bluuebunny/crossref_metadata_2025 huggingface dataset:
Exosomes mediate immune activation post-myocardial infarction
Authors: Dasari Siddarth , Asalla Suman , Kumar Vinay , Bansal Shyam
Published: May 2023

Myocardial infarction (MI) and ischemic heart failure (HF) are associated with immune activation in the bone-marrow and the lymphoid tissues. Our studies also show that spleens lose 58% of their weight by 1-day post-MI as compared to sham-operation. Flow cytometric analysis further showed that, as compared to sham-operated mice, spleens of 1d post-MI mice contained 2-3-fold lower numbers of leukocytes (16.9X106 vs. 47.7X106), CD11c+ dendritic cells (DCs; 0.22X106 vs. 0.56X106), Ly6G+ neutrophils (0.45X106 vs. 1.08X106), CD11b+ myeloid cells (1.2X106 vs. 2.8X106), and Ly6C+ monocytes (0.4X106 vs. 1.0X106). Activation of antigen-presenting innate immune cells would also suggest activation of adaptive immune responses. Thus, using flow cytometry we also measured CD4+ and CD8+ T-cells in the spleens of 1d MI and sham mice and found that both of these cell types (2.2X106 vs. 5.7X106 for CD4+ T-cells and 1.5X106 vs. 3.8X106 for CD8+ cytotoxic T-cells) were also decreased. These data suggest that damage-associated signals from the infarcted heart are communicated to the splenic innate and adaptive immune cells leading to their efflux and trafficking to the heart. However, the communication vehicles that relay these damage-associated signals from the heart to the spleen are unknown. We further hypothesized that these signals are carried by the exosomes, small (100-200 nm) extracellular vesicles released by all cells. To test our hypothesis, we isolated exosomes from the plasma of 1d MI and sham mice, injected them into the tail-veins of naïve mice and using echocardiography measured their cardiac function at 8 weeks post-injection to assess if they developed HF-like systolic dysfunction. We found that, at 8 weeks, mice injected with 1d MI exosomes developed significant systolic dysfunction with increased end-systolic and end-diastolic volumes, and decreased ejection fraction. These changes were not seen in mice injected with 1d sham exosomes. Further, using flow cytometry we also measured different splenic immune cells in the recipient mice. Our data show that spleens of mice injected with 1d MI exosomes, as compared to 1d sham exosomes, contained higher numbers of activated immune cells and showed increased levels of proinflammatory monocytes (without any changes in anti-inflammatory monocytes), CD11c+ conventional and monocyte-derived DCs and helper T-cell subsets such as T-regs, Th1, Th2, Th17. Our results suggest that exosomal signals could be a novel potential mechanism by which cardiac injury is communicated to the lymphoid tissues to affect immune cell recruitment following MI.

This work was supported by the National Institutes of Health grants to. Dr Shyam S. Bansal (R00 HL132123 and R01 HL153164).

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.
Endothelial Nrf2 modulates blood pressure and vasoconstrictor responses in conscious mice
Authors: Dhyani Neha , Rudebush Tara , Gao Li , Zucker Irving
Published: May 2023

Introduction: Excessive oxidant stress has been shown to mediate a variety of cardiovascular pathological conditions, including hypertension. The level of reactive oxygen species (ROS) in tissues is regulated by the balance between the production of ROS and the ability to scavenge superoxide and hydrogen peroxide by a myriad of antioxidant enzymes and small molecules. One of the most important regulators of antioxidant activity is the ubiquitous transcription factor Nuclear factor erythroid 2-related factor 2 (Nrf2). Nrf2 binds to antioxidant response elements of multiple genes following its release from its inhibitory protein Keap1 and part of the E3 ubiquitin ligase complex. In this study we hypothesized that endothelial Nrf2 is a critical factor in blood pressure regulation following vasoconstrictor stimuli. Methods: Transgenic mice were created using the endothelial Tie2 promoter expressing Cre recombinase (Tie2-Cre) crossed with either Nrf2 floxed or Keap1 floxed mice so that Nrf2 would be deleted or upregulated respectively, and selectively in the endothelium. Cre negative mice were used as wild type (WT) controls. Mice were instrumented with radiotelemetry transducers to chronically record arterial blood pressure and heart rate in the conscious state. Blood pressure and heart rate were recorded for 24 hours once per week. After baseline recordings, animals were challenged with an osmotic minipump infusion of Angiotensin II (Ang II) (600ng/Kg/min) for 7 days or L-nitroarginine methyl ester (L-NAME) (100 mg/kg/d in drinking water). Results: Baseline mean arterial pressure (MAP) was 101.6 ± 2.5 mm Hg (mean±SEM; n=7) in WT mice (daytime =95.4±1.5; nighttime =106.1±1.0). In mice with endothelial Nrf2 deletion baseline MAP was significantly higher(p<0.01) during nighttime (112.1±1.1). In addition, the response to Ang II was significantly augmented (p < 0.001) in Nrf2 KO mice compared to WT mice (148.9±5.3 vs 116.8±6.7 mm Hg). Both day and nighttime values were higher in Nrf2 KO mice. Furthermore, Nrf2 KO mice exhibited a delayed return to baseline upon pump emptying. In contrast, in one Keap1 KO mouse there was a response to Ang II that was similar to WT (92.1 mm Hg baseline to 116.3 mmHg post Ang II). In Nrf2 deleted mice, the hypertensive response to L-NAME was no different than that of WT mice. Conclusion: In summary, these preliminary data suggest that endothelial Nrf2 plays an important role in modulation of blood pressure at rest and during oxidative stress induced by Ang II. The L-NAME data suggests that upregulation of Nrf2 increases the bioavailability of nitric oxide to blunt Ang II mediated vasoconstriction.

Supported by NHLBI P01-62222

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.
Wheat Germ Supplementation Attenuates Obesity-Induced Neuroinflammation in the RVLM of IL-10 KO mice
Authors: Di Agostino Ashley , Plakkot Bhuvana , Monteiro Raisa , Sivasubramanian Mahesh , Alake Sanmi , Lucas Edralin , Subramanian Madhan
Published: May 2023

Obesity is known to be a major factor in the increase of cardiovascular risk and inflammatory responses in the brain. It has been found that high-fat diet (HFD)-induced obesity increases sympathetic nerve activity (SNA) through a cascade of chronic metabolic processes such as an increase in oxidative stress and inflammation. The rostral ventrolateral medulla (RVLM) is a specific part of the brainstem that regulates SNA. Wheat germ (WG), a rich source of bioactive compounds including polyunsaturated fatty acid, has been shown to have an anti-inflammatory effect on the gut environment of HFD-fed mice. A healthy gut microbiota has been shown to have a regulatory effect on the gut-brain axis, especially under stress. In this study, we investigated the effects of WG supplementation on RVLM inflammation in high-fat high cholesterol diet (HFHC)-fed IL-10 KO mice. Six-week-old female C57BL/6 (WT) and IL-10 KO mice ( n = 4-5/group) were randomly assigned to treatment groups: WT and IL-10 KO fed a control diet (10% fat kcal), IL-10 KO fed HFHC (43.4% fat kcal, 1% cholesterol) diet with or without 10% WG for 12 weeks. At the end of 12 wks of treatment, animals were sacrificed and RVLM mRNA expression of proinflammatory cytokines and chemokine were analyzed using qPCR. HFHC increased the expression of IL1b, TNFa, and MCP-1 in the RVLM. WG supplementation was able to reverse the mRNA expression of IL1b (p<0.05), TNFa (p<0.05), and MCP1 (p<0.05). These results indicate that HFHC diet-induced obesity increases inflammation in the RVLM and WG supplementation could reverse these effects. Future studies would focus on understanding the mechanisms by which WG reverses the inflammation in the brain.

NIH R15HL148844, AHA AIREA959725, and Oklahoma Agriculture Experiment Station 2153907

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.
Oral Administration of Amiloride Does Not Affect Spontaneous Cardiac Baroreflex Sensitivity in Humans
Authors: Fernandes Igor , Sabino-Carvalho Jeann , Mattos Joao , Campos Monique , Mansur Daniel , Vianna Lauro , Nobrega Antonio
Published: May 2023

Pre-clinical models suggested that administration of Amiloride at the carotid sinus antagonizes the acid-sensing ion channels (ASICS), an essential component of the mechanoelectrical transducer in the arterial baroreceptors, reducing cardiac baroreflex sensitivity and perturbing the homeostatic beat-to-beat blood pressure (BP) regulation. However, it is still undetermined whether these findings could be translated into humans. Therefore, we determined whether oral administration of Amiloride reduces the spontaneous cardiac baroreflex (cBRS) sensitivity and perturbs BP regulation in humans. Heart rate (HR; electrocardiography) and beat-to-beat BP (finger photoplethysmography) were continuously measured in 5 healthy men (32 ± 6 yrs) during at least 5 min in two different conditions: (1) Control (CTL) and (2) 3 h after of oral administration of Amiloride (AMD – 5 mg + 50 mg of hydrochlorothiazide), a non-selective antagonist of ASICs. Spontaneous cardiac BRS was determined for all sequences (three or more consecutive beats with corresponding increases or decreases in systolic BP and R-R interval) combined (cBRSall) and separately for up (cBRSup – increases in BP and R-R interval) and down (cBRSdown – decreases in BP and R-R interval) sequences. HR (AMD 65 ± 7 vs. CLT 63 ± 12 beats/min, P = 0.543), mean (AMD 88 ± 9 vs. CLT 88 ± 7 mmHg, P = 0.687), and systolic BP (AMD 126 ± 14 vs. 128 ± 11 mmHg, P = 0.717) were similar in both conditions. Likewise, spontaneous cBRSall (AMD 20 ± 9 vs. CTL 22 ± 6 ms/mmHg, P = 0.543), cBRSup (AMD 19 ± 10 vs. CTL 22 ± 6 ms/mmHg, P = 0.495,) and cBRSdown (AMD 21 ± 9 vs. CTL 21 ± 8 ms/mmHg, P = 0.937) were preserved after AMD administration. In summary, our preliminary results demonstrated that the non-selective antagonism of ASICS via oral administration of AMD did not affect spontaneous cardiac baroreflex sensitivity and BP regulation in humans.

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.
Parental obesity exacerbates immunometabolism and renal dysfunction in offspring exposed to acute kidney injury
Authors: do Carmo Jussara , Mouton Alan , Aitken Nikaela , Wang Zhen , Dai Xuemei , Frierson Timia , Walker Xavier , Oppedal Kari , Larson kaylie , Omoto Ana , Li Xuan , Hall John , da Silva Alexandre
Published: May 2023

Kidney disease is estimated to occur in 37 million people in the US, approximately 15% of the adult population. Obesity is a major driver of kidney disease via mechanisms that are still unclear, and pre-conception obesity in the parents may program kidney dysfunction and inflammation in their offspring. Using a model of parental obesity where male and female mice are fed high fat (H) diet prior to mating, with dams maintained on the H diet during gestation and lactation, we found that male offspring exhibit hypertension and initial signs of kidney injury in adulthood (24 weeks of age). We examined whether offspring from obese parents exhibit worse outcome after acute kidney injury (AKI), and whether this is associated with an immunometabolic shift in macrophages (Mɸ) toward glycolytic and pro-inflammatory phenotypes, as assessed by flow cytometry and Seahorse extracellular flux analysis to measure extracellular acidification rate (ECAR) as a measure of glycolysis. The following groups of male mice were submitted to bilateral renal AKI by clamping the left and right pedicles for 30 min: 1) lean offspring fed normal (N) diet born from N diet lean parents (NN, n=5); 2) obese offspring fed H diet born from N diet parents (NH, n=6); 3) lean offspring fed N diet born from H diet obese parents (HN, n=6); and 4) obese offspring fed H diet born from H diet obese parents (HH, n=7). Compared to NH and NN offspring from lean parents, HH and HN offspring from obese parents exhibited worse kidney dysfunction with higher urinary renal osteopontin (OPN) levels (93.8±8.7 and 73.7±16.6 vs. 213.8±24.0 and 151.1±24.6 μg/24h). We also found increased CD45+ Mɸ-like cells which play a role in initiating and maintaining kidney inflammation, as well as worse kidney injury score, increased tubular cell death, tubular blood clot, and glomerular tubularization in offspring from obese parents. Additionally, HH and HN offspring exhibited expansion of renal M1 Mɸ phenotype 4 days after AKI assessed by ECAR (glycolysis: 5.9±0.8 and 4.1±0.5 vs. 9.5±0.9 and 13.5±0.8 ΔmpH/min). Thus, parental obesity is associated with a shift towards M1 inflammatory Mɸ phenotype and increased risk of worse renal dysfunction in their offspring after AKI, even when offspring remain lean.

NIDDK 1R01121411, NIGMS P20GM104357 and NIGMS U54GM115428)

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.
CAMP stimulated NKCC2 phosphorylation at Thr-96,101 is in part independent from SPAK: a Role for TNIK
Authors: Granados Jessica , Caceres Paulo S. , Ortiz Pablo
Published: May 2023

Abnormally enhanced NaCl reabsorption by the thick ascending limb of the loop of Henle (TAL) contributes to the development of salt-sensitivity. NaCl reabsorption by the TAL is mediated by the apical Na/K/2Cl cotransporter NKCC2. NKCC2 activity can be stimulated by phosphorylation at Thr 96,101 by upstream kinases SPAK or OSR1. We found that Thr 96,101 phosphorylation is enhanced in TALs of Dahl Salt Sensitive rats (Dahl SS) on normal or high salt diet. Genetic deletion of SPAK in Dahl SS lowers NKCC2 phosphorylation by 40% and blunts salt-sensitive hypertension but does not completely restore these to baseline. Other kinases may be involved in NKCC2 phosphorylation. Using a targeted proteomics approach, we identified TNIK (Traf2 and NCK interacting kinase) as a kinase that binds and phosphorylates Thr 96,101 in NKCC2 in rats and mice (manuscript under revision). In addition, we and others found that cAMP is a potent stimulus for NKCC2 Thr-96,101 phosphorylation. We hypothesize that TNIK is in part responsible for cAMP-stimulated NKCC2 phosphorylation at Thr-96,101. First, we tested the ability of a novel TNIK inhibitor (NCB-0846) to decrease baseline NKCC2 phosphorylation in Dahl SS rats. Treating TALs from Dahl SS with NCB-0846 (0.1 μM) for 25 min decreased NKCC2 Thr 96,101 phosphorylation by 21 ± 2% compared to the vehicle group (p<0.0001, n=3). Next, we measured NKCC2 phosphorylation in rats with genetic deletion of SPAK in a Dahl SS background. In SPAK KO rats, the cAMP analogue db-cAMP (500 μM) increased NKCC2 phosphorylation at Thr 96,101 by 626 ± 46% (p<0.0001, n=3) and the TNIK inhibitor NCB-0846, blunted cAMP-stimulated NKCC2 phosphorylation by 56 ± 4.3% (p=0.0013, n=3). There was no significant decrease in phosphorylation at phospho-Ser 126 NKCC2 (cAMP=46 ± 7-fold vs cAMP+NCB= 31±5-fold, n.s). These data indicate that in the absence of SPAK, TNIK mediates cAMP-stimulated NKCC2 phosphorylation at Thr-96,101. To further support this, we studied TALs from whole animal TNIK knock-out mice (TNIK KO). db-cAMP (500 μM) increased NKCC2 phosphorylation by 1330 ± 268% in wild-type mice (p<0.01) whereas cAMP-stimulated NKCC2 phosphorylation was blunted by 49 ± 15% in TNIK KO TALs (p<0.05, n=3). No significant difference was found in cAMP-stimulated NKCC2-Ser 126 phosphorylation in TNIK KO (WT=23 ± 12-fold vs TNIK KO=15 ± 4-fold, n=3). We conclude that enhanced NKCC2 phosphorylation at baseline in Dahl SS TALs is in part caused by TNIK and that cAMP-stimulated NKCC2 Thr-96,101 phosphorylation in TALs is in part independent from SPAK and involves TNIK. Our data point to potential role for TNIK in renal ion transport, blood pressure regulation and urine concentration.

AHA15GRNT25710369

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.
Differential Response to Whole-Body Metabolic vs. Respiratory Acidosis in Angiotensin Receptor Type 1A Knockout Mice
Authors: Gilker Eva , Xu Kui , Chen Xiaoqin , Boron Walter , Moss Fraser
Published: May 2023

Disturbances in the homeostatic mechanisms that control acid base status lead to alterations in arterial pH (pHa) and in turn cause cell dysfunction and therefore have devastating effects on whole-body physiology. Many major acid-base transporter proteins have been identified; however, there is only a rudimentary understanding of how cells sense changes in acid-base status and the signaling pathways that lead to a response. The CO2/HCO3 − buffering system is in equilibrium with pH, thus changes in [CO2] or [HCO3 −] will lead to disturbances in acid-base homeostasis. The respiratory and renal systems modulate the [CO2] and [HCO3 −] respectively, working in tandem to maintain pHa between 7.35-7.45 and compensatory mechanisms exist to defend against acid-loads. For example, during respiratory acidosis (RAc) when there is an elevation in PCO2 causing a drop in pH, the kidneys (primarily the proximal tubules) will increase HCO3 − reabsorption and generation in an effort to rescue the pHa. Similarly, during metabolic acidosis (MAc) when an acid load is brought on by a decrease in [HCO3 −], the respiratory system will decrease [CO2] to bring pHa back to a physiological level. Using isolated perfused proximal tubules (PTs), our lab has established that changes in [CO2] at the basolateral membrane evoked a change in H+ secretion ( J H) that required endogenously secreted ANGII to bind angiotensin receptor type 1a (AT1A), at the apical membrane. In addition, pharmacological block of apical AT1A or knockout (KO) of the gene lowers baseline J H and eliminates the increased J H in response to elevated [CO2]. Here, we aim to determine the whole-body response to both RAc and MAc in wild-type (WT) vs global AT1A KO animals by monitoring arterial blood gases and urinary acid excretion. Our results indicate that during induced MAc (1% NH4Cl in drinking water), AT1A KO mice present a more pronounced pHa acidification than WTs after 2 days, but are able to recover after 7 days of exposure. On the contrary, when we expose animals to RAc (8% inspired CO2), there is no difference between WTs or AT1A KOs, indicating that the AT1A receptor may be important early in the defense against MAc, but may play little to no role in the defense against RAc.

R01-DK113197 and R01-DK128315

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.
Lack of Kidney Injury Sex Differences in Unilateral Ureter Obstructed Mice
Authors: Goorani Samaneh , Goorani Samaneh , Imig John
Published: May 2023

Background: Renal fibrosis is a critical pathophysiological event in chronic kidney disease. Unilateral ureteral obstruction (UUO) is a well-established and widely used model for investigating renal fibrosis. Because the UUO surgery is difficult to conduct in female mice, there are limited studies to evaluate UUO kidney injury sex differences in mice.The goal of this experimental study is to compare renal injury caused by UUO in wild-type and renal hypoplasia low nephron number mice. Method: Male and female C57Bl/J6 wild-type (WT) and ROP/Le- Os Ces1ca/+ Ces1ca/J oligosyndactyl (ROP Os/+) mice were subjected to UUO or sham surgery, and kidney tissue collected at the end of the 10-day experimental period. Kidney histological analysis and mRNA expression determined renal fibrosis, tubular injury, collagen deposition, extracellular matrix proteins, and immune cell infiltration. Results: Our data demonstrated marked renal injury, fibrosis, and extracellular matrix production in the kidneys of male and female UUO WT and ROP Os/+mice. Renal cortical fibrosis was 16.8 ± 2.3% in male and 17.3 ± 1.6% in female UUO WT mice. ROP Os/+ UUO kidney cortical fibrosis averaged 9.1 ± 1.6% in male and 10.3 ± 1.3% in female mice. Kidney immunohistochemical staining for α-smooth muscle actin (a-SMA) averaged 4.7 ± 0.8% in male and 4.1 ± 0.5% in female UUO WT mice and 3.1 ± 0.4% in male and 2.8 ± 0.3% in female UUO ROP Os/+ mice. Kidney mRNA expression of a-SMA, fibronectin (Fn1), and ZEB-1 was increased to a similar degree in male and female WT and ROP Os/+ mice following UUO. UUO increased renal expression of a-SMA, Fn1, and ZEB-1 to a greater degree in UUO ROP Os/+ compared to UUO WT mice. Kidney immunohistochemical analysis of MAC2 was conducted in male and female WT and ROP Os/+ mice following UUO. Renal MAC2 levels were increased to a similar extent in male and female UUO WT, and male and female UUO ROP Os/+ mice compared to their respective sham operated control groups. Interestingly, renal inflammation phenotypes following UUO were 30-45% greater in WT compared ROP Os/+ mice. Summary: These findings demonstrate UUO induced kidney fibrotic injury did not exhibit sex difference in WT or ROP Os/+ mice.

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.
A potential role for GPR107 in hypoglycemia unawareness
Authors: Hagena Lisa , Ennis Teresa , Kolar Grant , Samson Willis , Yosten Gina
Published: May 2023

Hypoglycemia unawareness (HU) is a major barrier to achieving glycemic control in diabetic patients and remains a severe complication for type 1 and many type 2 diabetics. Due to the limited availability of animal models and significant variations between existing models, our understanding is limited regarding the cellular mechanisms and neural pathways responsible for this loss of metabolic control in the setting of recurrent hypoglycemia. Our group has recently identified GPR107 as a receptor for neuronostatin (NST), a 13 amino acid peptide derived from the somatostatin prepro-hormone. In our novel rat model of HU, we show that GPR107 expression is dramatically increased in the hypothalamus of rats exhibiting loss of hypoglycemia-induced food intake. Using a GPR107-flox rat model, we evaluated the effect of loss of GPR107 in the hypothalamus on the development of HU. Our preliminary data indicate that knockdown of hypothalamic GPR107 protects rats against HU induced by exposure to recurrent hypoglycemia. Future work will focus on the mechanisms underlying the role of GPR107 in the development of this dangerous clinical entity.

National Institute of Health (NIH)

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.
Defining the role of endoplasmic reticulum in the initiation of paligenosis
Authors: Nguyen Thanh , Cho Charles , Radyk Megan , Mills Jason
Published: May 2023

Reprogramming is crucial for cellular renewal in adult organs that lack dedicated stem cells for replacement of cell loss after injury. Because such cell plasticity is likely to be executed by a conserved cellular program, we have begun to identify the conserved cellular-molecular features of the process in which differentiated cells are recruited as progenitors. We have characterized three successive stages by which this occur with differentiated cells: 1) the cells downscale their organelle contents, 2) cells activate a progenitor-like gene network, and 3) cells reenter the cell cycle. We have given this conserved process the name paligenosis. Here, we investigate upstream triggers of paligenosis. Using a high-dose tamoxifen (HD-TAM) injury model to induce paligenosis in zymogenic chief cells of the murine stomach corpus, we observed ultrastructural changes in the endoplasmic reticulum (ER) during paligenosis initiation (e.g., swelling of ER lamellae, liberation of ribosomes from rough ER, and overall loss of ER). This led us to hypothesize that dynamic changes in ER were an upstream event in paligenosis. ER functioning is in part monitored by the Integrated Stress Response (ISR) with the paramount ER stress sensor being PERK, which then autophosphorylates and in turn phosphorylates the α subunit of the eukaryotic initiation factor 2 (eIF2α). Phospho-eIF2α halts global translation while upregulating a specific set of genes to restore homeostasis. Here, we go on to show that HD-TAM activates the ISR in paligenotic gastric chief cells, triggering global attenuation of protein synthesis. However, this “ER stress” ISR pathway must occur in a critical, temporally limited window early in paligenosis, as pharmaceutically-induced prolonged ER stress caused paligenosis to halt in early stages with cells failing to proliferate. Blocking the ISR also halted paligenosis in early stages. Ongoing experiments include the use of the glycosylation-inhibiting drug tunicamycin to determine if inducing the ISR via ER stress is sufficient to initiate paligenosis; and deleting PERK in chief cells during paligenosis using Perkflox/flox; Mist1CreERT2 mice. Our results provide initial insights on the dynamic functionality of the ER and its role and necessity during cellular regeneration.

This work was supported by funding from the NIH through the National Institute of Diabetes and Digestive and Kidney Diseases and the National Cancer Institute.

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.
Blockade of leptin receptor (LepR) in the Carotid Body area attenuated obesity-induced Hypertension
Authors: Shin Mi Kyung , Kim Lenise , Tang Wan-Yee , Polotsky Vsevolod
Published: May 2023

RATIONALE: Obesity leads to high cardiovascular morbidity and mortality via multiple mechanisms including obstructive sleep apnea (OSA) and hypertension. Obesity leads to high levels of leptin which is produced in adipocytes. Increased leptin levels have also been implicated in increased sympathetic activity and the pathogenesis of hypertension in obesity. However, mechanisms for the effects of leptin on blood pressure are unclear. The carotid bodies (CB) express leptin receptor (LepR), but the mechanisms and consequences of leptin action in CB are unknown. We have previously discovered that high level of leptin enhances hypertension acting on Transient receptor potential melastatin 7 (Trpm7) channel in the CB (Shin et al., 2019). Here, we investigate the relevance of Leptin receptor (LepR) in CB for the pathogenesis of hypertension by obesity. We hypothesize that obesity causes hypertension action on LepR in CB. METHODS: C57BL/6J lean male mice (n = 16) and diet-induced obese male mice (DIO, n = 11) were implanted with telemetry in the left femoral artery for continuous blood pressure monitoring. In DIO mice, the adenoviral vectors carrying anti-sense RNA, Ad-LepR shRNA (n = 6) or Ad-scrambled control shRNA (n = 5) , were applied locally to the CB region. Blood pressure measurements were performed at baseline and, in DIO mice, 9-11 days after CB infection with the adenoviral vector. RESULT: DIO increased mean arterial pressure by 14 mm Hg during the day ( p<0.0 001) and by 9 mm Hg at night ( p <0.001) compared to the lean mice. Ad-LepR shRNA abolished obesity-induced hypertension ( p<0.05). LepR knockdown was particularly effective during the day, when animals were predominantly asleep, decreasing mean arterial pressure by 8.5 mmHg. Control shRNA had no effect on DIO-induced hypertension. CONCLUSION: Our study has shown that inhibition of LepR in the carotid bodies abolished obesity-induced hypertension.

19CDA34700025, HL 133100, HL128970

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.
Relationship between serum ß-hydroxybutyrate and hepatic fatty acid oxidation in individuals with obesity and NAFLD
Authors: Shryack Grace , Moore Mary , Meers Grace , Wieschhaus Nicole , Alessi Isabella , Johnson Sarah , Wheeler Andrew , Ibdah Jamal , Parks Elizabeth , Rector Scott
Published: May 2023

Nonalcoholic fatty liver disease (NAFLD), the most common liver disease worldwide, is characterized by the presence of excess lipid (>5%) accumulation which can progress to lobular inflammation (NASH), ballooning, and fibrosis. Lipid breakdown, or mitochondrial fatty acid oxidation (FAO), typically leads to two disparate pathways, ketogenesis or the tricarboxylic acid cycle. In a healthy state, FAO and ketogenesis occur proportionally. Additionally, serum ketones levels, specifically ß-hydroxybutyrate (ß-HB), are often utilized as a surrogate marker for hepatic FAO; yet, whether this relationship exists under conditions of NAFLD is not clearly established. Here, we compared fasting serum ketone levels with direct measurement of mitochondrial palmitate (long chain fatty acid) oxidation from liver biopsies obtained from patients undergoing bariatric surgery stratified based on liver disease severity (n=100). NAFLD progression to NASH was associated with reductions in liver HMGCS2 content and gene expression (P < 0.05) (the rate limiting enzyme in ketosis), while this was not accompanied by differences in serum β-HB (P > 0.05). Worsening liver mitochondrial complete palmitate oxidation corresponded with lower liver Hmgcs2 gene expression (r = 0.30, P = 0.01) but not incomplete or total (complete + incomplete) palmitate oxidation (P > 0.05). Likewise, serum β-HB did not correspond with incomplete, complete, or total palmitate oxidation (P > 0.05). Interestingly, we found that liver Hmgcs2 gene expression and serum β-HB significantly correlated with liver mitochondrial β-HAD activity (r = 0.35, P = 0.007) (r = 0.26, P = 0.04) and the mitochondrial fatty acid transporter CPT1a mRNA (r = 0.50, P < 0.0001) (r = 0.42, P < 0.0001) These results suggest that regulators of ketogenesis may meditate markers in mitochondrial FAO, but serum ß-HB may not a suitable measure of FAO in the context of NAFLD.

This work was supported by an NIH R01 DK113701-01 (R.S.R., E.J.P., J.A.I) and was partially supported by VA-Merit Grant I01BX003271-01 (R.S.R.). This work was supported with resources and the use of facilities at the University of Missouri and Harry S. Truman Memorial Veterans Hospital in Columbia, Missouri.

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.
Neuronal Membrane Repair Defect as a Contributor to Amyloid Beta Pathogenesis in Alzheimer's Disease
Authors: Bulgart Hannah , Lopez Perez Miguel , Tucker Alexis , Usmani Maham , Weisleder Noah
Published: May 2023

Alzheimer’s Disease (AD) is a common neurodegenerative disease that displays two hallmark protein accumulations in the brain – extracellular amyloid beta (Aβ) and intracellular hyperphosphorylated tau. Aβ has been shown to directly penetrate the neuronal cell membrane and damage the membrane. This compromises membrane integrity, increases membrane permeability that alters membrane conductance and elevates intracellular calcium concentrations which contribute to reactive oxygen species production and mitochondrial dysfunction. This damage to the plasma membrane requires a quick and efficient repair mechanism to restore the barrier function of the membrane and avoid neuronal cell death. However, a defect in membrane repair has not been examined as a contributor to AD. Here, we aimed to assess membrane repair capacity in AD in vitro models as a potential contributor to neuronal death. To assess the effect of Aβ on membrane repair kinetics, we exposed primary mouse hippocampal neurons to 1μM recombinant human Aβ42 (rhAβ42) for 16 hours prior to experimentation. Additionally, we treated wildtype neuronal cells with healthy and AD patient with elevated levels of Aβ (PSEN1 M146L mutation) cortical neuron iPSC conditioned media. We also exposed primary mouse hippocampal neurons to cerebrospinal fluid (CSF) from non-AD and AD patients. Treated cells were challenged with an established laser damage assay where a two-photon laser is used to damage the cell membrane in the presence of FM4-64 dye. Dye fluorescence at the injury site is used as a proxy for membrane repair kinetics. Lastly, we investigated autoantibody production targeting proteins involved in membrane repair in non-AD and AD sera and CSF via custom ELISAs. Overall, we observed an induced membrane repair defect when wildtype neuronal cells are treated with rhAβ42, AD iPSC conditioned media, and AD CSF. We also observed a significant increase in autoantibody production in AD biofluids as compared to the non-AD controls. Autoantibody levels were inversely correlated with efficient membrane repair capacity and positively correlated with Aβ concentrations. These results illustrate Aβ’s role in impairing neuronal membrane repair capacity, and the potential contribution of autoantibodies targeting membrane repair proteins in AD.

This work is supported by The Ohio State University Dean's Discovery grant and Dept. of Physiology & Cell Biology Nishikawara Student Research grant.

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.
Chronic intermittent hypoxia induced increases in central angiotensin II release in the MnPO
Authors: Farmer George , Bachelor Martha , Little Joel , Cunningham J.
Published: May 2023

The brain is sensitive to Angiotensin II (ANG II) and we have previously shown that ANG II is released by brain slices containing the median preoptic nucleus (MnPO) and subfornical organ (SFO) in an activity dependent manner. In addition, angiotensin II type 1 receptors and angiotensin converting enzyme 1 are upregulated in the MnPO following chronic intermittent hypoxia (CIH), an animal model of obstructive sleep apnea. Here, we investigate changes in ANG II release in the SFO to MnPO pathway following 7-day CIH treatment.Using isoflurane (2-3%) anesthesia, male Sprague-Dawley rats (250-350g) received microinfusions (0.4 μL) of AAV2-hSyn-ChR2(E123A)-eYFP-WPRE in the SFO. After recovery, rats were subjected to 7 days of CIH (0800-1600 hrs) or Normoxia. The CIH protocol consisted of 6 min cycles (3 min 21% O2, 3 min 10% O2) repeated 10x/hr for 8 hours (during the normal inactive/sleep phase) on 7 consecutive days. For sniffer cell measurements, rats were anesthetized with isoflurane (2-3%) and sagittal slices (300 μm) containing the MnPO and SFO were cut using standard in vitro slice procedures. Sniffer cells consisted of Chinese Hamster Ovary cells transfected with a commercially available plasmid for the angiotensin type 1a (AT1a) receptor (Origene Tech.) and R-GECO (Addgene #32462) as previously described. Sniffer cells were placed on the median preoptic nucleus (MnPO) of sagittal in vitro brain slices. Both spontaneous and evoked (SFO stimulation) changes in fluorescent intensity of sniffer cells on the MnPO were measured.Sniffer cell recordings were made from 5 normoxia and 8 CIH treated rats. The frequency of spontaneous sniffer cell activity in slices from normoxic rats (n = 92) was comparable to rates previously reported in Sprague-Dawley rats. Spontaneous sniffer cell activity in slices from CIH treated rats (n = 156) was increased compared to normoxic treated rats (p < 0.05). Furthermore, bath application of TTX (1 uM) reduced, but did not abolish, spontaneous sniffer cell activity in slices from both normoxic (n = 95, p < 0.001) and CIH (n = 108, p < 0.001) treated rats. Incidence of optogenetic evoked responses in sniffer cells were limited in slices from both the normoxic and CIH treated rats and there were no differences detected in these evoked responses.The current study indicates that spontaneous ANG II release in the MnPO 1) is not entirely action potential dependent, 2) is enhanced following 7-days CIH treatment, and 3) the CIH induced increase is action potential dependent. While optogenetic stimulation of SFO increased sniffer cell activity, CIH had no effect evoked release of ANG II in these preliminary studies.

Work supported by: P01 HL088052, R01 HL155977

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.
Human serum fibrinogen induces brain microvascular endothelial dysfunction and blood-brain barrier dysregulation via dynamin related protein 1 dependent pathway
Authors: Chandra Partha , Rutkai Ibolya , Castorena-Gonzalez Jorge , Busija David
Published: May 2023

Background: Fibrinogen (Fgn) is one of several pathogenic factors in the brain microvasculature. It is upregulated in the blood of the aged, Alzheimer’s disease, and other neurological disease patients. We explored the possible role of a human serum Fgn (hFgn) mediated mechanism for endothelial and blood-brain barrier (BBB) dysregulation in primary human brain microvascular endothelial cells (HBMEC). Methods: To assess the endothelial and BBB dysregulation ex vivo, we measured protein expression change by western blot and immunofluorescence in hFgn-treated HBMEC. Cell viability was determined by NAD(P)H-dependent 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)-based assay, and in vitro BBB-permeability was assessed by transwell migration assay. Stable DRP1 knockdown cells were developed by transduction of shRNA(h) lentivirus particles in HBMEC. Results: Clinical studies have reported that Fgn is an abundant protein in human blood plasma at concentrations ranging from 1.5–4 mg/mL with a normal half-life of 3–5 days, and the plasma concentration of Fgn increased progressively with age in healthy subjects. We observed that 4 mg/mL of hFgn induced significant cytotoxicity and generated mitochondrial reactive oxygen species in HBMEC within 24 h. With the same concentration of Fgn, we observed that most of the mitochondria were elongated or hyperfused, possibly due to significant reduction of phosphorylated dynamin related protein 1 (pDRP1[S616]) in hFgn-treated HBMEC. Moreover, mitochondrial oxidative phosphorylation (OXPHOS)-related mitochondrial complex-II, -III, and -V proteins were also decreased in hFgn-treated HBMEC. Interestingly, autophagy-related, microtubule-associated protein light chain 3B-II (LC3B-II) and lysosomal membrane-associated protein 2 (LAMP2) decreased, while a negative regulator of autophagy, p62, increased in hFgn-treated HBMEC, suggesting hFgn-mediated induction of defective autophagy. Importantly, hFgn induced BBB permeability, possibly due to loss of several BBB-related proteins such as JAM-A, occludin, ZO-1/-2, and PECAM-1 in HBMEC. DRP1 knockdown by shRNA(h) resulted in reduction of OXPHOS and BBB-related proteins as well as defective autophagy induction in HBMEC. Conclusions: Our results suggest that detrimental effects of elevated Fgn downregulates DRP1, leading to endothelial and BBB dysfunction in the brain microvasculature, which affects neuronal health/function and adds vulnerability to strokes and neurological diseases.

HL148836, AG063345, NS114286, AG047296, and AG074489.

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.
Angiotensin II-mediated neuroinflammation in the hippocampus contributes to neuronal deficits and cognitive impairment in heart failure rats
Authors: Campos-Lira Elba , Althammer Ferdinand , Roy Ranjan , Kirchner Matthew , Whitley Kathryn , Davis Steve , Montanez Juliana , Ferreira-Neto Hildebrando , Zafar Usama , Parent Marise , Stern Javier
Published: May 2023

Background: Chronic hypertension can lead to the development of heart failure (HF), a debilitating disease affecting ~65 million people worldwide. In addition to impaired cardiovascular performance and associated systemic complications, most patients with HF suffer from depression and/or cognitive decline. Although neuroinflammation and brain hypoperfusion occur in humans and rodents with heart failure (HF), the underlying neuronal substrates, mechanisms, and their relative contribution to cognitive deficits in HF remains unknown. Methods: To address this critical gap in our knowledge, we used a well-established HF rat model that mimics clinical outcomes observed in humans, along with a multidisciplinary approach combining behavioral, electrophysiological, neuroanatomical, molecular and systemic physiological approaches. Results: Our studies support neuroinflammation, hypoperfusion/hypoxia and neuronal deficits in the hippocampus of HF rats, which correlated with the progression and severity of the disease. An increased expression of angiotensin II AT1a receptors (AngII-AT1aRs) in hippocampal microglia preceded the onset of neuroinflammation. Importantly, blockade of AT1Rs with a clinically used therapeutic drug (losartan) efficiently reversed neuroinflammatory endpoints (but not hypoperfusion/hypoxia ones), resulting in turn in improved cognitive performance in HF rats. Finally, we show than circulating AngII leaks and access the hippocampal parenchyma in HF rats, constituting a likely source initiating the neuroinflammatory cascade. Conclusions: We provide compelling evidence for a novel mechanism that contributes to cognitive deficits in HF. We identified a neuronal substrate (hippocampus), a novel mechanism (Angiotensin II-driven neuroinflammation) and a potential novel neuroprotective therapeutic target (AngII-AT1aRs) for the treatment of cognitive deficits in HF.

DFG Postdoc Fellowship AL 2466/1-1 to FA; National Heart, Lung, and Blood Institute Grant NIH HL090948 to JES, National Institute of Neurological Disorders and Stroke Grant NIH NS094640 to JES and funding provided by the Center for Neuroinflammation and Cardiometabolic Diseases (CNCD) at Georgia State University.

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 intrusion during N2 sleep is negatively associated with nocturnal heart rate variability in healthy adults
Authors: Migliaccio Gianna , Greenlund Ian , Barkstrom Elyse , Bigalke Jeremy , Nicevski Jennifer , Tikkanen Anne , Carter Jason
Published: May 2023

Increased cognitive hyperarousal is a proposed mechanism associated with clinical poor sleep including insomnia. An objective marker of nocturnal hyperarousal is high frequency electroencephalography during sleep, including increased beta frequency during stage two (N2) sleep. Acute and chronic poor sleep quality are associated with autonomic dysfunction, but little is known regarding the relationship between nocturnal beta frequency intrusion and nocturnal autonomic function. The purpose of this study was to explore the association between beta spectral power during stage two (N2) sleep and nocturnal heart rate variability (HRV) in healthy female and male adults. We hypothesized that heightened beta spectral power during N2 sleep would be associated with reduced nocturnal HRV. Twenty-four participants (12 males, 12 females; age, 25 ± 1 years; BMI 27 ± 1 kg/m2) underwent an 8-hour overnight sleep opportunity with continuous polysomnography and 5-lead electrocardiogram monitoring. Periods of 5 – 10 minutes of stable N2 sleep absent of cortical arousals, respiratory events, and limb movement-associated arousals were identified (one per cycle) for analysis. Periods from the first two cycles of N2 sleep from the full night were selected for analysis. Beta spectral power from the central and frontal regions and nocturnal HRV data were analyzed and averaged across each hemisphere. Time-domain HRV was quantified as the root mean squared of successive differences (RMSSD) and proportion of R-R intervals (RRI) varying by greater than 50 ms (pNN50). An inverse correlation was observed between central beta power and RRI (R= -0.473, p=0.019), pNN50 (R= -0.450, p=0.028), and RMSSD (R= -0.406, p=0.049). This was paralleled by a similar relationship between frontal beta and RRI (R= -0.487, p=0.016) and pNN50 (R= -0.423, p=0.039). Our results indicate a negative association between N2 beta power and nocturnal HRV measures. These findings suggest that nocturnal hyperarousal may be associated with poor cardiac autonomic function and has implications for individuals with insomnia in whom cognitive hyperarousal is frequently reported.

Support is provided by the National Institutes of Health (AA-024892, U54GM115371; P20GM103474, 2TL1 TR002318).

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.