Daryl O. Schwenke

Regulator of G-Protein Signaling Subtype 4 Mediates Antihypertrophic Effect of Locally Secreted Natriuretic Peptides in the Heart

Background— Mice lacking guanylyl cyclase-A (GC-A), a natriuretic peptide receptor, have pressure-independent cardiac hypertrophy. However, the mechanism underlying GC-A-mediated inhibition of cardiac hypertrophy remains to be elucidated. In the present report, we examined the role of regulator of G-protein signaling subtype 4 (RGS4), a GTPase activating protein for Gq and Gi, in the antihypertrophic effects of GC-A. Methods and Results— In cultured cardiac myocytes, treatment of atrial natriuretic peptide stimulated the binding of guanosine 3′,5′-cyclic monophosphate-dependent protein kinase (PKG) I-&agr; to RGS4, PKG-dependent phosphorylation of RGS4, and association of RGS4 and G&agr;q. In contrast, blockade of GC-A by an antagonist, HS-142-1, attenuated the phosphorylation of RGS4 and association of RGS4 and G&agr;q. Moreover, overexpressing a dominant negative form of RGS4 diminished the inhibitory effects of atrial natriuretic peptide on endothelin-1–stimulated inositol 1,4,5-triphosphate production, [3H]leucine incorporation, and atrial natriuretic peptide gene expression. Furthermore, expression and phosphorylation of RGS4 were significantly reduced in the hearts of GC-A knockout (GC-A-KO) mice compared with wild-type mice. For further investigation, we constructed cardiomyocyte-specific RGS4 transgenic mice and crossbred them with GC-A-KO mice. The cardiac RGS4 overexpression in GC-A-KO mice significantly reduced the ratio of heart to body weight (P<0.001), cardiomyocyte size (P<0.01), and ventricular calcineurin activity (P<0.05) to 80%, 76%, and 67% of nontransgenic GC-A-KO mice, respectively. It also significantly suppressed the augmented cardiac expression of hypertrophy-related genes in GC-A-KO mice. Conclusions— These results provide evidence that GC-A activates cardiac RGS4, which attenuates G&agr;q and its downstream hypertrophic signaling, and that RGS4 plays important roles in GC-A-mediated inhibition of cardiac hypertrophy.

Early Ghrelin Treatment after Myocardial Infarction Prevents an Increase in Cardiac Sympathetic Tone and Reduces Mortality

Acute myocardial infarction (MI) initiates an increase in cardiac sympathetic nerve activity (CSNA), which ultimately exacerbates chronic cardiac dysfunction. Ghrelin (Ghr), a GH-releasing peptide, is an effective treatment for improving cardiac function in chronic heart failure. Ghr also suppresses renal sympathetic nerve activity (SNA) and, therefore, may have important therapeutic benefits in the early stages of acute MI: by reducing CSNA. In this study we hypothesized that early Ghr administration may prevent an increase in CSNA in the acute phase after MI. CSNA was continuously recorded in urethane-anaesthetized rats before and for 5 h after acute MI (or sham). MI was induced by ligation of the left anterior descending coronary artery. Rats received an injection of either saline or Ghr (150 microg/kg, sc) 1 min, or 2 h, after the infarct. CSNA remained stable during the 5-h recording duration in sham rats. MI induced a maximal 110% increase in SNA, which was prevented in rats that received Ghr 1 min after infarct. When Ghr was injected 2 h after MI (SNA had increased by approximately 85%), SNA decreased to pre-MI activity. Importantly, early Ghr administration significantly reduced the high mortality rate associated with MI (61% mortality in untreated MI rats cf. approximately 23% in Ghr-treated MI rats). These results show that early Ghr treatment prevents the increase in CSNA after MI, which may contribute to the improved chances of survival. Whether these early beneficial effects of Ghr also have long-term benefits for improving cardiac function is an area that requires further investigation.

Synchrotron Radiation Imaging for Advancing Our Understanding of Cardiovascular Function

Synchrotron radiation (SR) is increasingly being used for micro-level and nano-level functional imaging in in vivo animal experiments. This review focuses on the methodology that enables repeated and regional assessment of vessel internal diameter and flow in the resistance vessels of different organ systems. In particular, SR absorption microangiography approaches offer unique opportunities for real-time in vivo vascular imaging in small animals, even during dynamic motion of the heart and lungs. We also describe recent progress in the translation of multiple phase-contrast imaging techniques from ex vivo to in vivo small-animal studies. Furthermore, we also review the utility of SR for multiple pinpoint (dimensions 0.2×0.2 mm) assessments of myocardial function at the cross-bridge level in different regions of the heart using small-angle X-ray scattering, resulting from increases in SR flux at modern facilities. Finally, we present cases for the use of complementary SR approaches to study cardiovascular function, particularly the pathological changes associated with disease using small-animal models.

Pressurization of vertebral bodies during vertebroplasty causes cardiovascular complications: an experimental study in sheep.

Study Design. An experimental study of cardiovascular complications arising during vertebroplasty (VP) of multiple levels in sheep. Objectives. To investigate the effect of pressurizing vertebral bodies during VP using different materials in the development of fat embolism (FE) and any associated cardiovascular changes. Summary of Background Data. Polymethylmethacrylate (PMMA) is the material of choice for VP. However, PMMA has several disadvantages, such as toxicity, exothermic curing, uncertain long-term biomechanical effects, and biocompatibility. Alternative materials are being developed for VP; however, there is the concern that an increase in intraosseous pressure and displacement of bone marrow contents could lead to FE and hypotension during VP regardless of what type of materials is used. Methods. In 20 sheep, four vertebral bodies were augmented either with PMMA or bone wax. Heart rate; arterial, central venous, and pulmonary artery pressure; cardiac output; and blood gas values were recorded. Postmortem the lungs were subjected to histologic evaluation. Results. The consecutive augmentation of four vertebral bodies with PMMA induced a cumulative FE that gradually deteriorated baseline mean arterial blood pressure (MABP) and blood gas values. The augmentation with bone wax resulted in similar cardiovascular changes and amount of intravascular fat in the lungs. Conclusion. There are potential cardiovascular complications during VP of multiple levels regardless of the augmentation material used. The deteriorating baseline MABP during VP is associated with the pressurization of the vertebral bodies rather than with the use of polymethylmethacrylate.

Role of Rho-kinase signaling and endothelial dysfunction in modulating blood flow distribution in pulmonary hypertension

Rho-kinase-mediated vasoconstriction and endothelial dysfunction are considered two primary instigators of pulmonary arterial hypertension (PAH). However, their contribution to the adverse changes in pulmonary blood flow distribution associated with PAH has not been addressed. This study utilizes synchrotron radiation microangiography to assess the specific role, and contribution of, Rho-kinase-mediated vasoconstriction and endothelial dysfunction in PAH. Male adult Sprague-Dawley rats were injected with saline (Cont-rats) or monocrotaline (MCT-rats) 3 wk before microangiography was performed on the left lung. We assessed dynamic changes in vessel internal diameter (ID) in response to 1) the Rho-kinase inhibitor fasudil (10 mg/kg iv); or 2) ACh (3 μg · kg⁻¹ · min⁻¹), sodium nitroprusside (SNP, 5 μg · kg⁻¹ · min⁻¹), and N(ω)-nitro-l-arginine methyl ester (l-NAME, 50 mg/kg iv). We observed that MCT-rats had fewer vessels of the microcirculation compared with Cont-rats. The fundamental result of this study is that fasudil improved pulmonary blood flow distribution and reduced pulmonary pressure in PAH rats, not only by dilating already-perfused vessels (ID > 100 μm), but also by restoring blood flow to vessels that had previously been constricted closed (ID < 100 μm). Endothelium-dependent vasodilation was impaired in MCT-rats primarily in vessels with an ID < 200 μm. Moreover the vasoconstrictor response to l-NAME was accentuated in MCT-rats, but only in the 200- to 300-μm vessels. These results highlight the importance of Rho-kinase-mediated control and endothelial control of pulmonary vascular tone in PAH. Indeed, an effective therapeutic strategy for treating PAH should target both the smooth muscle Rho-kinase and endothelial pathways.

Synchrotron-based angiography for investigation of the regulation of vasomotor function in the microcirculation in vivo.

1 Real‐time imaging of the vascular networks of any organ system in vivo is possible with synchrotron radiation (SR) angiography. In this review, we discuss the advantages of SR angiography over clinical X‐ray imaging and other non‐ionizing imaging modalities. Current limitations are also described. 2 The usefulness of dual‐energy and temporal subtraction approaches to K‐edge iodine imaging are compared. 3 High‐resolution images of the microcirculation in small animals are now being collected routinely by multiple research groups through public access research programmes at synchrotrons worldwide. Such images are permitting unrivalled insights into vasomotor regulation deep within intact organ systems, such as the brain, kidney, lung and heart. For example, recent observations indicate changes in vascular control mechanisms in pulmonary hypertension that are specific to certain branching segments of the pulmonary circulation. 4 New possibilities for non‐iodinated contrast agents in SR angiography are briefly described. 5 High‐resolution angiography in vivo using SR will now allow us to identify vessels with localized or non‐uniform vasoconstriction in states such as diabetes or to characterize the extent of endothelial dysfunction in the circulation following hypertension or ischaemic–reperfusion injury. In the near future, this research is expected to reveal the contribution of resistance vessel dysfunction to diverse pathophysiological states, such as stroke, hypertension and ischaemic heart disease.

Degradation of hydroxyapatite coating on a well-functioning femoral component.

We carried out a histological study of a proximally hydroxyapatite (HA)-coated femoral component, retrieved after 9.5 years of good function. The HA coating had completely degraded. Bone was in direct contact with the titanium surface in all the areas which had been coated, with no interposing fibrous tissue. There were no signs of particles, third-body wear, adverse tissue reactions or osteolysis. Bone remodelling was evident by the presence of resorption lacunae; tetracycline labelling showed bone laid down six years after implantation. The loss of the HA-coating had no negative effect on the osseo-integration of the stem. We conclude that the HA coating contributes to the fixation of the implant and that its degradation does not adversely affect the long-term fixation.

Dynamic Synchrotron Imaging of Diabetic Rat Coronary Microcirculation In Vivo

Objective—In diabetes, long-term micro- and macrovascular damage often underlies the functional decline in the cardiovascular system. However, it remains unclear whether early-stage diabetes is associated with in vivo functional impairment in the coronary microvasculature. Synchrotron imaging allows us to detect and quantify regional differences in resistance microvessel caliber in vivo, even under conditions of high heart rate. Methods and Results—Synchrotron cine-angiograms of the coronary vasculature were recorded using anesthetized Sprague-Dawley rats 3 weeks after treatment with vehicle or streptozotocin (diabetic). In the early diabetic state, in the presence of nitric oxide and prostacyclin, vessel diameters were smaller (P<0.01) and endothelium-dependent vessel recruitment was already depressed (P<0.05). Endothelium-dependent and -independent vasodilatory responses in individual coronary vessels were not different in vivo. Inhibition of NO and PGI2 production in diabetes uncovered early localized impairment in dilation. Diabetic animals displayed focal stenoses and segmental constrictions during nitric oxide synthase/cyclooxygenase blockade, which persisted during acetylcholine infusion (P<0.05), and a strong trend toward loss of visible microvessels. Conclusion—Synchrotron imaging provides a novel method to investigate coronary microvascular function in vivo at all levels of the arterial tree. Furthermore, we have shown that early-stage diabetes is associated with localized coronary microvascular endothelial dysfunction.

One Dose of Ghrelin Prevents the Acute and Sustained Increase in Cardiac Sympathetic Tone after Myocardial Infarction

Acute myocardial infarction (MI) increases sympathetic nerve activity (SNA) to the heart, which exacerbates chronic cardiac deterioration. The hormone ghrelin, if administered soon after an MI, prevents the increase in cardiac SNA and improves early survival prognosis. Whether these early beneficial effects of ghrelin also impact on cardiac function in chronic heart failure has not yet been addressed and thus was the aim of this study. MI was induced in Sprague Dawley rats by ligating the left coronary artery. One bolus of saline (n = 7) or ghrelin (150 μg/kg, sc, n = 9) was administered within 30 min of MI. Two weeks after the infarct (or sham; n = 7), rats were anesthetized and cardiac function was evaluated using a Millar pressure-volume conductance catheter. Cardiac SNA was measured using whole-nerve electrophysiological techniques. Untreated-MI rats had a high mortality rate (50%), evidence of severe cardiac dysfunction (ejection fraction 28%; P < 0.001), and SNA was significantly elevated (102% increase; P = 0.03). In comparison, rats that received a single dose of ghrelin after the MI tended to have a lower mortality rate (25%; P = NS) and no increase in SNA, and cardiac dysfunction was attenuated (ejection fraction of 43%; P = 0.014). This study implicates ghrelin as a potential clinical treatment for acute MI but also highlights the importance of therapeutic intervention in the early stages after acute MI. Moreover, these results uncover an intricate causal relationship between early and chronic changes in the neural control of cardiac function in heart failure.

Exercise mediated protection of diabetic heart through modulation of microRNA mediated molecular pathways

Hyperglycaemia, hypertension, dyslipidemia and insulin resistance collectively impact on the myocardium of people with diabetes, triggering molecular, structural and myocardial abnormalities. These have been suggested to aggravate oxidative stress, systemic inflammation, myocardial lipotoxicity and impaired myocardial substrate utilization. As a consequence, this leads to the development of a spectrum of cardiovascular diseases, which may include but not limited to coronary endothelial dysfunction, and left ventricular remodelling and dysfunction. Diabetic heart disease (DHD) is the term used to describe the presence of heart disease specifically in diabetic patients. Despite significant advances in medical research and long clinical history of anti-diabetic medications, the risk of heart failure in people with diabetes never declines. Interestingly, sustainable and long-term exercise regimen has emerged as an effective synergistic therapy to combat the cardiovascular complications in people with diabetes, although the precise molecular mechanism(s) underlying this protection remain unclear. This review provides an overview of the underlying mechanisms of hyperglycaemia- and insulin resistance-mediated DHD with a detailed discussion on the role of different intensities of exercise in mitigating these molecular alterations in diabetic heart. In particular, we provide the possible role of exercise on microRNAs, the key molecular regulators of several pathophysiological processes.