bluuebunny/crossref_metadata_2025 huggingface dataset:
CD8+ T cells mediate kidney fibrosis in diet-induced obese mice
Authors: Erickson John , Edell Claudia , Walker Savannah , Colson Jackson , Pollock Jennifer
Published: May 2024

Diet-induced obesity (DIO) is an epidemic in the United States and globally. Obesity is well known to exacerbate and/or promote kidney disease. Inflammation is a major factor in DIO and kidney disease. Utilizing a 20-week DIO mouse model (45% fat) and a normal diet (ND; 10% fat), our lab previously showed greater kidney medullary interstitial fibrosis as well as increased kidney medullary CD3+ T cells in DIO mice when compared to ND mice. Specifically, we found increased kidney CD8+ T cells, but similar CD4+ T cells, in DIO mice compared to ND mice. We hypothesized that CD8+ T cells in DIO mice are a primary factor in promoting kidney interstitial fibrosis. DIO mice were treated with anti-CD8 antibody or control anti-IgG antibody and ND mice were treated with control anti-IgG antibody for the final two weeks of the DIO protocol (n=7-9). DIO/anti-IgG and DIO/anti-CD8 mice had significantly higher body weight than ND/anti-IgG mice as expected (one-way ANOVA; p<0.0001). Administration of anti-CD8 treatment in DIO mice had null effects on body weight (one-way ANOVA, p=0.509) and kidney weight (one-way ANOVA; p=0.067). Utilizing flow cytometry, we verified that anti-CD8 treatment significantly reduced CD8+ T cells compared to anti-IgG in the kidney (one-way ANOVA; p=0.0077; DIO/anti-IgG vs DIO/anti-CD8, p=0.0066) and in the blood (one-way ANOVA; p=0.0034; DIO/anti-IgG vs DIO/anti-CD8, p=0.0048). Using Picrosirius Red histological staining to assess kidney fibrosis, we found that anti-CD8 treatment significantly blunted kidney medullary fibrosis (one-way ANOVA, p=0.0177, ND/anti-IgG vs DIO/anti-IgG, p=0.0329; DIO/anti-IgG vs DIO/anti-CD8, p=0.0318), whereas glomerular fibrosis was not affected by anti-CD8 treatment in DIO mice (one-way ANOVA, p=0.31). Urine was collected in 12-hour increments and analyzed for markers of kidney damage. All groups of mice showed similar urinary KIM-1 excretion (one-way ANOVA; p=0.62) and protein excretion (one-way ANOVA; p=0.69). DIO mice with anti-CD8 or anti-IgG treatment had significantly increased urinary NGAL excretion compared to ND mice (one-way ANOVA; p=0.0203). In summary, anti-CD8 treatment reduced circulating and kidney CD8+ T cells, as well as kidney medullary interstitial fibrosis but not urinary NGAL excretion. These results indicate that CD8+ T cells mediate kidney medullary interstitial fibrosis in a 20 week DIO mouse model.

Funding: R01 DK134562, R25 DK115353, F31HL167626.

This is the full abstract presented at the American Physiology Summit 2024 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.
Fenoldopam-mediated increase in autophagy associates with recruiting WIPI2 protein in early endosomes in vascular smooth muscle cells
Authors: Polzin Jacob , Lu Amy , Amatya Bibhas , Armando Ines , Jose Pedro , Lee Hewang
Published: May 2024

Autophagy, a crucial cellular homeostatic process, has protective roles in the pathogenesis of hypertension. Dopamine, a monoamine neurotransmitter, regulates autophagy in the kidney and vascular system. However, the exact roles and mechanisms of dopaminergic system on the regulation of autophagy is not well known. In rat aortic vascular smooth muscle cells, fenoldopam (FEN, 25 nM/15 min), a D1-like receptor agonist, increased the ATG5 and LC3-II (a late autophagosome maker) protein expression. Consistently, FEN decreased p62/SQSTM1 protein expression. The FEN-mediated increase in ATG5 was attenuated by SCH39166 (1 μM/15 min), a D1- like receptor antagonist, indicating that FEN increased autophagy in the autophagosome formation stage through its D1-like receptors. FEN increased the protein expression of WD Repeat Domain, Phosphoinositide Interacting 2 (WIPI2) in a concentration- and time-dependent manner, which is important in the early steps of autophagosome formation. FEN (25 nM, 15 min) increased the WIPI2 puncta fluorescence intensity and the colocalization of WIPI2 with EEA1 (Veh: 7.12±1.1% vs FEN: 9.96±0.9%, N=5, P < 0.001, Student’s t test), a typical phosphatidylinositol-3-phosphate binding protein in early endosomes, indicating that WIPI2 recruitment in early endosomes is important in the early stage of FEN-mediated autophagic process. Taken together, FEN increases autophagy with WIPI2 recruitment in the early endosomes in vascular smooth muscle cells.

This work was supported by US National Institutes of Health grants R01DK134574 and R01DK119652.

This is the full abstract presented at the American Physiology Summit 2024 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.
Availability of receptors for advanced glycation end-products (RAGE) influences differential transcriptome expression in mice exposed to chronic secondhand smoke
Authors: Van Slooten Ryan , Curtis Katrina , Kirkham Maddie , Chang Ashley , Arroyo Juan , Reynolds Paul
Published: May 2024

The receptor for advanced glycation end-products (RAGE) has a central function in orchestrating inflammatory responses in multiple disease states. RAGE is a transmembrane pattern recognition receptor with particular interest in lung disease due to its naturally abundant pulmonary expression. Our previous research demonstrated an inflammatory role for RAGE following acute exposure to secondhand smoke (SHS). However, chronic inflammatory mechanisms associated with RAGE remain unclear. In this study, we assessed transcriptional outcomes in mice exposed to chronic smoke in the context of RAGE expression. RAGE knockout (RKO) and wild type (WT) mice were exposed to SHS five times weekly via a nose-only delivery system (Scireq Scientific, Montreal, Canada) for six months and compared to mice exposed to room air (RA) only. Total lung RNA was isolated using the Direct-zol RNA MiniPrep kit (Zymo Research, Irvine, CA) and mRNA was purified using poly-T oligo-attached magnetic beads. Synthesis of cDNA, library construction, and sequencing was performed using standard approaches. We specifically compared the phenotypic and environmental conditions from WT+RA, WT+SHS, and RKO+SHS mice. Preprocessing and analysis of RNA-sequencing gene expression data included read trimming, mapping and quantifying the reads to transcripts, and calculating significant differentially expressed genes. The results of these analyses were summarized and compared via Venn diagrams, volcano plots, and functional gene cluster enrichment analysis. Notable gene clusters were specific to cytoskeletal elements, inflammatory signaling, and ciliogenesis. Finally, gene ontologies (GO) demonstrated significant biological pathways that were differentially impacted by the presence of RAGE. These data collectively identify several opportunities to further dissect RAGE signaling in the context of SHS exposure and foreshadow possible therapeutic modalities.

This work was supported by funding from the National Institutes of Health (NIH 1R15-HL152257).

This is the full abstract presented at the American Physiology Summit 2024 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.
The multi-omic response to exercise training across rat tissues: interactive tools for data dissemination through the MoTrPAC data hub
Authors: Lindholm Malene E , Zhen Jimmy , Amar David , Gay Nicole R , Jean Beltran Pierre M , Yu Jay , Samdarshi Mihir
Published: May 2024

The Molecular Transducers of Physical Activity Consortium (MoTrPAC) aims to map the molecular responses to exercise and training to elucidate how exercise improves health and prevents disease. The first MoTrPAC data provides an extensive temporal map of the dynamic multi-omic response to endurance training across multiple rat tissues. All results can be viewed, interrogated, and downloaded in a user-friendly, publicly accessible data portal ( https://motrpac-data.org ). The MoTrPAC data compendium includes transcriptomics, proteomics, metabolomics, phosphoproteomics, acetylproteomics, ubiquitylproteomics, DNA methylation, chromatin accessibility, and multiplexed immunoassay data. This compilation of 211 datasets across 19 tissues, 25 molecular assays, and 4 training time points in adult male and female rats constitutes 19 terabytes of data. Over 35,000 analytes were found to be differentially regulated in response to endurance training, with many displaying sexual dimorphism. We observed a male-specific recruitment of immune cells to adipose tissues and an anticorrelated transcriptional response in the adrenal gland related to the stress response. Temporal multi-omic and multi-tissue integration demonstrated similar temporal responses in the heart and skeletal muscle, reflecting a concerted adaptation of mitochondrial biogenesis and metabolism. Integrative multi-omic network analysis revealed connections between the heat shock-mediated stress response and mitochondrial biogenesis. Training increased phospholipids and decreased triacylglycerols in the liver, and there were extensive changes to mitochondrial protein acetylation. Many changes were relevant for human health conditions, such as non-alcoholic fatty liver disease, inflammatory bowel disease, cardiovascular wellness, and tissue damage and repair. Altogether, this MoTrPAC resource provides an unprecedented view of the effects of exercise across an organism, revealing mechanistic details of how exercise impacts mammalian health. The MoTrPAC data hub is the primary online resource to disseminate this large scale multi-omics data.

The MoTrPAC Study is supported by NIH grants U24OD026629 (Bioinformatics Center), U24DK112349, U24DK112342, U24DK112340, U24DK112341, U24DK112326, U24DK112331, U24DK112348 (Chemical Analysis Sites), U24AR071113 (Consortium Coordinating Center), U01AG055133, U01AG055137 and U01AG055135 (PASS/Animal Sites).

This is the full abstract presented at the American Physiology Summit 2024 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.
Differential Distribution and Expression of α1-adrenergic Receptors in Low Shear Stress Induced Neointimal Formation
Authors: Cooper Kathryn , Taylor Mark
Published: May 2024

Low fluid shear stress in conduit arteries has emerged as a determinant of neointimal formation—an accumulation of vascular smooth muscle cells on the surface of the intima—the most challenging component of atherosclerosis to treat. α1-adrenergic receptor (α1-AR) antagonists have been well evaluated in vascular injury models yielding reduced neointimal formation. However, the α1-AR subtypes’ (α1A-AR, α1B-AR, α1D-AR) role in neointimal formation have not been determined under low shear stress conditions. α1-AR subtypes are expressed in various tissues and evoke different responses to the same ligand resulting in both adaptive and maladaptive responses. This study was designed to identify and characterize the α1-AR subtypes in low shear stress induced neointimal formation. An in vivo partial ligation mouse model was utilized to actuate low fluid shear stress and create neointimal formation in the left carotid artery in 14 days. The right carotid artery remained uninjured as the control. For tissue collection and processing, the left and right carotid arteries were harvested, snap-frozen in liquid nitrogen, and cryosectioned. Immunofluorescence staining was performed using primary antibodies for α1B-AR or α1A-AR and CD31 (endothelial cell marker). Confocal microscopy combined with processing and analysis was utilized for the detection, location, and distribution of cell markers. Histomorphometric analysis revealed significant structural changes, including the medial area and neointimal area, between the injured and non-injured arteries at the 14-day timepoint. Moreover, the injured arteries showed an increase in percent area stenosis, compared to the non-injured control. Histological observation of fluorescently labeled images revealed α1B-AR is densely distributed in the neointimal and medial area of the injured carotid arteries compared to the non-injured control. NucBlue (nuclear) staining indicated an increase in cell number in the medial and neointimal areas of the injured artery. Together, these results offer insight into changes in vascular α1-ARs under low shear stress conditions suggesting that α1B-AR may be upregulated in response to injury and contribute to neointimal formation.

NIH HL155288.

This is the full abstract presented at the American Physiology Summit 2024 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.
Low Hepatic Cardiolipin Levels Promote Increased Mitochondrial Electron Leak and Metabolic Dysfunction-associated Steatohepatitis
Authors: Lang Marisa , Shahtout Justin , Poss Annelise , Siripoksup Piyarat , Patil Mallikarjun , Peterlin Alek , Johnson Jordan
Published: May 2024

Mitochondria adapt to changing energetic demands of the cell. We recently reported that mitochondrial membrane lipid composition represents a regulatory mechanism by which effciency of oxidative phosphorylation becomes altered. Metabolic-associated fatty liver disease (MAFLD) is a progressive disorder characterized by oxidative stress and accumulation of fat in hepatocytes that can lead to metabolic dysfunction-associated steatohepatitis (MASH). We hypothesized that MAFLD alters mitochondrial membrane lipids to induce mitochondrial electron transfer ineffciency and promotes oxidative stress. Lipid mass spectrometric analyses in four models of MAFLD (high-fat diet, ob/ob, carbon tetrachloride administration (CCI4), and Gubra-Amylin-NASH® diet), revealed that low mitochondrial cardiolipin (CL) is a common signature associated with all MAFLD models. In hepa1-6 cells, lentivirus-mediated knockdown of cardiolipin synthase (CLS, generates CL in mitochondria) was suffcient to reduce respiration and increase electron leak. Liver mitochondria from mice with a hepatocyte-specific knockout of CLS exhibited defects in mitochondrial bioenergetics including increased electron leak, concomitant to pathogenesis of MASH. Together, these findings provide evidence that a reduction in hepatic cardiolipin may drive the progression of MAFLD to induce MASH pathology.

R01GM144613-03R01AG074535-02S2R01DK107397 07.

This is the full abstract presented at the American Physiology Summit 2024 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.
Progranulin and Lysosomal pH: Implications for the Proinflammatory Response in Neurodegenerative Diseases
Authors: Yang Andrew Z. , Harrison Alisha M. , Uppalapati Chandana K. , Pascual Agnes S. , Biparva Pegah , Leyva Kathryn J. , Hull Elizabeth E.
Published: May 2024

Progranulin (PGRN) deficiency and lysosomal dysfunction have been independently linked to Alzheimer's disease. Several lines of evidence suggest that there is a link between PGRN and lysosomal function. PGRN is a pleiotropic signaling molecule whose activity depends upon differential proteolytic processing. The different PGRN subunits generated during processing can regulate inflammation, lysosomal function, and/or growth. Not only is the lysosome a site of PGRN processing, the resulting PGRN subunits promote the function of several lysosomal proteases. In addition, in models of PGRN insuffciency, increasing the levels of PGRN restores lysosomal function and reduces inflammation. As microglial inflammation is key to the development of neurodegenerative diseases, this work investigates the link between lysosomal function, PGRN, and inflammation utilizing two cell lines, each for a distinct set of experiments. The SW13 human adrenal carcinoma cell line that exists in two epigenetically distinct subtypes was used to focus on differences in lysosomal pH and PGRN processing. The HMC3 human microglial cell line was used to measure microglial activation and cytokine production. We hypothesize that PGRN, when appropriately processed, will restore lysosomal function to reduce pro-inflammatory cytokine production. Results suggest that differential processing of PGRN is correlated to both differences in rate of growth, inflammatory cytokine production, and lysosomal pH as PGRN expression and processing varies when cells are stimulated which may be related to observed changes in lysosomal pH and proteases. Ongoing experiments address the possibility that decreasing lysosomal pH may shift the processing of PGRN, influencing the biological activities associated with this versatile signaling molecule. As mutations in the PGRN gene are implicated in both dysfunctional lysosomal pH and a variety of neurodegenerative diseases, these results may provide a novel approach to regulate proper intracellular functions to slow or prevent the development of disease.

Midwestern University Biomedical Sciences Program Funds awarded to AZY and PB; Kenneth A. Suarez Research Fellowship Funding awarded to PB; Arizona Alzheimer's Consortium Pilot Grant Funding awarded to EEH and KJL; Midwestern Arizona Alzheimer’s Consortium (MAAC) Grant Funding awarded to EEH and KJL; Midwestern University Intramural Funding awarded to EEH and KJL.

This is the full abstract presented at the American Physiology Summit 2024 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.
Insulin-Dendrimer Nanocomplex for Multi-Day Glucose-Responsive Insulin Therapy in Ossabaw Miniature Swine with Type 1 Diabetes
Authors: Xian Sijie , Xiang Yuanhui , Liu Dongping , Fan Bowen , Mitrová Katarína , Ollier Rachel , Alloosh Muhammad Ali
Published: May 2024

Background: Therapy for type 1 diabetes resulting from immunogenic destruction of the insulin-producing pancreatic beta cells has been a challenge for over 100 years. Delivering exogenous insulin to maintain euglycemia is a delicate balance between plasma glucose transport to insulin-responsive tissues, glucose absorption in the gut, and the pharmacodynamics of insulin. The synthesis of short- and long-acting insulins and use of insulin pumps have made great strides in the standard of diabetes care. Despite advances, hypoglycemia is a major limitation. There is no approved therapy that offers glucose-responsive insulin function. Toward this end we engineered a nanoscale complex combining both electrostatic and dynamic-covalent interactions between a synthetic dendrimer carrier and an insulin analogue modified with a high-affnity glucose-binding motif. Hypothesis: This insulin nanocomplex will yield an injectable insulin depot providing both glucose-sensitive and long-lasting insulin availability in a translation preclinical model. Methods: Ossabaw minature swine (41-59 kg; N=6) were made diabetic with alloxan, which was confirmed by stable hyperglycemia (228±43 mg/dL). Oral glucose tolerance tests (OGTT) were conducted at the diabetic baseline and each of the following 6 days after one subcutaneous injection of the insulin nanocomplex. Results: The basline OGTT elicted blood glucose of 350-450 mg/dL for 90 min of the 150 min OGTT, while serum insulin remained at ~5 μU/mL, nearly the detection limit. In contrast, 3 h after injection of insulin nanocomplex (~0.8-0.9 mg/kg) blood glucose decreased to 71±12 mg/dL. Subsequent OGTTs on days 1-6 increased blood glucose from normoglycemia to ~200 mg/dL and serum insulin increased to ~60-220 μU/mL, followed by return to nearly normolgycemia hours after the OGTT. Hypoglycemia was avoided in all pigs. Conclusions: The subcutaneous nanocomplex of a modified insulin and a synthetic dendrimer carrier produced a glucose-responsive insulin depot for week-long glycemic control following a single routine injection. These findings offer great promise for overcoming the hypoglycemia risk of intensive glucose control. Future studies should include additional physiological challenges of exercise and nutritional states.

Support: Juvenile Diabetes Research Foundation 5-CDA-2020-947-A-N, Helmsley Charitable Trust 2019PG-T1D016 and 2102-04994, American Diabetes Association Pathway 1-19-ACE-31, National Science Foundation BMAT1944875, European Union-Next Generation EU LX22NPO5104, Czech Academy of Sciences RVO 6138963.

This is the full abstract presented at the American Physiology Summit 2024 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.
Daunorubicin Treatment Impairs Cardiac Function in The Ts65Dn Mouse Model Of Down Syndrome
Authors: Buckman Michelle , Vasileva Anastasiia , Vasilyev Mikhail , Jedlicka Charles , DeRuisseau Lara , Tomasson Michael , Bates Melissa
Published: May 2024

Anthracycline treatment for leukemia is effective but has toxic side effects on the cardiovascular system. Few studies regarding the impact of anthracyclines on the heart in individuals with Down syndrome have been addressed although they are at a higher risk of developing leukemia. It is unknown if established mechanisms apply to individuals with Down syndrome. We hypothesize that anthracycline cardiotoxicity will lead to the enhanced impairment of cardiac function and initiate pronounced vascular dysfunction in a mouse model of Down syndrome. We used the Ts65Dn mouse model of Down syndrome to investigate the impact of daunorubicin chemotherapy on survival, arterial stiffness and left ventricular function. Four-month-old Ts65Dn (n=12) and wild-type control Ts65Dn (WT, n=8) mice were treated with 2mg/kg of daunorubicin or saline. Following treatment, left ventricular catheterization was performed to assess cardiac function. Serum cardiac troponin levels were evaluated using ELISA. Histological analysis was conducted on the heart and aorta of mice to assess the accumulation of elastin and collagen. We found decreased end-diastolic pressure (p=0.016) and ejection fraction (p=0.029) in Ts65Dn daunorubicin-treated mice compared to WT control. However, contractility (end-systolic pressure-volume relationship, p=0.221) and diastolic stiffness coeffcient (end-diastolic pressure-volume relationship, p=0.418) remained unchanged. Ts65Dn mice had a higher cardiac troponin concentration (p=0.02). Histological assessment showed no significant difference in collagen (p=0.841) and elastin (p=0.768) content. Daunorubicin negatively impacted end-diastolic pressure and ejection fraction of the Ts65Dn mouse model of Down syndrome. We demonstrated that we could recapitulate the findings of anthracycline cardiotoxicity in Ts65Dn mice indicating this is a good model to investigate toxicity. Continued work with this model will better elucidate the mechanisms underlying anthracycline cardiotoxicity in Down syndrome. This shows the need for constant screening after anthracycline treatment which will improve the survival of this population.

NIH R21 HD099573.

This is the full abstract presented at the American Physiology Summit 2024 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 Tissue Distribution of Microplastics and the Systemic Metabolic Changes After Gastrointestinal Exposure in Mice
Authors: Romero Aaron
Published: May 2024

Marcus M. Garcia, Aaron S. Romero, Seth D. Merkley, Jewel L. Meyer-Hagen, Charles Forbes, Matthew J. Campen, Julie G. In, and Eliseo F. Castillo Microplastics (MP) are slated to be the next environmental threat to humans. It is estimated humans consume approximately 5 grams of MP per week. To date, the systemic impact of MP ingestion on metabolic pathways in systemic organs have not been studied utilizing environmentally relevant doses and mixtures. To investigate the impact of ingested MPs on target metabolic pathways, mice were exposed to either polystyrene (5 μm) microspheres or a mixture of MPs consisting of polystyrene (5 μm), polyethylene (1-4 μm), and the biodegradability and biocompatible plastic, poly-(lactic-co-glycolic acid) (5 μm). Exposures were performed twice a week for four weeks at a dose of either 0, 2, or 4 mg/week via oral gastric gavage. Our findings demonstrate that, in mice, ingested MPs can pass through the gut barrier and accumulate in distant tissues. Additionally, we report on the metabolic changes that occur in the colon, liver and brain which show differential responses that are dependent on dose and type of MPs exposure. This study provides critical insight into the pervasive issues of plastic pollution and highlights dose-dependent and polymer type specific metabolic changes in the colon, liver and brain caused by MPs exposure.

NIH/NIEHS 1R01 ES032037-01A1.

This is the full abstract presented at the American Physiology Summit 2024 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.
Chronic intermittent hypoxia alters CD8+ expression in the bone marrow of C57BL/6 mice
Authors: Edwin Immaculate , Jedlicka Charles , Zhang Zishan , Vasilyev Mikhail , Vasileva Anastasiia , Tomasson Michael , Bates Melissa
Published: May 2024

Cytotoxic CD8+ cells are critical in anticancer immune response and have become prevalent in immunotherapies over the past decade. Anti-LAG3/anti-PD1 combined immunotherapies increase the number of CD8+ cells. Moderate to severe sleep apnea increases the risk of cancer-related mortality. Chronic intermittent hypoxia, a feature of sleep apnea, decreases the percentage of perforin-expressing CD8+ cells, impairing cytotoxicity. Our objective is to determine the distribution of Prf1(perforin), LAG3(lymphocyte-activation gene 3), and PDCD1(programmed cell death 1) in C57BL/6 mice exposed to chronic intermittent hypoxia (CIH). We hypothesize that Prf1, PDCD1, and LAG3 will be expressed on three discrete clusters with a higher expression of LAG3 and PDCD1 and lower expression of Prf1 between C57BL/6 mice exposed to chronic intermittent hypoxia and normoxia. Bone marrow from the femur of C57BL/6 mice exposed to chronic intermittent hypoxia or normoxia was harvested. Bone marrow mononuclear cells were isolated, RNA was extracted, and single-cell RNA sequencing was performed. Loupe browser was used to re-cluster and quantify expression for genes of interest. Prf1 and PDCD1 are expressed in CD3+CD8−CD4− cluster and LAG3 is expressed in CD3−CD8+CD4+ cluster. LAG3 and Prf1 are present among the top ten genes, while PDCD1 is not in those clusters. Further analysis of the top ten genes of CD3+CD8−CD4− has demonstrated they are a group of natural killer cells. Contradictory to our expectation, Prf1 and PDCD1 are present in CD3+CD8−CD4− cluster while LAG3 is present in CD3−CD8+CD4+ cluster. Further analysis is necessary to determine the underlying mechanisms causing these changes.

5R01CA244271-02.

This is the full abstract presented at the American Physiology Summit 2024 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.
E-Cigarette Aerosol Alters Pulmonary Surfactant Function
Authors: Graham Emma , McCaig Lynda , Abu Al-Burak Salem , Veldhuizen Ruud
Published: May 2024

Introduction: E-cigarette (EC) and vaping use continue to remain popular, particularly in young populations despite several clinical reports of vaping associated lung injury. One of the first compounds that EC aerosols comes into contact within the lungs during a deep inhalation is pulmonary surfactant. A complex mixture of lipids and proteins, surfactant reduces surface tension in the lungs maintaining compliance and preventing collapse. Currently, information on how EC aerosols impacts pulmonary surfactant remains limited, particularly in the in vivo scenario. We hypothesized that exposure to EC aerosol will impair the surface tension reducing ability of surfactant. Methods: In vitro EC aerosol exposure : Bovine Lipid Extract Surfactant (BLES) was used as a model surfactant in a direct exposure syringe system. BLES (2ml) was placed in a syringe (30ml) attached to an EC. The generated aerosol was drawn into the syringe and then expelled, repeated 30 times. In vivo EC aerosol exposure : Male and female Wistar rats were exposed to vehicle or menthol flavoured EC aerosol. Rats were exposed for 15 minutes for either a single or multiple (up to 8) exposures, with control animals receiving room air. Animals were euthanized and bronchoalveolar lavage was performed to collect surfactant. Analyzing surfactant function: Samples underwent dynamic compression and expansion, and biophysical analysis after exposure, completed using a constrained drop surfactometer (CDS). Results: Significant increases in minimum surface tensions were observed after exposure to EC aerosol across 20 compression/expansion cycles. Surfactant function was altered in vitro independent of vaping device, temperature, or inclusion of nicotine. Vehicle aerosol exposure produced alterations in surfactant properties, however addition of menthol flavouring significantly increased surface tensions compared to control and vehicle exposure. Preliminary in vivo studies indicate that aerosol exposure is well-tolerated by both male and female animals. Further experiments will analyze the function of surfactant isolated from these animals. Discussion: EC aerosols alter surfactant function through increases in minimum surface tension in vitro. This effect was more pronounced with menthol flavoured EC aerosol. It is anticipated that our in vivo experiments will corroborate these findings. Ultimately, impairment of surfactant due to EC aerosol exposure may contribute to lung dysfunction and susceptibility to further injury.

Western University, NSERC, OGS.

This is the full abstract presented at the American Physiology Summit 2024 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.
The consequences of environment-induced heat stress on the kidney in female mice
Authors: Roths Missey , Rudolph Tori , Krishna Swathy , Michael Alyona , Selsby Joshua
Published: May 2024

Environment-induced heat stress (EIHS) is caused by a sustained elevation in body temperature, due to prolonged exposure to excess heat and humidity. Given the present and increasing frequency of EIHS and the gap in knowledge regarding EIHS-mediated impacts to the kidney, the purpose of this investigation was to determine the extent to which EIHS alters kidney health. We hypothesized EIHS would cause kidney injury and cellular dysfunction. To test this hypothesis, 7 wk old C57 female mice were assigned to either EIHS (n=14; 36.7 ± 0.16 °C, 36.3 ± 0.5 % humidity) or thermoneutral (TN) conditions (n=12, 31.2 ± 1.01 °C, 35 ± 0.7% humidity) for 24 h, kidneys were removed, weighed, and were either fixed for histology or frozen for biochemical measures. Environment-induced HS increased rectal temperature by 2.1 °C (p< 0.001), decreased body weight by 10% (p=0.03), and decreased absolute kidney weight (10%; p=0.02) compared to TN. Environment-induced HS increased relative protein abundance of heat shock proteins 70 (48%, p<0.01), 90 (37%, p<0.01), and 60 (36%, p=0.02), as well as heat shock factor 1 (2-fold, p=0.02) compared to TN. Histological inspection revealed EIHS animals had increased vacuolation in the proximal convoluted tubules (p<0.01) compared to TN animals. Using a western blot approach, we measured biomarkers of kidney tubular health and discovered EIHS increased protein abundance of NGAL (88%, p<0.01), IL-10 (36%, p=0.05), IL-6 (46%, p<0.01), MCP-1 (74%, p=0.04), and L-FABP (99%, p<0.01), which are suggestive of kidney injury. Environment-induced HS increased markers of inflammatory signaling including TLR4 (44%, p=0.04), phosphorylated (p-) NFκB (93%, p<0.01), p-JNK (54 kD; 51%, p=0.02), p-JNK (46 kD, 35%, p=0.02), AP-1 (33%, p=0.02), and TNFα (39%, p=0.05). Endoplasmic reticulum stress was increased by EIHS as Bip, p-IRE1α, Perk, XBP1s, ATF4, and CHOP were increased 27-190% (p<0.05-0.01) compared to TN animals. Markers of mitochondrial abundance remained similar between groups; however, EIHS may alter mitochondrial fusion and fission as Opa1 (71%, p=0.01), MFN2 (29%, p=0.04) and DRP1 (59%, p=0.04) were increased compared to TN; however, MFN1, FIS1, and p-DRP1 were similar between groups. Collectively, these data suggest that 24 h of EIHS causes kidney injury, but may also be suffcient to initiate changes that contribute to acclimation.

This study was supported by USDA 2020-02716.

This is the full abstract presented at the American Physiology Summit 2024 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.
Sympathetic Transduction to Blood Pressure in Patients with Chronic Kidney Disease
Authors: Trotter Claire E. , Young Benjamin E. , Conley Rie V. , Kaur Jasdeep , Mohan Ponnaiah C. , Fadel Paul J.
Published: May 2024

Chronic kidney disease (CKD) is a major health problem affecting more than 35 million Americans. Notably, more patients with CKD die of cardiovascular complications than progress to end-stage renal disease. An overactive sympathetic nervous system is a well-known cardiovascular risk factor. Previous research has suggested elevated resting sympathetic nerve activity and a heightened α-adrenergic receptor sensitivity in patients with CKD, both of which may predispose this population to greater sympathetic transduction. Therefore, we tested the hypothesis that patients with stage III CKD have an augmented sympathetic transduction to blood pressure (BP) compared to controls without CKD. In 7 patients with stage III CKD and 8 controls muscle sympathetic nerve activity (MSNA; peroneal microneurography) and beat-to-beat BP (finometer) were continuously recorded during 10 minutes of quiet supine rest. Modelflow was used to estimate stroke volume and calculate total vascular conductance (TVC). Sympathetic transduction was quantified as the peak change in mean arterial pressure (MAP) and the nadir reduction of TVC over 10 cardiac cycles following spontaneous MSNA bursts using signal averaging. Patients with CKD and controls had similar resting MAP (CKD: 102 ± 14 mmHg; CON: 95 ± 8 mmHg; P = 0.27), MSNA burst frequency (CKD: 33 ± 10 bursts/min; CON: 31 ± 14 bursts/min; P = 0.78) and MSNA burst incidence (CKD: 53 ± 15 bursts/100 heartbeats; CON: 47 ± 24 bursts/100 heartbeats; P = 0.60). Peak increases in MAP following all bursts of MSNA were not different between patients with CKD and controls (CKD: 2.1 ± 0.7 mmHg; CON: 2.2 ± 0.9 mmHg; P = 0.83). Likewise, nadir reductions in TVC following all bursts of MSNA were not different in patients with CKD (CKD: -1.2 ± 0.6 ml/min/mmHg; CON: -1.4 ± 0.5 ml/min/mmHg; P = 0.46). Similar results were observed when considering burst patterning. MSNA bursts occurring in multiples (adjacent to other bursts) produced comparable peak MAP (CKD: 2.6 ± 0.6 mmHg; CON: 2.8 ± 1.2 mmHg; P = 0.67) and nadir TVC (CKD: -1.5 ± 0.7 ml/min/mmHg; CON: -1.9 ± 0.7 ml/min/mmHg; P = 0.25) responses between groups. There were also no differences in peak MAP and nadir TVC responses following single bursts of MSNA (bordered by cardiac cycles lacking bursts; both P > 0.30). These preliminary data suggest that, in contrast to our hypothesis, sympathetic transduction to BP is not augmented in patients with stage III CKD.

This project was supported by the National Heart, Lung, and Blood Institute R01 HL-127071 (PJF).

This is the full abstract presented at the American Physiology Summit 2024 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.
Antioxidant-induced modulation of sympathetic transduction to arterial blood pressure in humans
Authors: Veach Jodi , Kovarik Jack , Gangwish Megan , Clonch Davina , Thyfault John , Miles John , Holwerda Seth
Published: May 2024

Purpose: Excess plasma free fatty acids (FAA) promote oxidative stress and an increase in peripheral resistance. Therefore, we tested the hypothesis that excess FAA limits sympathetic control of arterial blood pressure (BP) and that sympathetic control would be restored during co-administration of ascorbic acid, which is an antioxidant with vasodilatory properties. Methods: A 20% IV fat emulsion (Intralipid) was administered for 2 hours to 5 lean, healthy adults with and without co-infusion of ascorbic acid (separate visits) in a double-blinded, crossover study design. Sympathetic control of BP was examined by determining the BP response to individual bursts of muscle sympathetic nerve activity (MSNA) (microneurography). Briefly, beat-to-beat mean arterial pressure (MAP) was measured continuously (finger cuff) and signal-averaged over 15 cardiac cycles following each detected burst of MSNA over a 10-min resting period. Results: In contrast to our hypothesis, no change was observed in sympathetic transduction during lipid infusion (1.5 ± 0.4 vs. 1.4 ± 3 mmHg, P=0.75). However, co-infusion of ascorbic acid significantly increased sympathetic transduction compared with baseline (1.5 ± 0.4 vs. 2.0 ± 3 mmHg, P=0.04), in addition to reducing peripheral resistance (0.73 ± 0.11 vs. 0.53 ± 12 mmHg·min·mL−1, P=0.07). MSNA and MAP were not significantly changed during the 2-hr infusions. Conclusions: These preliminary findings suggest that excess FAA do not modulate sympathetic transduction in lean, healthy adults. However, ascorbic acid infusion reduces peripheral resistance and increases sympathetic transduction, demonstrating an inverse relation between peripheral resistance and sympathetic transduction in healthy adults.

Funding and support: R01HL159370-01 (S.W.H.). This work was also supported by a CTSA grant from NCATS awarded to the University of Kansas for Frontiers: University of Kansas Clinical and Translational Science Institute (# UL1TR002366) The contents are solely the responsibility of the authors and do not necessarily represent the offcial views of the NIH or NCATS.

This is the full abstract presented at the American Physiology Summit 2024 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.
Diversity Outbred Mouse Model Recapitulates Human Heterogeneity in the Development of Western Diet-Induced Complications
Authors: Chittimalli Kishore , Tolle Andrew , Rahman Mansib , Jarajapu Yagna
Published: May 2024

Background: Diet-induced obesity (DIO) increases risk for cardiovascular disease largely due to compromised gut barrier integrity and systemic inflammation. Animal models for understanding the pathophysiology of complications associated with DIO do not take address the genetic factors, which determine the individual variation in the susceptibility to develop complications. Hypothesis: DO mice recapitulate the heterogeneity of western diet-induced metabolic syndrome, gut barrier disruption and systemic inflammation. Methods: DO mice were fed with either control (CD) or western diet (WD) (Teklad) (n = 10) for 16 weeks. Blood glucose levels, glucose tolerance and the intestinal permeability were determined. Inflammatory cells in the bone marrow or blood were characterized in basal conditions or in response to hindlimb ischemia (HLI). Blood flow recovery following HLI was measured by using Laser Doppler Imaging system and the intestinal-stem-cells (ISCs) were characterized by using markers, Lgr5, Olfm4 and Wnt3. Results: Wide variation in the weight gain, hyperglycemia and glucose tolerance in DO mice was observed following WD (n = 10). Similarly, wide variation was observed in Wnt3 expression and in the density of ISCs in colon of DO mice on WD. Heterogeneity in the ISC layer correlated with the variation in the intestinal permeability induced by WD compared to the CD ( p < 0.05, n = 10). The number of inflammatory cells (monocytes – CD45+Ly6G−Ly6C+CD115+, neutrophils - CD45+Ly6G+Ly6C+CD115−, pro-inflammatory macrophages - CD45+CD11b+Ly6G−Ly6C+F4/80+CCR2+CD80+ and anti-inflammatory macrophages - CD45+CD11b+Ly6G−Ly6C+F4/80+CX3CR1+CD206+) in the BM or blood showed heterogeneity in WD-DO mice in accordance with variability in the glucose tolerance. Accordingly, blood flow recovery following HLI showed wide variation with three mice on WD showing full recovery, which correlated with variability in the mobilization of inflammatory cells (n = 9). Conclusion: DO mouse model depicts human diversity in response to WD-induced changes in gut barrier integrity, inflammation and ischemic vascular repair therefore serves as a suitable model for evaluating pharmacological agents that would be effective in a diverse population.

Jackson Laboratory Diversity Outbred Pilot Award National Institute of Aging (AG056881).

This is the full abstract presented at the American Physiology Summit 2024 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.
Sox9 links biliary maturation to branching morphogenesis
Authors: Hrncir Hannah , Bombin Sergei , Goodloe Brianna , Hogan Connor , Gracz Adam
Published: May 2024

During liver development, bipotential epithelial progenitor cells, termed hepatoblasts, give rise to hepatocytes and biliary epithelial cells (BECs). BECs line the intrahepatic bile ducts which are essential for liver function. The adult intrahepatic bile duct (IHBD) network is hierarchical in structure with smaller, peripheral “ductules” draining into larger “ducts” that eventually exit the liver. Despite proposed models from histological and genetic studies, mechanisms differentiating local “duct” or “ductule” hierarchies and driving the global IHBD architecture are poorly understood. Previous studies of IHBD development have focused on mechanisms of BEC specification/ de novo bile duct formation by tubulogenesis. However, the contribution of pre-existing BECs to IHBD expansion and morphogenesis is unclear. Previous studies on liver specific Sox9 knockout (Sox9cKO) mice showed that Sox9 is important for timing of BEC specification during development and prevents cystogenesis in aged mice. We hypothesize that SOX9 is an important regulator of IHBD local and global architecture during development by promoting IHBD branching morphogenesis. Using whole-lobe 3D imaging and iDISCO+, we find that adult, embryonic, and postnatal Sox9cKO mice demonstrate ductal paucity with bias towards ductule, rather than duct, loss. Bulk RNA-sequencing reveals that Sox9cKO BECs are transcriptionally “immature” and have gene expression signatures consistent with a failure to fully acquire BEC identity. scRNA-sequencing identifies ductule-like cells in a subset of Sox9cKO BECs correlating with elevated Inhba, the gene precursor of Activin A. Sox9cKO biliary organoids exhibit non-cystic morphology that is reproduced in Activin A treated WT organoids. In vitro and in vivo, Activin A signaling inhibition experiments partially rescue BEC morphology. Together, our data demonstrate that SOX9 drives BEC maturation and promotes ductule formation, in part through inhibiting Activin A. Our data support a model for IHBD development involving distinct mechanisms governing Sox9-independent large duct tubulogenesis versus Sox9-dependent small ductule branching morphogenesis.

F31DK134199; PI: Hannah Hrncir T34GM149511; PI: Anita Corbett, PhD; R01DK132653; PI: Adam Gracz, PhD.

This is the full abstract presented at the American Physiology Summit 2024 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.
Loss of AC10 Impairs Neutrophil Recruitment and Alters Bacterial Colonization in the Lung During Bacterial Pneumonia
Authors: Stone Madeline , Choi Chung-Sik , Sayner Sarah
Published: May 2024

Rationale: Pseudomonas aeruginosa is a gram-negative bacterium and a common cause of nosocomial pneumonia that can progress to acute respiratory distress syndrome (ARDS), characterized by hypoxia, pulmonary edema, elevated bicarbonate, and widespread inflammation. Neutrophil recruitment is a critical line of defense against P. aeruginosa; however, excessive, or dysregulated neutrophil infiltration can exacerbate tissue damage and contribute to the severity of ARDS. Recently, AC10 has been implicated in neutrophil transendothelial migration via CD99. Despite the growing recognition and significance of AC10 in various physiological processes, its role in neutrophil recruitment during ARDS remains poorly understood. Thus, herein we hypothesize that loss of AC10 attenuates neutrophil recruitment during pneumonia and increase bacterial colonization in the lung. Methods: Wild-type and AC10 knockout mice were intratracheally inoculated with increasing doses of P. aeruginosa. Twenty fours after infection, bronchoalveolar lavage fluid (BALF) was collected and the percent of PMNs in the BALF determined. In separate experiments, mice were intratracheally inoculated with P. aeruginosa for 6 or 24 hours and organs (lungs, kidney, liver, spleen and brain) harvested, homogenized and the homogenate cultured overnight on Pseudomonas isolation agar for quantification of colony forming units (CFUs). Results: Our data confirmed CD99 expression in rat pulmonary microvascular endothelial cells and whole lung of both wild type and AC10 knockout mice. While there was no difference in the percentage of PMNs in the BALF between male wild type and knockout animals at any of the bacterial doses tested, in the female AC10 knockout mice, there was a decrease in the percentage of cells PMNs in the BALF compared to female wild type litter mates at several doses tested (1x103 up to 1x105, p<0.05). At a higher dose of P. aeruginosa (2.5x106), there was no difference in bacterial clearance between male WT and AC10 knockout mice, but a trend toward an increase in the CFU harvested from lung tissue 6 hours post infection in female KO mice. Conclusion: Thus, CD99 is expressed in the pulmonary endothelium. Further, in female mice, AC10 plays a role in neutrophil recruitment into the airspace. Further studies are required to resolve whether AC10 plays a role in bacterial clearance in female mice in a model of P. aeruginosa pneumonia.

HL121513.

This is the full abstract presented at the American Physiology Summit 2024 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.