Two-photon microscopy for imaging of the (atherosclerotic) vascular wall: a proof of concept study

BACKGROUND: Understanding atherogenesis will benefit significantly from simultaneous imaging, both ex vivo and in vivo, of structural and functional information at the (sub)cellular level within intact arteries. Due to limited penetration depth and loss of resolution with depth, intravital and confocal fluorescence microscopy are not suitable to study (sub)cellular details in arteries with wall thicknesses above 50 microm. METHODS: Using two-photon laser scanning microscopy (TPLSM), which combines 3D resolution and large penetration depth, we imaged mouse carotid arteries. RESULTS: In thin slices, (sub)cellular structures identified using histochemical techniques could also be identified using TPLSM. Ex vivo, structural experiments on intact atherosclerotic arteries of Apo-E(-/-) mice demonstrated that in contrast to confocal or wide-field microscopy, TPLSM can be used to visualize (sub) cellular structural details of atherosclerotic plaques. In vivo, pilot experiments were carried out on healthy arteries of wild-type C57BL6 and atherosclerotic arteries of Apo-E(-/-) mice. As an example of functional measurements, we visualized fluorescently labeled leukocytes in vivo in the lumen. Additionally, detailed morphological information of vessel wall and atherosclerotic plaque was obtained after topical staining. CONCLUSIONS: Thus, TPLSM potentially allows combined functional and structural studies and can therefore be eminently suitable for investigating structure-function relationships at the cellular level in atherogenesis in the mouse.     

Ultrastructural changes in mesenteric arteries from spontaneously hypertensive rats. A morphometric study

Morphometric measurements at the electron microscope level were carried out on three categories of mesenteric arteries representing elastic (superior mesenteric), muscular and arteriolar vessels, from 10- to 12-week-old spontaneously hypertensive rats (SHR) and age-matched Wistar-Kyoto normotensive rats (WKY). Changes were observed only in muscular and arteriolar vessels of SHR, mainly as thickening of the vessel wall due to hypertrophy of the media. In muscular arteries, hypertrophy of the endothelial cells, widening of the subendothelial space, increased volume of the internal elastic lamina (IEL), and both hyperplasia and hypertrophy of the smooth muscle cells (SMC) in the media contributed to the wall thickening. In arteriolar vessels, increase in the subendothelial space and IEL, and hyperplasia of the SMC in the media were involved in the increased thickness of the vessel wall. There was no difference in the collagen content in all vessels, but elastin was increased in the muscular and arteriolar vessels of SHR. Nerve density was also increased in arteriolar vessels of SHR. These changes, especially the increase of SMC in muscular and arteriolar vessels, may be related to the elevated blood pressure in SHR.     

Mechanisms underlying hypertrophic remodeling and increased stiffness of mesenteric resistance arteries from aged rats

The mechanisms associated with structural and mechanical alterations of mesenteric resistance arteries from aged rats were investigated by using pressure myography, confocal microscopy, immunofluorescence, and picrosirius red staining. Arteries from old rats showed: (i) increased wall and media thickness, greater number of smooth muscle cell (SMC) layers but decreased density of SMC; (ii) increased number of adventitial cells; (iii) hypertrophy of nuclei of SMC and endothelial cells; (iv) increased stiffness associated with increased total collagen content and collagen I/III deposition in the media; and (v) similar content but changes in elastin structure in the internal elastic lamina. Hypertrophic outward remodeling in aged rat resistance arteries involve adventitial cells hyperplasia, reorganization of the same number of hypertrophied SMC in more SMC layers leading to thickened media and endothelial cell hypertrophy. Fibrosis associated with collagen deposition and changes in elastin structure might be responsible for the increased stiffness of resistance arteries from aged rats.     

In vitro high resolution intravascular imaging in muscular and elastic arteries.

High resolution (125-microns lateral, 55-microns axial) images of 16 muscular (femoral) and 15 elastic (common carotid) human arteries were made in vitro with use of a prototype 45-MHz intravascular imaging system. Four distinct regions of scattering, excluding plaque, were identified in the ultrasound images corresponding histologically to the adventitia, media, thickened intima and elastic laminae, both internal and external. Arterial samples imaged under pressure and in a collapsed state underwent dimensional changes but exhibited similar levels of backscatter amplitude. All the elastic arteries displayed a prominent echogenic media, whereas all the muscular arteries displayed an echolucent media. Scattering from the internal elastic lamina in muscular arteries provided an excellent landmark for defining the location and extent of intimal thickening or plaque. In elastic arteries the internal elastic lamina could not be distinguished from the echogenic media; consequently, the boundary between the media and intimal layer was indistinct. Differences in the relative concentration and organization of collagen and elastin were found to provide a consistent explanation for the differences in scattering that were observed between individual layers within an artery as well as between muscular and elastic arteries.     

Morphometric study of mesenteric arteries from genetically hypertensive Dahl strain rats

Morphometric measurements on different arteries at the light-microscopic level and ultrastructural studies of the mesenteric arteries were carried out in salt-sensitive (DS) and salt-resistant (DR) Dahl rats given a high-salt (8%) or low-salt (0.4%) diet for 6-7 weeks. Hypertension was produced in DS rats given high-salt diet (DS-H), while only moderate hypertension was produced in DS rats given low-salt diet (DS-L). Blood pressure in DR rats given high salt (DR-H) and low salt (DR-L), however, was normal. Cross-sectional area of the media was increased significantly in the superior mesenteric artery (an elastic artery), large mesenteric arteries (muscular arteries) and small mesenteric arteries (small muscular arteries or arterioles) from DS-H rats. In all the vessel types, this increase was positively correlated with the increase in blood pressure. In the superior mesenteric artery, medial wall increase was probably due to an increase in intercellular space, and/or hypertrophy of the smooth muscle cells. Similarly, increase in the media of small mesenteric arteries was probably due to hypertrophy of the smooth muscle cells. In contrast, increase in the media of large mesenteric arteries was related to hyperplasia of the smooth muscle cells. Damage to endothelial cells was noted in the 3 vessel types from DS-H. Intimal lesions composed of myointimal cells were found in the superior mesenteric arteries of all the rat groups. Our results showed that the incidence of these lesion formations was higher in the following order: DS-H greater than DS-L greater than DR-H greater than DR-L, suggesting that the degree of hypertension (DS vs. DR rats) and the amount of salt in the diet (DR-H vs. DR-L) may be some of the factors contributing to the development of these lesions. We conclude that hyperreactivity of the arteries due to increase in medial smooth muscle mass (e.g. muscular arteries), and/or probably impaired relaxation capability of the arteries in the DS-H rats due to endothelial cell damage, may contribute to the elevation of BP in the Dahl model of genetic hypertension.     

Relation of intra-acinar arteries and their regression with changes of pulmonary artery pressure and blood gas values

Pulmonary hypertension (PAH) induced by monocrotaline in thirteen rhesus monkeys, seven of them treated by hydralazine to reduce PAH. In hypertensive animals the PAP and PaCO2 increased, the PaO2 decreased. The structural changes in intra-acinar artery manifested mainly by an increase in number of muscular artery resulting from muscularization of precursor cells (pericytes and intermediate cells) located within partially muscular and nonmuscular arterial wall to smooth muscle cell, and by the medial wall thickened due to hypertrophy and hyperplasia of smooth muscle cells as well as accumulation of a large amount of collagen, especially the type 1 collagen. In treated animals the PAPm dropped concomitantly with blood gas values reversing to normal level. The remodelled arteries showed clearly structural regressions: The medial wall thickness was decreased in which the hypertrophied smooth muscles became slender or disappeared, the amount of extracellular matrix, especially the volume density of collagen, decreased. Most of muscularized arteries reversed to nonmuscular arteries leading to an increase in number of nonmuscular artery and a decrease in number of muscular artery. It is concluded that the intra-acinar artery remodelling and their regression closely correlated with the changes of PAP and blood gas values.     

Conducting pulmonary arteries: structural adaptation to extrauterine life in the pig

A quantitative light microscopical and ultrastructural study of the elastic and large muscular pulmonary arteries of 30 Large White pigs aged from birth to 6 months yielded light microscopical measurements on 120 arteries and 62,560 ultrastructural assessments, which composed a computerised database. After birth mean arterial medial thickness and mean smooth muscle cell diameter decreased during the first 4 days (p less than 0.01). Mean interlamellar distance decreased, reaching its nadir at 1-3 weeks (p less than 0.01). All these features increased between 3 weeks and adulthood (p less than 0.01). In the smooth muscle cells synthetic rather than contractile organelles were dominant during the first 3 weeks. Between 3 weeks and adulthood, however, smooth muscle cell myofilament volume density increased (p less than 0.0001). At all ages the smooth muscle cells of the outer lamellar units of elastic arteries had a greater myofilament volume density than those of adluminal units (p less than 0.0001). The amount of connective tissue increased between 3 weeks and adulthood, collagen and basement membrane increasing preferentially (p less than 0.0001, p less than 0.05, respectively). Thus the conducting pulmonary arteries, like the peripheral resistance arteries, adapt structurally to extrauterine life. Remodelling occurred rapidly after birth, and then gradually during growth, as connective tissue was deposited and smooth muscle cells matured.     

Hyperreactive arterial endothelial cells in atherogenesis and cyclic AMP phosphodiesterase inhibitor in prevention and treatment of atherosclerotic disorders.

The hyperreactive arterial endothelial cells have been introduced in this paper. They are characterized by their ability to transport particles too large for the small holes of the internal elastic lamina locating underneath the endothelial cells, such as carbon particles with the similar size of LDL, floating beta-lipoprotein, Lp(a) and especially of VLDL, into the subendothelial space from the blood stream by their abnormally strong contracting and phagocytosis-like activity. Large particles such as carbon particles with a size of 200 to 700 angstrom are too large to penetrate further through holes of the internal elastic lamina from the subendothelial space to muscular layers of the arterial wall, resulting in stagnating for a long time in the subendothelial space, thus showing the atherogenic property of the hyperreactive arterial endothelial cells. Such endothelial cells appear spotty and streaky in the localized endothelial lining of predominantly susceptible parts to atherosclerosis in susceptible animal species such as rabbit, chicken, and rhesus monkey especially densely in their atheromatous lesions, but do not generally appear in non-susceptible animals to atherosclerosis like rate and dog. They are extremely few in infant rabbit, but increase by age.They appear in hypertensive rat, showing a characteristic distribution even in small groups of arteriessuch as the circle of Willis. Cyclic AMP, and especially dibutyryl cyclic AMP, exhibited an inhibitory effect on the hyperreactivity of those cells. Cyclic AMP phosphodiesterase inhibitors, EG467 and eg626, exhibited a powerful inhibitory effect on the contracting and phagocytosis-like activity of those cells, as in the case of pyridinolcarbamate, which enhances enzymes to produce ATP and inhibits slightly cyclic AMP phosphodiesterase, although its inhibitory effect on cyclic AMP phosphodiesterase is weaker than that of EF467 and EG626. The usefulness of the inhibitors on cyclic AMP phospodiesterase of arterial endothelial cells and platelets and on that of brain, such as EG467 AND EG626, has been suggested in the treatment of atherosclerotic disorders, especially of cerebral atherosclerosis. Some of the hitherto desperate mental disability of the aged seem to be a promising target for treatment with cyclic AMP phosphodiesterase inhibitors.     

缬沙坦涂层支架对支架术后再狭窄中胶原沉积的影响

目的评价缬沙坦涂层支架对支架置入后新生内膜中胶原沉积的影响及其预防支架内再狭窄的可行性。方法18只新西兰大白兔分为裸支架组,载体涂层支架组,缬沙坦涂层支架组3组。采用多层涂布技术制备缬沙坦涂层支架和载体涂层支架。分别将裸支架、载体涂层支架及缬沙坦涂层支架置入兔腹主动脉。术前、术后及支架置入3个月后分别行定量腹主动脉造影(QCA)测量血管直径。3个月后处死实验兔,分别测定3组支架血管段的管腔面积,内外弹力膜围绕面积,新生内膜面积及最大内膜厚度并做比较。将支架血管段进行MASSON染色,观察胶原沉积情况,天狼星红-苦味酸染色进一步观察胶原的类型。结果裸支架组(n=8),载体涂层支架组(n=8),缬沙坦涂层支架组(n=10),QCA测量的术前、术后及3个月后腹主动脉直径相似。缬沙坦组管腔面积最大,新生内膜面积最小,裸支架组、载体涂层支架组、缬沙坦涂层支架组平均管腔面积分别为(4345548±125822)μm2,(4302061±167952)μm2,(5016269±207934)μm2,平均新生内膜面积分别为(1119635±163503)μm2,(1135636±136555)μm2,(441577±74099)μm2,平均最大内膜厚度分别为(240±30)μm,(192±21)μm,(116±12)μm。MASSON染色可见新生内膜中主要是胶原沉积,天狼星红-苦味酸染色发现胶原沉积主要为Ⅲ型胶原,间或有Ⅰ型胶原。结论缬沙坦涂层支架主要是通过抑制胶原沉积抑制支架置入后血管内膜增生发挥其防治支架内再狭窄作用的。     

20例动脉粥样硬化性患者临床治疗的探讨

动脉粥样硬化是动脉硬化中常见、重要的类型。本病主要累及大型及中型的肌弹力型动脉,以主动脉,冠状动脉、脑动脉和四肢大动脉为多见,常导致管腔闭塞或管壁破裂出血等严重后果。其特点是动脉发生了非炎症性、退行性和增生性的病变,导致管壁增厚变硬,失去弹性和管腔缩,动脉粥样硬化的病理变化主要累及循环系统的大型弹力型动脉如主动脉、肺动脉、椎动脉等中型肌弹力型动脉如冠状动脉和脑动脉。受累动脉的病变从内膜开始,先后有多种病变同时存在,包括局部有脂质和复合糖类积聚、出血和血栓形成、纤维组织增生和钙质沉着,并有动脉中层的逐渐退化和钙化。现代细胞分子生物学技术显示,动脉粥样硬化病变均具有平滑肌细胞增生、大量胶原纤维、弹力纤维和蛋白多糖等结缔组织基质形成以及细胞内外脂质积聚的特点,由于在动脉内膜积聚的脂质外观呈现黄色粥样,因此称为动脉粥样硬化。     

Structure and mechanical properties of resistance arteries in hypertension: role of adhesion molecules and extracellular matrix determinants

Abnormalities of resistance arteries may play a role in the pathogenesis and pathophysiology of hypertension in experimental animals and humans. Vessels that, when relaxed, measure <400 microm in lumen diameter act as the major site of vascular resistance and include a network of small arteries (lumen approximately 100 to 400 microm) and arterioles (<100 microm). Because increased peripheral resistance is generated by a narrowed lumen diameter, significant effort has been focused on determining the mechanisms that reduce lumen size. Three important vascular components are clearly involved, including alterations of vascular structure, mechanics (stiffness), and function. Structural abnormalities comprise a reduced lumen diameter and thickening of the vascular media, resulting in an increased media-lumen ratio. Changes in the mechanical properties of an artery, particularly increased stiffness, may also result in a reduced lumen diameter. These vascular abnormalities may be caused or influenced by the expression and/or topographic localization of extracellular matrix components, such as collagen and elastin, and by changes in cell-extracellular fibrillar attachment sites, such as adhesion molecules like integrins. This article discusses the abnormalities of resistance arteries in hypertension and reviews the evidence suggesting an important role for adhesive and extracellular matrix determinants.     

Frequency of neuromuscular junctions on arteries of different dimensions in the rabbit, guinea pig and rat

The medio-adventitial border of a variety of perfusion-fixed arteries of young adult rabbits, guinea pigs and rats has been studied in the electron microscope. The arterial media in the different vessels ranged from 2 to 25 cells thick. Neuromuscular junctions, defined as axon varicosities containing synaptic vesicles closely apposed to the outer surface of smooth muscle cells, with only a single layer of basal lamina intervening between axon and muscle membranes, were identified in all three species. Junctions were found in most vessels less than 1 mm in diameter with a frequency ranging from 8,000-150,000 junctions per square millimeter of smooth muscle surface, the number generally increasing with decreasing arterial diameter. These small arteries were mostly, but not exclusively, muscular rather than elastic. In large arteries, such as abdominal aortae and some elastic arteries lying close to the heart (e.g. the carotid), no junctions were found. In a few vessels, such as guinea pig basilar (muscular) and rat and guinea pig superior mesenteric (elastic) arteries, junctions were found infrequently (1,000-4,000/mm2). The data indicate that all muscular arteries in rats and guinea pigs, and most muscular arteries in rabbits, receive innervation in the form of sympathetic neuromuscular junctions. Whilst a few elastic vessels are sparsely innervated with junctions, some are surrounded by axon bundles containing vesicle-filled varicosities. The physiological significance of the latter is obscure.     

Changes of elastic properties of magistral arteries in patients with ischemic heart disease during treatment with atorvastatin and simvastatin

AIM: To elucidate effect of atorvastatin and simvastatin on elastic properties and reactivity of large arteries in patients with ischemic heart disease. MATERIAL: Patients (n=38) with ischemic heart disease receiving atorvastatin (10 mg/day, n=20) and simvastatin (20 mg/day, n=18). METHODS: Elasticity of arteries was assessed by pulse wave propagation velocity measured by Coloson computer system. Test with reactive hyperemia was used for assessment of large muscular type vessels reactivity in response to blood flow augmentation was assessed by. RESULTS: Treatment with statins for 6 months was associated with positive changes of elasticity of arteries of elastic type (-10.05% and -4.66% in atorvastatin and simvastatin treated patients, respectively) and endothelium dependent reactivity of arteries of muscular type (-16.01%, p<0.001 and -11.1% p<0.001 compared with initial values in atorvastatin and simvastatin treated patients, respectively). CONCLUSION: Therapy with statins positively affects parameters of elasticity and reactivity of large arteries.     

Vascular changes at the prehypertensive phase in the mesenteric arteries from spontaneously hypertensive rats

Morphometric measurements of three categories of mesenteric vessels (representing elastic, muscular and arteriolar vessels) from prehypertensive spontaneously hypertensive rats (SHR) and age-matched Wistar-Kyoto rats (WKY) were carried out at the light and electron microscope levels. Structural alterations of the blood vessels were already present in the SHR, even though the blood pressure was not yet elevated as compared with age-matched WKY. No change was found in the elastic vessels (superior mesenteric artery). Among the muscular arteries (i.e. large mesenteric arteries), the increase in vessel wall cross-sectional area was due to the increase in the intima, media and adventitia. Increase in media was due to hyperplasia of the smooth muscle cells. The smooth muscle cells were not hypertrophied. Nerve density was also higher in the large mesenteric arteries of SHR. In the arteriolar vessels (i.e. small mesenteric arteries), wall to lumen ratio, as well as media to lumen ratio, were increased in the SHR. The number of smooth muscle cell layers was also increased. In all these vessel types, the cross-sectional area of the lumen under maximal relaxation was similar between SHR and WKY, except in small mesenteric arteries where the lumen was smaller in the SHR. Our results suggest that structural alteration of the blood vessels at the prehypertensive phase may be one of the contributing factors leading to the development of hypertension in the SHR.     

Adenovirus-mediated gene transfer of superoxide dismutase and catalase decreases restenosis after balloon angioplasty

BACKGROUND: Reactive oxygen species (ROS) production increases after injury and potentially contributes to restenosis after angioplasty. We therefore evaluated the effect of adenovirus-mediated gene transfer (Ad) of superoxide dismutase (SOD) and catalase (CAT) on ROS production and restenosis after balloon angioplasty. METHODS: O(2)(-) and H(2)O(2 )production was quantified in cultured cells after incubation with either LPS or CuSO(4). Angioplasty and gene transfer were performed in rabbit atherosclerotic iliac arteries. One artery was injected with AdSOD and AdCAT, while the contralateral artery was injected with an adenovirus carrying no transgene, and served as control. RESULTS: ROS production was significantly decreased after adenovirus-mediated gene transfer of SOD and CAT as compared with control. Treated arteries showed less restenosis (32 +/- 27 vs. 63 +/- 19%, p = 0.003) and less constrictive remodeling (1.2 +/- 0.3 vs. 0.9 +/- 0.2, p = 0.02) than control arteries. Arteries injected with AdSOD and AdCAT showed better vasoreactivity to acetylcholine (11 +/- 4 vs. -1 +/- 6%, p < 0.05), lower collagen density (43 +/- 16 vs. 53 +/- 23%, p = 0.03), and lower inflammatory cell infiltration (22 +/- 6 vs. 36 +/- 11%, p = 0.04) than control arteries. CONCLUSIONS: Our data suggest that adenovirus-mediated gene transfer of SOD and CAT reduced oxidative stress, restenosis, collagen accumulation, and inflammation and improved endothelial function after angioplasty.     

Characterization of pulmonary vascular remodelling in smokers and patients with mild COPD.

Intimal enlargement of pulmonary arteries is an early change in chronic obstructive pulmonary disease (COPD). The cellular and extracellular components that are involved in this enlargement are unknown. The present study was designed to characterize the structural changes occurring in pulmonary muscular arteries in the initial disease stages. Lung specimens from patients with moderate COPD (n=8; forced expiratory volume in one second (FEV1), 66 +/- 10% predicted) and smokers without airflow obstruction (n=7; FEV1, 86 +/- 6% pred), were investigated by histochemistry to characterize extracellular matrix proteins and by immunohistochemistry to identify intrinsic cells of the vascular wall. In both COPD patients and smokers, the majority of cells present in the enlarged intimas were stained by specific smooth muscle cell (SMC) markers. No staining with endothelial or fibroblast markers was shown. A proportion of SMCs did not stain with desmin, suggesting cellular heterogeneity in this population. Elastin was the most abundant extracellular matrix protein and collagen was seen in a lower proportion. The amount of collagen was related to the intimal thickness (p<0.001). The findings demonstrated smooth muscle cell proliferation, as well as elastin and collagen deposition, in the thickened intimas of pulmonary arteries in moderate chronic obstructive pulmonary disease patients and smokers, suggesting that these abnormalities may originate at an early stage in cigarette smoke-induced respiratory disease.     

Mechanical and functional predictive factors for restenosis and arterial remodeling after experimental angioplasty

Arterial remodelling plays an important part in post-angioplasty restenosis but the physiopathology of this process is not fully understood. Abundant collagen synthesis and endothelial dysfunction have been demonstrated after angioplasty, but their role in restenosis and remodelling has not been studied. The aim of this study was therefore to assess endothelial function and collagen with respect to the severity of restenosis and the type of arterial remodelling. Atherosclerosis was induced by an association of endothelial abrasion and a high cholesterol diet in the femoral arteries of 22 white New Zealand rabbits. Four weeks later, angioplasty was performed. The acetylcholine endothelium-dependant vasomotricity (expressed as % inhibition of contraction to phenylephrine), collagen and morphology were assessed 28 days after angioplasty. The change in acetylcholine endothelium-dependant vasomotricity was greater in severe restenosis (r = 0.61, p = 0.02). Endothelium-dependant relaxation was not significantly altered when remodelling was expansive and very abnormal when it was constrictive (35.5 +/- 13.0 vs 3.7 +/- 7.9%; p = 0.04). Restenosis was associated with an increase in collagen (r = 0.69, p = 0.004). The density of collagen was significantly higher in constrictive remodelling than in expansive remodelling (34.5 +/- 4.5 vs 18.2 +/- 4.7%; p = 0.03). Endothelial dysfunction and collagen accumulation are correlated with the severity of restenosis and with constrictive remodelling after angioplasty in an experimental model.     

Effects of antihypertensive therapy on hypertensive vascular disease

Hypertension is associated with alterations in the structure, function, and mechanical properties of large and small arteries. Changes in the endothelium, smooth muscle cell, extracellular matrix, and possibly the adventitia, contribute to complications of hypertension. In large arteries, vascular hypertrophy is found, often with increased stiffness of media components. In small arteries, particularly in mild hypertension, rearrangement of smooth muscle cells around a smaller lumen without changes in media volume (eutrophic remodeling) occurs; in more severe hypertension, hypertrophic remodeling with increased vascular stiffness can be found. Vascular remodeling is accompanied by an increase in the extracellular matrix, particularly collagen deposition. Recent studies have demonstrated that vascular remodeling and endothelial dysfunction of small and large vessels may be normalized by treatment with some antihypertensive agents (angiotensin converting enzyme inhibitors, angiotensin AT1 receptor antagonists, and long-acting calcium channel blockers). Angiotensin converting enzyme inhibitors have now been shown to improve outcomes in hypertensive patients, an effect that may in part be related to the vascular protective effects reviewed here.     

Vasopeptidase inhibition has potent effects on blood pressure and resistance arteries in stroke-prone spontaneously hypertensive rats

The antihypertensive agent omapatrilat represents a novel approach to antihypertensive therapy, namely vasopeptidase inhibition. Omapatrilat (BMS-186716) concomitantly inhibits neutral endopeptidase and angiotensin-converting enzyme, leading to protection from degradation of natriuretic and other hypotensive peptides in addition to interruption of the renin-angiotensin system. Although the potency of omapatrilat on reduction of blood pressure has been reported, its effects on resistance artery structure and function were unknown. We tested omapatrilat in stroke-prone spontaneously hypertensive rats (SHRSP), a malignant model of hypertension, with the hypothesis that it would improve the structure and endothelial function of mesenteric resistance arteries. Ten-week-old SHRSP were treated orally for 10 weeks with omapatrilat (40 mg/kg per day). Mesenteric arteries (lumen <300 microm) were studied on a pressurized myograph. After 10 weeks, untreated SHRSP had a systolic blood pressure of 230+/-2 mm Hg that was significantly reduced (P<0.05) by omapatrilat (145+/-3 mm Hg). Omapatrilat treatment improved endothelium-dependent relaxation of resistance arteries as elicited by acetylcholine (10(-5) mol/L) but had no significant effect on endothelium-independent relaxation produced by a nitric oxide donor (sodium nitroprusside). This suggested that there existed endothelial dysfunction in SHRSP that was corrected by vasopeptidase inhibition, probably in part caused by the potent blood pressure-lowering effect of omapatrilat. Media width and media/lumen ratio were significantly decreased (P<0.05) by omapatrilat, and a trend (P=0.07) to increase lumen diameter was observed. Vascular stiffness (slope of the elastic modulus versus stress curve) was unaltered by omapatrilat. In conclusion, omapatrilat, acting as a potent antihypertensive agent, may improve structure and endothelial function of resistance arteries in SHRSP, a severe form of genetic hypertension.     

Histomorphometric,biochemical and ultrastructural changes in the aorta of salt-loaded stroke-prone spontaneously hypertensive rats fed a Japanese-style diet

BACKGROUND AND AIM: It is demonstrated that dietary habits play a role in cardiovascular diseases. In stroke-prone spontaneously hypertensive rats (SHRsp), concomitant salt loading and a Japanese-style diet greatly accelerate hypertension and the appearance of cerebrovascular lesions by directly damaging arterial vessels. A number of studies have characterised medium and small vessel lesions in SHRsp, but little attention has been paid to the changes in the wall structure of large arteries induced by exposure to a salt-enriched diet. The aim of this study was to investigate the effects of a Japanese-style diet and salt loading on the thoracic aorta. METHODS AND RESULTS: Two-month-old SHRsp were kept on a Japanese-style diet with 1% sodium chloride solution replacing tap water. Two months later, they were sacrificed and compared with age-matched or two-month-old control SHRsp kept on a standard diet and tap water in terms of the histomorphometry, ultrastructure and biochemical composition of the thoracic aorta. The vessel was consistently thicker in the four-month-old SHRsp (+20%, p < 0.05 vs two-month-old rats) regardless of diet. The salt-loaded SHRsp showed a significant reduction in elastic fibre density (-20%, p < 0.05 vs two-month-old rats) and an increase in the other matrix components (%), whereas the four-month-old controls showed preserved elastic fibres and a significant increase in the other matrix components (+65%, p < 0.05 vs two-month-old rats). There was a considerable increase in the amounts of 4-OH-proline (+147%), 5-OH-lysine (+174%) and desmosines (+360%) in the four-month-old controls vs their two-month-old counterparts (p < 0.01), but not in the salt-loaded animals. Ultrastructural analysis revealed clear damage and accelerated aging in the thoracic aorta of the salt-loaded SHRsp. CONCLUSIONS: Salt loading and a Japanese-style diet destabilize thoracic aorta architecture in SHRsp after two months of treatment.