Ed VanBavel

Microparticles in cardiovascular diseases

Microparticles are membrane vesicles released from many different cell types. There are two mechanisms that can result in their formation, cell activation and apoptosis. In these two mechanisms, different pathways are involved in microparticle generation. Microparticle generation seems to be a well regulated process. Microparticles vary in size, phospholipid and protein composition. They have a potent pro-inflammatory effect, promote coagulation and affect vascular function. Since these processes are all involved in the pathogenesis of cardiovascular disease and circulating microparticle numbers are altered in many cardiovascular diseases, a role for microparticles in the pathogenesis of cardiovascular diseases is likely. Although hard evidence for a role of microparticles in cardiovascular diseases at present is still only limited, new evidence is accumulating rapidly to support this theory. Elucidation of the microparticle composition and the mechanisms involved in exertion of their effects will supply this evidence and enable us to develop additional intervention strategies for prevention and treatment of cardiovascular diseases.

Vascular function in preeclampsia.

Preeclampsia is a multisystem disorder peculiar to human pregnancy. It occurs in 4-5% of all pregnancies and remains a leading cause of maternal and neonatal mortality and morbidity. The pathophysiology of this syndrome is not fully understood. Two stages of vascular dysfunction seem to be involved. In the early stage suboptimal development of the placenta and a hemodynamic maladaptation to pregnancy exist. At this stage maternal constitutional factors such as genetic and immunological factors and pre-existing vascular diseases may play a role. Due to this defective placentation a factor is released from the placenta, supposedly under the influence of ischemia. This factor then results in the late vascular dysfunction characterised mainly by a generalised endothelial dysfunction, leading to the clinical syndrome of preeclampsia. This review attempts to unravel the mechanisms that may contribute to preeclampsia-associated changes in vascular function and to indicate the research needed to improve our understanding of this disease.

Small Artery Remodeling Depends on Tissue-Type Transglutaminase

Remodeling of small arteries is essential in the long-term regulation of blood pressure and blood flow to specific organs or tissues. A large part of the change in vessel diameter may occur through non–growth-related reorganization of vessel wall components. The hypothesis was tested that tissue-type transglutaminase (tTG), a cross-linking enzyme, contributes to the inward remodeling of small arteries. The in vivo inward remodeling of rat mesenteric arteries, induced by low blood flow, was attenuated by inhibition of tTG. Rat skeletal muscle arteries expressed tTG, as identified by Western blot and immunostaining. In vitro, activation of these arteries with endothelin-1 resulted in inward remodeling, which was blocked by tTG inhibitors. Small arteries obtained from rats and pigs both showed inward remodeling after exposure to exogenous transglutaminase, which was inhibited by addition of a nitric oxide donor. Enhanced expression of tTG, induced by retinoic acid, increased inward remodeling of porcine coronary arteries kept in organ culture for 3 days. The activity of tTG was dependent on pressure. Inhibition of tTG reversed remodeling, causing a substantial increase in vessel diameter. In a collagen gel contraction assay, tTG determined the compaction of collagen by smooth muscle cells. Collectively, these data show that small artery remodeling associated with chronic vasoconstriction depends on tissue-type transglutaminase. This mechanism may reveal a novel therapeutic target for pathologies associated with inward remodeling of the resistance arteries.

Intracellular Ca2+, Intercellular Electrical Coupling, and Mechanical Activity in Ischemic Rabbit Papillary Muscle: Effects of Preconditioning and Metabolic Blockade

During myocardial ischemia, electrical uncoupling and contracture herald irreversible damage. In the present study, we tested the hypothesis that an increase of intracellular Ca2+ is an important factor initiating these events. Therefore, we simultaneously determined tissue resistance, mechanical activity, pH(0), and intracellular Ca2+ (with the fluorescent indicator indo 1, Molecular Probes, Inc) in arterially perfused rabbit papillary muscles. Sustained ischemia was induced in three experimental groups: (1) control, (2) preparations preconditioned with two 5-minute periods of ischemia followed by reperfusion, and (3) preparations pretreated with 1 mmol/L iodoacetate to block anaerobic metabolism and minimize acidification during ischemia. In a fourth experimental group, intracellular Ca2+ was increased under nonischemic conditions by perfusing with 0.1 mmol/L ionomycin and 0.1 mumol/L gramicidin. Ca2+ transients and contractions rapidly disappeared after the induction of ischemia. In the control group, diastolic Ca2+ began to rise after 12.6 +/- 1.3 minutes of ischemia; uncoupling, after 14.5 +/- 1.2 minutes of ischemia; and contracture, after 12.6 +/- 1.5 minutes of ischemia (mean +/- SEM). Preconditioning significantly postponed Ca2+ rise, uncoupling, and contracture (21.5 +/- 4.0, 24.0 +/- 4.1, and 23.0 +/- 5.3 minutes of ischemia, respectively). Pretreatment with iodoacetate significantly advanced these events (1.9 +/- 0.7, 3.6 +/- 0.9, and 1.9 +/- 0.2 minutes of ischemia, respectively). In all groups, the onset of uncoupling always followed the start of Ca2+ rise, whereas the start of contracture was not different from the rise in Ca2+. Perfusion with ionomycin and gramicidin permitted estimation of a threshold [Ca2+] for electrical uncoupling of 685 +/- 85 nmol/L. In conclusion, the rise in intracellular Ca2+ is the main trigger for cellular uncoupling during ischemia. Contracture is closely associated with the increase of intracellular Ca2+ during ischemia.

Branching patterns in the porcine coronary arterial tree. Estimation of flow heterogeneity.

The aim of this study is to quantify the porcine coronary arterial branching pattern and to use this quantification for the interpretation of flow heterogeneity. Two casts of the coronary arterial tree were made at diastolic arrest and maximal dilation. The relation between length and diameter of arterial segments was quantified, as well as the area expansion ratio and diameter symmetry of vascular nodes. These relations were used to construct computer models of the coronary arterial tree, covering diameters between 10 and 500 microns. Topology of these simulated trees was analyzed using Strahler ordering: Bifurcation ratio, diameter ratio, and length ratio were constant along orders 2-8 and equal to 3.30, 1.51, and 1.63, respectively. In each order, the number of segments per Strahler vessel was almost geometrically distributed. For the lowest orders, these predictions were confirmed by direct observations. From the network model, local pressure and flow were also predicted: Pressure fell from 90 to 32 mm Hg at the 10-microns level. The coefficient of variation (CV) of flow in individual segments was dependent on the number of perfused terminal segments (Nt) according to the fractal relation CV(Nt) approximately Nt(1-D), where D is the fractal dimension (1.20). CV of flow in 1-g tissue units was predicted to be 18%. This study shows that the structure of the coronary arterial bed is an important determinant of the fractal nature of local flow heterogeneity.

Inward Remodeling Follows Chronic Vasoconstriction in Isolated Resistance Arteries

The hypothesis was tested that chronic vasoconstriction is followed by a structural reduction in lumen diameter, measured at full dilation. An in vitro model of pressurized rat skeletal muscle arterioles was used. During a 3-day experimental period, constriction of active vessels was achieved with fetal calf serum or endothelin-1 (ET-1). Maximal dilation revealed inward remodeling from 179 ± 6.5 µm lumen diameter on day 0 to 151 ± 6.3 µm on day 3 at 75 mm Hg in vessels incubated with serum (n = 8). Similarly, ET-1 induced inward remodeling from 182 ± 5.2 to 164 ± 3.7 µm (n = 6). When constriction during organoid culture was inhibited with papaverin or verapamil, inward remodeling was fully prevented: 184 ± 6.3 to 184 ± 5.8 µm for papaverin (n = 6) and 174 ± 5.5 to 177 ± 7.4 µm for verapamil (n = 6). A chronic reduction in diameter without tone was achieved in vessels that were kept at a low pressure (2–5 mm Hg; n = 6). Here, no remodeling was found, thereby ruling out that a chronic reduction in diameter alone is sufficient for inward remodeling. These data show that a persistent active reduction in lumen diameter is followed by inward remodeling of arterioles.

Isolated microparticles, but not whole plasma, from women with preeclampsia impair endothelium-dependent relaxation in isolated myometrial arteries from healthy pregnant women

OBJECTIVE This study was performed to establish whether microparticles from plasma of women with preeclampsia cause endothelial dysfunction, as described for isolated myometrial arteries in preeclampsia. STUDY DESIGN Myometrial arteries were isolated from biopsy specimens obtained at cesarean delivery from healthy pregnant women (n = 22) and mounted in a wire myograph. Bradykinin concentration-response curves were obtained before and after 1-hour incubation or after overnight incubation with one of the following preparations of plasma from individual women with preeclampsia (n = 16): Whole plasma, microparticle-free plasma, isolated microparticles resuspended in physiologic saline solution or physiologic saline solution. Overnight incubation was also performed with microparticles isolated from healthy pregnant women (n = 6). One-hour incubation was performed with 2% or 10% solution and overnight incubation with 5% solution. RESULTS No effect of preeclamptic plasma, with or without microparticles, on bradykinin-mediated relaxation was observed. Overnight, but not 1-hour, incubation with preeclamptic microparticles caused abolishment of bradykinin-mediated relaxation in contrast to healthy pregnant microparticles (P <.005). CONCLUSION Preeclamptic microparticles, but not healthy pregnant microparticles cause endothelial dysfunction in isolated myometrial arteries from healthy pregnant women after overnight incubation, whereas other preeclamptic plasma constituents protect the endothelium from this effect.

Flow-Dependent Remodeling of Small Arteries in Mice Deficient for Tissue-Type Transglutaminase: Possible Compensation by Macrophage-Derived Factor XIII

Chronic changes in blood flow induce an adaptation of vascular caliber. Thus, arteries show inward remodeling after a reduction in blood flow. We hypothesized that this remodeling depends on the crosslinking enzyme tissue-type transglutaminase (tTG). Flow-dependent remodeling was studied in wild-type (WT) and tTG-null mice using a surgically imposed change in blood flow in small mesenteric arteries. WT mice showed inward remodeling after 2 days of low blood flow, which was absent in arteries from tTG-null mice. Yet, after continued low blood flow for 7 days, inward remodeling was similar in arteries from WT and tTG-null mice. Studying the alternative pathways of remodeling, we identified a relatively high expression of the plasma transglutaminase factor XIII in arteries of WT and tTG-null mice. In addition, vessels from both WT and tTG-null mice showed the presence of transglutaminase-specific crosslinks. An accumulation of adventitial monocytes/macrophages was found in vessels exposed to low blood flow in tTG-null mice. Because monocytes/macrophages may represent a source of factor XIII, tTG-null mice were treated with liposome-encapsulated clodronate. Elimination of monocytes/macrophages with liposome-encapsulated clodronate reduced both the expression of factor XIII and inward remodeling in tTG-null mice. In conclusion, tTG plays an important role in the inward remodeling of small arteries associated with decreased blood flow. Adventitial monocytes/macrophages are a source of factor XIII in tTG-null mice and contribute to an alternative, delayed mechanism of inward remodeling when tTG is absent.

The influence of aging and aortic stiffness on permanent dilation and breaking stress of the thoracic descending aorta

OBJECTIVE To assess the influence of aging and aortic stiffness on the extent of irreversible deformation and breaking stress of the human thoracic aorta. METHODS From 14 human heart valve donors without aortic disease (mean age 35 years, range 8-59 years), 14 intact segments of the thoracic descending aorta were studied within 48 h after cardiac arrest. In an experimental setup, the segments were submitted to increasing hydrostatic pressure loads, both statically and dynamically, while radius and wall thickness were monitored echocardiographically. Pressure-radius curves were constructed. Radius and wall thickness were determined at a pressure of 100 mmHg. Radius at elastin resting length and collagen recruitment pressure (Pcol, mmHg) were derived from the pressure-radius relationship and stress-strain curves were constructed to yield Youngs moduli of elastin and collagen. Distensibility (D, mmHg-1) was determined while loading the segment with a sinusoidal pressure wave of 120/50 mmHg at both 0.5 and 1 Hz. Subsequently increasing static pressure loads of 400, 800, 1200 and 1600 mmHg were applied. After each pressure load, the increase in aortic radius at a pressure of 100 mmHg (Rinc) was determined. The experiment continued until rupture occurred and breaking stress (sigma break, N m-2) was calculated, donor age and aortic stiffness were correlated with Rinc and sigma break of the aortic segments. RESULTS Mean breaking stress of the 14 segments was 2.7 x 10(6) N m-2. Breaking stress was negatively correlated with age (r2 = 0.66) and positively with D (r2 = 0.44) and with Pcol (r2 = 0.18). Seven segments survived a pressure load of 800 mmHg, in these vessels, the extent of irreversible dilation was positively correlated with age (r2 = 0.42) and negatively with D (r2 = 0.40) and Pcol (r2 = 0.40). CONCLUSION Permanent deformation and rupture of the human thoracic aorta following pressure overload are influenced by age, distensibility and collagen recruitment pressure.

Myogenic Activation and Calcium Sensitivity of Cannulated Rat Mesenteric Small Arteries

Pressure-induced activation of vascular smooth muscle may involve electromechanical as well as nonelectromechanical coupling mechanisms. We compared calcium-tone relations of cannulated rat mesenteric small arteries during pressure-induced activation, depolarization (16 to 46 mmol/L K+), and alpha1-adrenergic stimulation (1 micromol/L phenylephrine). The intracellular calcium concentration was expressed as the fura-2 ratio, normalized to the maximal and minimal ratios. In order to compare activation levels at various pressures, tone was expressed as the ratio of active wall tension to the maximal active tension. The passive and maximal active pressure-diameter relations needed for the calculation of tone were determined in a separate set of experiments, using isometric loading of cannulated vessels. Pressure steps from 20 to 60 and then to 100 mm Hg caused a modest rise of calcium. Nifedipine (1 micromol/L) blocked both the calcium rise and the resulting myogenic responses. Electromechanical coupling could not fully account for the myogenic response: the calcium sensitivity, defined as the slope of the calcium-tone relation, was five times higher during pressure-induced activation compared with potassium stimulation and twice as high as the sensitivity during alpha1-adrenergic stimulation. We therefore conclude that the myogenic response involves a small but necessary rise in calcium due to influx through L-type calcium channels, as well as a nonelectromechanical coupling mechanism that greatly enhances the calcium sensitivity of the contractile machinery.