Systemic Vasculature Research Articles

A 4-Chamber Model Heart with 3D Printed Silicone Aorta and Peripheral Arteries to Simulate the Human Cardiovascular System in a Mock Circulatory Loop

Purpose Mock circulatory loops (MCL) provide an in vitro platform to evaluate hemodynamic performance of cardiovascular (CV) medical devices. Designing a tunable MCL that mimics patient-specific physiology could both accelerate evaluation of medical devices prior to in vivo studies, and provide patient-specific testing in vitro. We developed an MCL that mimics human CV physiology with a 4-chamber model heart and 3D printed silicone vasculature based on patient-specific images. Methods The model heart is composed of the left/right atria & ventricles, and atrioventricular and aortic/pulmonary valves. The atrial/ventricular septa are designed as circular silicone-membranes to simulate physiologic septal deformations which occur in the native heart with a pressure differential between chambers. Each of the four chambers is made of acrylic and driven by linear motor. A displacement transducer connected to each linear motor can measure the volume change in each chamber while pressure is simultaneously recorded. A systemic vasculature model is fabricated out of 3D printed silicone based on patient-derived images. The compliance of the silicone selected is designed to match the arterial stiffness of the human vascular system. Results The assembled model heart contains atria, ventricles, and systemic vasculature. Each atrial and ventricular chamber can accommodate a maximum volume of 98 mL and 192 mL, respectively. The area and thickness of the silicone septum between the atria are 890 mm2, and 1.8 mm. The ventricular septal area and thickness are 1590 mm2 and 4.5 mm. Changes in pressure differential between the ventricular chambers affect septal configuration to incorporate ventricle-ventricle interaction effects. The linear motors are driven based on the elastance functions derived from pressure-volume (PV) loops obtained in the 4 chambers to enable the 4-chamber heart to physiologically respond to preload and afterload changes -in a similar fashion to the native heart. Conclusion This is the first MCL system that mimics human heart size, complexity and physiology and contains a human vascular circuit that can be tuned to patient-specific parameters. By including a physiologic control system governing the linear motors we can evaluate patient-specific hemodynamics, including the Frank-Starling mechanism in this MCL.

Monocrotaline pyrrole induces pulmonary endothelial damage through binding to and release from erythrocytes in lung during venous blood reoxygenation.

Monocrotaline has been widely used to establish an animal model of pulmonary hypertension, most frequently in rats. An important feature of this model resides in the selectivity of monocrotaline injury toward the pulmonary vascular endothelium versus the systemic vasculature when administrated at standard dosage. The toxic metabolite of monocrotaline, monocrotaline pyrrole, is transported by erythrocytes. This study aimed to reveal whether partial pressure of oxygen of blood determined the binding and release of monocrotaline pyrrole from erythrocytes in rats with one subcutaneous injection of monocrotatline at the standard dosage of 60 mg/kg. Our experiments demonstrated that monocrotaline pyrrole bound to and released from erythrocytes at the physiological levels of partial pressure of oxygen in venous and arterial blood, respectively, and then aggregated on pulmonary artery endothelial cells. Monocrotaline pyrrole-induced damage of endothelial cells was also dependent on partial pressure of oxygen. In conclusion, our results demonstrate the importance of oxygen partial pressure on monocrotaline pyrrole binding to erythrocytes and on aggregation and injury of pulmonary endothelial cells. We suggest that these mechanisms contribute to pulmonary selectivity of this toxic injury model of pulmonary hypertension.

Tobacco Education: Reduced Risk for Peripheral Artery Disease

Peripheral arterial (vascular) disease (PAD/PVD) in its association with significant mortality and morbidity rates has become a significant public health concern. One of the most influential risk factors for PAD is tobacco use, which carries a 3-4-fold increase d risk for PAD often presenting as severe disease. The diagnosis of PAD is usually made a decade earlier in smokers than nonsmokers. The amputation rates in patients with PAD who smoke is twice higher than those that have never smoked. Smoking elevates the risk for PAD several folds and approximately 90% of persons with PAD have a history of smoking. Although the precise mechanism by which chronic smoking induces vascular disease is not entirely understood, growing evidence shows that impairment of endothelial morphology and function plays a crucial role in the pathogenesis of vascular disease. Oxidants, delivered by cigarette and deposited in pulmonary vessels through the systemic vasculature, activate superoxide producing enzymes within the vascular wall via oxidative stress and might be the cause of endothelial dysfunction and dysregulation of endothelial barrier. The World Health Organization (WHO) estimates that there are currently 1.1 billion tobacco smokers’ worldwide ages 15 years and older. Recently, smoking-related death have been said to account for 4.9 million persons per year worldwide. Tobacco use is considered the most important preventable vascular risk factor for PAD in men and women. The association between smoking and PAD is even stronger than that between smoking and coronary disease. The connection between smoking and PAD was identified in 1911 when Erb reported that intermittent claudication was three times more common in smokers and six times more common in heavy smokers in comparison with nonsmokers.

Loss of Estrogen Receptor Alpha (ERα) Exacerbates Experimental Pulmonary Arterial Hypertension (PAH)

Background and Hypothesis: PAH is a sexually dimorphic cardiopulmonary disease characterized by excessive vasoconstriction and pulmonary artery remodeling, leading to right ventricular (RV) failure and death. While women are more likely to develop PAH, they exhibit more favorable hemodynamics and increased survival compared to men. These improved outcomes in women with PAH have been linked to protective effects of the sex steroid 17ß-estradiol (E2). While E2’s receptor ERα is protective in the systemic vasculature, its function in the cardiopulmonary system has not been explored. We hypothesized that loss of ERα exacerbates PAH.
 Experimental Design: Studies were performed in male and female wild type (WT) or ERα loss-of-function mutant (ERαmut) rats with monocrotaline (MCT)-induced PAH as well as disease-free controls. We quantified hemodynamics (RV catheterization), RV structure and function (echocardiography) and pulmonary artery remodeling (Verhoff-van Giesson staining). Lung tissues were analyzed for expression of pulmonary vascular homeostatic regulators BMPR2 and apelin and pro-survival regulator ERK (Western blot). P<0.05 (ANOVA) was considered significant.
 Results: ERαmut rats did not differ hemodynamically from WT controls. However, after MCT administration, ERαmut rats exhibited more severe disease than WT MCT rats (demonstrated by increased RV hypertrophy, RV systolic pressure, total pulmonary resistance index, as well as decreased cardiac index and stroke volume index (p<0.05). Interestingly, female ERαmut MCT rats exhibited more severe disease than their male counterparts. Apelin expression decreased in ERαmut MCT lungs compared to WT and ERαmut controls (p<0.05). Furthermore, female WT MCT lungs exhibited preserved apelin expression compared to male WT MCT (p<0.05). BMPR2 expression in ERαmut MCT lungs decreased compared to WT and ERαmut controls, as well as WT MCT (p<0.05).
 Conclusion: Loss of ERα aggravates MCT-PAH, indicating that ERα exerts protective effects in the cardiopulmonary system. Harnessing ERα signaling may represent a novel treatment strategy for women and men with PAH.

365. Maternal and fetal consequences of preeclampsia

Vascular dysfunction in women with preeclampsia is complex. Placenta-derived circulating factors and/or the maternal response to these factors contribute to vascular endothelial cell dysfunction. Risk factors for preeclampsia include many conditions such as maternal aging, pre-existing hypertension, obesity and diabetes. The heterogeneity of this population, including the diversity of circulating factors such as oxLDL, TNF α , and advanced glycation end products, has contributed to the overall complexity of targeting a specific pathway for therapeutic prevention and/or intervention. Interestingly, these circulating factors are also among the factors known to upregulate and/or activate the lectin-like oxidized LDL receptor-1 (LOX-1). Our laboratory has shown that LOX-1 is increased in the systemic vasculature of women with preeclampsia and contributes to vascular oxidative stress. Moreover, our rat model of reduced uteroplacental perfusion (RUPP), which has preeclampsia-like signs, demonstrates increased LOX-1 expression, increased vascular oxidative stress and ultimately vascular dysfunction. Since LOX-1 has multiple ligands, it may be a point of convergence for many plasma factors known to be elevated in preeclampsia; including placental-derived microparticles; a current area of investigation. Notably, pregnancy has an important influence on both short- and long-term cardiovascular outcomes for women and their offspring. One risk factor for preeclampsia is maternal age, an important factor considering that the age at which women experience their first pregnancy has steadily increased through the decades. We use aged female rats (9.5 months; equivalent to ∼35 yr old human), to investigate the impact of maternal aging on later-life maternal and offspring cardiovascular health. Our data show that advanced maternal age worsens postpartum vascular function and that offspring born from aged dams have an altered cardiovascular risk profile that is sex-specific. These data further illustrate pregnancy as a window of opportunity to assess both maternal and offspring future cardiovascular risk.

249. BKCa channels; a new target to treat maternal hypertension?

Introduction Severe hypertension is a common indication for delivery amongst women with pre-eclampsia (PE), contributing to iatrogenic prematurity and its associated complications. Large-conductance calcium-activated potassium channels (BKCa), which promote vasorelaxation, are downstream targets of many vasoactive agents, including nitric oxide and angiotensin II. Estrogen-mediated increases in uterine artery BKCa activity are an important physiological adaptation to normal pregnancy. However, the role of BKCa has not been assessed in the wider systemic vasculature in either normal pregnancy or pregnancy pathologies. Outwith pregnancy, BKCa activity is reduced in association with PE risk factors, including age, hypoxia and diabetes. Furthermore, 5,6-EET, a BKCa inhibitor, is reported to be increased in the serum of women with PE. Objectives To determine the effect of BKCa activation on systemic resistance arteries of pregnant women and whether this is different in PE. Methods Resistance arteries ( Results NS11021 induced a substantial relaxation of omental resistance arteries obtained from women with either NP (N0; final relaxation Control 23 ± 8% Vs. NS11021 76 ± 4%; p Discussion Systemic arteries from women with both NP and PE relax in response to activation of BKCa channels. Direct BKCa activation provides an opportunity to bypass the inherent vascular dysfunction of PE to achieve comparable vasorelaxation in women with PE. On-going work will determine whether BKCa-activating compounds can offer a new treatment for maternal hypertension in PE.

Spontaneous pulmonary hypertension in genetic mouse models of natural killer cell deficiency.

Natural killer (NK) cells are cytotoxic innate lymphoid cells with an established role in the regulation of vascular structure in pregnancy and cancer. Impaired NK cell function has been identified in patients with pulmonary arterial hypertension (PAH), a disease of obstructive vascular remodeling in the lungs, as well as in multiple rodent models of disease. However, the precise contribution of NK cell impairment to the initiation and progression of PAH remains unknown. Here, we report the development of spontaneous pulmonary hypertension in two independent genetic models of NK cell dysfunction, including Nfil3−/− mice, which are deficient in NK cells due to the absence of the NFIL3 transcription factor, and Ncr1-Gfp mice, which lack the NK activating receptor NKp46. Mouse models of NK insufficiency exhibited increased right ventricular systolic pressure and muscularization of the pulmonary arteries in the absence of elevated left ventricular end-diastolic pressure, indicating that the development of pulmonary hypertension was not secondary to left heart dysfunction. In cases of severe NK cell impairment or loss, a subset of mice failed to develop pulmonary hypertension and instead exhibited reduced systemic blood pressure, demonstrating an extension of vascular abnormalities beyond the pulmonary circulation into the systemic vasculature. In both mouse models, the development of PAH was linked to elevated interleukin-23 production, whereas systemic hypotension in Ncr1-Gfp mice was accompanied by a loss of angiopoietin-2. Together, these results support an important role for NK cells in the regulation of pulmonary and systemic vascular function and the pathogenesis of PAH.

Effects of aging on liver microcirculatory function and sinusoidal phenotype.

The socioeconomic and medical improvements of the last decades have led to a relevant increase in the median age of worldwide population. Although numerous studies described the impact of aging in different organs and the systemic vasculature, relatively little is known about liver function and hepatic microcirculatory status in the elderly. In this study, we aimed at characterizing the phenotype of the aged liver in a rat model of healthy aging, particularly focusing on the microcirculatory function and the molecular status of each hepatic cell type in the sinusoid. Moreover, major findings of the study were validated in young and aged human livers. Our results demonstrate that healthy aging is associated with hepatic and sinusoidal dysfunction, with elevated hepatic vascular resistance and increased portal pressure. Underlying mechanisms of such hemodynamic disturbances included typical molecular changes in the cells of the hepatic sinusoid and deterioration in hepatocyte function. In a specific manner, liver sinusoidal endothelial cells presented a dysfunctional phenotype with diminished vasodilators synthesis, hepatic macrophages exhibited a proinflammatory state, while hepatic stellate cells spontaneously displayed an activated profile. In an important way, major changes in sinusoidal markers were confirmed in livers from aged humans. In conclusion, our study demonstrates for the first time that aging is accompanied by significant liver sinusoidal deregulation suggesting enhanced sinusoidal vulnerability to chronic or acute injuries.

From pregnancy to cardiovascular disease: Lessons from relaxin-deficient animals to understand relaxin actions in the vascular system.

Early maternal vascular adaptations to pregnancy are predominantly driven by changes in vascular tone, reactivity, and remodeling. Failure of the maternal systemic vasculature to adapt sufficiently can lead to serious complications of pregnancy. The hormone relaxin is widely recognized for its contribution to the essential renal and systemic hemodynamic adaptations in early pregnancy through direct actions on the maternal vasculature. Studies in relaxin gene knockout mice revealed that endogenous relaxin is not only a "pregnancy hormone" but has pleiotropic actions in various tissues in males and non-pregnant females. There is strong interest in relaxin's actions in the vasculature and its utility in the treatment of vascular diseases. Relaxin treatment in rodents for 2-5days or acute intravenous injection enhances endothelium-dependent relaxation and decreases myogenic tone in resistance arteries. These vascular actions are prolonged, even in the absence of circulating relaxin, and are underpinned by the production of endothelium-derived relaxing factors including nitric oxide, endothelium-derived hyperpolarization, and prostacyclin. Relaxin is also capable of remodeling the vascular wall in a variety of blood vessels in disease conditions. Lessons learned in pregnancy research have aided studies investigating the potential therapeutic potential of relaxin in cardiovascular disease.

Vascular type 1 angiotensin receptors control blood pressure by augmenting peripheral vascular resistance in female mice.

Angiotensin II (ANG II) is a major mediator of hypertension pathogenesis. In addition, there are well-documented differences in expression of the renin-angiotensin system (RAS) components and ANG II responses between males and females, which may explain sex differences in blood pressure (BP) and hypertension epidemiology. We previously showed that type 1A angiotensin (AT1A) receptors in vascular smooth muscle cells (VSMCs) play a critical role in BP regulation and hypertension pathogenesis, but these studies were carried out in male mice. Therefore, the major goal of the current studies was to examine the impact of VSMC AT1A receptors on BP and hypertension pathogenesis in female mice. We found that elimination of VSMC AT1A receptors in female mice reduced (≈8 mmHg) baseline BP without altering sodium sensitivity. The severity of ANG II-induced hypertension was diminished (≈33% reduction in BP), particularly during the last 2 wk of chronic ANG II infusion, compared with controls, but natriuresis was not altered during the first 5 days of ANG II infusion. Urinary norepinephrine levels were enhanced in female SMKO compared with control mice. There was a virtually complete elimination of ANG II-induced kidney hemodynamic responses with attenuation of acute vasoconstrictor responses in the systemic vasculature. These findings demonstrate that direct vascular actions of AT1A receptors play a prominent role in BP control and hypertension pathogenesis in female mice.

Relaxin as a Therapeutic Target for the Cardiovascular Complications of Diabetes.

Cardiovascular complications are the major cause of mortality in patients with diabetes. This is closely associated with both macrovascular and microvascular complications of diabetes, which lead to organ injuries in diabetic patients. Previous studies have consistently demonstrated the beneficial effects of relaxin treatment for protection of the vasculature, with evidence of antioxidant and anti-remodeling actions. Relaxin enhances nitric oxide, prostacyclin and endothelium-derived hyperpolarization (EDH)-type-mediated relaxation in various vascular beds. These effects of relaxin on the systemic vasculature, coupled with its cardiac actions, reduce pulmonary capillary wedge pressure and pulmonary artery pressure. This results in an overall decrease in systemic and pulmonary vascular resistance in heart failure patients. The anti-fibrotic actions of relaxin are well established, a desirable property in the context of diabetes. Further, relaxin ameliorates diabetic wound healing, with accelerated angiogenesis and vasculogenesis. Relaxin-mediated stimulation of vascular endothelial growth factor (VEGF) and stromal cell-derived factor 1-α, as well as regulation of metalloproteinase expression, ameliorates cardiovascular fibrosis in diabetic mice. In the heart, relaxin is a cardioprotective molecule in several experimental animal models, exerting anti-fibrotic, anti-hypertrophy and anti-apoptotic effects in diabetic pathologies. Collectively, these studies provide a foundation to propose the therapeutic potential for relaxin as an adjunctive agent in the prevention or treatment of diabetes-induced cardiovascular complications. This review provides a comprehensive overview of the beneficial effects of relaxin, and identifies its therapeutic possibilities for alleviating diabetes-related cardiovascular injury.

Smooth Muscle PPARγ Mutation Causes Impaired Renal Blood Flow and Salt‐Sensitive Hypertension

Failure to vasodilate in systemic and renal vasculature in response to salt loading is a hallmark of salt sensitivity in hypertensive humans and experimental models. Human subjects carrying dominant negative mutations in PPARγ exhibit hypertension while global PPARγ activation consistently lowers blood pressure (BP). To determine whether tissue‐specific actions of PPARγ regulate vascular function, salt sensitivity and BP, we studied mice specifically expressing a human hypertension‐causing mutation in PPARγ (P467L) selectively in smooth muscle (S‐P467L). S‐P467L transgenic mice and non‐transgenic littermate controls (NT) were fed regular diet (0.4% salt) or high salt (HS) diet containing 4% salt for 4 weeks. Salt induced marked elevations in systolic BP in S‐P467L mice (136±4 mmHg salt n=6 vs 121±3 mmHg regular diet n=7, p<0.01, two‐way ANOVA), but not in NT mice (121±4 mmHg salt n=4 vs 116±4.0 mmHg regular diet n=5). Because salt induced similar increases in cardiac output in S‐P467L and NT animals, the pressor effect of salt intake in S‐P467L mice was not due to a larger fluid volume, but likely higher peripheral vascular resistance. In keeping with this, HS‐fed S‐P467L mice, but not NT controls, exhibited severe impairment in acetylcholine (ACh)‐ and sodium nitroprusside (SNP)‐induced vasorelaxation in carotid artery (Maximal relaxation at 30 μM ACh: S‐P467L 31±4.9% n=4 vs. NT 90±1.8% n=6, p<0.01; Maximal relaxation to 30 μM SNP: S‐P467L 38±2.8% n=4 vs. NT 89±2.6% n=6, p<0.01, two‐way ANOVA Repeated Measurements) and in cerebral basilar artery (Maximal relaxation at 100 μM ACh: S‐P467L ‐3.2±9.3% n=7 vs. NT 57±5.9% n=5, p<0.01). Importantly, salt‐induced vascular dysfunction rapidly developed in S‐P467L mice after receiving HS diet for only 3 days, preceding BP elevations. The impaired vasorelaxation in HS‐fed S‐P467L mice was associated with smaller renal artery diameter (S‐P467L 322±21 μm n=6 vs NT 389±22 μm n=5, p<0.05 two‐way ANOVA) and blunted renal blood flow (S‐P467L 36±3.6 μL/min/g n=6 vs. NT 50±6.4 μL/min/g n=5, p<0.05). These changes were not caused by differences in circulating renin‐angiotensin levels or renal sympathetic nerve activity between the two strains. Preliminary data indicated that during the 3rd week of HS diet, S‐P467L mice produced 36% less nitrate/nitrite in 24 hour urine compared to NT controls, which suggested blunted renal bioavailability of nitric oxide, a potent inhibitor of Na‐K‐2Cl cotransporter (NKCC2). This was associated with a declined capacity of HS‐fed S‐P467L mice to excrete an acute volume challenge, which was rescued by an NKCC2 inhibitor furosemide, but not by the Na‐Cl‐cotransporter inhibitor hydrochlorothiazide. Our data support the novel concept that smooth muscle PPARγ regulates systemic vascular resistance, renal perfusion and tubular sodium transport, and loss of these protective actions of PPARγ predisposes to salt sensitivity and hypertension.Support or Funding InformationDr. Jing Wu is supported by an American Heart Association Postdoctoral Fellowship (17POST33660685). This project is funded by R01 HL125603 and R01 HL131689 to Dr. Curt Sigmund.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Endothelial-dependent dilation following chronic hypoxia involves TRPV4-mediated activation of endothelial BK channels

Following chronic hypoxia (CH), the systemic vasculature exhibits blunted vasoconstriction due to endothelial-dependent hyperpolarization (EDH). Previous data demonstrate that subsequent to CH, EDH-mediated vasodilation switches from a reliance on SKca and IKca channels to activation of the endothelial BKca channels (eBK). The mechanism by which endothelial cell stimulation activates eBK channels following CH is not known. We hypothesized that following CH, EDH-dependent vasodilation involves a TRPV4-dependent activation of eBK channels. ACh induced concentration-dependent dilation in pressurized gracilis arteries from both normoxic and CH rats. Inhibition of TRPV4 (RN-1734) attenuated the ACh response in arteries from CH rats but had no effect in normoxic animals. In the presence of L-NNA and indomethacin, TRPV4 blockade attenuated ACh-induced vasodilation in arteries from CH rats. ACh elicited endothelial TRPV4-mediated Ca2+ events in arteries from both groups. GSK1016790A (GSK101, TRPV4 agonist) elicited vasodilation in arteries from normoxic and CH rats. In arteries from normoxic animals, TRAM-34/apamin abolished the dilation to TRPV4 activation, whereas luminal iberiotoxin had no effect. In CH rats, only administration of all three Kca channel inhibitors abolished the dilation to TRPV4 activation. Using Duolink®, we observed co-localization between Cav-1, TRPV4, and BK channels in gracilis arteries and in RAECs. Disruption of endothelial caveolae with methyl-β-cyclodextrin significantly decreased ACh-induced vasodilation in arteries from both groups. In gracilis arteries, endothelial membrane cholesterol was significantly decreased following 48 h of CH. In conclusion, CH results in a functional coupling between muscarinic receptors, TRPV4 and Kca channels in gracilis arteries.

Effect of histidine on sorafenib-induced vascular damage: Analysis using novel medaka fish model

BackgroundSorafenib (SFN) is an anti-angiogenic chemotherapeutic that prolongs survival of patients with hepatocellular carcinoma (HCC); its side effects, including vascular damages such as hand-foot syndrome (HFS), are a major cause of therapy discontinuation. We previously reported that maintenance of peripheral blood flow by intake of dried bonito broth (DBB) significantly prevented HFS and prolonged the administration period. The amino acids contained in DBB probably contribute to its effects, but the mechanism has not been clarified. We hypothesized that histidine, the largest component among the amino acids contained in DBB, has effects on SFN-induced vascular damage, and evaluated this possibility using a novel medaka fish model. MethodsThe fli::GFP transgenic medaka fish model has a fluorescently visible systemic vasculature. We fed the fish with SFN with and without histidine to compare blood flow and vascular structure among the differently fed models. The vascular cross-sectional area of each fish was measured to determine vascular diameter changes. ResultsOur results demonstrated that SFN-fed medaka developed a narrower vascular diameter. In addition, this narrowing was counteracted by addition of histidine to the medaka diet. We observed no positive effect of histidine on regeneration of cut vessels or on cell growth of endothelial cells and HCC cell lines. ConclusionWe proved the efficacy of the medaka model to assess vascular changes after administration of specific chemicals. And our results suggest that SFN causes vascular damage by narrowing peripheral vessel diameter, and that histidine effectively counteracts these changes to maintain blood flow.

Oxygen-sensing in the Lung is Regulated by Mitochondrial Complex I Ndufs2: Role of Mitochondria-derived Hydrogen Peroxide in Hypoxic Pulmonary Vasoconstriction

Resistance pulmonary arteries (PA) constrict in response to alveolar hypoxia; conversely the systemic vasculature dilates. Hypoxic pulmonary vasoconstriction (HPV) matches ventilation to perfusion, optimizing systemic oxygen delivery. The core mechanism of HPV is intrinsic to pulmonary artery smooth muscle cells (PASMC) and involves mitochondrial electron transport chain (ETC)-mediated redox regulation of oxygen-sensitive ion channels culminating in increased cytosolic calcium ([Ca2+]i). However, the molecular identity of the mitochondrial O2-sensor and the mechanism by which it regulates vascular tone remains unknown. We hypothesize that the O2-sensor resides in ETC complex I and initiates HPV by varying H2O2 in proportion to PO2. The primary objective was to identify the mitochondrial redox O2-sensor. A secondary goal was identify the ETC-derived reactive oxygen species (ROS) that initiates HPV. Mitochondria were isolated from lungs or kidneys of adult male Sprague-Dawley rats, housed under conditions of normoxia or chronic hypoxia (FiO2 0.1 for 2-3 weeks). Mitochondria-conditioned media (MCM) was then infused into an isolated, perfused lung to examine the effects on HPV. The effects of acute hypoxia on mitochondrial-derived H2O2 and [Ca2+]i was assessed in PASMC using with a dynamic, mitochondria-targeted H2O2 probe and FURA2-AM, respectively. Immunocaptured ETC complex I from pulmonary and renal arterial SMCs was assessed by gel electrophoresis to identify differentially expressed subunits. HPV was attenuated by MCM from normoxic lung (nLM), but not MCM from chronically hypoxic lung or kidney. Catalase attenuated the effects of nLM. Acute hypoxia (~40 mmHg) decreased H2O2 within 112±7 seconds, followed, within 205±34 seconds, by increased [Ca2+]i. The expression of a Complex I subunit, NADH dehydrogenase [ubiquinone] iron-sulfur protein 2 (Ndufs2), was increased in PA versus renal artery SMC. siRNA knockdown of Ndufs2 (siNdufs2) decreased normoxic H2O2 levels and prevented hypoxia-induced increases in [Ca2+]i. Right heart catheterization and intravital microscopy demonstrated that in vivo nebulization of siNdufs2 selectively attenuated HPV in rats and mice. Mitochondrial-derived H2O2 originating from ETC Complex I in PASMC acts as a normoxic vasodilator. Acute hypoxia decreases this vasodilator, leading to HPV. Ndufs2 is a requisite component of the PASMC’s mitochondrial O2-sensor, and acts through its dynamic ability to regulate H2O2 production.

The Use of High Dose Insulin Therapy in a Case of Diltiazem (Calcium Channel Blocker) Poisoning

SESSION TITLE: Critical Care 1 SESSION TYPE: Affiliate Case Report Poster PRESENTED ON: Tuesday, October 31, 2017 at 01:30 PM - 02:30 PM INTRODUCTION: In cases of calcium-channel blocker (CCB) poisoning patients develop bradycardia, hypotension, hyperglycemia, metabolic acidosis and cardiogenic shock. Often conventional therapy fails and leads to mortality. We report a case of CCB poisoning and use of High-dose insulin (HDI) therapy with marked improvement. CASE PRESENTATION: This is a 47-year-old male with a history of CAD, hypertension presented to the emergency department (ED) with complaints of chest pain. In ED, the patient became combative with the staff and had to be chemically restrained with Haldol and Ativan. He was admitted to medical ICU for bradycardia and hypotension. As a suicide attempt, he had overdosed on Diltiazem (CCB). He was intubated to protect his airway. He was persistently hypotensive with bradycardia and junctional rhythm on ECG and received IV atropine, glucagon, and calcium with no improvement. Subsequently required four vasopressors. He was initially placed in transcutaneous pacing and later temporary pacemaker was placed for symptomatic bradycardia. He was started on High Dose Insulin (HDI) therapy for calcium channel blocker overdose. He continued to have hyperglycemia (blood glucose 500-600 mg/dl) which is a known complication of CCB poisoning, insulin infusion was continued. He was found to have metabolic acidosis (CCB poisoning) for which he was started initially on Bicarbonate infusion and later continuous renal replacement therapy (CRRT). He was transferred to a tertiary center, due to acute respiratory distress syndrome (ARDS), where aggressive diuresis and ARDS protocol were used. The patient was extubated and after supportive therapy and Psychiatry clearance was discharged home. DISCUSSION: The proposed mechanism for HDI in CCB poisoning and cardiogenic shock: 1. increased inotropy, 2. increased intracellular glucose transport, and 3. vascular dilatation. HDI is not a vasopressor. To the contrary, insulin is a vasodilator of the systemic, coronary, and pulmonary vasculature. Insulin enhances microvascular perfusion at the capillary and pre-capillary concentration to achieve perfused capillary density similar to that of exercising muscle. Decreasing vascular resistance by these mechanisms (independent of inotropy) results in enhanced cardiac output.[1] Initial recommended dose of insulin, a bolus of 1 U/kg followed by 0.5-1 U/kg/h continuous infusion. The infusion rate increased by 2 U/kg/h every 10 min to a maximum of 10 U/kg/h if no increase in cardiac output or clinical improvement is seen. The most common side effects of HDI is hypokalemia and hypoglycemia, close monitoring is required. CONCLUSIONS: The current case illustrates how there could be a delay in therapy for drug poisoning and failure of conventional therapy, in severe cases. We should be aware of the alternative treatments. In this case, prompt action and aggressive intervention saved this patient’s life. Reference #1: Engebretsen, K.M., et al., Clin Toxicol (Phila), 2011. 49(4): p. 277-83. DISCLOSURE: The following authors have nothing to disclose: Rana Wajahat, Wajahat Lodhi, Narendrakumar Alappan No Product/Research Disclosure Information

Abstract P413: Vascular AT1 Receptors Control Blood Pressure by Augmenting Peripheral Vascular Resistance in Female Mice

Angiotensin (Ang) II is a major mediator of hypertension pathogenesis and end organ damage. Our understanding of how Ang II elicits these effects on a cellular level continues to evolve. However, the majority of these studies utilize male subjects, despite the awareness that males and females differ in mechanisms of BP regulation. Here we investigated the role of vascular smooth muscle cell (VSMC) AT1A receptors to BP control in female mice. Using Loxp technology and Cre transgenes, we created female mice with cell-specific deletion of AT 1A receptors in smooth muscle cells (SMKO mice). We found that elimination of these receptors led to a significant (8 mmHg) reduction in baseline BP in female SMKO mice (similar to male SMKO mice) compared with controls (108±2 versus 116±1 mmHg; P =0.004) with no effect on modulating sodium sensitivity in female SMKO mice (unlike male SMKO mice in which sodium sensitivity was enhanced). Over a 4-week Ang II infusion, the severity of Ang II - dependent hypertension was minimally affected during the first 2 weeks of Ang II infusion, corresponding to no difference in sodium excretion. However, female SMKO mice had a 35% less BP elevation compared to controls over the final 2 weeks of Ang II infusion (SMKO, 20±2 versus control, 30±3 Change in SBP mmHg; P =0.01). The acute vasoconstrictor responses to Ang II in the systemic vasculature were reduced (by approximately 50-75%) in female SMKO mice (0.1 μg/kg- SMKO, 3.6±1.2 mmHg versus control, 7.3±0.7 mmHg; P =0.01, 0.3 μg/kg- SMKO, 3.3±1.8 mmHg versus control, 12.1 ± 1.3 mmHg; P =0.0008, 1.0 μg/kg- SMKO, 8.5±1.9 mmHg versus control, 19.4±2.0 mmHg; P =0.002). In addition, the acute Ang II reduction in renal tissue perfusion was nearly eliminated in female SMKO mice. In contrast, we previously reported that in male SMKO mice have a much larger (50%) reduction in BP during chronic Ang II infusion associated with increased natriuresis, but relatively preserved vasoconstrictor response (25% reduction) during acute Ang II infusions. These findings suggest that in female mice, direct actions of AT 1A receptors in VSMCs are essential for regulation of basal BP and Ang II-dependent hypertension. However, this effect is mediated by reductions in peripheral vascular resistance rather than sodium retention.

Abstract P503: Role of TRPV4 and Endothelial BK Channels in Endothelial-dependent Dilation Following Chronic Hypoxia

Following chronic hypoxia (CH) the systemic vasculature exhibits blunted vasoconstriction due to endothelial cell-dependent hyperpolarization (EDH). Previous data demonstrate that after CH, EDH-mediated dilation switches from a reliance on SK ca and IK ca channels to activation of endothelial BK ca channel (eBK). The mechanism by which endothelial cell stimulation activates eBK channels following CH is not known. We hypothesized that following CH, EDH-dependent dilation involves a transient receptor potential V4 (TRPV4)-dependent activation of eBK channels. ACh induced concentration-dependent dilation in pressurized gracilis arteries from normoxic and CH rats. TRPV4 blockade only attenuated ACh-induced vasodilation in arteries from CH rats (Fig. 1). ACh elicited an increase in endothelial TRPV4-dependent Ca 2+ events in arteries from both groups. Direct activation of TRPV4 elicited dilation in arteries from normoxic and CH rats (97 ± 2 % and 95 ± 2 % for normoxic and CH, respectively). In arteries from normoxic rats, inhibition of SK ca and IK ca abolished dilation to TRPV4 activation (6 ± 5 %), whereas, luminal iberiotoxin had no effect (89 ± 7 %). In CH rats, only administration of all three K ca channel inhibitors abolished dilation to TRPV4 activation (3 ± 10%). Disruption of endothelial caveolae with methyl-β-cyclodextrin significantly decreased ACh-induced dilation in arteries from both groups. Using a proximity ligation assay, we observed co-localization between caveolin-1, TRPV4, and eBK channels in the endothelium of gracilis arteries. In conclusion, CH results in functional coupling between muscarinic receptors, TRPV4 and K ca channels in gracilis arteries.

Cardiovascular effects of histamine in three widely diverse species of reptiles.

The cardiovascular system of vertebrates is regulated by a vast number of regulatory factors, including histamine. In pythons, histamine induces a strong tachycardia and dilates the systemic vasculature, which resembles the cardiovascular response to the elevated metabolic rate during digestion. In fact, there is an important role of increased histaminergic tone on the heart during the initial 24h of digestion in pythons. Whilst the cardiovascular effects of histamine are well studied in pythons, little is known about the effects in other groups of reptiles. The histaminergic effects on the heart vary among species and histamine may exert either pressor and depressor effects by causing either constrictive or dilatory vascular responses. Here, we investigated the cardiovascular effects of histamine in three species of reptiles with very different cardiovascular and pulmonary morphologies. Experiments were performed on both anesthetized and recovered animals. We show a species-dependent effect of histamine on the systemic vasculature with dilation in rattlesnakes and constriction in turtles and caimans but no effect on the pulmonary circulation. The histamine-induced dilation in rattlesnakes was mediated through an activation of H2-receptors, whereas the histamine-induced constriction in caimans was mediated through both adrenergic signaling and H1-receptors activation. In all three species, histamine-induced tachycardia by direct stimulation of histaminergic receptors as well as an indirect activation of adrenoreceptors. This finding highlights a more complex mechanism underlying the action of histamine than previously recognized in reptiles.

Relationship between Vascular Resistance and Sympathetic Nerve Fiber Density in Arterial Vessels in Children With Sleep Disordered Breathing.

BackgroundSleep disordered breathing in children is associated with increased blood flow velocity and sympathetic overactivity. Sympathetic overactivity results in peripheral vasoconstriction and reduced systemic vascular compliance, which increases blood flow velocity during systole. Augmented blood flow velocity is recognized to promote vascular remodeling. Importantly, increased vascular sympathetic nerve fiber density and innervation in early life plays a key role in the development of early‐onset hypertension in animal models. Examination of sympathetic nerve fiber density of the tonsillar arteries in children undergoing adenotonsillectomy for Sleep disordered breathing will address this question in humans.Methods and ResultsThirteen children scheduled for adenotonsillectomy to treat sleep disordered breathing underwent pupillometry, polysomnography, flow‐mediated dilation, resting brachial artery blood flow velocity (velocity time integral), and platelet aggregation. The dorsal lingual artery (tonsil) was stained and immunofluorescence techniques used to determine sympathetic nerve fiber density. Sympathetic nerve fiber density was correlated with increased resting velocity time integral (r=0.63; P<0.05) and a lower Neuronal Pupillary Index (r=−0.71, P<0.01), as well as a slower mean pupillary constriction velocity (mean, r=−0.64; P<0.05). A faster resting velocity time integral was associated with a slower peak pupillary constriction velocity (r=−0.77; P<0.01) and higher platelet aggregation to collagen antigen (r=0.64; P<0.05). Slower mean and peak pupillary constriction velocity were associated with higher platelet aggregation scores (P<0.05; P<0.01, respectively).ConclusionsThese results indicate that sympathetic activity is associated with change in both the function and structure of systemic vasculature in children with sleep disordered breathing.