bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelial Cell Dysfunction is Associated with the Development of Hypertension and Cardiac Hypertrophy in Npr1 Gene‐disrupted Mice
Authors: Das Subhankar , Krazit Stephen T , Pandey Kailash N
Published: November 2023

Cardiac hypertrophy is a major predictor of progressive heart disease and an adverse prognosis. Atrial and brain natriuretic peptides (ANP and BNP) bind to guanylyl cyclase/natriuretic peptide receptor‐A (GC‐A/NPRA) and exert the beneficial effects against hypertension and cardiac hypertrophy. The objective of the present study was to examine the effect of Npr1 (coding for NPRA) gene‐disruption on the process of endothelial cell dysfunction in the development of hypertension and cardiac hypertrophy. We used six‐month old mice having different copies of Npr1 gene (0‐copy, 1‐copy, and 2‐copy). The blood pressure and echocardiographic measurements were performed in each group. Blood was collected by direct cardiac puncture and cardiac tissues were harvested immediately and snap frozen in liquid nitrogen. The 0‐copy (Npr1−/−) mice showed 35–45 mmHg higher systolic blood pressure (SBP) as compared to 2‐copy (Npr1+/+) mice. Echocardiographic measurements of Npr1 gene‐disrupted 1‐copy (Npr1+/−) and 0‐copy mice showed significant increased LVEDD (27% in 0‐copy and 10% in 1‐copy mice), intraventricular septal wall thickness (IVSTDD, 30% in 0‐copy and 15% in 1‐copy mice), and reduced fractional shortening by 24% in 0‐copy and 12% in 1‐copy mice as compared with control mice. There was also a significant increase in plasma levels of plasminogen activator inhibitor‐1 (PAI‐1; 2.2‐fold), soluble intracellular adhesion molecule (sICAM; 1.8‐fold), soluble vascular cell adhesion molecule (sVCAM; 25%), E‐selectin (4‐fold) in Npr1 gene‐disrupted (0‐copy) animals as compared with wild‐type mice. Cardiac collagen content was significantly increased by 3‐fold in 0‐copy mice as compared to control mice and was confirmed by histological study with Masson's trichrome staining of cardiac tissues. In the present study, increased plasma levels of sICAM, sVCAM, PAI‐1 and E‐selectin suggest the involvement of endothelial cell dysfunction in producing cardiac hypertrophy in Npr1 null mutant mice. Our results implicate that ANP/NPRA signaling exerts an inhibitory effect on the agents, which cause endothelial dysfunction in the heart leading to cardiac hypertrophy and cardiac remodeling.

Support or Funding Information

This work was supported by NIH grants (HL057531 and HL062147) and by an Institutional Development Award (IDeA) from the NIGMS.
Sex Differences in Sympathetic Neuronal Control of Mesenteric Arteries from Obese Dahl SS Hypertensive Rats
Authors: Gebremedhin Kibrom , Galligan James , Xu Hui
Published: November 2023

Obesity‐associated hypertension is a risk factor for stroke, type 2 diabetes and kidney failure. Hypertension is less common in premenopausal women than in age‐matched men possibly due estrogen induced vasodilation in women. Sympathetic nerve activity increases in obesity‐related hypertension via the elevated norepinephrine (NE) and ATP release and increased constriction in resistance mesenteric arteries (MA). We hypothesized that the causes of obesity‐related hypertension differ between male and female Dahl salt‐sensitive (ss) rats. Rats were fed control and high fat (HF) diet beginning at 3 weeks of age. Body weight and mean arterial pressure (MAP) were measured until they were sacrificed at 24–26 weeks. Males (450 g) gained more weight than females (300 g) irrespective of diet and all HF fed rats became hypertensive (MAP>140 mmHg). The estrus cycle stage in female rats was determined cytologically by vaginal lavage before euthanasia. Mesenteric arteries (Inner diameter; male=275–350 μm, female=245–315 μm) were harvested and mounted on a pressure (60 mmHg) myograph and inner diameter was measured continuously. MA contractile responses to electrical field stimulation (30 stimuli at 0.2–30Hz) and drug applications were measured. MA neurogenic constriction was greater in HF‐fed male (n=9) compared to HF‐fed female arteries (n=3). Neurogenic constriction was similar in control animals while HF‐fed males had greater constriction than females. Neurogenic constriction did not vary with the estrus cycle in female rats regardless of diet. Neurogenic constrictions were blocked by combined application of prazosin (0.1 μM, α1‐adrenergic receptor antagonist) and PPADS (10 μM, P2X‐purinergic receptor antagonist) as well as suramin (100μM, P2X and P2Y‐purinergic receptor antagonist) in arteries from male and female rats. PPADS/Suramin inhibited ATP‐induced MA contraction. Constrictions caused by exogenous NE and ATP were similar in MA from HF‐fed male and female rats. Immunostaining for tyrosine hydroxylase (NE marker) and vesicular nucleotide transporter (ATP marker) revealed greater nerve distribution in HF‐fed MA from male and female rats. NE HPLC shows similar levels in male and female MA; however females have greater NE level in the perivascular fat. Our data indicate that the mechanisms of neurogenic vasoconstriction differ in MA from obese male and female rats. Differences in neurotransmission and/or nerve fiber distribution in obese male and in female rats might explain the sex differences in hypertension prevalence between human males and females.

Support or Funding Information

NIH PPG: P01HL070687
Evaluation of Cardiac Autonomic Balance and Sympathetic Vasomotor Activation to Different Targets in a Model of Hypertension Induced by Nitric Oxide Synthase Blockade
Authors: Zambrano Lysien Ivania , Pontes Roberto B , Oliveira Tales L , Garcia Michelle L , Nishi Erika E , Campos Ruy R , Bergamaschi Cassia T
Published: November 2023

High blood pressure is a global public health problem. Several factors are involved in the generation and maintenance of hypertension, such as increased sympathetic nerve activity (SNA), although controversies have been reported. Our aims were to evaluate cardiac autonomic balance and sympathetic activation of the renal (rSNA) and the splanchnic nerve (sSNA), in animals with hypertension induced by nitric oxide synthase blockade (L‐NAME). Wistar rats were treated with L‐NAME (20mg/kg/day, once a day, for 8 days). L‐NAME treated group presented increased blood pressure (152 ± 02 mmHg, n=17), compared with control group (101±01 mmHg, n= 15).Plasma nitric oxide (NO) measurement presented a significantly reduction in L‐NAME (6.8 ±0.4 pg/L n=15), compared to CTL group (9.9 ± 1.2 pg/L n=06), both rSNA (147 ± 10spikes/s, n=16) and sSNA (137 ± 13 spike/s n=14) were higher in L‐NAME treated group, compared with control group (114 ± 5, n=10 and 74 ± 13spikes/s, n=09, respectively). The spectral analysis of the pulse interval (PI) demonstrated that the L‐NAME group presented increased low frequency component (LF) (19.75±1.75, n=07 vs CTL: 6.75±1,10, n=07), reduced high frequency component (HF) (81.29 ± 1.80 n=07 vs CTL: 92.50±1.20, n=07) leading to a sympatho‐vagal imbalance (LF/HF) (0.24 ± 0.02, n=07 vs CTL: 0.08 ± 0.01, n=07).

Taken altogether, our data show that the hypertension induced by nitric oxide synthase blockade is characterized by cardiac autonomic imbalance and sympathoexcitation to more than one territory and that can be involved in the generation and/or maintenance of high blood pressure.

Support or Funding Information

CAPES, CNPq, UNAH.
Physiological changes in intracranial pressure affect arterial pressure
Authors: Guild Sarah‐Jane , Saxena Utkarsh Anand , Malpas Simon , McBryde Fiona , Ramchandra Rohit
Published: November 2023

A number of cardiovascular diseases are characterised by increased levels of sympathetic nerve activity. However, the driving force behind this increase remains unclear. Studies have suggested that, in some of these diseases, reduced cerebral perfusion may cause a sympathetically mediated increase in arterial pressure in an effort to rectify the reduced cerebral perfusion e.g. Selfish Brain Hypothesis (Paton et al 2009). The objective of this research was to determine a) if physiological increases in intracranial pressure (ICP) will cause a reflex increase in arterial blood pressure to maintain cerebral perfusion pressure and b) if the response is driven by increased sympathetic nerve activity.

Using an instrumented conscious sheep model, we measured intracranial pressure (subdural) and arterial pressure (carotid) using solid‐state Millar catheters. Saline was infused into the lateral ventricle through an intracerebroventricular catheter to increase intracranial pressure in a ramp‐like fashion. The ICP ramp was repeated after 2 hours of ganglionic blocker hexamethonium.

The ramp increase in ICP led to a reflex linear increase in arterial pressure even when ICP remained within normal physiological levels (0–20 mmHg) (n=6, p<0.01). For example: a 10 mmHg increase in ICP lead to a 6.5 ± 1.4mmHg (n=6) increase in arterial pressure. Ganglionic blockade significantly reduced or abolished this increase in arterial pressure, suggesting mediation by increased sympathetic nerve activity. This is supported by preliminary direct renal sympathetic nerve recordings.

We believe that this increase in arterial pressure is via an active control mechanism to try to correct reduced cerebral perfusion caused by the increased intracranial pressure. We speculate that alterations in this response may lead to increased sympathetic nerve activity and worsen prognosis in cardiovascular disease.

Our preliminary data indicate that this response is active at physiological levels of intracranial pressure and not just at extreme levels where other reflexes such as the Cushing's response would be active. We propose that this response is an important novel addition to other known mechanisms protecting the brain from under‐perfusion.

Support or Funding Information

Supported by the Health Research Council of New Zealand
Significance of Endothelial ENaC in NO Synthesis and Blood Pressure
Authors: Ydegaard Rikke , Lærkegaard Hansen Pernille B. , Svenningsen Per , Oxlund Christina Stolzenburg , Jacobsen Ib Abildgaard , Jensen Boye L
Published: November 2023

The epithelial sodium channel, ENaC, is located in thevascular endothelial cells and inhibits eNOS and nitric oxide (NO) synthesis in single cells. Aldosterone‐MR stimulates activity of ENaC. We hypothesized: 1) Spironolactone (MR blocker) and amiloride (ENaC blocker) increase NO formation in hypertensive patients 2) ENaC‐antagonists amiloride and benzamil lead to an acute NO‐dependent blood pressure reduction in mice.

Methods

1) Nitrite/nitrate(surrogate for NO) was measured in urine/plasma before and after spironolactone (25–50 mg, 8 weeks (n=119)) or placebo and amiloride treatment (5–10 mg, 8 weeks n=60). 2) Indwelling catheters were implanted in the femoral artery and vein in wild type and eNOS deficient mice for undisturbed chronic blood pressure and heart rate recordings and infusion of drugs. 3 doses of amiloride (0.005–0.5 mg/kg) were administered in boli and blood pressure was recorded with high resolution for 10 minutes at each level. In wild type mice, acute effect of benzamil (0.05 mg/kg) was investigated two days after administration of amiloride.

Result

1) NOx/creatinineratio in urine decreased after spironolactone (P=0.01, n=58) with no difference after placebo. No differences in cGMP or NOx levels in plasma and urine were detected after treatment with amiloride 2) A significant acute and transient fall in blood pressure and heart rate was observed in mice receiving amiloride compared with vehicle (p=0.04, n=10), but not after benzamil. In order to investigate if nitric oxide was a part of the observed response, the effect of amiloride was investigated in eNOS deficient mice. Here, 0.05 and 0.5 mg/kg amiloride also significantly lowered blood pressure (n=9).

Conclusion

Administration of amiloride leads to an acute and transient fall in BP and HR in mice, indicating such rapid effects that other mechanisms than renal sodium excretion could be relevant. The observed response on BP was not dependent on eNOS and likely not mediated by ENaC. Amiloride and spironolactone are not likely to have antihypertensive action through endothelial ENaC.

Support or Funding Information

The Danish Heart Foundation
Hydrogen Sulfide Decreases Blood Pressure by Influencing the Excitability of Area Postrema Neurons
Authors: Smith Pauline M , Wang Susan J , Ferguson Alastair V
Published: November 2023

Hydrogen sulfide (H2S), an endogenous gaseous transmitter, has been shown to act in the central nervous system (CNS) to influence cardiovascular (CV) function. Microinjection of H2S into central autonomic control centers such as the paraventricular nucleus and subfornical organ has been shown to increase blood pressure (BP) and electrophysiological studies have demonstrated that H2S depolarizes neurons in these same regions. Depolarizing effects of H2S have also been demonstrated in the nucleus tractus solitarius, a medullary autonomic control center with reciprocal connections to the immediately adjacent area postrema (AP). The AP is a circumventricular organ (CVO) with well documented roles in cardiovascular regulation.

The present study was undertaken to examine the cardiovascular effects of microinjection of the H2S donor, sodium hydrogen sulfide (NaHS), into the AP of urethane anesthetized male Sprague Dawley rats and the cellular consequences of bath application of NaHS on the excitability of dissociated AP neurons. Microinjection (0.5 μl) of NaHS (10 μM) caused rapid, short duration (<90 sec) decreases in BP (mean area under the curve (AUC) =−389.2 ± 82.6 mmHg*sec, n=6) and HR (mean AUC = −6.6 ± 1.4 beats, n=6) while whole cell perforated patch electrophysiology revealed that bath application of NaHS (1mM) elicited solely hyperpolarizing responses (mean Δ in membrane potential = −3.6 ± 0.8 mV, n=8) in the majority (n=8/13) of AP neurons tested. These observations identify the AP as a CNS location at which H2S may act to influence cardiovascular regulation.

Support or Funding Information

Supported by Canadian Institutes of Health Research
Myeloid‐Derived Suppressor Cells in Adipose Tissue Homeostasis
Authors: Stivers Katlin Brooke , Chilton Paula , Beare Jason , Dale Jacob , Kaufman Christina L , Hoying James B
Published: November 2023

Under steady‐state conditions, adipose tissue (AT) is highly metabolic, using free fatty acid (FFA) oxidation reactions to maintain an appropriate energy balance within the body. Even at relatively low concentrations, the by‐products of these reactions, such as reactive oxygen species (ROS), can lead to tissue damage and have been implicated as being partially responsible for inducing the chronic AT inflammation seen in obesity. This inflammation is also known to increase insulin resistance in the adipose, leading to diabetes. A network of adipose‐resident immune cells copes with this ever present threat of FFA oxidation and ROS induced inflammation with potent immunosuppressive mechanisms which help to maintain immune homeostasis within the tissue. During obesity, this homeostasis is disrupted by the infiltration of inflammatory cells into the adipose. This abject phenotypic switch of adipose‐resident immune cells has brought to light the importance of understanding how these cells work to maintain this initial homeostasis in lean adipose and what signals lead to the degradation of this immunosuppressive network during obesity.

Initial flow cytometry data, from our lab's previous studies, revealed a functionally important population of CD11b+ Gr1+ stromal cells in the adipose. This surface marker combination, along with the anti‐inflammatory nature of lean adipose in general, led us to hypothesize that these cells were myeloid‐derived suppressor cells (MDSCs). MDSCs are a heterogeneous collection of immunosuppressive cells typically associated with cancer or chronic inflammatory diseases. We used flow cytometry to determine the surface marker expression profiles of the resident myeloid cells in the SVF, while CFSE and MTT assays were used to test the ability of SVF subsets to suppress proliferation of activated T cells. We found that approximately 2–3% of the adipose stroma expressed the classic CD11bhi Ly6Chi Ly6G− SSClow MDSC surface marker phenotype. Co‐culturing this population with activated T cells revealed their capacity to suppress T cell proliferation. Our findings suggest that there is a population of MDSCs resident in healthy adipose tissue. These potent immunosuppressive, endogenous adipose cells could represent a new therapeutic target for diabetes and obesity, as studies have already demonstrated that injections of culture‐derived MDSCs into obese mice increases insulin sensitivity and delays the development of metabolic syndromes.

Support or Funding Information

Kosair Charities
Wfs1 mutation makes mice more susceptible to metabolic changes caused by antipsychotics
Authors: Toots Maarja , Narvik Jane , Ottas Aigar , Plaas Mario , Vasar Eero
Published: November 2023

Mutations in WFS1 gene cause autosomal recessive disorder Wolfram syndrome (WS). While homozygous mutations in Wfs1 cause WS, studies have shown that heterozygous Wfs1 mutation carriers have increased risk for psychiatric disorders, type 2 diabetes and metabolic syndrome. Therefore, in this study, the effect of chronic administration of haloperidol (first generation antipsychotic) and olanzapine (second generation antipsychotic) on different metabolic parameters was investigated, using targeted mass‐spectrometry. Results show that heterozygous Wfs1 mutation mice have changes in their metabolism and are more receptive to changes caused by antipsychotics, which may indicate increased susceptibility to metabolic syndrome accompanying antipsychotic treatment.
Catecholamine Metabolism via Monoamine Oxidase Disrupts Oxidative Phosphorylation Efficiency in Glutathione Peroxidase‐4 Deficient Mouse and Human Myocardium
Authors: Nelson Margaret , Anderson Ethan J
Published: November 2023

Glutathione peroxidase‐4 (GPx4) is a selenoenzyme expressed in mitochondria and nuclear compartments which exclusively neutralizes lipid peroxides. Monoamine oxidase (MAO), an outer mitochondrial membrane‐bound enzyme responsible for catecholamine metabolism, produces a reactive catechol‐aldehyde and H2O2 upon substrate deamination. Current findings in human myocardial samples from our lab show that obese/diabetic patients exhibit greater maximal MAO activity and content, but lower GPx4 compared with nondiabetic patients, suggesting that both increased MAO and decreased GPx4 act to compound levels of reactive aldehyde species in diabetic myocardium. Our preliminary experiments using nitro blue tetrazolium staining have revealed protein modification by catechol‐aldehydes derived from norepinephrine (NE) and dopamine (DA) in whole heart lysate and isolated cardiac mitochondria from mice and humans, an effect mitigated by MAO inhibitors. We further examined the effects of catecholamine metabolism via MAO on ATP production and O2 consumption (OxPhos) in isolated cardiac mitochondria prepared from WT and GPx4‐deficient (GPx4+/−) mice, and in permeabilized myofibers prepared from human atrial myocardium. Both DA and NE significantly decreased ATP production and O2 consumption in GPx4+/− mice, but not WT, by 15% and 13%, respectively (p<0.05). Furthermore, the ATP/O ratio, an indicator of OxPhos efficiency, was also reduced in the GPx4+/− mice. In human atrial myocardium (n=11), NE significantly decreased ATP/O by ~35% (p<0.05), which was attenuated by MAO inhibitors. This translational study provides evidence that 1) catechol‐aldehydes are a unique form of oxidative post‐translational modification in the heart, and 2) hearts with diminished lipid peroxide‐scavenging capacity (e.g., obese/diabetic patients) are more susceptible to catecholamine‐induced disruption of mitochondrial OxPhos via MAO.

Support or Funding Information

R01HL122863
Xanthine oxidase inhibition attenuates arterial stiffness and prevents macrophage recruitment and M1 polarization in western diet fed male mice
Authors: Aroor Annayya , Jia Guanghong , Habibi Javad , Sun Zhe , Barron Brady J , Dongqing Chen , Martinez‐Lemus Luis A , Manrique Camila , Nistala Ravi , Whaley‐Connell Adam T , Meininger Gerald A , Sowers James R
Published: November 2023

Vascular stiffness associated with obesity is emerging as an independent risk factor for cardiovascular disease but the mechanisms and mediators of obesity associated vascular stiffness are not clearly understood. In this regard, hyperuricemia caused by enhanced xanthine oxidase activity is receiving more attention as an independent risk factor for cardiovascular disease under the setting of obesity caused by consumption of a western diet (WD) rich in fat and refined sugars. We hypothesized that both xanthine oxidase and uric acid are associated with vascular stiffness and that inhibition of xanthine oxidase activity would prevent arterial stiffness in a clinically relevant model of WD‐induced obesity and cardiovascular dysfunction. Four week‐old C57BL6/J male mice were fed a WD with excess fat (46%) and fructose (17.5%) with or without xanthine oxidase inhibitor, allopurinol (125mg/L), a potent xanthine oxidase inhibitor, for 16 weeks. Xanthine oxidase inhibition had no effect on increases in body weight, and fat mass but significantly (P<0.05) improved systemic insulin resistance. Xanthine significantly attenuated WD‐induced increase in plasma uric acid levels and xanthine oxidase activity in aorta. Endothelial stiffness and extracellular matrix/vascular smooth muscle stiffness as determined by atomic force microscopy were significantly increased (P<0.05) in WD fed mice and were significantly decreased by xanthine oxidase inhibition. WD induced increases in MCP‐1, CD11b, CD86, IL‐6 and oxidative stress, were prevented by xanthine oxidase inhibition. These results suggest xanthine oxidase inhibition suppresses macrophage recruitment and M1 polarization in WD fed mice. Taken together, these findings support the notion that increased xanthine oxidase activity in the vasculature and increased production of uric acid with intake of a WD promotes vascular stiffness, vascular inflammation and maladaptive immune response that lead to vascular injury.

Support or Funding Information

K08HL129074‐01 (to C.M.), VA I01 BX001981/BX/BLRD, R01‐HL073101 and R01‐HL107910 (to J.R.S).
Does Type 2 Diabetes Modulate Peripheral Cutaneous Vascular Function?
Authors: Akbari Pegah , Fujii Naoto , Boulay Pierre , Sigal Ronald J. , Kenny Glen P.
Published: November 2023

Aging is associated with impairments in cutaneous vasodilation during heat stress (Holowatz et al. Am J Physiol, 284:1662, 2003) which is exacerbated in individuals with type 2 diabetes mellitus (Charkoudian. Mayo Clin Proc, 78:603, 2003). This diabetes‐related modulation of cutaneous vasodilation may be due to attenuated peripheral sensitivity of skin vessels to receptor activation, though this has never been directly investigated. Thus, in this preliminary study, we aimed to evaluate the influence of type 2 diabetes mellitus on the modulation of cutaneous vasodilation in response to activation of the following receptors: cholinergic, purinergic, nicotinic and β‐adrenergic. In this preliminary study, seven older adults (59±9 years) with well‐controlled T2D (hemoglobin A1c, 7.5% ± 1.0, duration 8±5 years) and seven age‐matched counterparts without T2D (Controls) (61±8 years) participated in the study. Subjects were placed in a semi‐recumbent position in a thermoneutral room (~23 °C) during which time four intradermal microdialysis fibers were implanted on the dorsal side of the left forearm within the dermal layer of the skin. Approximately 90 min after the completion of the fiber insertion, cutaneous vascular conductance (CVC, measured as laser‐doppler flowmetry/mean arterial pressure) was then continuously monitored at the four forearm skin sites perfused via intradermal microdialysis with the following receptor agonists in a dose dependent manner: 1) methacholine (cholinergic receptor agonist, 5 doses: 0.0125, 0.25, 5, 100, 2000 mM), 2) ATP (purinergic receptor agonist, 5 doses: 0.03, 0.3, 3, 30, 300 mM), 3) nicotine (nicotinic receptor agonist, 5 doses: 1.2, 3.6, 11, 33, 100 mM), and 4) isoproterenol (β‐adrenergic receptor agonist, 3 doses: 1, 10, 100 μM). Increased CVC in response to all doses of methacholine administration were similar between groups (all P ≥ 0.15). Further, observed increases in CVC resulting from the five dose dependent infusion of ATP were similar in both Controls and T2D (all P ≥ 0.51), as was the case in all five doses of nicotine infusion (all P ≥ 0.32). Finally, similar increases in CVC were observed in both groups following the infusion of all three doses of isoproterenol (all P ≥ 0.20). Our preliminary findings show no attenuations in the sensitivity of the examined cholinergic and non‐cholinergic receptors in older adults with well‐controlled type 2 diabetes.

Support or Funding Information

Funding support provided by the Canadian Institutes of Health Research (grant no. 286363, funds held by Dr. Glen P. Kenny).
The Role of GPR40 (FFAR1) in Pharmacological vs Physiological Control of Incretin Secretion
Authors: Chen Yanyun , Song Min , Riley Jonathan , Reynolds Nichole , Hamdouchi Chafiq , Miller Anne
Published: November 2023

G protein‐coupled receptor 40 (GPR40), also known as free fatty acid receptor 1 (FFAR1), is a member of a family of lipid‐activated receptors that is highly expressed in the pancreas and enteroendocrine cells with lower levels expressed in the brain. GPR40 mediates medium and long chain fatty acid stimulated insulin secretion in the presence of elevated glucose levels. This glucose‐dependent insulin secretion property has established GPR40 agonists as attractive therapeutic agents for the treatment of type 2 diabetes.

We have shown previously that synthetic, small molecule GPR40 agonists normalize glucose levels during oral glucose tolerance tests performed in mice after acute oral administration. In addition, incretin levels, total and active GLP‐1 and GIP, are significantly elevated immediately following oral administration of the GPR40 agonists. These findings suggest that GPR40 may play an important role in the incretin secretion following a meal. The following studies were performed to address this hypothesis.

A synthetic GPR40 agonist, liquid mixed meal, or vehicle alone was administered orally at t = 0 minutes to wild type (WT) or GPR40 knockout (KO) mice and blood was collected for incretin determination at 0.5, 1.5 and 3 hours. Administration of the GPR40 agonist in WT mice led to a robust increase in active GLP‐1 with peak concentrations reaching 12 pg/ml at the 0.5 hour time point followed by levels falling to 5 pg/ml for the remainder of the study (3 hours). The changes in active GIP levels were similar. In contrast, elevations in GLP‐1 and GIP levels were absent in GPR40 KO mice demonstrating that the incretin effect following administration of the GPR40 agonist was GPR40 mediated.

The same experimental design was performed in mice orally administrated a liquid mixed meal. Active GLP‐1 levels peaked at the 1 hour time point (5.5 pg/ml) and fell to levels equivalent to those seen with the vehicle control at 6 hours. Changes in active GIP levels were mild demonstrating a peak concentration at 0.5 hour (0.75 pg/ml) and rapidly returning to normal levels. Interestingly, the same changes in active GLP‐1 and GIP were seen in the GPR40 KO mice compared to those in the WT mice. These findings demonstrate that the incretin secretion following a liquid mixed meal does not require GPR40, suggesting that other receptors are responsible for the meal‐stimulated elevation in active GLP‐1 and GIP.
Physiologic Mechanisms of Insulin Resistance During a Critical Postnatal Window of Development
Authors: Ortiz Rosa Isela , Chávez‐Maldonado Juan Pablo , Velasco Myrian , Larqué Carlos Alfonso , Hiriart Marcia
Published: November 2023

Introduction

Insulin resistance is a condition in which the insulin‐sensitive tissues cannot respond appropriately to hormone effects. Insulin resistance has been associated with pathologies however, insulin resistance is present in physiologic states such as pregnancy and adolescence. Our work group analyzed changes on glucose and insulin levels during postnatal development. We found that males and females rats near the end of lactation show insulin resistance, characterized by hyperglycemia and hyperinsulinemia, which disappears later. This state associates with a critical window in pancreatic development.

Objective

To determine the mechanisms involved in the insulin resistance observed in the weaning period in males and females rats.

Methods

We used male and female Wistar rats at 20 postnatal days (20 pd) and healthy adult rats (250–280 g). We analyzed fasting glucose and insulin levels and made glucose tolerance test (GTT) and insulin sensitivity test (ITT). We performed insulin‐stimulated glucose uptake assays in adipose tissue and analyzed P70S6K‐specific inhibitory sites of serine and threonine phosphorylation of IRS‐1 in adipose tissue and skeletal muscle.

Results

We observed increased fasting glucose concentrations and insulin levels in both female and male at 20 pd, compared to adult rats. The blood glucose levels in GTT in male and female rats of 20 pd were higher to those of adults, along with the test. Systemic insulin response was less in male and female at 20 pd rats, compared to adults. Insulin‐stimulated glucose uptake in adipose tissue was lower in 20 pd, compared to adults. P70S6K phosphorylation in Thr389 and IRS‐1 phosphorylation in Ser1101 were higher in adipose tissue at 20 pd in males and females rats than in adult rats.

Conclusion

Wistar males and females rats at 20 pd have high plasma levels of glucose and insulin compared with adult rats. These data together with the lack of response in the ITT, a higher phosphorylation of P70S6K kinase and IRS1 at a inhibitory sites and lower insulin‐stimulated glucose uptake could be involved in the development of physiological insulin resistance.

Support or Funding Information

This work was partially financed by CONACYT CB‐253222 and DGAPA IN213114
SIRT3 Inactivation Impairs Endothelial Function and Promotes Vascular Calcification in Hypercholesterolemic Mice
Authors: Roos Carolyn M , Zhang Bin M , Fujimoto Hirokazu , Hagler Michael A , Miller Jordan D
Published: November 2023

Recent work by other investigators have shown that SIRT3 is an important regulator of endothelial function in obese patients or animals, as reductions in SIRT3 drive increased mitochondrial ROS (mtROS) production. The role of SIRT3 in atherosclerotic mice, however, remains unclear. Therefore, we hypothesized genetic inactivation of SIRT3 in hypercholesterolemic mice will impair vascular function, reduce antioxidant gene expression, and increase pro‐osteogenic signaling. We used Ldlr−⁄−/ApoB100/100 mice that were either wild‐type (LA‐SIRT3+/+) or null for SIRT3 (LA‐SIRT3−⁄−). After 12 weeks of western diet feeding, we assessed endothelial function of aorta using isolated organ bath chambers and measured changes in gene expression using qRT‐PCR. Consistent with our hypothesis, loss of SIRT3 significantly impaired relaxation to acetylcholine (Ach) (LA‐SIRT3−⁄−: 48.2%±3.0, LA‐SIRT3+/+: 57.6%±1.7). Incubation with Mito‐Tempol significantly improved endothelial‐dependent relaxation in LA‐SIRT3−⁄− mice (57.0%±2.9), but not in LA‐SIRT3+/+ mice. Interestingly, losses of SIRT3 did not alter expression of MnSOD, ecSOD, or CuZnSOD. Gene expression of the late‐stage osteoblast marker, osterix, tended to be increased in LA‐SIRT3−⁄− mice compared to wild‐type mice (p = 0.06). Collectively, losses of SIRT3 impair endothelial function and tend to increase osteogenic signaling in severe hypercholesterolemia. Importantly, these functional and molecular changes and may result from increased mtROS, which may serve as a viable therapeutic target to slow pathological processes during progression of atherosclerosis.

Support or Funding Information

NHLBI R01‐HL111121
Loss of endothelial Fto is protective against metabolic disease
Authors: Krüger Nenja , Biwer Lauren A , Good Miranda E , Rüther Ulrich , Isakson Brant E
Published: November 2023

Genetic variations within the fat mass and obesity associated gene (FTO) are known to correlate with obesity and hypertension in humans, demonstrating a possible key role in vascular health. Therefore, we hypothesized that FTO may be important at the vascular level. Because endothelial cells (EC) are known to have a central role in the development of obesity‐induced metabolic changes and hypertension, we wanted to investigate if murine loss of endothelial Fto may influence obesity‐induced pathophysiological changes. Therefore, EC‐specific Fto deficient mice were generated using a tamoxifen‐inducible Cre system. To analyze obesity‐dependent effects, mice were either fed a normal chow (NC) or a high fat diet (HFD) for 12 weeks. Regardless of the endothelial deletion of Fto, the mice still gained a significant amount of weight and were obese, indicating the Fto gene in endothelium did not inhibit weight gain like global Fto deficiency in mice. Next we determined whether loss of endothelial Fto affected obesity‐induced metabolic changes in HFD‐fed mice. While serum lipid analysis showed no differences in dependence of endothelial Fto, glucose and insulin tolerance was significantly improved in HFD‐fed mice without endothelial Fto. Due to the importance of EC in vascular tone of resistance arteries, we isolated 3rd order mesenteric arteries from NC‐ and HFD‐fed mice with and without endothelial Fto and performed pressure curves to analyze myogenic tone. While loss of endothelial Fto did not influence myogenic tone in resistance arteries of NC‐fed mice, remarkably, endothelial Fto deletion provided protection from HFD‐induced changes on myogenic tone. Subsequently, blood pressure was measured in HFD‐fed mice with and without endothelial Fto using radiotelemetry and it revealed that loss of endothelial Fto significantly reduced blood pressure. Collectively, these data suggest that loss of endothelial Fto was protective against HFD‐induced changes on myogenic tone in mesenteric arteries and subsequently reduced blood pressure in vivo without influencing body weight. Furthermore, glucose homeostasis, but not serum lipid levels, were specifically improved in HFD‐fed mice suggesting that endothelial Fto is not only important in the development of obesity‐induced hypertension, but also in the development of insulin resistance and hyperglycemia.

Support or Funding Information

AHA, NHLBI and IRTG1902
Comorbidity Between Metabolic Syndrome and Depression Exacerbates Oxidative Stress and Inflammation in the Brain
Authors: Branyan Kayla , DeVallance Evan , Asano Shinichi , Lemaster Kent , Allen Colton , Perry Sydney , Frisbee Jefferson , Chantler Paul
Published: November 2023

Major cardiovascular disease risk factors, such as metabolic syndrome (MetS) and depression, are associated with a chronic inflammatory state and increased oxidative stress. As in other peripheral microvessels, inflammation and oxidative stress impair the microvasculature of the brain, leading to increased risk of stroke. It is unclear how the comorbidity between MetS and depression affect the inflammatory and oxidative stress conditions in the brain. To determine if depression and MetS together further exacerbate cerebral inflammation and oxidative stress compared to either disease alone, markers of oxidative stress were assessed through dihydroethidium (DHE) and DAF‐FM assays along with PCR and ELISA to evaluate the inflammatory response in the brains of obese Zucker rats (OZRs), a model of MetS, and lean Zucker rats (LZRs), healthy controls. Reactivity of the middle cerebral artery (MCA) was used as a measure of vascular function. The MCA was isolated and then hung in a pressurized vessel myobath to be exposed to increasing doses of acetylcholine (Ach) to test endothelium‐dependent dilation. OZRs had increased superoxide production in the basilar arteriole (p<0.05), which was exacerbated with UCMS. Similarly, the OZRs and OZR‐UCMS had a significant decrease in nitric oxide production as compared to both LZR groups. Concentrations of anti‐inflammatory cytokines, IL‐4 and IL‐10, were reduced in OZRs and further reduced in OZR‐UCMS. Inflammatory marker (TNFα and CD68) mRNA abundance was elevated in OZRs and to a higher extent in OZR‐UCMS. This oxidative stress and inflammation contributed to decreased MCA reactivity in OZR controls from that of LZR controls in response to Ach (p<0.01). In both LZRs and OZRs, MCA dilation was further attenuated with UCMS as compared to their respective control values. These data show that the comorbidity of MetS and depression exacerbates the oxidative stress and inflammatory states associated with each disease separately. Our results suggest that the cerebral environment is drastically changed during this comorbidity and leads to vascular dysfunction, which can cause an increased risk of poor cardiovascular outcomes.

Support or Funding Information

National Institute of Health (5P20GM109098)
Amyloidogenic Medin Induces Endothelial Dysfunction and Vascular Inflammation Through the Receptor for Advanced Glycation Endproducts
Authors: Migrino Raymond Q. , Davies Hannah , Truran Seth , Karamanova Nina , Franco Daniel A. , Serrano Geidy , Beach Thomas , Madine Jillian
Published: November 2023

Aims

Medin is a common amyloidogenic protein in humans, especially in older individuals. It has been implicated in the development of arterial degenerative changes associated with vascular aging. The mechanisms underlying medin‐induced vascular injury remain unknown. The aims are to assess medin's effects on human peripheral and central arteriole endothelial function, nitrative stress and vascular inflammation and identify potential mechanisms of injury.

Methods and Results

Ex‐vivo human adipose and leptomeningeal arterioles were exposed (1 hour) to medin (0.1, 1 or 5 μM) without or with FPS‐ZM1 (100 μM, receptor for advanced endglycation products [RAGE]‐specific inhibitor) and endothelial function (acetylcholine dilator response) was compared to baseline control. Medin's effect on adipose arteriole smooth muscle function was tested by comparing post‐exposure dilator response to DETA NONOate (a nitric oxide [NO] donor) compared to baseline response. Human umbilical vein endothelial cells were exposed to medin (20 hours) without or with FPS‐ZM1 and oxidative and nitrative stress, cell viability and pro‐inflammatory signaling measures were obtained. Medin caused impaired endothelial function (versus baseline response to 10−4 M acetylcholine: −38.6±6.7% and −35.0±8.9% in adipose and leptomeningeal arterioles, respectively, each p<0.05) in human adipose and leptomeningeal arterioles. Medin did not affect adipose arteriole dilator response to DETA NONOate. Medin‐induced endothelial dysfunction was reversed by RAGE inhibitor. Medin decreased endothelial cell NO production and cell viability, increased peroxynitrite, superoxide production and profoundly increased gene and protein expression of interleukin‐6 and interleukin‐8 via activation of nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NFκB). These effects were reversed by RAGE inhibitor.

Conclusions

Medin causes human microvascular endothelial dysfunction, oxidative and nitrative stress and pro‐inflammatory signaling mediated via RAGE. The discovery of this novel mechanism of injury may have implications in our understanding of and development of treatments to reverse vascular aging.

Support or Funding Information

The work was supported by National Institutes Health (NIA R21AG044723, NINDS U24NS072026, NIA P30AG19610, NIA RO1AG019795); Veterans Affairs Merit grant (I01BX007080); British Heart Foundation (FS/12/61/29877); the Arizona Department of Health Services (contrast 211002); the Arizona Biomedical Research Commission (4001, 0011, 05‐901 and 1001); Michael J. Fox Foundation for Parkinson's Research and Amyloidosis Foundation. The study was based on work supported by the Department of Veterans Affairs and VA employment.
Dynamics of mitochondrial function changes in the cerebral vasculature of type 2 diabetic rats
Authors: Merdzo Ivan , Rutkai Ibolya , Sure Venkata N. L. R. , McNulty Catherine A. , Katakam Prasad V. , Busija David W.
Published: November 2023

Mitochondrial dysfunction has been suggested as a potential underlying cause of pathological conditions associated with type 2 diabetes (T2DM). We have previously shown that mitochondrial respiration levels assessed by the Seahorse XFe24 analyzer in large cerebral arteries, and mitochondrial protein levels measured by western blot in cerebral arteries and microvessels, were similar in insulin resistant, non‐diabetic, Zucker obese rats and lean control rats (Am J Physiol 310:H830‐838, 2016). In the present study we extended our investigations into mitochondrial dynamics and superoxide production of the cerebral vasculature of 14 week old Zucker diabetic fatty obese rats (ZDFO) with early T2DM.

Body weight and blood glucose levels were significantly higher in the ZDFO compared to the control, lean (ZDFL) rats (347.2 ± 10.18 g vs. 320.4 ± 3.9 g, n = 20, P < 0.05; and 26.33 ± 1.56 mmol/L vs. 9.18 ± 0.46 mmol/L, n = 20, P < 0.05; respectively). In addition, basal mitochondrial respiration and proton leak were significantly decreased in the large cerebral arteries of the ZDFO rats compared with ZDFL (181.4 ± 10.93 pM/min/μg protein vs. 129.7 ± 8.211 pM/min/μg protein, n = 59, P < 0.05; and 149.3 ± 10.41 pM/min/μg protein vs. 100.3 ± 7.56 pM/min/μg protein, n = 59, P < 0.05; respectively). Furthermore, superoxide production was significantly increased in the microvessels of the ZDFO group (2355 ± 628 vs. 208 ± 48, n = 10, P < 0.05). The expression of mitochondrial proteins total MnSOD and VDAC were significantly lower in the cerebral microvessels (90.35 ± 9.13% vs. 123.1 ± 12.77%, n = 13, P < 0.05; and 23.63 ± 2.62% vs. 30.54 ± 2.07%, n = 12, P < 0.05; respectively), and levels the acetylated or active form of MnSOD were significantly reduced in large arteries of ZDFO group (89.72 ± 3.17% vs. 52.23 ± 6.01%, n = 12, P < 0.05). Despite evidence of increased oxidative stress in ZDFO, exogenous superoxide dismutase was not able to restore mitochondrial respiration in ZDFO rats.

Our results show for the first time that mitochondria of large arteries and microvessels are adversely affected in the cerebral vasculature during the early stages of T2DM.

Support or Funding Information

This work was supported by National Institutes of Health grants (DWB: HL‐077731and HL093554), Louisiana Board of Regents Support Fund‐Research Competitiveness Subprogram (PVK: LEQSF(2014‐17)‐RD‐A‐11), American Heart Association National Center NRCP Scientist Development Grant (PVK: 14SDG20490359), and American Heart Association Post‐Doctoral Fellowship Grant (IR: 15POST23040005). This research was also supported by the Louisiana Board of Regents Endowed Chairs for Eminent Scholars program (DWB).
Can exercise protect against chronic stress induced cerebral microvessel changes in Lean and Obese Zucker rats?
Authors: Sheets Whitney Jordan , Asano Shinichi , Dawson Mariah , Ashman Melissa , Clayton Stuart , Brooks Steven , Frisbee Jefferson , Chantler Paul
Published: November 2023

Background

Chronic stress‐induced depression is a major cardiovascular risk factor, and metabolic Syndrome (MetS) is linked with depression. The relationship among chronic stress/depression (UCMS), MetS, CVD, and cerebrovascular dysfunction is poorly understood. We determined how UCMS and Exercise training affected cerebral microvessels density (MVD), and the pro (nitric oxide, VEGF, STAT3) and anti (TSP‐1) angiogenic factors. We hypothesize that UCMS will decrease MVD in Obese Zucker Rats (OZR = MetS), and exercise training will prevent this rarefaction by increasing pro‐angiogenic mediators, while reducing anti‐angiogenic factors including oxidative stress.

Methods

Nine‐week old lean rats (LZR) and OZR were randomly assigned (n=6/group) to: Control(C), Exercise (EX), subjected daily to an UCMS, and EX+UCMS for 8 weeks. At 17 weeks, brains were removed and snap frozen in Tissue Tek medium. Cortex MVD were measured by fluorescence IHC (rat anti‐CD31 antibody). Oxidative stress was assessed in cerebral tissue by ELISA technique (anti 3‐Nitrotyrosine antibody) and by DHE. Cerebral inflammatory and angiogenic markers for STAT3, VEGF and TSP‐1 were measured by western blot or ELISA.

Results

MVD was decreased (−31%, p<0.05) in the cortex from OZR‐UCMS vs. OZR control. Exercise training attenuated UCMS induced decreased MVD in OZR by 41%. In contrast, there were no significant differences (8% decrease to 4% increase) in MVD among LZR groups. In both LZR and OZR UCMS increased oxidative stress, and decreased nitric oxide in the cerebral vessels. Although VEGF results were not yet significant, a slight decrease was seen in OZR UCMS that was recovered with exercise training in UCMS. Initial data also suggests that TSP‐1, which is higher in OZR vs. LZR control, is also influenced by UCMS, and Ex. Compared to LZR‐C, LZR‐UCMS had an 74% increase in STAT3. Although both OZR‐C and OZR UCMS exhibited significantly increased STAT3 phosphorylation compared to LZR‐C, there was no difference between OZR‐C and OZR UCMS. Both LZR‐Ex and OZR‐Ex attenuated UCMS induced elevated STAT3 activity.

Conclusion

Taken together, UCMS drives the development of decreased brain MVD in OZR. Chronic stress and obesity both independently increased STAT3 signaling in the brain, which was then abrogated by exercise in both lean and obese animals. Exercise also prevented the onset of cortical rarefaction seen in OZR‐UCMS. We are further exploring TSP1 and VGEF pathways.

Support or Funding Information

Acknowledgements: American Physiological Society Short‐Term Experience for Underrepresented Persons Fellowship and the National Institute of Diabetes and Digestive and Kidney Diseases for helping to fund this project. National Institute of Health (5P20GM109098), and National Institute of General Medical Sciences Grant U54 GM‐104942.