bluuebunny/crossref_metadata_2025 huggingface dataset:
Measurement of hippocampal arteriolar diameter and blood velocity in awake head-fixed mice
Authors: Williams Alexandra , White Carl
Published: May 2023

Cerebral hypoperfusion is associated with neurodegeneration. However, the influence of disease on the microcirculation of specific brain regions is poorly understood. In the current study, we validated a technical approach that allows us to simultaneously image arteriolar diameter and blood velocity in the hippocampi of awake, head-fixed mice. The approach involved implanting a gradient-Index (GRIN) lens into the hippocampus and cementing it in place together with a steel head-fixation bar. Imaging was performed after 7-14 days recovery. On the day of imaging, we intravenously administered Alexa Fluor 633 hydrazide (1.25 mg/kg) to label arterioles, and after 90 minutes, secured the mice on a custom-built stage of a widefield fluorescence microscope. We measured blood velocity in parallel with vessel diameter by imaging fluorescent latex beads infused by tail vein cannulation. Vessel diameter and blood velocity were assessed in 14 vessels from 6 mice. Hypercapnia induces cerebral vasodilation and increases cerebral blood flow. In our experiments, inhalation of 10% CO2 caused a 12.1 ± 6% (mean ± SEM) increase in vessel diameter and a significant (P<0.01, student’s t-test) increase in blood velocity from 1.09 ± 0.1 to 1.73 ± 0.2 cm/s (mean ± SEM). These data demonstrate the feasibility of this approach to assess microvascular function and perfusion in highly localized brain regions.

NIH grant (HL142906)

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 the cofilin pathway play a role in closure of the avian ductus arteriosus (Gallus gallus)?
Authors: Rippamonti Jessica , Dzialowski Ed
Published: May 2023

The ductus arteriosus (DA) is an oxygen-sensitive embryonic blood vessel present in all developing amniotes. The DA connects the pulmonary artery to the aorta to shunt blood away from nonfunctioning lungs and toward the fetal respiratory organ. Increased arterial PO2 after birth or hatch initiates permanent closure of the DA. Permanent closure of this vessel upon birth or hatch establishes proper separation of the pulmonary and systemic circuits in the mammalian and avian neonate. Smooth muscle contraction is governed by myosin light chain kinase (MLCK), which stimulates increased cross-bridge cycling, and through increases in actin polymerization. Actin polymerization occurs in smooth muscle cells when globular actin (G actin) polymerizes to form filamentous actin (F actin), allowing for greater contractile strength. Regulation of actin polymerization in some blood vessels involves the cofilin pathway. P-21 activated kinase (PAK) regulates Lim kinase, which inhibits the actin depolymerizer cofilin. Activation of cofilin results in the severing of filamentous actin, leading to decreased tension of vessels. In this study we examined the role of the cofilin pathway in maintaining DA patency in the embryonic stage and subsequent constriction and closure during hatching in the chicken. Following Isoflurane inhalation, embryonic DA were excised from developing chickens and vessel tension was evaluated using wire myography. The DA were maintained in physiological saline solution and low oxygen conditions before being exposed to inhibitors of the PAK pathway followed by exposure to elevated oxygen. To examine the role of the cofilin pathway in DA physiology, we used IPA3 to inhibit P-21 activated kinase, and LIMKi3 and SR7826 to inhibit Lim kinase. Exposure to IPA3 did not alter baseline vessel tension or contraction in response to elevated oxygen. Inhibition of Lim kinase did not disrupt vessel tension or responsiveness to elevated oxygen levels. Our findings suggest that the cofilin pathway is not involved in regulating tension dynamics during the embryonic development and subsequent closure of the DA at hatching.

This research was funded by Grant R15HL14887 from NIH-NHLBI.

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.
17-Hydroxyprogesterone caproate lowers angiotensin II type 1 receptor agonistic autoantibody and improves hypertension in early-onset preeclampsia
Authors: Amaral Lorena , McCray Morgan , Evans Kymberlee , Hines Dorianne , Campbell Nathan , Deer Evangeline , Herrock Owen , Turner Ty , Wallace Kedra , Ibrahim Tarek , Lawson Frances , Harrell Ariel , LaMarca Babbette
Published: May 2023

Preeclampsia (PE), new onset hypertension and significant end-organ dysfunction with or without proteinuria after 20 weeks of gestation, affects 8% of all pregnancies in the U.S. and it is associated with progesterone deficiency, chronic inflammation, elevated angiotensin II type 1 receptor agonistic autoantibody (AT1-AA) and endothelial dysfunction. Currently, the early delivery of the feto-placental remains the best treatment for PE. Elevations in progesterone and T helper 2 cytokines favor the immunotolerance of the fetus during healthy pregnancy. Progesterone, through its receptors, stimulates anti-inflammatory and vasodilatory pathways. Therefore, this study was designed to test the hypothesis that progesterone, in the form of 17-hydroxyprogesterone caproate (17-OHPC), lowers AT1-AA and inflammation while reducing maternal blood pressure in women with early-onset preeclampsia (EO-PE). PE women received 17-OHPC (250 mg, I.M.) and blood draws were collected before and after 17-OHPC treatment. Placentas were collected at the delivery. Progesterone and PIBF levels were 15.4 +/- 45.6 pg/mL (n=12), 14.4 +/- 1.0 ng/mL in PE (n=10), which significantly increased to 155+/-57.0 pg/mL, 16.39 +/-0.74 ng/mL (p< 0.05 vs. PE) in PE+17-OHPC (n=6). TNF-alpha and IL-6 were 41.3 +/- 3.9 pg/mL (n=8), 20.1 +/-7.9 (n=11) in PE, which decreased to 21.1+/-5.5 (n=6, p< 0.05 vs. PE), 5.2+/-2.1 in PE+17-OHPC (n=5). AT1-AA levels were 23+/-5.2 ΔBPM in PE (n=5), which significantly reduced to 2.1 ΔBPM in PE+17-OHPC (n=8, p< 0.05). 17-OHPC prolonged time of delivery beyond 72h on average and maternal blood pressure was 148 +/- 3.7 mmHg in PE (n=19), which significantly decreased to 136+/-4 in PE+17-OHPC (n=14, p< 0.05). In conclusion, 17-OHPC reduces AT1-AA and inflammation while improved blood pressure and maternal outcomes in response to early-onset preeclampsia.

Supported by NIH grants RO1HD067541, 1U54GM115428, P20GM121334

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.
Oaks to arteries: The Physiology Core Concept of flow down gradients supports transfer of student reasoning
Authors: Doherty Jennifer , McFarland Jenny , Wenderoth Mary Pat
Published: May 2023

The Physiology Core Concept of flow down gradients is a major concept in physiology as pressure gradients are the key driving force for the bulk flow of fluids in biology. However, students struggle to understand this principle is foundational to the mechanism governing bulk flow across diverse physiological systems (e.g., blood flow, phloem sap flow). In this study, we explore if bulk flow items that differ in context (i.e., taxa, amount of scientific terminology, living or non-living system) or which aspect of the pressure gradient is kept constant (i.e., starting pressure or pressure gradient) influences undergraduate students’ reasoning. We found item context did not impact the type of reasoning students used. However, students were more likely to use the Physiology Core Concept of “flow down [pressure] gradients” when the pressure gradient was kept constant and less likely to use this concept when the starting pressure was kept constant. We also investigated if item context or which aspect of the pressure gradient is kept constant impacted how consistent students were in the type of reasoning they used across two bulk flow items on the same homework. We found most students were consistent across item contexts (76%) and aspects of the pressure gradient kept constant (70%). Students who reasoned using “flow down gradients” on the first item were the most consistent (86, 89%) while students using “pressures indicate (but don’t cause) flow” were the least consistent (43, 34%). Students who are less consistent know pressure is somehow involved or indicative of fluid flow but do not have a firm grasp of the concept of a pressure gradient as the driving force for fluid flow. These findings are the first empirical evidence to support the claim that using Physiology Core Concept reasoning supports transfer of knowledge across different physiological systems.

National Science Foundation award DUE 1661263

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 adults who had a concussion, cardiac autonomic function is not impaired beyond three months from injury
Authors: Garza Nicole , Young Benjamin , Skow Rachel , Nandadeva Damsara , Stephens Brandi , Lockette Warren , Fadel Paul
Published: May 2023

Introduction: Concussion is a form of traumatic brain injury (TBI) which temporarily disrupts normal brain function. It is well established that cardiac autonomic function is impaired immediately following concussion. However, studies investigating cardiac autonomic function as measures of spontaneous cardiac baroreflex sensitivity (BRS) and heart rate variability (HRV) beyond three months from the event are limited, and the results are inconsistent. Therefore, the purpose of this study was to investigate the lasting impact of concussion on cardiac BRS and HRV in participants who are beyond 3 months from their last concussion. We hypothesized that cardiac BRS and HRV would be impaired in adults who previously had a concussion compared to healthy controls with no known history of concussion. Methods: Five adults (3 females, 27 ± 6 years; mean ± standard error) with a diagnosed concussion prior to assessment (6.2 ± 0.5 months) and six controls (3 females, 27 ± 5 years) without a known history of concussion were studied. Heart rate (HR; ECG) and beat-to-beat blood pressure (BP; finger photoplethysmography) were recorded continuously during a five-minute supine rest period. Spontaneous cardiac BRS was estimated using the Sequence Method for overall BRS gains, then separately for up gains (increase systolic BP: increase R-R interval) and down gains (decrease systolic BP: decrease R-R interval). HRV was measured in the time-domain using the root mean square of successive differences between normal heartbeats (RMSSD), and in the frequency-domain using normalized low frequency (LF) and high frequency (HF) power, and as the LF/HF ratio. Results: There were no differences in resting BP or HR between groups (P > 0.050 for both). Overall cardiac BRS gains was also not different between controls (20.3 ± 4.3 ms/mmHg) and participants with TBI (28.6 ± 9.3 ms/mmHg, P = 0.4116). Likewise, we found no group differences for up and down gains (P > 0.050 for both). For HRV measures, no differences between groups were observed for RMSSD (control: 63 ± 20 ms; TBI: 63 ± 21 ms, P = 0.890), LF power (control: 40 ± 9 n.u.; TBI: 40 ± 14 n.u.; P > 0.999), HF power (control: 56 ± 9 n.u.; TBI: 57 ± 14 n.u.; P = 0.940), or the LF/HF ratio (control: 1.08 ± 0.50; TBI: 1.49 ± 0.85; P = 0.792). Conclusion: These preliminary findings suggest that, in contrast to our hypothesis, adults at six months post-concussion do not demonstrate impairments in any indices of cardiac autonomic function.

This project was supported by the Henry M. Jackson Foundation HU0001-18-2-0016.

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.
XX Sex Chromosome Complement in Female Mice is Obligatory for Vascular Function
Authors: Sakamuri Anil , Visniauskas Bruna , Kilanowski-Doroh Isabella , McNally Alexandra , Imulinde-Sugi Ariane , Lindsey Sarah , Ogola Benard
Published: May 2023

Biological sex plays an important role in cardiovascular disease (CVD) predisposition and incidence. Arterial stiffening assessed by pulse wave velocity is an independent marker for CVD and contributes to early vascular aging. Men have stiffer arteries than women, a sex difference that is diminished with aging. Sex hormones are indicated to play a role in stiffening arteries but the role for sex chromosomes is unknown. To assess the role for sex hormones, sex chromosomes or their interaction we used four core genotype (FCG) mice (N=5-8/group). 2-Way ANOVA and Sidak’s multiple comparison test was used for statistical analysis and P<0.05 was considered significant. In this mouse model, males (M) with testes and female (F) with ovaries have either XX or XY sex chromosome complement. Mice were either left gonadal intact or gonadectomized at 8 weeks followed by PWV assessment, blood pressure, and pressure myography at 16 weeks. In gonadal intact mice, males were significantly stiffer than females (P=0.002) with XYM (3.5 m/s) having greater PWV than XXM (3 m/s), XXF (2.8 m/s), and XYF (2.8 m/s), respectively without significant difference in blood pressure (BP). Gonadectomy increased PWV in all groups removing the sex difference in XYM (4.6 m/s), XXM (4.3 m/s), XXF (4.6 m/s), XYF (4.4 m/s) while BP remained unchanged. Pressure diameter relationship was greater in XYM than XXM (P<0.001) in both gonadal intact and gonadectomized mice. Aortic distensibility was greater in XXF than XYF (P=0.008) and XYM than XXM (P=<0.001). Stress-strain curves indicated significant leftward shift (P<0.05) XYM, XXM, and XYF mice after gonadectomy. Masson’s trichrome staining was increased (P<0.05) in mice with XX than XY sex chromosome complement after gonadectomy. In conclusion, we found that decline in sex hormones increases arterial stiffening, removes sex differences in distensibility, and shifts elastic modulus to the left in all groups but XXF mice.

R00HL155841 and R25HL105400

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.
The length-tension characteristics of small coronary arteries vary with transmural origin
Authors: Essajee Sal , Warne Cooper , Tucker Selina , Tune Johnathan , Dick Gregory
Published: May 2023

There are transmural differences in the structure of arteries across the left ventricular wall. For example, for arteries of the same size, wall thickness is greater in arteries of the epicardium than those from the endocardium. This observation suggests that there could be differences in their passive and active length-tension relationships, as different amounts of connective tissue or smooth muscle would be expected to alter these characteristics. We tested this hypothesis by studying similarly sized porcine coronary arteries from opposite transmural locations. Endocardial arteries had a diameter of 389 ± 33 μm (n = 8), while epicardial arteries measured 388 ± 50 μm (n = 6). A wire myograph was used to study the mechanical properties of these arteries under isometric conditions in Krebs-Henseleit buffer at 37 °C. Arteries were cut into rings with an axial length of 2 mm. Rings were repetitively stimulated to contract at increasing lengths with the addition of high extracellular K+ (80 mM). Coronary arteries developed active tension to a plateau level over approximately 3-5 min and K+-induced contractions readily washed out. Arteries from the epicardium were stiffer, as the passive-length tension curve of these vessels was elevated over arteries from the endocardium. Passive tensions at optimal length were 3.2 ± 0.4 vs. 5.6 ± 1.5 mN/mm (p < 0.05). The active tension developed in response to K+ depolarization was greater in epicardial arteries. Active tensions at optimal length were 3.4 ± 1.1 vs. 2.4 ± 0.3 mN/mm (p < 0.05). Our results represent the first comparison of transmural differences in coronary arteries under isometric tension. Our findings support the hypothesis that differences exist in the passive and active length-tension relationships of epicardial and endocardial arteries that correlate with wall thickness.

R01HL158723

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.
T3 Transection as a Mouse Model of Bowel Dysfunction after Spinal Cord Injury
Authors: Wireman Olivia , Hammers Gabrielle , Sams Ellie , Marino Felicia , Bailey William , Patel Samir , Gensel John
Published: May 2023

Bowel dysfunction, in the form of constipation and/or incontinence, is one of the most prevalent and life-impacting co-morbidities associated with spinal cord injury (SCI) with no long- term treatment available. The extent to which SCI-induced changes in the colon (e.g. inflammation, fibrosis, and reduced enteric neuron density) cause progressive dysfunction is understudied. There remains a significant unmet need to develop strategies to prevent or reverse bowel dysfunction after SCI. This need remains, in part, due to the lack of a mouse model which recapitulates the human condition. We hypothesized that a T3 spinal transection in mice would trigger bowel dysfunction alongside early colonic inflammation and downstream remodeling of the colon musculature as seen in both humans and rats after SCI. Bowel dysfunction was defined as increased number of fecal pellets within the colon, smaller pellet size, and decreased water content in the stool. The number of fecal pellets within the colon increased significantly in animals receiving SCI compared to sham (laminectomy only) injuries by 4 days post injury (dpi) and persisted to 21dpi. Furthermore, fecal pellet size was significantly decreased in the SCI animals at 21dpi and trended toward decreased water content. Together, these functional measurements are indicative of chronic bowel dysfunction after T3 transection. We next molecularly interrogated whole colon tissue for inflammatory cytokine levels and found increased levels of IFN-γ at 4 dpi but not at 21dpi and found increased collagen deposition in the colonic musculature at both 4 and 21 dpi in the SCI versus sham animals. The acute spike in INF-γ and deposition of collagen after injury are consistent with our hypothesis that T3 transection causes acute inflammation followed by remodeling of the colon wall. Establishing this mouse model will enable further interrogation of cell-type specific responses and signaling pathways using transgenic models.

T32 Fellowship in Physiology (GM118292) and T32 Fellowship Neurobiology of CNS Injury & Repair (5T32 NS077889)

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.
The role of Glycoprotein Nonmetastatic Melanoma Protein B in vascular inflammation during infectious challenge
Authors: Lamb Celia , Nguyen Hong-Ngan , Stark Ryan
Published: May 2023

Glycoprotein nonmetastatic melanoma protein B (GPNMB) is a transmembrane glycoprotein that has been shown to play an important role in inflammation. GPNMB levels are increased in association with pro-inflammatory conditions, such as lipopolysaccharide exposure. Further, GPNMB is increased in aged endothelial cells, and reduction of GPNMB in mice has been shown to reduce aging and increase lifespan. Thus, we sought to test the hypothesis that reduction in GPNMB would provide a protective phenotype to endothelial cells undergoing bacterial challenge. In order to test this, GPNMB was knocked down in either immortalized or primary human dermal microvascular endothelial cells (HMVECs) using siRNA. The cells were then treated with heat-killed Escherichia coli (HKEC, 3x108 cells/ml), one of the most common pathogens encountered in severe, systemic infections. With immortalized (i)HMVECs, siRNA-treated GPNMB knockdown produced cells appearing more compact and organized when exposed to HKEC as compared to siRNA controls. GPNMB knockdown iHMVECs also showed decreased IL-6 cytokine production after HKEC exposure (93.8 ± 8.1 pg/ml in siRNA controls versus 4.3 ± 0.03 pg/ml in siGPNMB, p < 0.005). Under similar conditions, primary (p)HMVECs exposure to siGPNMB showed no specific morphologic changes. Because GPNMB is also expressed by other cells, we also tested whether exogenous, secreted GPNMB from a non-endothelial source could impact HMVECs during infection. Interestingly, exogenous GPNMB exposure also significantly reduced pHMVEC IL-6 secretion (1372 ± 89 in control cells treated with HKEC versus 672 ± 36 in GPNMB treated cells followed by HKEC, p < 0.0001) during exposure to HKEC. Thus, our data suggests that endogenous GPNMB may be detrimental to endothelial cells depending on their age. Alternatively, exogenous GPNMB may offer protection to endothelial cells under certain conditions. Overall, the role of GPNMB in vascular health appears both age and source specific. Understanding the dynamic role of GPNMB may provide an opportunity to intervene in a GPNMB-specific manner to provide protection to endothelial cells during acute infectious challenge.

Grant: R35 GM138191

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.
Fibroblast growth factor-21 attenuates olanzapine induced hyperglycemia
Authors: Jeromson Stewart , Medak Kyle , Bellucci Annalaura , Arbeau Meagan , Wright David
Published: May 2023

Impaired glucose and lipid metabolism are known side effects of the anti-psychotic drug olanzapine. A single bolus of olanzapine induces a rapid rise in blood glucose, circulating fatty acids, and ketones. Strategies to offset these impairments in metabolism are of great interest. Our lab has demonstrated that interventions such as exercise, fasting, and ketogenic diets are effective at reducing olanzapine induced hyperglycemia. Fibroblast growth factor 21 (FGF-21) is a secreted protein that is induced by these interventions. We hypothesized that increased FGF-21 may be a common protective factor in reducing the hyperglycemic response to olanzapine. Our aim was to test if altered FGF-21 concentrations impacted the glycemic response to olanzapine. To increase FGF-21, male mice were fed a low-fat diet (10% Kcal) supplemented with Fenofibrate (0.2% w/w) or a matched control diet for 7 days prior to injection of Olanzapine or vehicle control. The fenofibrate diet significantly increased circulating FGF-21 concentrations (LFD: 744±404pg/ml, Fen: 3187±1050pg/ml, p=0.001, N=14). Olanzapine increased glycemia and circulating fatty acids but mice on the fenofibrate diet had an attenuated glycemic response to olanzapine and reduced circulating fatty acids compared to mice fed the control diet (Area under curve LFD: 1321±354 AU, Fen:1783±70 AU, P=0.0014, N=7). We also investigated if the loss of FGF-21 would lead to an increased glycemic response to olanzapine. In FGF-21 knockout mice, olanzapine induced a larger hyperglycemic response compared to olanzapine treated controls (Area under curve FGF-21 KO: 1788±280 AU, Con:1413±373 AU, p=0.00072, N=12). FGF-21 knockout, however, did not reduce the effect of olanzapine on serum non-esterified fatty acids or beta-hydroxybutyrate. These data show that FGF-21 may determine the extent of the hyperglycemic response to olanzapine and increasing FGF-21 may also offset the impairments in lipid metabolism.

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.
Beta-arrestin-biased protease-activated receptor-2 antagonists preferentially limit allergen-induced responses in vivo.
Authors: Schiff Hillary , Gillman Samuel , Pederson William , Dussor Gregory , Price Theodore , Vagner Josef , DeFea Kathryn , Boitano Scott
Published: May 2023

Biased signaling in G protein-coupled receptors (GPCRs) has emerged as a target for drug development. Protease-activated receptor-2 (PAR2) is a GPCR present in the airway epithelium with biased signaling that has been shown to trigger both detrimental effects [airway hyperresponsiveness (AHR), inflammation, mucus overproduction] and beneficial effects (bronchorelaxation) associated with allergen-induced asthma. These dual effects have been shown to be dictated by the two primary signaling pathways downstream of PAR2 activation: Gαq/Ca 2+ signaling is associated with bronchorelaxation, and β-arrestin/MAPK signaling is associated with AHR, inflammation and mucus overproduction. We have developed full (C391 which antagonizes Gαq/Ca 2+ and β-arrestin/MAPK signaling) and biased (C781 which antagonizes β-arrestin/MAPK signaling) antagonists of PAR2 and tested their efficacy in acute allergen-induced asthma mouse models. Both full and β-arrestin/MAPK signaling compounds attenuated AHR and inflammation in mouse models, and to a lesser extent, mucus overproduction. In a human bronchial model, the full PAR2 antagonist C391 prevented PAR2- dependent bronchial relaxation while the biased PAR2 antagonist C781 did not. Antagonism by C781 and other biased PAR2 antagonists can fine tune and effectively improve safety profiles for drug development in asthma treatment.

Grants from the National Institute of Health (NS098826, AI140257, HL16024, HL152942)

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.
Understanding the role of ZO-1 in cell phenotype and gene expression through myosin inhibition
Authors: Turbow Benjamin , Doran John , Uwaezuoke Brittany , Wilkinson Nancy , Peng Lindsey , Cheng Kwan , King Jonathan
Published: May 2023

Zonula occludens-1 (ZO-1) is a tight junction protein contributing to junctional formation as well as mediating interactions with junctional proteins and the actomyosin cytoskeleton. To investigate the role of ZO-1, Madin-Darby canine kidney (MDCK) II knockout cells lacking ZO-1 were created utilizing a CRISPR gene editing platform. These ZO-1 KO cells display characteristic phenotypic changes including actin accumulation and junction linearization. We evaluated membrane tension and found ZO-1 loss leads to increased membrane tension and nuclear density. We hypothesized that due changes in cytoskeletal organization and membrane tension the ZO-1 KO cells would spread less following plating. It is suspected that phenotypic changes related to actin accumulation may be the result of myosin dysregulation in the absence of ZO-1. ZO-1 KO cells that are exposed to blebbistatin, a myosin II inhibitor, resemble wild type cells in cell height and junction linearization. RNAseq and qRT-PCR were used to investigate gene expression changes related to ZO-1 loss to further interrogate ZO-1’s role in regulating membrane tension. This study highlights and expands our understanding of how ZO-1 regulates the mechanical properties of epithelial cells.

Trinity University, Mach Research Fellowship

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.
Investigating the contribution of cytokinesis to population-based measurements of epithelial Leak Pathway permeability
Authors: Monaco Ashley , Amsler Kurt
Published: May 2023

The mechanism mediating the paracellular movement of macromolecules across epithelial cell layers, the Leak Pathway, remains poorly understood. Two recent studies have demonstrated that epithelial cell cytokinesis allows the transient paracellular passage of macromolecules (Stephenson et al. 2019. Dev Cell 48:445-459, Richter et al. 2022. Ann NY Acad Sci 1516:151- 161). It is unclear, however, if this cytokinesis-mediated transepithelial passage of macromolecules can contribute significantly to the Leak Pathway permeability measured in a post-confluent epithelial cell population. The renal epithelial cell line, Madin Darby Canine Kidney (MDCK II), develops tight junctions and forms a typical paracellular permeability barrier upon reaching a confluent density. MDCK II cells have been reported to maintain a basal rate of cell proliferation, even in post-confluent cell populations. To investigate the role of cytokinesis in contributing to population-based measurements of Leak Pathway permeability, we examined the effect of arresting and releasing cytokinesis in post-confluent populations of MDCK II cells on Leak Pathway permeability. We confirmed that post-confluent populations of MDCK II cells exhibit a low level of cell proliferative activity. Leak Pathway permeability was determined by measuring the flux of 20 kDa fluorescein-dextran across post-confluent populations of MDCK II cells grown on permeable membrane filters. Inhibiting cytokinesis of MDCK II cells by overnight treatment with RO-3306, a CDK-1 inhibitor, decreased modestly the paracellular flux of 20 kDa fluorescein-dextran compared to untreated cell populations. Washing out RO-3306 1 hour prior to the flux assay produced an increase in paracellular flux of 20 kDa fluorescein-dextran to a level higher than that observed in untreated cell populations. Our results support the working hypothesis that cytokinesis can contribute to Leak Pathway permeability measured in epithelial cell populations. They suggest that more proliferative epithelial cell populations will exhibit higher rates of Leak Pathway permeability compared to epithelial cell populations with very low levels of proliferative activity. The significant Leak Pathway permeability measured in MDCK II cell populations maintained in the presence of RO-3306 suggests that cytokinesis is not the sole mechanism mediating Leak Pathway permeability across epithelial cell populations.

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.
TUDCA treatment restores anticontractile function of perivascular adipose tissue (PVAT) in post-weaning protein-restricted mice
Authors: Davel Ana , Freitas Israelle , Victorio Jamaira , Carneiro Everardo
Published: May 2023

Healthy perivascular adipose tissue (PVAT) has anticontractile, anti-inflammatory and antioxidant actions that can be disrupted by cardiometabolic risk factors. In the present study we hypothesized that protein restriction in the early phases of development could result in PVAT dysfunction and tauroursodeoxycholic acid (TUDCA) could be beneficial by inhibiting endoplasmic reticulum stress (ERS). To test this hypothesis, post-weaning male and female mice fed a normoprotein (NP, 14% protein) or a low protein (LP, 6% protein, isocaloric) diet for 15 weeks. In the last 2 weeks, some NP and LP animals received TUDCA (300 mg/kg/day) or vehicle. At the end of treatment, thoracic aorta was isolated to assess vascular responses with or without adjacent PVAT, as well as PVAT gene expression and histology. As results, the presence of PVAT reduced the phenylephrine-induced contraction in aorta of female NP and LP and male NP but not in male LP, suggesting LP-induced PVAT dysfunction in males only. In males, aortic PVAT from LP group showed less lipid content and increased collagen deposition. Decreased expression of PRDM-16, PPAR-γ, PGC-1α and leptin was observed in PVAT of LP group. These functional and morphological PVAT alterations in males were reversed by TUDCA treatment. Because TUDCA inhibits ERS, next we evaluated gene expression of ERS markers. There was an increased expression of GRP78, ATF4/6, IRE1α, and CHOP in PVAT from LP group, which was normalized by TUDCA. Also, TUDCA treatment normalized the reduced PVAT expression of the TUDCA receptor FXR in LP group. In conclusion, our data suggest that post-weaning protein-restriction impairs PVAT function and structure in males. These changes were associated with decreased expression of adipogenesis and adipocyte differentiation factors, and increased ERS markers. TUDCA reversed protein-restriction-induced PVAT abnormalities. Therefore, TUDCA emerges as a potential therapy for vascular complications associated with early undernutrition.

The authors have nothing to disclose.

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.
Primary Isolated Mouse Aortic Smooth Muscle Cells Phenotypic Characterization and Mitochondrial Respiration
Authors: Sakamuri Anil , Kamau Anne , Ogola Benard
Published: May 2023

Sexual dimorphism is demonstrated in cardiovascular disease (CVD). Vascular dysfunction is a critical player in CVD involving mitochondrial respiration in aortic smooth muscle cells. In this study, we characterized contractile and synthetic gene expression, and mitochondrial respiration by sex in aortic smooth muscle cells (ASMC). We hypothesize that female than male ASMC have increased contractile genes and better mitochondrial energetics. Adult male and female C57BL6J mice were euthanized, and thoracic aorta excised. Aortic tissue sections were subject to enzymatic digestion and cultured for subsequent experiments. The 2-delta delta CT method using beta actin as housekeeping gene was used for analysis. For mitochondrial respiration, confluent (70%) cells were trypsinized, enumerated, and seeded in Seahorse mini-culture plates (20,000 cells/well). After overnight incubation mitochondrial stress test was performed to measure basal respiration, maximal respiration, spare capacity, ATP production, proton leak, and non-mitochondrial respiration. All mitochondrial respiratory parameters were expressed as oxygen consumption rate normalized with protein levels (OCR, picomoles/L of O2 per microgram protein, n=8-10 wells/group). Unpaired t-test was used for statistical analysis and P<0.05 was considered significant. To characterize the sex of isolated cells (n=6-8/group), we used sex specific primers that showed significantly higher Sry (P<0.001) and Xist (P<0.001) gene expression of male and female ASMC, respectively. Next, we assessed contractile and synthetic genes including myocardin, alpha actin, and myosin heavy chain 11 that were higher in female than male ASMC (P<0.05). However, myosin heavy chain 10 was higher (P=0.002) in male than female cells. The basal respiration and ATP production were significantly high in female ASMC by 79% and 77%, respectively, when compared with male ASMC (2.48 ± 0.13 vs. 1.38 ± 0.15, P= 0.0001; 2.11 ± 0.11 vs. 1.19 ± 0.11, P= 0.0001). Female cells indicated increased spare respiratory capacity compared to male ASMC (1.67 ± 0.31 vs. 1.1 ± 0.23, p= 0.01). The maximal respiration and proton leak were significantly higher in female than male cells by 69% and 109% (4.21 ± 0.33 vs. 2.49 ± 0.31, P= 0.001: 0.42 ± 0.02 vs. 0.2 ± 0.04, P= 0.001). Female and male cells did not show any significant difference in non-mitochondrial respiration. Based on these results, female than male ASMC exhibit increased contractile gene expression and better mitochondrial energetics at basal and stressed conditions. The sex-specific differences in mitochondrial function may attribute to sex bias in CVD.

R00HL155841 and R25HL105400

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.
High-resolution temporal transcriptomic profiling of the distal convoluted tubule identifies an early adaptive response to furosemide
Authors: Jung Hyun Jun , Tahaei Ebrahim , Welling Paul
Published: May 2023

The distal convoluted tubule (DCT) undergoes hypertrophy and hyperplasia to increase salt absorption in response to physiologic and pathological stresses, and has been implicated in the development of loop-diuretic resistance. However, the molecular mechanisms underlying DCT remodeling are unknown. We investigated progressive changes in the transcriptome of the early DCT (DCT1) in a time series (12, 24, 48, and 96 hours) after the administration of furosemide to identify initiating gene regulatory pathways. For high-resolution coverage, RNAseq was performed on isolated DCT1 segments, identified using a DCT1-specific Cre-driven fluorescent protein, tdTomato. An average of 17 million sequences were read per sample, providing genome-wide coverage of the DCT transcriptome (15499 genes). Sequence reads aligned on the mouse transcripts were quantified using Salmon, and differential gene expression analysis was performed using edgeR. All samples were enriched with known DCT-specific genes and lacked other nephron segment specific markers, indicating uniform purification of DCT tubules. Pathway and pattern clustering analysis of the time-resolved DCT transcriptome profiles revealed the early-induced genes were highly enriched in gene sets associated with cell cycle progression and cell proliferation. Meta-analysis of transcription factor binding enrichment at promoters of the early-induced genes revealed enrichment of genomic binding sites of Hypoxia-inducible factor 1 (HIF-1) signaling components (HIF1A, STAT3, RELA) across the early-induced gene promoters. Immunoblot analysis in kidney cortical tissues confirmed rapid activation of the HIF-1 signaling components following furosemide treatment. In summary, a core transcriptional regulatory axis in the DCT is activated rapidly after loop-diuretic treatment to drive a program of cell proliferation in the DCT. The results identify HIF1 signaling as a proximal pathway in the initiation of proliferative DCT remodeling.

NIDDK, LeDucq Foundation

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.
Sex-specific impact of angiotensin II on arterial pressure and LV mass during angiotensin converting enzyme inhibition in spontaneously hypertensive rats
Authors: Floyd Dana , Garvin Alexandra , Hale Taben
Published: May 2023

Sex differences in arterial pressure responses to angiotensin II (AngII) have been noted, particularly in normotensive animals. In addition, sex-specific changes in renin angiotensin system (RAS) components have been implicated in differential regulation of arterial pressure between males and females. The present study evaluated the sex-specific responses to AngII infusion, angiotensin converting enzyme inhibitor (ACEi) treatment, and AngII infusion during ACEi treatment on mean arterial pressure (MAP) and left ventricular hypertrophy (LVH) in spontaneously hypertensive rats (SHR). Young adult male and female SHR were treated with the ACEi, enalapril (30mg/kg per day, p.o.) alone or in combination with AngII at a low (200 ng/kg per min) or high (400 ng/kg per min) dose for two weeks. MAP was measured by radiotelemetry and data presented as percent change from pre-treatment baseline. At time of sacrifice, left ventricles were excised and weighed and normalized to body weight. ACEi induced equivalent percent reductions in MAP between male (-21 ± 1.2%) and female (-23 ± 1.9%) SHR. AngII (high) infusion significantly increased MAP, with a tendency toward a greater effect in male compared to female SHR (male: 29 ± 4.6% vs. female: 17 ± 3.6%, p=0.09). During ACEi, low dose Ang II did not significantly impact the antihypertensive effect of ACEi in males or females. However, administration of the high dose of AngII during ACEi resulted in a marked sex difference whereby the male SHR exhibited an increase in arterial pressure (ACEi: -21.22% ± 1.21, ACEi+AngII high: +39.49% ± 1.39, p<0.05), while in females AngII had no impact on the depressor response to ACEi alone (ACEi: -22.59% ± 1.89, ACEi+AngII high: -18.14% ± 0.88, p>0.05). Evaluation of left ventricular hypertrophy (LVH) revealed a similar sex difference. The low dose of AngII prevented the reversal of LVH induced by ACEi in male (Con: 2.54 ± 0.09, ACEi: 1.99 ± 0.04*, ACEi+AngII: 2.34 ± 0.13†, *p<0.05 vs. Con, †p<0.05 vs. ACEi), but not female rats (Con: 2.95 ± 0.02, ACEi: 2.43 ± 0.04*, ACEi+AngII: 2.46 ± 0.09*, *p<0.05 vs. Con). In summary, ACEi produces equivalent reductions in arterial pressure and LVH in male and female SHR when the endogenous RAS is intact. However, inhibition of the RAS reveals a marked sex difference in the responsiveness to AngII both with respect to arterial pressure and cardiac hypertrophic responses. Gaining a greater understanding of sex differences in blood pressure control will ultimately lead to improved antihypertensive cardioprotective therapies for women.

R01HL153112-02

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.
Social Anxiety, Academic Self-Efficacy, and Course Performance in Pre-Nursing Students Enrolled in Human Anatomy and Physiology I and II
Authors: Flotte Elizabeth
Published: May 2023

This study investigates the effects of social anxiety and academic self-efficacy on course structure preference, exam scores, and final grades. The study included 25 students enrolled in accelerated Human Anatomy and Physiology I and II college courses offered consecutively within the Fall 2022 semester. Students were surveyed three times throughout the semester and responses regarding anxiety, self-efficacy, and preference for course structure (lecture-based or active learning instruction) were compared to academic performance. Levels of social anxiety did not seem to affect overall course structure preference or course performance, although students indicated a variety of course activities that cause them anxiety. Students with higher levels of academic self-efficacy, however, displayed increased academic performance. The relationship between academic self-efficacy and performance was strongest on lecture exams with the greatest emphasis on physiology. This study supports the importance of building self-efficacy for success in courses covering complex topics and demonstrates a need to better understand the role of student anxiety.

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.
Loss of tissue inhibitor of proteinase 3 (TIMP3), but not TIMP4, worsens abdominal and thoracic aortic aneurysm
Authors: Hu Mei , Hu Mei , Meganathan Ilamaran , Kassiri Zamaneh
Published: May 2023

Background: Aorta exhibits regional heterogeneity (structural and mechanical), while different etiologies for thoracic and abdominal aortic aneurysm (TAA, AAA) are recognized. Tissue inhibitor of metalloproteinases (TIMPs) regulate vascular remodeling through different mechanisms. TIMP3 and TIMP4 have shown region-dependent functions in aortic pathologies. We investigated the region-specific function of these TIMPs in development of TAA versus AAA. Methods and Results: TAA or AAA was induced in male or female mice lacking TIMP3 ( Timp3-/-), TIMP4 ( Timp4-/-) or wildtype (WT) mice by peri-adventitial elastase application. Loss of TIMP3 exacerbated TAA and AAA severity in males and females. Timp3-/- mice exhibited a greater increase in proteinase activity, smooth muscle cell phenotypic switching post-AAA and -TAA whereas inflammation increased in the media post-AAA, but in the adventitia post-TAA. Timp3-/- mice showed impaired intimal barrier integrity ( in vivo) following AAA, but a greater adventitial vasa-vasorum branching post-TAA, which could explain the site of inflammation in TAA versus AAA. Timp4-/- mice responded similarly to TAA and AAA induction as the WT. In vitro, Timp3 knockdown in human aortic EC more severely compromised the EC monolayer permeability compared to Timp4 knockdown and control groups following elastase treatment. In human aorta specimens, TIMP3 showed a region-specific expression pattern in TAA versus AAA consistent with the impact of its loss in the impaired intimal barrier in AAA, and the adventitial region in TAA. Conclusions: TIMP3 plays a protective role in both AAA and TAA, whereas loss of TIMP4 has minimal impact on the severity of aneurysm formation. TIMP3 loss in the aneurysmal aorta could be responsible for the inflammation, adverse remodeling and aortic dilation in AAA and TAA, through different mechanisms. Support: CIHR, Heart and Stroke.

CIHR, Heart and Stroke.

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.