Richardson, Darcy M.

AbstractOver the past several decades, molecular genetic testing volumes have grown and testing has expanded from single‐gene assays to multigene panels, exome sequencing, and genome sequencing. The number of molecular genetic variants that require manual interpretation has grown simultaneously, resulting in an increased demand for education on molecular variant evaluation (MVE). To meet this growing need, a team of genetic counselors and educational experts undertook a quality improvement (QI) initiative with the objectives of assessing, standardizing, and scaling access to MVE education, without increasing instructor time to deliver the education. Using the Six Sigma define–measure–analyze–improve–control (DMAIC) framework, a flipped learning course with a series of standardized online modules was developed to deliver MVE education in an enduring and accessible format for a diverse group of learners. Outcome measures included the number of online modules developed, the number of individual learners and unique learner groups accessing MVE education, and direct instruction time required to deliver MVE education. Countermeasures to ensure maintenance of educational quality included post‐course learner satisfaction scores and performance on competency assessments. Both the total number of learners and the number of unique learner groups accessing MVE education increased, while instructor time required to deliver content per learner decreased. Learner satisfaction scores remained constant and performance on competency assessments improved. The QI initiative successfully scaled MVE education to a diverse group of learners without decreasing learner outcomes or satisfaction. The flipped learning format provides a scalable and flexible educational model for instructors and learners in a rapidly changing environment that often includes remote work and education.

Abstract —Insulin and insulin-like growth factor (IGF) 1 affect coronary vasoactivity. Experimental hypercholesterolemia is associated with coronary atherogenesis and altered vasomotor regulation. Because the IGF axis is altered during atherogenesis, we postulated that experimental hypercholesterolemia is associated with an altered coronary vasoactive response to IGF-1 in vitro. Coronary arteries and arterioles from pigs fed either a normal or high-cholesterol diet for 10 weeks were contracted with endothelin-1 and relaxed with cumulative concentrations of insulin or IGF-1 (10 −12 to 10 −7 mol/L). Control arterioles were also incubated with the nitric oxide synthase inhibitor 10 −4 mol/L N G -monomethyl- l -arginine (L-NMMA) or the potassium channel blocker 10 −2 mol/L tetraethylammonium (TEA), contracted with endothelin-1, and relaxed with insulin or IGF-1. Experimental hypercholesterolemia (1) increased serum cholesterol (9.5±1.0 versus 1.9±0.08 mmol/L; P <0.0001), (2) caused coronary arterial and arteriolar endothelial dysfunction in vitro (attenuated vasorelaxation to bradykinin), (3) did not alter the epicardial response to either insulin ( P =0.80) or IGF-1 ( P =0.12), and (4) significantly attenuated the arteriolar response to IGF-1 (maximal relaxation of 79±6% versus 42±8%; P =0.01) but not insulin (43±6% versus 53±7%; P =0.99). Control arteriolar vasorelaxation to IGF-1 was attenuated by both L-NMMA ( P <0.001) and TEA ( P =0.01), whereas only L-NMMA attenuated insulin ( P <0.001). Staining for IGF-1 and IGF binding protein 2 was increased ( P <0.05) in arterioles of cholesterol-fed pigs. IGF-1 and insulin are therefore coronary arteriolar vasorelaxants through different mechanisms. Experimental hypercholesterolemia is associated with resistance to the coronary arteriolar vasorelaxing effects of IGF-1 but not insulin, in conjunction with increased ligand and binding-protein expression. The IGF axis may contribute to the altered coronary vasoactivity in hypercholesterolemia.

Abstract—Insulin and insulin-like growth factor-I (IGF-I) may play a role in the modulation of coronary artery tone, yet there are few data regarding their vasoactive effects on the coronary vascular bed. We evaluated the vasorelaxation effects of insulin and IGF-I on porcine coronary epicardial vessels in vitro and elucidated possible mechanisms. Porcine epicardial arteries were contracted with 10−7mol/L endothelin-1 and relaxed with cumulative concentrations of either insulin or IGF-I (10−12to 10−7mol/L). The above experiments were repeated in vessels without endothelium. Vessels were also incubated with the nitric oxide synthase inhibitorNG-monomethyl-l-arginine (L-NMMA; 10−4mol/L) with and without 10−3.5mol/Ll-arginine, the potassium channel blocker tetraethylammonium (TEA; 10−2mol/L), and the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-α]quinoxalin-1-one (ODQ; 10−5.5mol/L); vessels were then contracted with endothelin-1 and relaxed with insulin or IGF-I. Insulin and IGF-I were also added after contraction with 60 mmol/L KCl. Insulin and IGF-I caused a similar decrease in coronary epicardial tension after contraction with endothelin-1 (relaxation of 28±4% [n=7] and 25±3% [n=8] with insulin and IGF-I, respectively;P<0.0001 for both peptides). Removal of the endothelium did not affect these responses. Incubation with L-NMMA, but not ODQ, attenuated the vasorelaxation response to insulin and IGF in vessels without endothelium.l-Arginine did not reverse this effect of L-NMMA. KCl and TEA attenuated the vasorelaxation effect of both insulin and IGF-I. Thus, both insulin and IGF-I caused non–endothelium-dependent coronary vasorelaxation in vitro, probably through a mechanism involving the activation of potassium channels. These findings suggest that insulin and IGF-I participate in the regulation of coronary vasomotor tone.

Abstract—Hypercholesterolemia is associated with increased circulating and tissue endothelin-1 immunoreactivity, decreased nitric oxide (NO) activity, and altered endothelial function. We tested the hypothesis that chronic endothelin receptor antagonism preserves endothelial function and increases NO in experimental porcine hypercholesterolemia. Pigs were randomized to 3 groups: Group 1, a 2% high-cholesterol (HC) diet alone (n=7); group 2, RO-48-5695, a combined endothelin receptor antagonist, and an HC diet (n=8); and group 3, ABT-627, a selective endothelin-A receptor antagonist, and an HC diet (n=8). Coronary epicardial and arteriolar endothelial function was determined by a dose-response relaxation to bradykinin (10−11to 10−6mol/L),in all groups and in pigs maintained on a normal diet. Plasma total oxidized products of NO (NOx) were determined by chemiluminescence at baseline and after 12 weeks. Bradykinin-stimulated coronary epicardial and arteriolar relaxation in group 1 was attenuated compared with normal-diet controls. This relaxation was normalized with endothelin receptor antagonism. Plasma NOxdecreased after 12 weeks in group 1 (−74.8±5.5%). This decrease was attenuated in the endothelin receptor antagonist groups (group 2, −28.2±15.0%; group 3, −38.9±20.6%). Chronic endothelin receptor antagonism preserves coronary endothelial function and increases NO in hypercholesterolemia. This study supports a role of endothelin-1 in the regulation of NO activity and suggests a possible therapeutic role for endothelin receptor antagonists in hypercholesterolemia.

Erythropoietin (EPO) fosters tissue oxygenation by stimulating erythropoiesis. More recently, EPO has been recognized as a tissue-protective cytokine. In this study, we tested the hypothesis that endothelial NO synthase (eNOS) plays a key role in the vascular protective effect of EPO. A murine model of wire-induced injury of carotid artery was used to examine the effect of EPO on endothelial repair and arterial wall architecture. Recombinant human EPO (1000 U/kg, SC, biweekly) was administered for 2 weeks in wild-type and eNOS-deficient mice after which reactivity of isolated carotid arteries was studied in vitro, and the vasculature was histologically assessed. Injured arteries exhibited impairment of endothelium-dependent relaxations to acetylcholine ( P <0.05). This was associated with increased medial cross-sectional area ( P <0.05). EPO upregulated expression of phosphorylated Ser1177-eNOS and normalized the vasodilator response to acetylcholine ( P <0.05). Furthermore, EPO prevented the injury-induced increase in medial cross-sectional area ( P <0.05). The vascular protective effects of EPO were abolished in eNOS-deficient mice. Most notably, EPO significantly increased systolic blood pressure and enhanced medial thickening of injured carotid arteries in eNOS-deficient mice ( P <0.05). Our results demonstrate that EPO prevents aberrant remodeling of the injured carotid artery. The protective effects of EPO are critically dependent on activation of eNOS.

In cultured endothelial cells, the antioxidant, l -ascorbic acid (vitamin C), increases nitric oxide synthase (NOS) enzyme activity via chemical stabilization of tetrahydrobiopterin. Our objective was to determine the effect of vitamin C on NOS function and tetrahydrobiopterin metabolism in vivo. Twenty-six to twenty-eight weeks of diet supplementation with vitamin C (1%/kg chow) significantly increased circulating levels of vitamin C in wild-type (C57BL/6J) and apolipoprotein E (apoE)–deficient mice. Measurements of NOS enzymatic activity in aortas of apoE-deficient mice indicated a significant increase in total NOS activity. However, this increase was mainly due to high activity of inducible NOS, whereas eNOS activity was reduced. Significantly higher tetrahydrobiopterin levels were detected in aortas of apoE-deficient mice. Long-term treatment with vitamin C restored endothelial NOS activity in aortas of apoE-deficient mice, but did not affect activity of inducible NOS. In addition, 7,8-dihydrobiopterin levels, an oxidized form of tetrahydrobiopterin, were decreased and vascular endothelial function of aortas was significantly improved in apoE-deficient mice. Interestingly, vitamin C also increased tetrahydrobiopterin and NOS activity in aortas of C57BL/6J mice. In contrast, long-term treatment with vitamin E (2000 U/kg chow) did not affect vascular NOS activity or metabolism of tetrahydrobiopterin. In vivo, beneficial effect of vitamin C on vascular endothelial function appears to be mediated in part by protection of tetrahydrobiopterin and restoration of eNOS enzymatic activity.