 20-min after A+H. Neither MK-801 nor vehicle impacted pMF magnitude (2-way RM ANOVA, treatment effect, P=0.6). We conclude that the induction, but not maintenance, of A+H-induced pMF requires spinal NMDA receptor activation in contrast with AIH-induced phrenic long-term facilitation, which requires NMDA receptor activation for its maintenance (but not initiation). We reveal a novel mechanism of phrenic motor plasticity akin to hippocampal activity-induced synaptic plasticity, which requires NMDA receptor activation for initiation, but not maintenance. We suggest that ampakine pretreatment may have value in boosting the therapeutic impact of hypoxia-based neurorehabilitation strategies (Gonzalez-Rothi et al., J. Appl. Physiol., 2015). R01 HL139708 02 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Shorter hypoxic and inter-hypoxic intervals enhance acute intermittent hypoxia-induced phrenic long-term facilitation
Authors: Burrowes Kayla , Marciante Alexandria , Mitchell Gordon 
Published: May 2023

Phrenic long-term facilitation (pLTF) is a form of respiratory motor plasticity evoked by repeated exposure to low oxygen (acute intermittent hypoxia; AIH). AIH has emerged as a promising therapeutic approach to restore lost respiratory and non-respiratory function in people with spinal cord injury and other clinical disorders that compromise movement. Optimal AIH protocols ( e.g. episode severity/duration and number of hypoxic episodes) to elicit plasticity (and therapeutic benefit) have not yet been established. Three, 5-minute moderate hypoxic episodes with 5-minute intervals have been used most often to study pLTF. New data suggest that the same cumulative duration of hypoxia ( i.e. 15 min), when presented in shorter episodes, enhances pLTF by minimizing hypoxia-evoked adenosine release. Although the “standard” AIH protocol can be improved by reducing the duration of hypoxic episodes, little attention has yet been given to the duration of inter-hypoxic intervals. Thus, we began investigations concerning the impact of interval duration and episode number in moderate AIH protocols (mAIH; arterial PO2 = 40-55mmHg). pLTF was assessed in anesthetized, paralyzed, vagotomized and ventilated male Sprague-Dawley rats exposed to mAIH consisting of: 1) 3, 1 minute hypoxic episodes (3x1) with either 5-, 2- or 1-minute inter-hypoxic intervals; or 2) 2, 5-minute hypoxic episodes with a single 15-minute interval, compared with the conventional protocol (3, 5 minute hypoxic episodes, 5 minute intervals). mAIH consisting of 3x1, 5-min intervals elicit similar pLTF versus 3, 5 min episodes with 5-min intervals (64 ± 7% and 64% ± 1%, respectively; both n=3) at 60 minutes post-hypoxia. Thus, shorter intervals in an otherwise similar mAIH protocol consisting of 3x1 minute hypoxic episodes significantly enhance pLTF (1-minute: 146 ± 15%, n=5; 2 minute: 92 ± 17%, n = 3). Two, 5-minute hypoxic episodes with a single 15-minute inter-hypoxic interval still elicited pLTF of 51 ± 26% (n=4). These preliminary findings suggest both the hypoxic episode and interval duration are important determinants of mAIH-induced pLTF. Optimizing AIH-induced motor plasticity is necessary as we harness its therapeutic potential to treat severe neuromuscular disorders that compromise breathing. Supported by: NIH HL147554, HL148030, T32HL134621 (ABM) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Inhibition of Streptococcus pneumoniae colonization with deoxycholic acid, a secondary bile acid metabolite
Authors: Wier Meagan , McDevitt Erin , Vidal Ana , Alibayov Babek , Scasny Anna , McDaniel Larry , Vidal Jorge 
Published: May 2023

Streptococcus pneumoniae (pneumococcus) colonizes the upper respiratory tract, including the nasopharynx, of children and the elderly. Despite antibiotics and vaccines readily available, pneumococcus kills millions worldwide yearly. We have found that the secondary bile acid metabolite and endogenous salt deoxycholic acid (DoC) affects the viability of this human pathogen while also playing multiple roles in host physiology. Via in vitro studies, we assessed the anti-microbial activity of DoC and investigated its potential to eradicate S. pneumoniae colonization using a model of human nasopharyngeal colonization and an in vivo mouse model. At a physiological concentration, DoC (0.5 mg/ml; 1.27 mM) killed all tested strains of S. pneumoniae (n=48) 2 hours post-inoculation. Our model of nasopharyngeal colonization showed that DoC eradicated colonization by S. pneumoniae strains as quickly as 10 minutes post-exposure. The mechanism of action did not involve activating autolysis since autolysis-defective double mutant strains (Δ lytAΔ lytC and Δ spxBΔ lctO) were found to be as susceptible to DoC as the wild-type strain (WT). Oral streptococcal species (n=20) were, however, not as susceptible to DoC at 0.5 mg/ml. Unlike trimethoprim, whose spontaneous resistance frequency (srF) for TIGR4 or EF3030 was ≥ 1 x 10-9, no spontaneous resistance was observed with DoC. Finally, the efficacy of DoC to eradicate S. pneumoniae colonization was assessed in vivo using an intranasal topical route (i.n.) and separately as prophylactic treatment. Mice that were challenged with S. pneumoniae EF3030 carried a median of 4.05 x 105 CFU/ml four days post-inoculation compared to 6.67 x 104 CFU/ml for mice treated with DoC. Furthermore, mice in the prophylactic DoC group had approximately a 99% reduction of pneumococcal density illustrating the possibility of deoxycholic acid as a safe, endogenous human bile salt with therapeutic potential against increasingly-resistant S. pneumoniae strains. This study was in part supported by grants from the National Institutes of Health (NIH; 1R21AI144571-01 and 1R21AI151571-01A1 to J.E.V.). B.A. was supported by a Fulbright scholarship awarded by the U.S. Department of State. The content is solely the responsibility of the authors and does not necessarily represent the official view of the NIH or the U.S. Department of State. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
In vivo determination of direction of blood flow in the newly discovered SCN-OVLT vascular portal system in rat
Authors: Roy Ranjan , Yao Yifan , Silver Rae , Stern Javier 
Published: May 2023

By transporting products directly from the capillary bed of one region to the capillary bed of another region, vascular portal pathways enable minute amounts of important secretions to reach their specialized targets in high concentrations, without dilution in the systemic circulatory system. For decades there has been only one known portal system in the mammalian brain - that of the pituitary gland, first identified in 1933 (Popa and Fielding, J. Anatomy 1933). This year, we described a second portal pathway in the mouse linking the capillary vessels of the brain's clock suprachiasmatic nucleus (SCN) to those of the organum vasculosum of the lamina terminalis (OVLT), a circumventricular organ (Yao et al., Nat. Comm. 2021). A caveat in this initial work was that the direction of blood flow was unknown. To determine whether the SCN signaled the OVLT or vice-versa, we performed in vivo 2-photon imaging in anesthetized eGFP-vasopressin (VP) rats using a recently developed approach (Roy et al., Cell Report 2021) to study blood flow in this portal system. To delineate the SCN microvasculature in vivo, we intravenously infused fluorescent dextrans in anesthetized rats. The SCN-OVLT portal system was identified as Alexa 633 (an artery/arteriole specific dye)-negative vessels originating from a dense SCN capillary network that run rostrally towards the OVLT. These vessels displayed a mean diameter of ~20 μm. Blood flow in the portal vessels was measured by monitoring red blood cell (RBC) movement after intravenous injections with Rho70 kDa. Using kymographs, we found that in all cases, RBCs flowed rostrally, from the SCN towards the OVLT. Importantly, we found than blood flow was significantly higher at night (ZT17-19) compared to daylight (ZT5-7) (p< 0.001), while directionality remained the same (SCN→OVLT). Taken together, our results support the presence of a functional SCN-OVLT portal system in the rat in which blood flows unidirectionally from the SCN towards the OVLT. Moreover, our studies support the notion that blood flow in this system can be regulated. This clock portal system points to entirely new routes and targets for secreted signals from the SCN, restructuring our understanding of its output pathways. Support: NIH HLBI R01HL162575 to JES, AHA916907 to RKR and NSF 1749500 to RS. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
LRRC8A Anion Channel Knockout Stimulates the Nrf2 Antioxidant Response
Authors: Panja Sourav , Choi Hyehun , Nguyen Hong-Ngan , Lamb Fred 
Published: May 2023

Leucine Rich Repeat Containing 8A (LRRC8A) is an essential component of volume‐regulated anion channels (VRACs), which regulate cell volume. These anion channels are activated by pro-inflammatory mediators such as angiotensin II and tumor necrosis factor-α (TNFα). We previously demonstrated that VRAC inhibition or LRRC8A knockdown reduced Nox1 activation and impaired the inflammatory response to TNFα in cultured vascular smooth muscle cells. We hypothesized that loss of LRRC8A would also modify the antioxidant status of VSMCs. Vascular smooth muscle-specific (Sm22alpha-Cre) LRRC8A KO mice were created. Primary VSMC cultures were prepared from aortae and mesenteric arteries from male mice. Western blotting of protein from WT and LRRC8A KO mesenteric artery VSMCs revealed significant upregulation of proteins that are known to be under the control of the Nrf-2 antioxidant response; heme oxygenase 1 (2.1 fold), thioredoxin 1 (1.8 fold), and superoxide dismutase 1 (1.3 fold) superoxide dismutase 3 (1.8 fold). There was no change in the abundance of catalase.To further explore the role of LRRC8A in intracellular signaling, LRRC8A was immunoprecipitated from cultured WT and LRRC8A KO VSMCs and proteins were analyzed by Mass Spectrometry (MS). Reads were sorted for proteins identified in wild type but not in KO cells. This approach identified the Trim21 E3 ubiquitin ligase, a known regulator of the antioxidant response, as an LRRC8A-associated protein. Trim21 ubiquitinates and thereby promotes degradation of p62 (Sequestosome-1, SQSTM1). p62 sequesters Kelch-like ECH-associated protein 1 (Keap1), preventing it from promoting ubiquitination and destruction of Nrf-2 by the proteosome. Consistent with this, TRIM21 KO mice have an augmented antioxidant status and are protected from myocardial inflammation in response to doxorubicin or ligation of the left coronary artery. We considered the possibility that association of TRIM21 with LRRC8A (directly or indirectly) regulates its activity. Co-immunoprecipitation and western blotting confirmed association of LRRC8A with TRIM21. Pulldown of ubiquitinated proteins from WT and KO VSMCs failed to provide evidence that LRRC8A is ubiquitinated, however, p62 ubiquitination was markedly reduced in KO VSMCs. Thus, reduced p62 turnover in LRRC8A KO cells may enhance Keap1 sequestration and increase Nrf2 activity. The ability of LRRC8A knockdown to both impair the inflammatory response to TNFα and to enhance the antioxidant status of VSMCs identify this protein as an appealing target for the treatment of vascular inflammation. This work was supported by our both grant from NIDDK (1R01DK132948) and NIH (1 R01 HL160975-01A1). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Does cardio-hepatic communication contribute to pregnancy-induced cardiac growth and its reversal?
Authors: Schulman Emily , Collins Helen 
Published: May 2023

Introduction: During pregnancy, the maternal heart undergoes physiological cardiac growth, which reverses following parturition; however, the underlying processes that contribute are largely unknown. Changes in both systemic and cardiac metabolism appear to be associated with cardiac growth, which could also contribute during pregnancy. We have shown that several cardiac metabolic transcripts and proteins and liver-derived metabolites are elevated during pregnancy and postpartum and are associated with cardiac growth. Therefore, the goal of this study was to examine the extent to which the cardio-hepatic signaling axis contributes to pregnancy-induced cardiac growth and reversal. Methods and Results: Timed pregnancy studies in 12-week-old FVB/NJ mice revealed that both heart weight (HW:TL; p < 0.0001) and liver weight (LIV:TL; p <0.0001) were highest in lactating 1-week post-birth (PB) mice versus mid-pregnant (8 d), late-pregnant (LP; 16 d), and non-pregnant (NP) mice. Interestingly, we also found that HW:TL ( p < 0.01) and LIV:TL ( p < 0.0001) were significantly reduced in non-lactating 1-week post-birth mice (PBNL) compared with PB mice. Hearts and livers both remained significantly larger, irrespective of lactation status, following parturition through 6 weeks post birth (PB) versus NP. Plasma proteomics revealed significant increases in prolactin at LP in addition to increases in liver-derived proteins including hepatocyte growth factors; however, these levels returned to NP levels by PB. Examination of the liver proteome from the same mice revealed significant increases in fatty acid and ketone body metabolism proteins. Cardiac and liver immunoblot analysis revealed significant reductions in Pdk4 between 1- and 3-week PB and PBNL groups that was reversed by 6-weeks postpartum and a significant increase in Pdk4 expression at 6 weeks postpartum, respectively. Interestingly, we see a sustained increase in cardiac Bdh1 expression that persisted at 6-weeks postpartum which could contribute to the sustained increases in HW:TL versus NP. Conclusion: Changes in both lactation status and metabolism are likely contributing to similar patterns of growth seen in the heart and liver during and following pregnancy. NIGMS COBRE Pilot project grant P30 GM127607; NIH R01 HL147844 (Hill/ Jones); Jewish Heritage Fund for Excellence Faculty Support; Jewish Heritage Fund for Excellence Research Enhancement Award; UofL School of Medicine Basic Science award; NIGMS IDeA state funded proteomics voucher. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Folate malnutrition as metabolic stressor to promote advanced type 2 diabetes mellitus by switching bioenergetic fluxes and metabolic flexibility
Authors: Chen Wan-Jing , Huang Chih-Yang 
Published: May 2023

Type 2 diabetes mellitus (T2DM) is a group of metabolic disorder disease, including abnormal bioenergetic fluxes and glucose-related genetic regulation, eventually leads to severe diabetes complications. Clinical studies have shown that folate malnutrition positively correlated with the progression of T2DM, but the causal relationship and mechanism are still unclear. Aims of the study were to investigate whether the molecular mechanisms by which metabolic folate stress may promote diabetic memory metabolism switched bioenergetic fluxes and metabolic flexibility to mediate advanced T2DM. Primary Human Skeletal Muscle Cells (HSkMC) were cultured with high glucose for 4, 8, 12, and 16 days as the experiment models to test the hypothesis. As compared with the controls, cultivation of HSkMC cells with clinically low serum folate levels (LF) for 4-16 days gradually and significantly promoted (1) morphological alteration, (2) insulin resistance, (3) expressions of bioenergetic markers (NADP/NADPH, Lactate), and (4) mRNA expressions of late on-set T2DM molecules (INS, INSR, SLC2A2, SLC30A8, ABCC8). Energetic metabolic switch in the LF- HSkMC cells, but not the controls, was evident by reduced lactate release, pyruvate dehydrogenase E1 alpha (PDHA) expression and altered NADP/NADPH, NAD+/NADH ratios from 3.4 to1.8 and 2.0 to 1.2, respectively. Blockage of mTOR pathways, epigenetic acetylation (Sirt1) abolished LF-promoted metabolic disorder, regulated mRNA expressions of glucose and insulin related molecules (INSR, SLC2A2 and ABCC8). Collectively, metabolic folate stress promotes metabolic memory switched bioenergetic fluxes and metabolic flexibility to mediate advanced type 2 diabetes. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.