Non-responsiveness to cardioprotection by ischaemic preconditioning in Ossabaw minipigs with genetic predisposition to, but without the phenotype of the metabolic syndrome.

The translation of successful preclinical and clinical proof-of-concept studies on cardioprotection to the benefit of patients with reperfused acute myocardial infarction has been difficult so far. This difficulty has been attributed to confounders which patients with myocardial infarction typically have but experimental animals usually not have. The metabolic syndrome is a typical confounder. We hypothesised that there may also be a genuine non-responsiveness to cardioprotection and used Ossabaw minipigs which have the genetic predisposition to develop a diet-induced metabolic syndrome, but before they had developed the diseased phenotype. Using a prospective study design, a reperfused acute myocardial infarction was induced in 62 lean Ossabaw minipigs by 60 min coronary occlusion and 180 min reperfusion. Ischaemic preconditioning by 3 cycles of 5 min coronary occlusion and 10 min reperfusion was used as cardioprotective intervention. Ossabaw minipigs were stratified for their single nucleotide polymorphism as homozygous for valine (V/V) or isoleucine (I/I)) in the γ-subunit of adenosine monophosphate-activated protein kinase. Endpoints were infarct size and area of no-reflow. Infarct size (V/V: 54 ± 8, I/I: 54 ± 13% of area at risk, respectively) was not reduced by ischaemic preconditioning (V/V: 55 ± 11, I/I: 46 ± 11%) nor was the area of no-reflow (V/V: 57 ± 18, I/I: 49 ± 21 vs. V/V: 57 ± 21, I/I: 47 ± 21% of infarct size). Bioinformatic comparison of the Ossabaw genome to that of Sus scrofa and Göttingen minipigs identified differences in clusters of genes encoding mitochondrial and inflammatory proteins, including the janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway. The phosphorylation of STAT3 at early reperfusion was not increased by ischaemic preconditioning, different from the established STAT3 activation by cardioprotective interventions in other pig strains. Ossabaw pigs have not only the genetic predisposition to develop a metabolic syndrome but also are not amenable to cardioprotection by ischaemic preconditioning.

Specificity of Ca2+-activated K+ channel modulation in atherosclerosis and aerobic exercise training.

Vascular smooth muscle cells express several isoforms of a number of classes of K+ channels. Potassium channels play critical roles in the regulation of vascular smooth muscle contraction as well as vascular smooth muscle cell proliferation or phenotypic modulation. There is ample evidence that it is Ca2+ that enables these two seemingly disparate functions to be tightly coupled both in healthy and disease processes. Because of the central position that potassium channels have in vasocontraction, vasorelaxation, membrane potential, and smooth muscle cell proliferation, these channels continue to possess the potential to serve as novel therapeutic targets in cardiovascular disease. While there are questions that remain regarding the complete interactions between K+ channels, vascular regulation, smooth muscle cell proliferation, and phenotypic modulation in physiological and pathophysiological conditions, a broad understanding of the contributions of each class of K+ channel to contractile and proliferative states of the vasculature has been reached. This brief review will discuss the current understanding of the role of K+ channels in vascular smooth muscle cells in health and disease using the porcine vascular smooth muscle cell model with particular attention to new scientific discoveries contributed by the authors regarding the effect of endurance exercise on the function of the K+ channels.

Swine Disease Models for Optimal Vascular Engineering.

Swine disease models are essential for mimicry of human metabolic and vascular pathophysiology, thereby enabling high-fidelity translation to human medicine. The worldwide epidemic of obesity, metabolic disease, and diabetes has prompted the focus on these diseases in this review. We highlight the remarkable similarity between Ossabaw miniature swine and humans with metabolic syndrome and atherosclerosis. Although the evidence is strongest for swine models of coronary artery disease, findings are generally applicable to any vascular bed. We discuss the major strengths and weaknesses of swine models. The development of vascular imaging is an example of optimal vascular engineering in swine. Although challenges regarding infrastructure and training of engineers in the use of swine models exist, opportunities are ripe for gene editing, studies of molecular mechanisms, and use of swine in coronary artery imaging and testing of devices that can move quickly to human clinical studies.

Alloxan-induced diabetes exacerbates coronary atherosclerosis and calcification in Ossabaw miniature swine with metabolic syndrome.

BACKGROUND: There is a preponderance of evidence implicating diabetes with increased coronary artery disease (CAD) and calcification (CAC) in human patients with metabolic syndrome (MetS), but the effect of diabetes on CAD severity in animal models remains controversial. We investigated whether diabetes exacerbates CAD/CAC and intracellular free calcium ([Ca2+]i) dysregulation in the clinically relevant Ossabaw miniature swine model of MetS. METHODS: Sixteen swine, eight with alloxan-induced diabetes, were fed a hypercaloric, atherogenic diet for 6 months. Alloxan-induced pancreatic beta cell damage was examined by immunohistochemical staining of insulin. The metabolic profile was confirmed by body weight, complete blood panel, intravenous glucose tolerance test (IVGTT), and meal tolerance test. CAD severity was assessed with intravascular ultrasound and histology. [Ca2+]i handling in coronary smooth muscle (CSM) cells was assessed with fura-2 ratiometric imaging. RESULTS: Fasting and post-prandial blood glucose, total cholesterol, and serum triglycerides were elevated in MetS-diabetic swine. This group also exhibited hypoinsulinemia during IVGTT and less pancreatic beta cell mass when compared to lean and MetS-nondiabetic swine. IVUS analysis revealed that MetS-diabetic swine had greater percent wall coverage, percent plaque burden, and calcium index when compared to lean and MetS-nondiabetic swine. Fura-2 imaging of CSM [Ca2+]i revealed that MetS-nondiabetic swine exhibited increased sarcoplasmic reticulum Ca2+ store release and Ca2+ influx through voltage-gated Ca2+ channels compared to lean swine. MetS-diabetic swine exhibited impaired Ca2+ efflux. CONCLUSIONS: Diabetes exacerbates coronary atherosclerosis and calcification in Ossabaw miniature swine with MetS, accompanied by progression of [Ca2+]i dysregulation in advanced CAD/CAC. These results recapitulate increased CAD in humans with diabetes and establish Ossabaw miniature swine as an animal model for future MetS/diabetes comorbidity studies.

18F-NaF and 18F-FDG as molecular probes in the evaluation of atherosclerosis.

The early detection of atherosclerotic disease is vital to the effective prevention and management of life-threatening cardiovascular events such as myocardial infarctions and cerebrovascular accidents. Given the potential for positron emission tomography (PET) to visualize atherosclerosis earlier in the disease process than anatomic imaging modalities such as computed tomography (CT), this application of PET imaging has been the focus of intense scientific inquiry. Although 18F-FDG has historically been the most widely studied PET radiotracer in this domain, there is a growing body of evidence that 18F-NaF holds significant diagnostic and prognostic value as well. In this article, we review the existing literature on the application of 18F-FDG and 18F-NaF as PET probes in atherosclerosis and present the findings of original animal and human studies that have examined how well 18F-NaF uptake correlates with vascular calcification and cardiovascular risk.

Long-term spironolactone treatment reduces coronary TRPC expression, vasoconstriction, and atherosclerosis in metabolic syndrome pigs.

Coronary transient receptor potential canonical (TRPC) channel expression is elevated in metabolic syndrome (MetS). However, differential contribution of TRPCs to coronary pathology in MetS is not fully elucidated. We investigated the roles of TRPC1 and TRPC6 isoforms in coronary arteries of MetS pigs and determined whether long-term treatment with a mineralocorticoid receptor inhibitor, spironolactone, attenuates coronary TRPC expression and associated dysfunctions. MetS coronary arteries exhibited significant atherosclerosis, endothelial dysfunction, and increased histamine-induced contractions. Immunohistochemical studies revealed that TRPC6 immunostaining was significantly greater in the medial layer of MetS pig coronary arteries compared to that in Lean pigs, whereas little TRPC6 immunostaining was found in atheromas. Conversely, TRPC1 immunostaining was weak in the medial layer but strong in MetS atheromas, where it was predominantly localized to macrophages. Spironolactone treatment significantly decreased coronary TRPC expression and dysfunctions in MetS pigs. In vivo targeted delivery of the dominant-negative (DN)-TRPC6 cDNA to the coronary wall reduced histamine-induced calcium transients in the MetS coronary artery medial layer, implying a role for TRPC6 in mediating calcium influx in MetS coronary smooth muscles. Monocyte adhesion was increased in Lean pig coronary arteries cultured in the presence of aldosterone; and spironolactone antagonized this effect, suggesting that coronary mineralocorticoid receptor activation may regulate macrophage infiltration. TRPC1 expression in atheroma macrophages was associated with advanced atherosclerosis, whereas medial TRPC6 upregulation correlated with increased histamine-induced calcium transients and coronary contractility. We propose that long-term spironolactone treatment may be a therapeutic strategy to decrease TRPC expression and coronary pathology associated with MetS.

Effect of renal shock wave lithotripsy on the development of metabolic syndrome in a juvenile swine model: a pilot study.

PURPOSE: We performed a pilot study to assess whether renal shock wave lithotripsy influences metabolic syndrome onset and severity. MATERIALS AND METHODS: Three-month-old juvenile female Ossabaw miniature pigs were treated with shock wave lithotripsy (2,000 shock waves at 24 kV with 120 shock waves per minute in 2) or sham shock wave lithotripsy (no shock waves in 2). Shock waves were targeted to the upper pole of the left kidney to model treatment that would also expose the pancreatic tail to shock waves. Pigs were then instrumented to directly measure arterial blood pressure via an implanted radiotelemetry device. They later received a hypercaloric atherogenic diet for about 7 months. Metabolic syndrome development was assessed by the intravenous glucose tolerance test. RESULTS: Metabolic syndrome progression and severity were similar in the sham treated and lithotripsy groups. The only exception arterial blood pressure, which remained relatively constant in sham treated pigs but began to increase at about 2 months towards hypertensive levels in lithotripsy treated pigs. Metabolic data on the 2 groups were pooled to provide a more complete assessment of metabolic syndrome development and progression in this juvenile pig model. The intravenous glucose tolerance test revealed substantial insulin resistance with impaired glucose tolerance within 2 months on the hypercaloric atherogenic diet with signs of further metabolic impairment at 7 months. CONCLUSIONS: These preliminary results suggest that renal shock wave lithotripsy is not a risk factor for worsening glucose tolerance or diabetes mellitus onset. However, it appears to be a risk factor for early onset hypertension in metabolic syndrome.

Perivascular adipose tissue potentiates contraction of coronary vascular smooth muscle: influence of obesity.

BACKGROUND: This investigation examined the mechanisms by which coronary perivascular adipose tissue (PVAT)-derived factors influence vasomotor tone and the PVAT proteome in lean versus obese swine. METHODS AND RESULTS: Coronary arteries from Ossabaw swine were isolated for isometric tension studies. We found that coronary (P=0.03) and mesenteric (P=0.04) but not subcutaneous adipose tissue augmented coronary contractions to KCl (20 mmol/L). Inhibition of CaV1.2 channels with nifedipine (0.1 µmol/L) or diltiazem (10 µmol/L) abolished this effect. Coronary PVAT increased baseline tension and potentiated constriction of isolated arteries to prostaglandin F2α in proportion to the amount of PVAT present (0.1-1.0 g). These effects were elevated in tissues obtained from obese swine and were observed in intact and endothelium denuded arteries. Coronary PVAT also diminished H2O2-mediated vasodilation in lean and, to a lesser extent, in obese arteries. These effects were associated with alterations in the obese coronary PVAT proteome (detected 186 alterations) and elevated voltage-dependent increases in intracellular [Ca(2+)] in obese smooth muscle cells. Further studies revealed that the Rho-kinase inhibitor fasudil (1 µmol/L) significantly blunted artery contractions to KCl and PVAT in lean but not obese swine. Calpastatin (10 µmol/L) also augmented contractions to levels similar to that observed in the presence of PVAT. CONCLUSIONS: Vascular effects of PVAT vary according to anatomic location and are influenced by an obese phenotype. Augmented contractile effects of obese coronary PVAT are related to alterations in the PVAT proteome (eg, calpastatin), Rho-dependent signaling, and the functional contribution of K(+) and CaV1.2 channels to smooth muscle tone.

Shock wave lithotripsy targeting of the kidney and pancreas does not increase the severity of metabolic syndrome in a porcine model.

PURPOSE: We determined whether shock wave lithotripsy of the kidney of pigs with metabolic syndrome would worsen glucose tolerance or increase the risk of diabetes mellitus. MATERIALS AND METHODS: Nine-month-old female Ossabaw miniature pigs were fed a hypercaloric atherogenic diet to induce metabolic syndrome. At age 15 months the pigs were treated with 2,000 or 4,000 shock waves (24 kV at 120 shock waves per minute) using an unmodified HM3 lithotripter (Dornier MedTech, Kennesaw, Georgia). Shock waves were targeted to the left kidney upper pole calyx to model treatment that would also expose the pancreatic tail to shock waves. The intravenous glucose tolerance test was done in conscious fasting pigs before lithotripsy, and 1 and 2 months after lithotripsy with blood samples taken for glucose and insulin measurement. RESULTS: Pigs fed the hypercaloric atherogenic diet were obese, dyslipidemic, insulin resistant and glucose intolerant, consistent with metabolic syndrome. Assessments of insulin resistance, glucose tolerance and pancreatic ß cell function from fasting plasma glucose and insulin levels, and the glucose and insulin response profile to the intravenous glucose tolerance test were similar before and after lithotripsy. CONCLUSIONS: The metabolic syndrome status of pigs treated with shock wave lithotripsy was unchanged 2 months after kidney treatment with 2,000 high amplitude shock waves or overtreatment with 4,000 high amplitude shock waves. These findings do not support a single shock wave lithotripsy treatment of the kidney as a risk factor for the onset of diabetes mellitus.

Insulin-Dendrimer Nanocomplex for Multi-Day Glucose-Responsive Therapy in Mice and Swine.

The management of diabetes in a manner offering autonomous insulin therapy responsive to glucose-directed need, and moreover with a dosing schedule amenable to facile administration, remains an ongoing goal to improve the standard of care. While basal insulins with reduced dosing frequency, even once-weekly administration, are on the horizon, there is still no approved therapy that offers glucose-responsive insulin function. Herein, a nanoscale complex combining both electrostatic- and dynamic-covalent interactions between a synthetic dendrimer carrier and an insulin analogue modified with a high-affinity glucose-binding motif yields an injectable insulin depot affording both glucose-directed and long-lasting insulin availability. Following a single injection, it is even possible to control blood glucose for at least one week in diabetic swine subjected to daily oral glucose challenges. Measurements of serum insulin concentration in response to challenge show increases in insulin corresponding to elevated blood glucose levels, an uncommon finding even in preclinical work on glucose-responsive insulin. Accordingly, the subcutaneous nanocomplex that results from combining electrostatic- and dynamic-covalent interactions between a modified insulin and a synthetic dendrimer carrier affords a glucose-responsive insulin depot for week-long control following a single routine injection.

Adult Ossabaw Pigs Prefer Fermented Sorghum Tea over Isocaloric Sweetened Water.

Ossabaw pigs (n = 11; 5-gilts, 6-barrows; age 15.6 ± 0.62 SD months) were exposed to a three-choice preference maze to evaluate preference for fermented sorghum teas (FSTs). After conditioning, pigs were exposed, in four sessions, to choices of white FST, sumac FST, and roasted sumac-FST. Then, pigs were exposed, in three sessions, to choices of deionized H2O (-control; avoidance), isocaloric control (+control; deionized H2O and sucrose), and blended FST (3Tea) (equal portions: white, sumac, and roasted sumac). When tea type was evaluated, no clear preference behaviors for tea type were observed (p > 0.10). When the 3Tea and controls were evaluated, pigs consumed minimal control (p < 0.01;18.0 ± 2.21% SEM), and they consumed great but similar volumes of +control and 3Tea (96.6 and 99.0 ± 2.21% SEM, respectively). Likewise, head-in-bowl duration was the least for -control, but 3Tea was the greatest (p < 0.01; 5.6 and 31.9 ± 1.87% SEM, respectively). Head-in-bowl duration for +control was less than 3Tea (p < 0.01; 27.6 vs. 31.9 ± 1.87% SEM). Exploration duration was the greatest in the area with the -control (p < 0.01; 7.1 ± 1.45% SEM), but 3Tea and +control exploration were not different from each other (1.4 and 3.0 ± 1.45% SEM, respectively). Regardless of tea type, adult pigs show preference for FST, even over +control. Adult pigs likely prefer the complexity of flavors, rather than the sweetness alone.

A portable fiberoptic ratiometric fluorescence analyzer provides rapid point-of-care determination of glomerular filtration rate in large animals.

Measurement of the glomerular filtration rate (GFR) is the gold standard for precise assessment of kidney function. A rapid, point-of-care determination of the GFR may provide advantages in the clinical setting over currently available assays. Here we demonstrate a proof of principle for such an approach in a pig and dogs, two species that approximate the vascular access and GFR results expected in humans. In both animal models, a sub-millimeter optical fiber that delivered excitation light and collected fluorescent emissions was inserted into a peripheral vein (dog) or central venous access (pig) by means of commercial intravenous catheters. A mixture of fluorescent chimeras of a small freely filterable reporter and large non-filterable plasma volume marker were infused as a bolus, excited by light-emitting diodes, and the in vivo signals detected and quantified by photomultiplier tubes in both species in less than 60 min. Concurrent standardized 6-h iohexol plasma kidney clearances validated the accuracy of our results for both physiologic and a chronic kidney disease setting. Thus, our ratiometric technique allows for both measurement of plasma vascular volume and highly accurate real-time GFR determinations, enabling clinical decision making in real time.

Intracellular Ca2+ Dysregulation in Coronary Smooth Muscle Is Similar in Coronary Disease of Humans and Ossabaw Miniature Swine.

Intracellular free Ca2+ ([Ca2+]i) dysregulation occurs in coronary smooth muscle (CSM) in atherosclerotic coronary artery disease (CAD) of metabolic syndrome (MetS) swine. Our goal was to determine how CAD severity, arterial structure, and MetS risk factors associate with [Ca2+]i dysregulation in human CAD compared to changes in Ossabaw miniature swine. CSM cells were dispersed from coronary arteries of explanted hearts from transplant recipients and from lean and MetS swine with CAD. CSM [Ca2+]i elicited by Ca2+ influx and sarcoplasmic reticulum (SR) Ca2+ release and sequestration was measured with fura-2. Increased [Ca2+]i signaling was associated with advanced age and a greater media area in human CAD. Decreased [Ca2+]i signaling was associated with a greater number of risk factors and a higher plaque burden in human and swine CAD. Similar [Ca2+]i dysregulation exhibited in human and Ossabaw swine CSM provides strong evidence for the translational relevance of this large animal model.

Effect of diet-induced obesity and metabolic syndrome on skeletal muscles of Ossabaw miniature swine.

Ossabaw swine fed excess kilocalorie diet develop metabolic syndrome (MS) characterized by obesity, hypertension, insulin resistance, and glucose intolerance with/without dyslipidemia. The purpose of this study was to test the hypothesis that MS would have a detrimental effect on skeletal muscle structure and cause changes in the expression of myosin heavy chains (MHCs). Adult male Ossabaw swine were fed for 24 wk high-fructose or high-fat/cholesterol/fructose diets to induce normolipidemic MS (MetS) or dyslipidemic MS (DMetS), respectively, and were compared with the lean swine on control diet. MetS swine showed mild MS, lacking increases in total and low density lipoprotein (LDL) cholesterol, both of which were highly upregulated in DMetS swine. There was an ∼1.2-fold increase in the cross-sectional areas of muscle fibers in MetS and DMetS groups compared with control for biceps femoris and plantaris muscles. In plantaris muscles, DMetS diet caused an ∼2-fold decrease in slow MHC mRNA and protein expression and an ∼1.2- to 1.8-fold increase in the number of intramyocellular lipid (IMCL) droplets without large changes in the size of the droplets. There was a trend to the decrease in slow MHC expression in muscles of swine on MetS diet. The number of IMCL droplets in muscle fibers of the MetS group was comparable to controls. These data correlate well with the data on total plasma cholesterol (control = 60, MetS = 70, and DMetS = 298 mg/dl) and LDL (control = 29, MetS = 30, and DMetS = 232 mg/dl). We conclude that structural changes observed in skeletal muscle of obese Ossabaw swine correlate with those previously reported for obese humans.

Epicardial perivascular adipose-derived leptin exacerbates coronary endothelial dysfunction in metabolic syndrome via a protein kinase C-beta pathway.

OBJECTIVE: Factors released by perivascular adipose tissue (PVAT) disrupt coronary endothelial function via phosphorylation of endothelial NO synthase by protein kinase C (PKC)-beta. However, our understanding of how PVAT potentially contributes to coronary disease as a complication of obesity/metabolic syndrome (MetS) remains limited. The current study investigated whether PVAT-derived leptin impairs coronary vascular function via PKC-beta in MetS. METHODS AND RESULTS: Coronary arteries with and without PVAT were collected from lean or MetS Ossabaw miniature swine for isometric tension studies. Endothelial-dependent vasodilation to bradykinin was significantly reduced in MetS. PVAT did not affect bradykinin-mediated dilation in arteries from lean swine but significantly exacerbated endothelial dysfunction in arteries from MetS swine. PVAT-induced impairment was reversed by inhibition of either PKC-beta with ruboxistaurin (Eli Lilly and Company, Indianapolis, Ind) or leptin receptor signaling with a recombinant, pegylated leptin antagonist. Western blot and immunohistochemical analyses demonstrated increased PVAT-derived leptin and coronary leptin receptor density with MetS. Coronary PKC-beta activity was increased in both MetS arteries exposed to PVAT and lean arteries exposed to leptin. Finally, leptin-induced endothelial dysfunction was reversed by ruboxistaurin. CONCLUSIONS: Increases in epicardial PVAT leptin exacerbate coronary endothelial dysfunction in MetS via a PKC-beta-dependent pathway. These findings implicate PVAT-derived leptin as a potential contributor to coronary atherogenesis in MetS.

Serum proteomic analysis of diet-induced steatohepatitis and metabolic syndrome in the Ossabaw miniature swine.

We recently developed a nutritional model of steatohepatitis and metabolic syndrome in Ossabaw pigs. Here we describe changes in the serum proteome of pigs fed standard chow (control group; n = 7), atherogenic diet (n = 5), or modified atherogenic diet (M-ath diet group; n = 6). Pigs fed atherogenic diet developed metabolic syndrome and mildly abnormal liver histology, whereas pigs fed M-ath diet exhibited severe metabolic syndrome and liver injury closely resembling human nonalcoholic steatohepatitis (NASH). Using a label-free mass spectrometry-based proteomics approach, we identified 1,096 serum proteins, 162 of which changed significantly between any two diet groups (false discovery rate <5%). Biological classification of proteins with significant changes revealed functions previously implicated in development of NASH in humans, including immune system regulation and inflammation (orosomucoid 1, serum amyloid P component, paraoxonase 1, protein similar to alpha-2-macroglobulin precursor, beta-2-microglobulin, p101 protein, and complement components 2 and C8G), lipid metabolism (apolipoproteins C-III, E, E precursor, B, and N), structural and extracellular matrix proteins (transthyretin and endopeptidase 24.16 type M2), and coagulation [carboxypeptidase B2 (plasma)]. Several proteins with significant differential expression in pigs were also identified in our recent human proteomics study as changing significantly in serum from patients across the spectrum of nonalcoholic fatty liver disease, including apolipoproteins C-III and B, orosomucoid 1, serum amyloid P component, transthyretin, paraoxonase 1, and a protein similar to alpha-2-macroglobulin precursor. This serum proteomic analysis provides additional information about the pathogenesis of NASH and further characterizes our large animal model of diet-induced steatohepatitis and metabolic syndrome in Ossabaw pigs.

Metabolic syndrome reduces the contribution of K+ channels to ischemic coronary vasodilation.

This investigation tested the hypothesis that metabolic syndrome decreases the relative contribution of specific K(+) channels to coronary reactive hyperemia. Ca(2+)-activated (BK(Ca)), voltage-activated (K(V)), and ATP-dependent (K(ATP)) K(+) channels were investigated. Studies were conducted in anesthetized miniature Ossabaw swine fed a normal maintenance diet (11% kcal from fat) or an excess calorie atherogenic diet (43% kcal from fat, 2% cholesterol, 20% kcal from fructose) for 20 wk. The latter diet induces metabolic syndrome, increasing body weight, fasting glucose, total cholesterol, and triglyceride levels. Ischemic vasodilation was determined by the coronary flow response to a 15-s occlusion before and after cumulative administration of antagonists for BK(Ca) (penitrem A; 10 microg/kg iv), K(V) (4-aminopyridine; 0.3 mg/kg iv) and K(ATP) (glibenclamide; 1 mg/kg iv) channels. Coronary reactive hyperemia was diminished by metabolic syndrome as the repayment of flow debt was reduced approximately 30% compared with lean swine. Inhibition of BK(Ca) channels had no effect on reactive hyperemia in either lean or metabolic syndrome swine. Subsequent inhibition of K(V) channels significantly reduced the repayment of flow debt ( approximately 25%) in both lean and metabolic syndrome swine. Additional blockade of K(ATP) channels further diminished ( approximately 45%) the repayment of flow debt in lean but not metabolic syndrome swine. These data indicate that the metabolic syndrome impairs coronary vasodilation in response to cardiac ischemia via reductions in the contribution of K(+) channels to reactive hyperemia.

Contribution of BK(Ca) channels to local metabolic coronary vasodilation: Effects of metabolic syndrome.

This investigation was designed to examine the hypothesis that impaired function of coronary microvascular large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels in metabolic syndrome (MetS) significantly attenuates the balance between myocardial oxygen delivery and metabolism at rest and during exercise-induced increases in myocardial oxygen consumption (MVo(2)). Studies were conducted in conscious, chronically instrumented Ossabaw swine fed a normal maintenance diet (11% kcal from fat) or an excess calorie atherogenic diet (43% kcal from fat, 2% cholesterol, 20% kcal from fructose) that induces many common features of MetS. Data were collected under baseline/resting conditions and during graded treadmill exercise before and after selective blockade of BK(Ca) channels with penitrem A (10 microg/kg iv). We found that the exercise-induced increases in blood pressure were significantly elevated in MetS swine. No differences in baseline cardiac function or heart rate were noted. Induction of MetS produced a parallel downward shift in the relationship between coronary venous Po(2) and MVo(2) (P < 0.001) that was accompanied by a marked release of lactate (negative lactate uptake) as MVo(2) was increased with exercise (P < 0.005). Inhibition of BK(Ca) channels with penitrem A did not significantly affect blood pressure, heart rate, or the relationship between coronary venous Po(2) and MVo(2) in lean or MetS swine. These data indicate that BK(Ca) channels are not required for local metabolic control of coronary blood flow under physiological (lean) or pathophysiological (MetS) conditions. Therefore, diminished function of BK(Ca) channels does not contribute to the impairment of myocardial oxygen-supply demand balance in MetS.

Marvels, mysteries, and misconceptions of vascular compensation to peripheral artery occlusion.

Peripheral arterial disease is a major health problem and there is a significant need to develop therapies to prevent its progression to claudication and critical limb ischemia. Promising results in rodent models of arterial occlusion have generally failed to predict clinical success and led to questions of their relevance. While sub-optimal models may have contributed to the lack of progress, we suggest that advancement has also been hindered by misconceptions of the human capacity for compensation and the specific vessels which are of primary importance. We present and summarize new and existing data from humans, Ossabaw miniature pigs, and rodents which provide compelling evidence that natural compensation to occlusion of a major artery (i) may completely restore perfusion, (ii) occurs in specific pre-existing small arteries, rather than the distal vasculature, via mechanisms involving flow-mediated dilation and remodeling (iii) is impaired by cardiovascular risk factors which suppress the flow-mediated mechanisms and (iv) can be restored by reversal of endothelial dysfunction. We propose that restoration of the capacity for flow-mediated dilation and remodeling in small arteries represents a largely unexplored potential therapeutic opportunity to enhance compensation for major arterial occlusion and prevent the progression to critical limb ischemia in the peripheral circulation.

Adenosine receptor regulation of coronary blood flow in Ossabaw miniature swine.

Adenosine clearly regulates coronary blood flow (CBF); however, contributions of specific adenosine receptor (AR) subtypes (A(1), A(2A), A(2B), A(3)) to CBF in swine have not been determined. ARs generally decrease (A(1), A(3)) or increase (A(2A), A(2B)) cyclic adenosine monophosphate, a major mediator of vasodilation. We hypothesized that A(1) antagonism potentiates coronary vasodilation and coronary stent deployment in dyslipidemic Ossabaw swine elicits impaired vasodilation to adenosine that is associated with increased A(1)/A(2A) expression. The left main coronary artery was accessed with a guiding catheter allowing intracoronary infusions. After placement of a flow wire into the left circumflex coronary artery the responses to bolus infusions of adenosine were obtained. Steady-state infusion of AR-specific agents was achieved by using a small catheter fed over the flow wire in control pigs. CBF was increased by the A(2)-nonselective agonist 2-phenylaminoadenosine (CV1808) in a dose-dependent manner. Baseline CBF was increased by the highly A(1)-selective antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), but not changed by other AR-specific agents. The nonselective A(2) antagonist 3,7-dimethyl-1-propargylxanthine and A(2A)-selective antagonist 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM241385) abolished adenosine-induced CBF, whereas A(2B) and A(3) antagonism had no effect. Dyslipidemia and stenting decreased adenosine-induced CBF ∼70%, whereas A(1), A(2A), and A(2B) mRNA were up-regulated in dyslipidemic versus control >5-fold and there was no change in the ratio of A(1)/A(2A) protein in microvessels distal to the stent. In control Ossabaw swine A(1) antagonism by DPCPX positively regulated basal CBF. Impaired adenosine-induced CBF after stenting in dyslipidemia is most likely caused by the altered balance between A(1) and A(2A) signaling, not receptor expression.