bluuebunny/crossref_metadata_2025 huggingface dataset:
Band 3 and Glycophorin C as Potential Mediators of Erythrocyte Aggregation During the Onset of Thermal Burn Injury
Authors: Seidner Harrison S. , Gunter Geoffrey , Weber‐Fishkin Samantha , Frame Mary D.
Published: November 2023

Our goal is to understand aggregation behavior of flowing erythrocytes in the zone of stasis near thermal burn injury. In the temperature range of 42°C to 45°C erythrocytes are known to exhibit hyperaggregation of which the mechanism is unknown. First, we sought to understand the contribution of membrane proteins to aggregation in a vertical flow model of the onset of stasis where temperature was changed from 37°C to a test temperature. Erythrocytes from healthy humans were isolated from other blood components and re‐suspended in PBS, 0.5 g/dL Dextran 500 kDa, or autologous plasma. Erythrocyte solutions were prepared at 20% hematocrit and incubated with rabbit polyclonal antibodies (Abcam) to band 3 and Glycophorin C (GYPC) with human reactivity to test the contribution of the extracellular domains band 3 and GYPC to aggregation. To test for non‐specific binding, anti‐αvβ3 (Chemicon International) was used. The involvement of the intracellular domain of band 3 with aggregation was tested using 4,4′‐Diisothiocyano‐2,2′‐stilbenedisulfonic acid (DIDS), an intracellular‐binding band 3 inhibitor. Following monodispersion by higher Reynolds flow (Re = 0.254), flow was abruptly stopped and aggregates were allowed to form in a low shear environment where their subsequent center‐seeking behavior was observed. The sudden cessation of flow emulated the environment encountered by erythrocytes in the zone of stasis (Re < 0.01) immediately after the occlusion of microvessels in the zone of coagulation during the onset of thermal injury. To study the effects of hyperthermia on aggregation of erythrocytes in stasis the experimental procedure was performed for samples heated to 37 °C, 41 °C, 45 °C, and 49 °C. Temperature was controlled by a heated water chamber attached to the microchannel. Images were acquired for 3 minutes using a horizontally oriented Nikon microscope (20×) and intensified camera (Stanford Photonics, Inc.), passing through an acquisition board (EPIX, Inc.). Python‐based image processing software was used to quantify and analyze time‐dependent measures of aggregation rate and magnitude. Significant differences in aggregation magnitude and rate were observed between erythrocytes suspended in different solvents. Antibodies to band 3, and GYPC had minimal effects on aggregation parameters and temperature dependences were not observed. DIDS‐treated erythrocytes exhibited reduced aggregation that was temperature dependent. The lack of differences in aggregation in the presence of anti‐band 3, and anti‐GYPC indicate that the extracellular domains of these proteins are not mediators of aggregation but provide a strong basis for investigation of the intracellular domains of GYPC and band 3 as mediators of aggregation.
Influence of Light Sources and Photoperiods on Blood Physiological variables Broilers grown to heavy weights
Authors: Olanrewaju Hammed A. , Purswell Joseph L. , Collier Stephanie D. , Branton Scott L.
Published: November 2023

There are a number of studies on the effect of different light sources especially ICD light on the physiology of broilers grown to 2.1 kg, but fewer such studies with conflicting reports are available on growth performance and blood physiological parameters of broilers grown to heavy weights (greater than 3 kg). We evaluated the influence of light sources and photoperiod on blood physiological parameters in 2 trials. A total of 720 1‐d‐old Ross × Ross 708 chicks were randomly distributed into 12 rooms (30 male and 30 female chicks/room). The 3 light sources consisted of compact fluorescent (CFL), light emitting diode (LED), and poultry specific filtered LED (PSF‐LED) and incandescent (ICD, standard) from d 1 to d 56 and 3 photoperiods (long/continuous [23L:1D] from d8–d56; regular/intermittent [2L:2D], and short/non‐intermittent [8L:16D] from d8–d48 and 23L:1D from d49–d56, respectively) at 50 % RH. Similar feed and water were provided ad libitum to all chickens. Arterial blood samples were collected on d 14, 28, 42, and 56 of age and analyzed immediately. Light sources, age, and sex had significant (P ≤ 0.05) effects on BW, Hb, Hct, electrolytes, angap, and mOsm, but all these changes were still within the normal acid‐base homeostasis and physiological ranges. Also, the short photoperiod significantly (P ≤ 0.05) reduced BW, pH level, pO2, sO2, and Ca2+, while significantly (P ≤ 0.05) elevating pCO2, Hct, Hb, and McHc concentrations. Acid‐base regulation during short photoperiod exposure had not deteriorated despite higher pCO2 that consequently decreased blood pH, due to a respiratory acidosis. Plasma corticosterone concentrations were not affected by treatments, suggesting an absence of physiological stress. It was concluded that the 3 LED light bulbs evaluated could be suitable for replacement of ICD along with regular/intermittent photoperiod for commercial poultry facilities to reduce energy costs and optimize production efficiency without compromising the welfare of broilers grown to heavy weights.
HS‐FGF2‐FGFR1 signaling contributes to pulmonary endothelial glycocalyx reconstitution
Authors: Schmidt Eric P , Yang Yimu , Suflita Matthew , Zhang Fuming , Dailey Kyrie , Haeger Sarah , Ford Joshay , Linhardt Robert
Published: November 2023

The pulmonary endothelial glycocalyx (and its in vivo manifestation, the endothelial surface layer) is a heparan sulfate (HS)‐rich layer lining the vascular lumen. Degradation of ESL HS leads to collapse of ESL thickness, endothelial dysfunction and organ injury. Surprisingly little is known about the endothelial mechanisms underlying reparative HS synthesis and their contribution to ESL reconstitution. We treated C57BL/6 mice with heparinase‐III, a bacterial glucuronidase that specifically degrades ESL HS. Mouse lung intravital microscopy revealed that ESL reconstitution was nearly complete within 24 hours after heparinase‐III, coincident with pulmonary induction of the HS biosynthetic enzyme exostosin 1 (Ext1). ESL reconstitution was prevented by treatment with 4‐F‐GlcNAc, an inhibitor of Ext1 HS polymerization. Loss of ESL reconstitution was associated with the onset of lung edema. Similarly, Ext1 induction and ESL reconstitution did not occur mice with endothelial‐specific deletion of fibroblast growth factor receptors 1 (FGFR1); accordingly, these knockout mice demonstrated pulmonary edema. Using surface plasmon resonance and BAF3 cell arrays, we determined that HS fragments (released as a consequence of ESL degradation) may stimulate FGF2‐FGFR1 signaling, inducing Ext1 expression in a human pulmonary microvascular endothelial cell line. Our findings suggest that the products of ESL degradation (i.e. fragmented HS) initiate processes responsible for homeostatic pulmonary ESL reconstitution.

Support or Funding Information

R01 HL125371
The Role of Matrix Metalloproteinases (MMPs) in Platelet Endothelial Cell Adhesion Molecule‐1 (PECAM‐1) Loss in Diabetic Retinopathy: In Vivo and in Vitro Studies
Authors: Eshaq Randa Shafiq , Harris Norman
Published: November 2023

OBJECTIVE

The aim of this study is to understand the mechanisms involved in the diabetes‐induced loss of PECAM‐1 in the retina, and whether it can be attributed to PECAM‐1 cleavage by matrix metalloproteinases (MMPs).

INTRODUCTION

Platelet endothelial cell adhesion molecule‐1 (PECAM‐1) is expressed on the surface of endothelial cells of the vasculature, and has major roles in inflammation, permeability, and angiogenesis. In addition, PECAM‐1 functions in cell signaling, cell‐cell adhesion, and cell survival. However, its physiological role in the pathophysiology of diabetic retinopathy (DR), the leading cause of blindness among working age adults worldwide, has not yet been investigated. We have previously found a significant decrease (50–90%) in retinal PECAM‐1 levels in a diabetic model of type I diabetes in rats.

DESIGN AND METHODS

For our animal studies, we injected age‐matched male Wistar rats with either vehicle (control) or streptozotocin (STZ, diabetic), with the latter at 30 mg/kg/day for three consecutive days to elicit a model of type I diabetes. For our in vitro studies, primary rat retinal microvascular endothelial cells (RRMECs) were grown in either normal glucose (NG, 5mM) or high glucose (HG, 25 mM) media, and treated for 6 days with 0 or 10% plasma collected from control or diabetic rats 8 weeks post‐injection. PECAM‐1, and MMP levels or activity, were assessed using immunofluorescence staining, western blotting, or zymography.

RESULTS

For RRMECs grown in NG conditions, treatment with 10% diabetic plasma resulted in significantly lower levels of PECAM‐1 than treatment with 10% control plasma (27%, p<0.05). PECAM‐1 loss was further exaggerated when cells were grown under HG conditions and treated with 10% diabetic plasma (74%, p<0.01). In contrast, control plasma added to RRMECs in NG conditions increased the expression of PECAM‐1 in a dose‐dependent manner. These findings indicate that normal plasma contributes to the expression of PECAM‐1, but some factor(s) in diabetic plasma has the reverse effect. MMPs, which are calcium‐dependent zinc‐containing endopeptidases, are known to cleave PECAM‐1. Plasma samples from diabetic rats revealed a significant increase in MMP‐2 levels (2‐fold, p<0.05), and a significant increase in MMP‐7 activity (p<0.01). Moreover, RRMECs showed a significant increase in MMP2 levels (12‐fold, p<0.01) when grown in HG media.

CONCLUSION

These results indicate a possible role for MMPs in the diabetes‐induced loss of PECAM‐1. Further studies will be conducted to investigate the potential benefits of protecting the retinal expression of PECAM‐1 by targeting the actions of MMPs.

Support or Funding Information

NIH RO1 EY017599
Norepinephrine Induces Endothelial Albumin Uptake via ROS and Src‐Dependent Mechanisms
Authors: Chignalia Andreia Zago , Isbatan Ayman , Minshall Richard , Dull Randal
Published: November 2023

Hypertensive pulmonary edema often occurs acutely and is typically associated with normal left ventricular performance. Factors other than simple Starling forces may contribute to the molecular mechanisms by which increased blood pressure leads to acute endothelial barrier failure and lung edema.

Methods

To mimic an adrenergically‐mediated hypertensive crisis, we administered norepinephrine (NE) to rats and assessed indices of edema development. Rats were anesthetized with isoflurane and mechanically‐ventilated; subjects received an infusion of NE to target a MAP = 150mmHg. Lung wet/dry ratio, ΔPaO2, plasma hematocrit, lung MPO activity, BAL protein and cell analysis were assessed. Inhibition of nitric oxide synthase (NOS) by L‐NAME was used to test the contribution of endothelial mechanotransduction on changes in permeability. In parallel studies, rat lung microvascular endothelial cells were stimulated with norepinephrine in the presence or absence of propranolol (10−5M, β‐adrenoceptor antagonist), phentolamine (10−5M, α‐adrenoceptor antagonist), PP2 (10−5M, Src inhibitor), apocynin (3×10−5M, ROS scavenger), or L‐NIO (10−4M, NOS inhibitor). Reactive oxygen species (ROS) production was determined by DHE fluorescence. Albumin uptake was assessed by ELISA and albumin transport was measured by fluorimetry.

Results

NE‐treated rats had a decrease in PaO2, an increase in lung albumin content, MPO activity and wet/dry ratio. BAL fluid had increased protein content and number of macrophages indicative of inflammation. L‐NAME significantly attenuated NE‐induced pulmonary edema. In vitro, albumin uptake and transport was increased by NE in a time‐dependent manner. ROS production was augmented by NE. Phentolamine, PP2 and apocynin decreased NE‐induced albumin uptake and ROS generation, effects not altered by L‐NIO.

We describe a novel pathway whereby norepinephrine stimulates albumin uptake and potentially endothelial permeability which might be important in hypertensive pulmonary edema.

Support or Funding Information

Department of Anesthesiology, University of Illinois at Chicago
EET‐dependent attenuation of coronary myogenic constriction in response to deletion/downregulation of soluble epoxide hydrolase
Authors: Froogh Ghezal , Qin Jun , Kandhi Sharath , Jiang Houli , Sun Dong , Huang An
Published: November 2023

Soluble epoxide hydrolase (sEH) catabolizes epoxyeicosatrienoic acids (EETs) to form DHETs that lack, or have less, cardioprotective properties. Thus, we hypothesized that a sex‐different regulation of pressure‐induced myogenic constriction in coronary arteries involves sEH‐dependent signaling. Pressure‐diameter relationships were assessed in isolated and cannulated coronary arteries of male (M) and female (F) wild type (WT) and sEH‐knockout (KO) mice. All vessels constricted in response to increases in intraluminal pressure from 60 to 120 mmHg. Myogenic constriction was significantly attenuated, associated with higher cardiac EETs and lower DHETs in M‐KO, F‐WT and F‐KO mice compared to M‐WT controls. Blockade of EETs with 14,15‐EEZE prevented the attenuated myogenic constriction in sEH‐KO mice. In the presence of EEZE, pressure‐diameter curves of females presented an upward‐shift from those of males, manifesting a sex‐different phenotype. Additional administration of L‐NAME eliminated the sex‐difference, leading to four overlapped pressure‐diameter curves. Cardiac sEH was downregulated in F‐WT compared to M‐WT mice, whereas the expression of eNOS was comparable among the groups. In summary, in combination with NO, the increased EET bioavailability as a function of genetic deletion and/or downregulation of sEH accounts for the female‐favorable attenuation of pressure‐induced constriction.

Support or Funding Information

This work was supported by grant NIH HL070653 and HL34300
Comparison in young White European (WE) and South Asian (SA) men of endothelium‐dependent cutaneous vasodilator responses: contribution of cyclooxygenase (COX) products
Authors: Tsitoglou Kyriakos , Martin Una , Marshall Janice
Published: November 2023

It is established that cardiovascular disease is more prevalent in South Asians (SA) than White Europeans (WE). The prevalence of diabetes central obesity and metabolic syndrome is particularly high in SAs, disorders that are associated with endothelial dysfunction. It is not known whether early markers of endothelial dysfunction can be detected even in young healthy SAs. Thus, we have compared responses evoked in recreationally‐active WE and SA men (18–25 years; n=10 in each case). SAs were 1st or 2nd generation residents in the UK with mother and father originating from India, Sri Lanka, Bangladesh or Pakistan; WEs were UK residents with WE parents. By using laser Doppler fluximetry, we monitored changes in cutaneous red cell flux (RCF) evoked in the cutaneous circulation of the forearm by iontophoresis of the endothelium‐dependent dilator acetylcholine (ACh, 8 × 20s pulses: 7 at 100 μA, 8th at 200 μA, at 60s intervals) and by release of arterial occlusion for 3 min (reactive hyperaemia). Responses were tested before and after aspirin (600mg p.o.) to assess the role of COX products. Mean arterial blood pressure (ABP) was continuously monitored by photoplethysmography using a finger cuff. Resting ABP was similar in WEs and SAs (124±1.8/70±1.7 vs 116.6±2.0/65.4±1.6 mmHg) and did not change during the protocol. Baseline RCF was also similar in WEs and SAs: 19.7±1.9 vs 12.3±0.9 perfusion units (PU), respectively. Further, ACh evoked similar dilator responses in WEs and SA, maximum RCF reaching 192.2±5.5 and 182.83 ± 5.20 perfusion units (PU) respectively. Aspirin had no net effect on ACh‐induced dilatation in either group (WEs: 182.6±4.5 and SAs: 193.0± 7.7 PU) indicating any influence of COX dilator and vasoconstrictor products was balanced in both WEs and SAs. By contrast, before aspirin, reactive hyperaemia was substantially larger in WEs than SA, peak RCF being 92.9±2.5 vs 74.2±3.1*PU, *: P<0.01 RMANOVA). Further, aspirin attenuated reactive hyperaemia in WEs, peak RCF being only 75.1 ± 2.6§PU: §: P<0.001 vs control response, but enhanced reactive hyperaemia in SAs, peak RCF reaching 84.7±2.7§PU. These results suggest that in young SA men, who have no overt signs of cardiovascular disease and are normotensive, there are already signs of endothelial dysfunction. Agonist‐evoked endothelium‐dependent dilatation is apparently normal in SAs with no net contribution from COX dilator or vasoconstrictor products, suggesting that nitric oxide and/or Endothelium‐dependent hyperpolarising factor are the main contributors in both ethnicities. However, reactive hyperaemia, which is induced by the combination of shear‐stress and metabolic stimuli, is blunted in young SAs relative to WEs and this is attributable to a predominating influence of vasoconstrictor COX products in SAs, but vasodilator COX products in WEs.

Support or Funding Information

Alexander S. Onassis Public Benefit Foundation
Chronic Nicotine Exposure Exacerbates Transient Focal Cerebral Ischemia‐Induced Brain Injury
Authors: Li Chun , Sun Hong , Arrick Denise M , Mayhan William G
Published: November 2023

Tobacco smoking is a risk factor contributing to the development and progression of ischemic stroke. Among many chemicals in tobacco, nicotine may be a key contributor. We hypothesized that nicotine alters the balance between oxidant and antioxidant networks leading to an increase in brain injury following transient focal cerebral ischemia. Male Sprague‐Dawley were treated with nicotine (2 or 4 mg/kg/day) for 4 weeks via an implanted subcutaneous osmotic minipump and subjected to a 2‐hour middle cerebral artery occlusion (MCAO). Infarct size and neurological deficits were evaluated at 24 hours of reperfusion. Superoxide levels were determined by lucigenin‐enhanced chemiluminescence. Expression of oxidant and antioxidant proteins was measured using Western Blot analysis. We found that chronic nicotine exposure significantly increased infarct size and worsened neurological deficits. In addition, nicotine significantly elevated superoxide levels of cerebral cortex under basal conditions. Cerebral ischemia/reperfusion (I/R) produced an increase in superoxide levels of cerebral cortex in control group, but no further increase was found in the nicotine group. Furthermore, chronic nicotine exposure did not alter protein expression of NADPH oxidase, but significantly decreased uncoupling protein‐2 (UCP‐2) in the cerebral cortex, MnSOD in the cerebral cortex and cerebral arteries. Our findings suggest that nicotine‐induced exacerbation in brain damage following transient focal cerebral ischemia may be related to a pre‐existing oxidative stress via decreasing of UCP‐2 and MnSOD.

Support or Funding Information

Funded by a fellowship from Center for Cardiovascular Diseases and Sciences, LSU Health Sciences Center‐Shreveport
Differential Effects of Hookah vs. Cigarette Smoking on Endothelial Function
Authors: Rezk‐Hanna Mary , Doering Lynn , Robbins Wendie , Mason O'Neil , Choung Jennifer , Benowitz Neal L , Elashoff Robert M , Sarna Linda , Victor Ronald
Published: November 2023

Hookah (water pipe) smoking is a major new understudied epidemic of tobacco abuse particularly affecting youth. Hookah's rapidly growing popularity is due to unregulated expansion of hookah cafes near college campuses and social media marketing to young adults as a safer avant‐garde alternative to cigarettes. Whereas multiple studies have shown that smoking even a single cigarette acutely impairs endothelial function, the acute effect of hookah smoking on endothelial function is unknown. Because burning charcoal briquettes are used to heat the hookah tobacco product, hookah smoke differs from cigarette smoke by delivering a large acute exposure to carbon monoxide and other charcoal combustion products including fine and ultrafine particles that have been suggested by the air pollution and tobacco literature to constitute putative endothelial toxins. To determine if hookah smoking acutely impairs endothelial function, in 21 healthy young adult hookah smokers who do not smoke cigarettes (age 25±1 years, mean±SE; 6 women, 15 men; body mass index, BMI 23.6±0.6 kg·m2), we measured endothelium‐dependent flow‐mediated dilation (FMD) by brachial artery ultrasound before and immediately after 30 minutes of ad lib hookah smoking in a custom‐built smoking chamber. In contrast to what we had hypothesized, brachial artery FMD did not decrease with hookah smoking but, surprisingly, increased from 6.7±0.5% to 9.7±0.6%, P<0.001: a 53.1±9.5% relative increase. In contrast, in five cigarette smokers (matched with the hookah smokers by age, gender, and BMI), FMD decreased acutely by 28.7±3.4% (p=0.006) after smoking one cigarette, thus confirming previous reports. Shear rate (the stimulus to FMD) was unaffected by either form of smoking. Furthermore, the increases in systolic blood pressure (SBP) and heart rate (HR) were more than 50% smaller with hookah than with a cigarette (ΔSBP: +7±1 vs. +17±9 mmHg, p<0.05; ΔHR: +15±2 vs. 28±5 beats.min−1, p<0.05) but the exhaled carbon monoxide level was almost 3‐fold greater after smoking hookah than after smoking a cigarette: 29±2 vs. 11±1 ppm, p<0.01). Thus, whereas brachial artery flow‐mediated dilation is acutely impaired by cigarette smoking, it is acutely augmented by hookah smoking. Further studies are indicated to determine if the high carbon monoxide exposure constitutes the major mechanism underpinning the augmented flow‐mediated dilation.
Cross Tolerance and Potentiation of Toll‐like Receptors in Endothelial Cells
Authors: Stark Ryan J , Koch Stephen R , Lamb Fred S , Sherwood Edward R
Published: November 2023

Toll‐like receptors (TLR) are important regulators of inflammation in endothelial cells. Activation of TLRs can reduce the pro‐inflammatory response to a subsequent TLR challenge, inducing a state of “tolerance”. We hypothesized that activation of TLR2 and TLR3 would induce cross tolerance to TLR4 activation. We cultured human microvascular (HMVECs) and umbilical vein (HUVECs) endothelial cells and exposed them to the TLR4 agonist lipopolysaccharide (LPS), the TLR3 agonist poly I:C, or the TLR1/2 agonist Pam3CSK4 for 16 hours. Cells were washed and then allowed to rest in media for 30 hours. Afterwards, cells were re‐exposed to different TLR ligands for 16 hours. Inflammation was assessed by quantifying IL‐6 levels in the culture media. Initial LPS exposure rendered both HUVECs and HMVECs tolerant to subsequent LPS exposure with a 47% and 48% reduction in IL‐6, respectively (p <0.05). Contrary to this, initial exposure to poly I:C increased IL‐6 production after LPS re‐exposure compared to LPS alone by 25% in HMVECs and 35% in HUVECs (p <0.05). Interestingly, though Pam3CSK4 produced little IL‐6 in HMVECs compared HUVECs (11.18 pg/ml HMVECs vs. 2197 pg/ml in HUVECs), initial exposure of HMVECs to Pam3CSK4 reduced IL‐6 in cells re‐exposed to LPS by 43% compared to LPS alone(p <0.05). These studies provide new knowledge into the modulatory relationship between TLRs in inciting inflammation in endothelial cells.

Support or Funding Information

This work was supported by the National Institute of General Medical Sciences (K08‐GM117367, R.S., R01‐GM104306, E. S.)
Overweight was related with sympathetic activation in offspring of hypertensives
Authors: Viana Ariane Oliveira , Rosa Fernando Alves Santa , Nascimento Mario Cesar , Shimojo Guilherme Lemos , De Paiva Júlia , De Angelis Kátia
Published: November 2023

Hypertension is a clinical condition with high rates of cardiac events and mortality. This disease has been related to overweight/obesity, cardiovascular and autonomic dysfunctions. The objective of this study was to evaluate anthropometric, hemodynamic and autonomic parameters in sedentary men with a positive (HS n= 32) or negative (NS n= 30) familial history of hypertension. A cross‐sectional study was performed with 62 healthy sedentary men (20–48 years old). The body mass index (BMI) was calculated and the blood pressure (BP) was measured 3 times by the auscultatory method. The RR interval was recording during a 20 minute period at rest using a frequencimeter (Polar® RS800 model). The heart rate variability was evaluated in the time and frequency domain. The overweight subjects (n=45, BMI: 29.22 [IQ 26.96–30.87Kg/m2]) presented significantly higher systolic BP (116.66 [IQ 88–128mmHg] vs. 104 [IQ 76.33–118.33mmHg]) and diastolic BP (85.33 [IQ 80.83–93.94mmHg] vs. 82.88 [IQ 73.00–86.22mmHg]) as compared to the eutrophic group (n= 17, BMI: 24.30 [IQ 24.78–29.72Kg/m2]). In addition, positive correlations were observed between the sympathovagal balance with the systolic BP (r = 0.377, p = 0.011) and the mean BP (r = 0.406, p = 0.006). The BMI was also correlated with diastolic BP (r = 0.335, p = 0.025). Regarding the familial history of hypertension, we observed increased BP and sympathetic modulation in the HS group compared to the NS group. Moreover, BP was increased in overweight HS subjects as compared to overweight NS. Correlation analysis involving HS and NS groups, showed positive correlations between sympathovagal balance with mean BP (r = 0.257, p = 0.044) and systolic BP (r = 0.262, p = 0.039); BMI with diastolic BP (r = 0.389, p = 0.002) and systolic BP (r = 0.336, p = 0.008). In conclusion, the overweight was related with sympathetic activation and BP increase in general populations as well as in offspring of hypertensives.

Support or Funding Information

Cnpq, Capes, UNINOVE
Continuous Glucose Measurements Using Telemetry in Male and Female Dahl Salt‐Sensitive Rats (DSS) on a High‐Fat Diet
Authors: Gillis Ellen , Ralph Erica , Tiesma Scott , Lindquist Ryan , Sullivan Jennifer
Published: November 2023

The HD‐XG glucose telemetry implant from DSI provides the ability to obtain continuous, real‐time, blood glucose measurements in rodents. The goal of the current study was to compare spot blood glucose measurements with values obtained using telemetry in male and female DSS. Male and female rats were implanted with glucose telemeters at 11 weeks of age, and allowed 1 week to recover before being placed on receivers. Rats were maintained on a normal‐salt, normal‐fat (NF; 16% caloric intake from fat) diet for 1 week and then were switched to a high‐fat diet (HF; 65% caloric intake from fat) for 4 weeks (n=4–5/group sex). Additional rats without telemeter implants were maintained on normal‐salt, normal‐fat diet for the duration of the experiment. All rats gained weight over the 4 week treatment period, however, rats of both sexes on a HF diet exhibited a significantly greater increase in body weight compared to rats on NF (males: 15±1% on NF vs. 22±2% on HF, p<0.05; females: 12±1% on NF vs. 18±2% on HF, p<0.05). There was not a sex difference in weight gain in response to HF diet. Mean absolute relative differences in blood glucose as measured by the HD‐XG compared to spot glucose ranged from 0.1% up to 4.8% in males and 0.7% up to 8.9% in females, indicating that the 2 measurements were always within 10% of one another. Baseline blood glucose levels in male and female DSS measured by HD‐XG were comparable (109±2 vs. 111±2 mg/dL, respectively). HF diet for 4 weeks did not significantly alter blood glucose levels in either sex (male: 106±3 mg/dL; female: 104±4 mg/dL). Additionally, average weekly glucose for both sexes were statistically identical whether analyzed over 24 hour periods or limited to only the light or dark periods. From this study we can conclude the following: 1) glucose telemeters are a reliable methods to obtain continuous measurements of blood glucose in male and female rats, 2) there are no sex differences in blood glucose following a 4 week HF diet in DSS, and 3) DSS are able to tolerate a HF diet with minimal alterations in body weight or blood glucose.
Chronic Vagus Nerve Stimulation Attenuates Renal Inflammation in Autoimmune‐Induced Hypertension
Authors: Pham Grace S , Fairley Amber S , Maloy Charles I , Mathis Keisa W
Published: November 2023

Chronic renal inflammation has been implicated in the pathogenesis of hypertension, which identifies the autoimmune disease systemic lupus erythematosus (SLE) as a compelling disease model due to its key features of lupus nephritis and prevalent hypertension. The cholinergic anti‐inflammatory pathway is an endogenous, vagally‐mediated neuroimmune pathway that, when impaired, may influence the development of chronic inflammation. While stimulation of the cholinergic anti‐inflammatory pathway attenuates inflammation in models of sepsis and other inflammatory diseases, it is not clear if the pathway is impaired in SLE, nor whether its stimulation is protective in SLE. We hypothesized that pharmacological activation of the cholinergic anti‐inflammatory pathway at the level of the vagus nerve would decrease renal inflammation and therefore blood pressure in a murine model of SLE hypertension. Initially, female SLE (NZWF1) and control (NZW) mice were administered the efferent vagal stimulant, CNI‐1493 (Semapimod hydrochloride; 8 mg/kg IP) or saline twice per week for 4 weeks starting at 30 weeks of age. SLE mice had 1143±439% (p<0.001), 74±24% (p=0.006), and 225±124% (p=0.009) higher renal cortical expression of the inflammatory mediators TNF‐α, IL‐1, and high mobility group box (HMGB)‐1 than controls (n=3–5/group). CNI‐1493 reduced TNF‐α, IL‐1 and HMGB‐1 by 13±11% (p=NS), 29±11% (p=0.05), and 42±5% (p=NS) in SLE mice. We continued to test our hypothesis by stimulating the efferent vagus via administration of the centrally‐acting acetylcholinesterase inhibitor, galantamine (GAL; 4mg/kg IP), or saline to SLE and control mice for 14 consecutive days starting at 32 weeks of age. SLE mice had 502±332% (p=0.037) and 345±35% (p=0.002) higher renal cortical expression of TNF‐α and IL‐6 than controls (n=2–4/group). GAL lowered TNF‐α and IL‐6 by 56 ±16% (p=NS) and 61±2% (p=0.006) in SLE mice. Interestingly, when we assessed whether mean arterial pressure (MAP) was altered following vagal stimulation and subsequent reduction of renal cytokines, we found that MAP (n=3–6/group) was higher in SLE mice compared to controls (143±11 vs. 116±4; p=0.003) and that both CNI‐1493 and GAL prevented the rise in MAP in SLE mice (130±4 and 135±7, respectively). These preliminary results suggest that chronic vagal nerve stimulation attenuates renal inflammation and therefore prevents the development of hypertension in SLE. Ultimately, delineating the role of the cholinergic anti‐inflammatory pathway in SLE could support further investigation into the pathway's role in other autoimmune diseases, as well as essential hypertension, which has recently been shown to arise from an inflammatory etiology.

Support or Funding Information

Studies supported by the AHA (#14SDG18320033) and UNTHSC (Faculty Pilot Grant)
Does chronic inhibition of brain endoplasmic reticulum (ER) stress alter metabolic and cardiovascular function in obese melanocortin‐4 deficient rat?
Authors: do Carmo Jussara M. , da Silva Alexandre A. , Yoo Jung , Moak Sydney P. , Spradley Frank , Hall John E.
Published: November 2023

The central nervous system (CNS) melanocortin system plays a key role in regulating metabolic function and in mediating obesity‐induced hypertension. Although, endoplasmic reticulum (ER) stress has been suggested to alter hypothalamic pathways that regulate food intake, body weight and blood pressure (BP), its role in mediating cardiovascular and metabolic dysfunction in obesity is still unclear. In this study we examined the role of brain ER stress in controlling blood pressure (BP) and metabolic functions in obese melanocortin‐4 receptor (MC4R) deficient rats. MC4R knockout (MC4R−/−, n=5) and control wild‐type Wistar Hannover rats (WT, n=5) were implanted with blood pressure (BP) telemetry transmitters and an intracerebroventricular (ICV) cannula was inserted into the third ventricle at 22 weeks of age. After 10 days of recovery, food intake, BP and HR were measured 24‐hrs/day. After stable baseline measurements for 4 days, tauroursodeoxycholic acid (TUDCA, 7mg/hr, ICV) was infused via osmotic minipump for 7 consecutive days to inhibit brain ER stress. At baseline, MC4R−/− rats ate 42% more food and were 41% heavier than WT rats. MAP was higher (118±1 vs. 111±1 mmHg) and HR was lower (332±3 vs. 353±4 bpm) in MC4R−/− rats. Inhibition of brain ER stress significantly decreased cumulative net food intake (−26+4 vs. −16+5 g) while causing no changes in blood glucose levels (WT: 97±2 vs. 95±4 and MC4R−/−: 109±4 vs.106± 6 mg/dl) or body weight (WT: 446±11 vs. 448±12 and MC4R−/−: 629±23 vs.631± 30 g). Inhibition of brain ER stress also did not alter BP in either group (3 and 2 mmHg) but increased HR (8±2 and 19±4 bpm), respectively, in WT and MC4R−/− rats. These results suggest that chronic inhibition of brain ER stress reduced appetite but did not alter body weight, blood glucose or BP in WT or MC4R−/− rats.

Support or Funding Information

(NHLBI‐PO1HL51971, NIGMS‐P20GM104357 and AHA SDG5680016)
Sex Differences in Renal Sodium Transporter Gene Expression
Authors: Patel Ravi S. , Solocinski Kristen , Cheng Kit‐Yan , Gumz Michelle L.
Published: November 2023

Regulation of sodium balance and blood volume by the kidneys is an important determinant of blood pressure control. Although the causative mechanisms remain unclear, it is well established that male sex is associated with higher blood pressure and increased cardiovascular risk compared to female sex. In order to determine a possible role for renal sodium transporters in sex‐dependent blood pressure regulation, we examined a number of renal transport genes in the kidney cortex of male and female mice. Under normal conditions, the circadian clock protein Per1 is a positive regulator of sodium transport genes including αENaC (α subunit of the epithelial sodium channel), Wnk1(with‐no‐lysine kinase 1), and NCC (sodium‐chloride cotransporter). Using qPCR and western blot analysis, we found significant differences between the sexes in a number of renal sodium transport genes and clock genes. Interestingly, Per1 (Period 1), Sgk1 (serum and glucocorticoid kinase 1), αENaC, Edn‐1 (Endothelin‐1), and NCC expression was higher in females compared to males whereas Wnk1 expression was equivalent between the sexes. These data show significant sex‐dependent differences in gene expression of distinct sodium transport genes and circadian clock genes. Whether or not these differences are related to sex‐dependent differences in blood pressure remains to be determined.

Support or Funding Information

This work was supported by NIH DK085193 and DK098460, the ASN Foundation for Kidney Research to MLG and 2T32HL083810 to KS.
Cyclic‐AMP Regulates Glycogen Synthase Kinase‐3β by a Feed‐Forward Mechanism in Polycystic Kidney Disease
Authors: Kakade Vijayakumar R , Tao Shixin , Zhou Xia , Li Xiaogang , Pandey Pankaj , Rajagopal Madhumita , Rao Reena
Published: November 2023

Glycogen synthase kinase 3β (GSK3β) is a serine/threonine protein kinase that plays an important role in cell survival, proliferation, metabolism and differentiation. GSK3β is known to be regulated at the level of its activity. However, GSK3β expression is increased in diabetes, Alzheimer's disease and certain cancers, although the mechanism is not clear. We observed increased GSK3β expression in polycystic kidney disease (PKD), a disease characterized by progressive cyst expansion that leads to end stage renal disease. Since GSK3β plays a pathophysiological role in disease progression in PKD, the current studies examined the mechanism for the increased GSK3β expression and its significance in PKD. In mouse models of autosomal dominant and autosomal recessive PKD, increase in renal GSK3β expression corresponded with increase in renal cAMP levels and disease progression. Examination of the GSK3β promoter revealed five cAMP response elements (CREs), potential binding sites for the transcription factor, CRE‐binding protein (CREB). Studies using M1‐cortical collecting duct, PKD1‐ gene knockout (PKD1‐‐KO) and control cells demonstrated that GSK3β is a cAMP responsive gene and elevated cAMP levels, as seen in PKD, can increase GSK3β protein and mRNA levels. CREB was found to bind to the GSK3β promoter and to be essential for cAMP mediated regulation of GSK3β expression. Importantly, this regulation was demonstrated to be part of a feed‐forward loop in which cAMP through CREB regulates GSK3β expression, and GSK3β in turn regulate cAMP levels and CREB activity. Inhibition of CREB reduced forskolin induced cAMP generation, which could be rescued by overexpression of constitutively active GSK3β showing that the relation between cAMP, CREB and GSK3β is cyclic in nature. Moreover, inhibition of GSK3β, or CREB significantly reduced cyst expansion as well as trans‐epithelial fluid secretion in vitro. Thus, GSK3β is regulated by a cAMP and CREB dependent feed forward loop, and disruption at any point of this cycle may be therapeutically useful to reduce cyst expansion and preserve renal function in PKD.

Support or Funding Information

NIDDK Support
The α2‐Subunit of AMP‐activated Protein Kinase Is a Potent Modulator of Myocardial Diastolic Calcium Homeostasis
Authors: Pohl Ulrich , Fein Evelyn , Kameritsch Petra , Schneider Holger , Engelhardt Stefan , Dendorfer Andreas , Blodow Stephanie
Published: November 2023

Rationale

The heterotrimeric enzyme 5′‐adenosine monophosphate‐activated protein kinase (AMPK) is known to have cardioprotective effects which may be partly due to a participation in the control of intracellular calcium concentration. Recently, we described in vascular smooth muscle a dilator role for AMPK which is due to sarcoplasmic reticulum Ca2+‐ATPase (SERCA)‐dependent calcium decrease. The present study elucidates a possible role of AMPK for myocardial diastolic calcium decrease.

Objective

To investigate, whether AMPK contributes to the regulation of SERCA dependent myocardial diastolic calcium homeostasis via phosphorylation of phospholamban (PLN) and to identify which catalytic subunit of AMPK was involved.

Methods and Results

Freshly obtained heart slices of wildtype and AMPKα2‐gene‐deficient mice were analyzed by recording contraction amplitudes and relaxation velocities in the presence or absence of the AMPK activator A769662. A769662 (30 μM, 15 min) increased phosphorylation of AMPKThr172 showing effective activation of the enzyme. The AMPK stimulator induced positive inotropic (contraction amplitudes +75%) and lusitropic (relaxation velocity +71%) effects in isolated heart slices (p<0.05 vs controls, n=6–11). The latter corresponded to a significant shortening of the time constant of Ca2+‐transient decay (tau, −25%, Fura2 method, n=15–23) which was similar to the effect observed after stimulation with isoproterenol (100 nM). A second structurally different AMPK activator, PT1 (30 μM), virtually mimicked the A769662‐induced positive inotropy and lusitropy. Western Blots (n=3–9) of freshly isolated, A769662‐stimulated adult mouse cardiomyocytes showed an enhanced phosphorylation of PLN, but exclusively at the regulatory site Thr17, whereas isoproterenol led to increases in phosphorylation of both regulatory sites, PLNSer16 and Thr17. The A769662‐induced effects were blunted in slices and cells isolated from AMPKα2‐gene‐deficient mice while isoproterenol was still effective (n=4–5).

Conclusions

Our data suggest that AMPK, via its catalytic α2‐subunit, induces phosphorylation of PLNThr17 thereby accelerating diastolic Ca2+‐decrease and increasing relaxation velocity. This novel AMPK‐PLN‐ pathway could serve as a valuable new therapeutic target in failing hearts showing reduced responsiveness to b‐adrenergic stimulation.

Support or Funding Information

Supported by the DZHK (German Centre for Cardiovascular Research)
Avian osmoregulation in flight: unique metabolic adaptations present novel challenges
Authors: Gerson Alexander R , Guglielmo Christopher
Published: November 2023

Migratory birds face unique challenges as they routinely complete extremely long flights where rates of respiratory water losses are high, but the continual maintenance of ion homeostasis is crucial. Songbirds (order Passeriformes) typically complete migration through a series of long duration nocturnal flights, each followed by a brief stopover period where fuel reserves are replenished. In flight, migratory birds rely on endogenous fat and protein for energy, resulting in dramatic reductions in lean and fat masses of individuals. During flight, rates of respiratory water loss are extremely high due to the high breathing frequency and tidal volumes required for sustained aerobic exercise. Despite prolonged flights over inhospitable environments, no studies to date have documented dehydration in migratory birds after long flights in the wild. In order to investigate the osmoregulatory strategies used by migratory birds, we developed a method to measure glomerular filtration rate in flying birds using a single injection of FITC inulin, which was eliminated following second order exponential decay kinetics. Using this methodology we investigated GFR in flight, during fasting and in fed birds and found no difference among fed, fasted and flown birds in GFR, but fractional water reabsorption was increased during both fasting and flight. We found no influence of rate of water loss on the increase in FWR during flight. In separate experiments we investigated the metabolic response of thrushes to dehydrating conditions during flight and found that birds increase rates of protein catabolism as a means to increase endogenous water production to offset high rates of respiratory water loss. In these studies we show that migrating birds do not dynamically regulate GFR, as has been found for other environmental challenges, but instead dynamically regulate rates of metabolic water production in order to maintain water balance in flight. This strategy allows the completion of long duration flights, but at the cost of organ and muscle mass. Moving forward we are investigating the mechanisms regulating lean mass catabolism in response to dehydration stress and the functional consequences of organ and muscle catabolism during flight in migratory birds.
The effect of diet on the metabolic rate and microbiome of Gromphadorhina portentosa
Authors: Contreras Heidy L. , Lorenz Todd
Published: November 2023

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

Support or Funding Information

ULV Faculty Research Grant
α1G T‐type Ca2+ channel plays a dual role in control of pulmonary microvascular endothelial cell P‐selectin surface expression
Authors: Zheng Zhen , Hu Xingang , Chen Hairu , Sellak Hassan , Alexeyev Mikhail F , Wu Songwei
Published: November 2023

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

Support or Funding Information

HL‐066299