Saxagliptin Prevents Increased Coronary Vascular Stiffness in Aortic-Banded Mini Swine.

Increased peripheral conduit artery stiffness has been shown in patients with heart failure (HF) with preserved ejection fraction. However, it is unknown whether this phenomenon extends to the coronary vasculature. HF with preserved ejection fraction may be driven, in part, by coronary inflammation, and inhibition of the enzyme DPP-4 (dipeptidyl-peptidase 4) reduces inflammation and oxidative stress. The purpose of this study was to determine the effect of saxagliptin-a DPP-4 inhibitor-on coronary stiffness in aortic-banded mini swine. We hypothesized saxagliptin would prevent increased coronary artery stiffness in a translational swine model with cardiac features of HF with preserved ejection fraction by inhibiting perivascular adipose tissue inflammation. Yucatan mini swine were divided into 3 groups: control, aortic-banded untreated HF, and aortic-banded saxagliptin-treated HF. Ex vivo mechanical testing was performed on the left circumflex and right coronary arteries, and advanced glycation end product, NF-κB (nuclear factor-κB), and nitrotyrosine levels were measured. An increase in the coronary elastic modulus of HF animals was associated with increased vascular advanced glycation end products, NF-κB, and nitrotyrosine levels compared with control and prevented by saxagliptin treatment. Aortas from healthy mice were treated with media from swine perivascular adipose tissue culture to assess its role on vascular stiffening. Conditioned media from HF and saxagliptin-treated HF animals increased mouse aortic stiffness; however, only perivascular adipose tissue from the HF group showed increased advanced glycation end products and NF-κB levels. In conclusion, our data show increased coronary conduit vascular stiffness was prevented by saxagliptin and associated with decreased advanced glycation end products, NF-κB, and nitrotyrosine levels in a swine model with potential relevance to HF with preserved ejection fraction.

Chronic interval exercise training prevents BKCa channel-mediated coronary vascular dysfunction in aortic-banded miniswine.

Conventional treatments have failed to improve the prognosis of heart failure with preserved ejection fraction (HFpEF) patients. Thus, the purpose of this study was to determine the therapeutic efficacy of chronic interval exercise training (IT) on large-conductance Ca2+-activated K+ (BKCa) channel-mediated coronary vascular function in heart failure. We hypothesized that chronic interval exercise training would attenuate pressure overload-induced impairments to coronary BKCa channel-mediated function. A translational large-animal model with cardiac features of HFpEF was used to test this hypothesis. Specifically, male Yucatan miniswine were divided into three groups ( n = 7/group): control (CON), aortic banded (AB)-heart failure (HF), and AB-interval trained (HF-IT). Coronary blood flow, vascular conductance, and vasodilatory capacity were measured after administration of the BKCa channel agonist NS-1619 both in vivo and in vitro in the left anterior descending coronary artery and isolated coronary arterioles, respectively. Skeletal muscle citrate synthase activity was decreased and left ventricular brain natriuretic peptide levels increased in HF vs. CON and HF-IT animals. A parallel decrease in NS-1619-dependent coronary vasodilatory reserve in vivo and isolated coronary arteriole vasodilatory responsiveness in vitro were observed in HF animals compared with CON, which was prevented in the HF-IT group. Although exercise training prevented BKCa channel-mediated coronary vascular dysfunction, it did not change BKCa channel α-subunit mRNA, protein, or cellular location (i.e., membrane vs. cytoplasm). In conclusion, these results demonstrate the viability of chronic interval exercise training as a therapy for central and peripheral adaptations of experimental heart failure, including BKCa channel-mediated coronary vascular dysfunction. NEW & NOTEWORTHY Conventional treatments have failed to improve the prognosis of heart failure with preserved ejection fraction (HFpEF) patients. Our findings show that chronic interval exercise training can prevent BKCa channel-mediated coronary vascular dysfunction in a translational swine model of chronic pressure overload-induced heart failure with relevance to human HFpEF.

Chronic low-intensity exercise attenuates cardiomyocyte contractile dysfunction and impaired adrenergic responsiveness in aortic-banded mini-swine.

Exercise improves clinical outcomes in patients diagnosed with heart failure with reduced ejection fraction (HFrEF), in part via beneficial effects on cardiomyocyte Ca2+ cycling during excitation-contraction coupling (ECC). However, limited data exist regarding the effects of exercise training on cardiomyocyte function in patients diagnosed with heart failure with preserved ejection fraction (HFpEF). The purpose of this study was to investigate cardiomyocyte Ca2+ handling and contractile function following chronic low-intensity exercise training in aortic-banded miniature swine and test the hypothesis that low-intensity exercise improves cardiomyocyte function in a large animal model of pressure overload. Animals were divided into control (CON), aortic-banded sedentary (AB), and aortic-banded low-intensity trained (AB-LIT) groups. Left ventricular cardiomyocytes were electrically stimulated (0.5 Hz) to assess Ca2+ homeostasis (fura-2-AM) and unloaded shortening during ECC under conditions of baseline pacing and pacing with adrenergic stimulation using dobutamine (1 µM). Cardiomyocytes in AB animals exhibited depressed Ca2+ transient amplitude and cardiomyocyte shortening vs. CON under both conditions. Exercise training attenuated AB-induced decreases in cardiomyocyte Ca2+ transient amplitude but did not prevent impaired shortening vs. CON. With dobutamine, AB-LIT exhibited both Ca2+ transient and shortening amplitude similar to CON. Adrenergic sensitivity, assessed as the time to maximum inotropic response following dobutamine treatment, was depressed in the AB group but normal in AB-LIT animals. Taken together, our data suggest exercise training is beneficial for cardiomyocyte function via the effects on Ca2+ homeostasis and adrenergic sensitivity in a large animal model of pressure overload-induced heart failure. NEW & NOTEWORTHY Conventional treatments have failed to improve the prognosis of heart failure with preserved ejection fraction (HFpEF) patients. Our findings show chronic low-intensity exercise training can prevent cardiomyocyte dysfunction and impaired adrenergic responsiveness in a translational large animal model of chronic pressure overload-induced heart failure with relevance to human HFpEF.

Endothelial dysfunction occurs independently of adipose tissue inflammation and insulin resistance in ovariectomized Yucatan miniature-swine.

In rodents, experimentally-induced ovarian hormone deficiency increases adiposity and adipose tissue (AT) inflammation, which is thought to contribute to insulin resistance and increased cardiovascular disease risk. However, whether this occurs in a translationally-relevant large animal model remains unknown. Herein, we tested the hypothesis that ovariectomy would promote visceral and perivascular AT (PVAT) inflammation, as well as subsequent insulin resistance and peripheral vascular dysfunction in female swine. At sexual maturity (7 months of age), female Yucatan mini-swine either remained intact (control, n = 9) or were ovariectomized (OVX, n = 7). All pigs were fed standard chow (15-20 g/kg), and were euthanized 6 months post-surgery. Uterine mass and plasma estradiol levels were decreased by ∼10-fold and 2-fold, respectively, in OVX compared to control pigs. Body mass, glucose homeostasis, and markers of insulin resistance were not different between control and OVX pigs; however, OVX animals exhibited greater plasma triglycerides and triglyceride:HDL ratio. Ovariectomy enhanced visceral adipocyte expansion, although this was not accompanied by brachial artery PVAT adipocyte expansion, AT inflammation in either depot, or increased systemic inflammation assessed by plasma C-reactive protein concentrations. Despite the lack of AT inflammation and insulin resistance, OVX pigs exhibited depressed brachial artery endothelial-dependent vasorelaxation, which was rescued with blockade of endothelin receptor A. Together, these findings indicate that in female Yucatan mini-swine, increased AT inflammation and insulin resistance are not required for loss of ovarian hormones to induce endothelial dysfunction.

Loss of Female Sex Hormones Exacerbates Cerebrovascular and Cognitive Dysfunction in Aortic Banded Miniswine Through a Neuropeptide Y-Ca2+-Activated Potassium Channel-Nitric Oxide Mediated Mechanism.

BACKGROUND: Postmenopausal women represent the largest cohort of patients with heart failure with preserved ejection fraction, and vascular dementia represents the most common form of dementia in patients with heart failure with preserved ejection fraction. Therefore, we tested the hypotheses that the combination of cardiac pressure overload (aortic banding [AB]) and the loss of female sex hormones (ovariectomy [OVX]) impairs cerebrovascular control and spatial memory. METHODS AND RESULTS: Female Yucatan miniswine were separated into 4 groups (n=7 per group): (1) control, (2) AB, (3) OVX, and (4) AB-OVX. Pigs underwent OVX and AB at 7 and 8 months of age, respectively. At 14 months, cerebral blood flow velocity and spatial memory (spatial hole-board task) were lower in the OVX groups (P<0.05), with significant impairments in the AB-OVX group (P<0.05). Resting carotid artery ß stiffness and vascular resistance during central hypovolemia were increased in the AB-OVX group (P<0.05), and blood flow recovery after central hypovolemia was reduced in both OVX groups (P<0.05). Isolated pial artery (pressure myography) vasoconstriction to neuropeptide Y was greatest in the AB-OVX group (P<0.05), and vasodilation to the Ca2+-activated potassium channel α-subunit agonist NS-1619 was impaired in both AB groups (P<0.05). The ratio of phosphorylated endothelial nitric oxide synthase:total endothelial nitric oxide synthase was depressed and Ca2+-activated potassium channel α-subunit protein was increased in AB groups (P<0.05). CONCLUSIONS: Mechanistically, impaired cerebral blood flow control in experimental heart failure may be the result of heightened neuropeptide Y-induced vasoconstriction along with reduced vasodilation associated with decreased Ca2+-activated potassium channel function and impaired nitric oxide signaling, the effects of which are exacerbated in the absence of female sex hormones.

The protective role of sex hormones in females and exercise prehabilitation in males on sternotomy-induced cranial hypoperfusion in aortic banded mini-swine.

During cardiac surgery, specifically sternotomy, cranial hypoperfusion is linked to cerebral ischemia, increased risk of perioperative watershed stroke, and other neurocognitive complications. The purpose of this study was to retrospectively examine the effect of sex hormones in females and exercise prehabilitation in males on median sternotomy-induced changes in cranial perfusion in a large animal model of heart failure. Cranial blood flow (CBF) before and 10 and 60 min poststernotomy was analyzed in eight groups of Yucatan mini-swine: female control, aortic banded, ovariectomized, and ovariectomized + aortic banded; male control, aortic banded, aortic banded + continuous exercise trained, and aortic banded + interval exercise trained. A median sternotomy decreased cranial perfusion during surgery in all pigs (~24 ± 2% relative to baseline; P ≤ 0.05). CBF was 30 ± 7% lower across all time points in all females vs. all males (P ≤ 0.05) and sternotomy decreased cranial perfusion (P ≤ 0.05) independent of sex (females = 34 ± 3% and males = 14 ± 3%) and aortic banding (intact control = 31 ± 5% and intact aortic banded = 31 ± 4%). CBF recovery at 60 min tended to be better in females vs. males (relative to 10 min poststernotomy, females = 23 ± 13% vs. males = -1 ± 5%) and intact aortic banded vs. control pigs (relative to 10 min poststernotomy, aortic banded = 43 ± 20% vs. control = 6 ± 16%; P ≤ 0.05) at 60 min poststernotomy. Ovariectomy impaired CBF recovery during cranial reperfusion 60 min following sternotomy (relative to baseline, all intact females = -1 ± 9% vs. all ovariectomized females = -15 ± 4%; P ≤ 0.05). Chronic exercise training completely prevented significant sternotomy-induced cranial hypoperfusion independent of aortic banding (sternotomy-induced deficit, all sedentary males = -24 ± 6% vs. all exercise-trained males = -7 ± 3%; P ≤ 0.05). Female sex hormones protected against impaired CBF recovery during reperfusion, while chronic exercise training prevented sternotomy-induced cranial hypoperfusion despite cardiac pressure overload.NEW & NOTEWORTHY Our findings suggest a median sternotomy may predispose patients, possibly postmenopausal women and sedentary men, to perioperative cerebral ischemia, an increased risk of cardiac surgery-related stroke, and resulting neurocognitive impairments. Specifically, data from this common surgical procedure show: 1) median sternotomy independently decreases cranial perfusion; 2) female sex hormones improve cranial blood flow recovery following sternotomy; and 3) exercise prehabilitation prevents sternotomy-induced cranial hypoperfusion. Exercise prehabilitation before cardiac surgery may be advantageous for capable patients.

Carotid Artery Vascular Mechanics Serve as Biomarkers of Cognitive Dysfunction in Aortic-Banded Miniature Swine That Can Be Treated With an Exercise Intervention.

BACKGROUND: Cognitive impairment in the setting of heart failure with preserved ejection fraction remains poorly understood. Using aortic-banded miniature swine displaying pathological features of human heart failure with preserved ejection fraction, we tested the hypothesis that increased carotid artery stiffness and altered carotid blood flow control are associated with impaired memory independent of decreased cardiac output. Furthermore, we hypothesized that chronic exercise prevents carotid artery vascular restructuring and preserves normal blood flow control and cognition in heart failure with preserved ejection fraction. METHODS AND RESULTS: Yucatan pigs aged 8 months were divided into 3 groups: control (n=7), aortic-banded sedentary (n=7), and aortic-banded exercise trained (n=7). At 6 months following aortic-banded or control conditions, memory was evaluated using a spatial hole-board task. Carotid artery vascular mechanics and blood flow were assessed at rest, and blood flow control was examined during transient vena cava occlusion. Independent of decreased cardiac output, the aortic-banded group exhibited impaired memory that was associated with carotid artery vascular stiffening, elevated carotid artery vascular resistance, and exaggerated reductions in carotid artery blood flow during vena cava occlusion. Chronic exercise augmented memory scores, normalized blood flow control, and improved indices of carotid artery vascular stiffening. Indices of vascular stiffening were significantly correlated with average memory score. CONCLUSIONS: Carotid artery stiffness and altered vasomotor control correlate with impaired cognition independent of cardiac systolic dysfunction. Carotid artery vascular mechanics may serve as a biomarker for vascular cognitive impairment in heart failure with preserved ejection fraction. Chronic low-intensity exercise reduces vascular stiffening and improves cognition, highlighting the utility of exercise therapy for treating vascular cognitive impairment in heart failure with preserved ejection fraction.

Saxagliptin and Tadalafil Differentially Alter Cyclic Guanosine Monophosphate (cGMP) Signaling and Left Ventricular Function in Aortic-Banded Mini-Swine.

BACKGROUND: Cyclic guanosine monophosphate-protein kinase G-phosphodiesterase 5 signaling may be disturbed in heart failure (HF) with preserved ejection fraction, contributing to cardiac remodeling and dysfunction. The purpose of this study was to manipulate cyclic guanosine monophosphate signaling using the dipeptidyl-peptidase 4 inhibitor saxagliptin and phosphodiesterase 5 inhibitor tadalafil. We hypothesized that preservation of cyclic guanosine monophosphate cGMP signaling would attenuate pathological cardiac remodeling and improve left ventricular (LV) function. METHODS AND RESULTS: We assessed LV hypertrophy and function at the organ and cellular level in aortic-banded pigs. Concentric hypertrophy was equal in all groups, but LV collagen deposition was increased in only HF animals. Prevention of fibrotic remodeling by saxagliptin and tadalafil was correlated with neuropeptide Y plasma levels. Saxagliptin better preserved integrated LV systolic and diastolic function by maintaining normal LV chamber volumes and contractility (end-systolic pressure-volume relationship, preload recruitable SW) while preventing changes to early/late diastolic longitudinal strain rate. Function was similar to the HF group in tadalafil-treated animals including increased LV contractility, reduced chamber volume, and decreased longitudinal, circumferential, and radial mechanics. Saxagliptin and tadalafil prevented a negative cardiomyocyte shortening-frequency relationship observed in HF animals. Saxagliptin increased phosphodiesterase 5 activity while tadalafil increased cyclic guanosine monophosphate levels; however, neither drug increased downstream PKG activity. Early mitochondrial dysfunction, evident as decreased calcium-retention capacity and Complex II-dependent respiratory control, was present in both HF and tadalafil-treated animals. CONCLUSIONS: Both saxagliptin and tadalafil prevented increased LV collagen deposition in a manner related to the attenuation of increased plasma neuropeptide Y levels. Saxagliptin appears superior for treating heart failure with preserved ejection fraction, considering its comprehensive effects on integrated LV systolic and diastolic function.

A new twist on an old idea part 2: cyclosporine preserves normal mitochondrial but not cardiomyocyte function in mini-swine with compensated heart failure.

We recently developed a clinically relevant mini-swine model of heart failure with preserved ejection fraction (HFpEF), in which diastolic dysfunction was associated with increased mitochondrial permeability transition (MPT). Early diastolic function is ATP and Ca(2+)-dependent, thus, we hypothesized chronic low doses of cyclosporine (CsA) would preserve mitochondrial function via inhibition of MPT and subsequently maintain normal cardiomyocyte Ca(2+) handling and contractile characteristics. Left ventricular cardiomyocytes were isolated from aortic-banded Yucatan mini-swine divided into three groups; control nonbanded (CON), HFpEF nontreated (HF), and HFpEF treated with CsA (HF-CsA). CsA mitigated the deterioration of mitochondrial function observed in HF animals, including functional uncoupling of Complex I-dependent mitochondrial respiration and increased susceptibility to MPT. Attenuation of mitochondrial dysfunction in the HF-CsA group was not associated with commensurate improvement in cardiomyocyte Ca(2+) handling or contractility. Ca(2+) transient amplitude was reduced and transient time to peak and recovery (tau) prolonged in HF and HF-CsA groups compared to CON. Alterations in Ca(2+) transient parameters observed in the HF and HF-CsA groups were associated with decreased cardiomyocyte shortening and shortening rate. Cellular function was consistent with impaired in vivo systolic and diastolic whole heart function. A significant systemic hypertensive response to CsA was observed in HF-CsA animals, and may have played a role in the accelerated the development of heart failure at both the whole heart and cellular levels. Given the significant detriment to cardiac function observed in response to CsA, our findings suggest chronic CsA treatment is not a viable therapeutic option for HFpEF.

A new twist on an old idea: a two-dimensional speckle tracking assessment of cyclosporine as a therapeutic alternative for heart failure with preserved ejection fraction.

We recently reported that mitochondrial dysfunction, characterized by increased mitochondrial permeability transition (MPT), was present in a translational swine model of heart failure with preserved ejection fraction (HFpEF). Cyclophilin D is a key component of the MPT pore, therefore, the purpose of this study was to test the efficacy of a novel cyclosporine (CsA) dosing scheme as a therapeutic alternative for HFpEF. Computed tomography (CT), two-dimensional speckle tracking two-dimensional speckle tracking (2DST), and invasive hemodynamics were used to evaluate cardiac function. CT imaging showed 14 weeks of CsA treatment caused eccentric myocardial remodeling (contrasting concentric remodeling in untreated HF animals) and elevated systemic pressures. 2DST detected left ventricular (LV) mechanics associated with systolic and diastolic dysfunction prior to the onset of significantly increased LV end diastolic pressure including: (1) decreased systolic apical rotation rate, longitudinal displacement, and longitudinal/radial/circumferential strain; (2) decreased early diastolic untwisting and longitudinal strain rate; and (3) increased late diastolic radial/circumferential mitral strain rate. LV mechanics associated with systolic and diastolic impairment was enhanced to a greater extent than seen in untreated HF animals following CsA treatment. In conclusion, CsA treatment accelerated the development of heart failure, including dilatory LV remodeling and impaired systolic and diastolic mechanics. Although our findings do not support CsA as a viable therapy for HFpEF, 2DST was effective in differentiating between progressive gradations of developing HF and detecting diastolic impairment prior to the development of overt diastolic dysfunction.

Herkinorin analogues with differential beta-arrestin-2 interactions.

Salvinorin A is a psychoactive natural product that has been found to be a potent and selective kappa opioid receptor agonist in vitro and in vivo. The activity of salvinorin A is unusual compared to other opioids such as morphine in that it mediates potent kappa opioid receptor signaling yet leads to less receptor downregulation than observed with other kappa agonists. Our initial chemical modifications of salvinorin A have yielded one analogue, herkinorin ( 1c), with high affinity at the microOR. We recently reported that 1c does not promote the recruitment of beta-arrestin-2 to the microOR or receptor internalization. Here we describe three new derivatives of 1c ( 3c, 3f, and 3i) with similar properties and one, benzamide 7b, that promotes recruitment of beta-arrestin-2 to the microOR and receptor internalization. When the important role micro opioid receptor regulation plays in determining physiological responsiveness to opioid narcotics is considered, micro opioids derived from salvinorin A may offer a unique template for the development of functionally selective mu opioid receptor-ligands with the ability to produce analgesia while limiting adverse side effects.