Robert M.K.W. Lee

Modulation of vascular function by perivascular adipose tissue: the role of endothelium and hydrogen peroxide

Perivascular adipose tissue (PVAT) attenuates vascular contraction, but the mechanisms remain largely unknown. The possible involvement of endothelium (E) and hydrogen peroxide (H2O2) was investigated.

Morphology of cerebral arteries

A comparison of the major cerebral arteries between humans and rats shows many similarities, including anomalies in their general organization, the structure of these vessels at the light and electron microscope levels and their morphological changes associated with cerebral vascular diseases. The general organization of the major cerebral arteries shows the following main differences between humans and rats. In rats, the internal carotid arteries have become an integral part of the circle of Willis. In the anterior cerebral arteries, a common variation in humans is the underdevelopment of one of the two arteries, whereas in rats, buttonhole-like structures are common in one or both arteries. The anterior communicating artery present in humans is absent in rats. The olfactory artery is prominent in rats, but absent in humans. The posterior communicating artery in humans is the most variable component of the circle of Willis, being asymmetric in its origin, diameters and branches. Similarly, the posterior cerebral arteries in rats often exhibit asymmetrical origin from the basilar artery. There was some confusion in the literature regarding the name of the posterior cerebral arteries in rats, but this was caused mainly by misquotations and incorrect interpretations of the papers. In humans, most aneurysms occur in the anterior half of the circle of Willis, and the incidence is higher in females than males; the middle cerebral artery is most often the one to become occluded, and the vertebral arteries are common sites for thrombosis. The various channels that constitute collateral circulation in humans provide a margin of safety, so that in case of cerebral occlusion due to thrombosis, atherosclerosis, or vasospasm related to hemorrhage, blood supply to the affected area can be maintained through these collaterals. Collateral circulation is also present in rats. However, in rats, information on the presence of various types of aneurysms, their location and frequency in normal and experimental models of hypertension and stroke is still lacking. Cerebral arteries from humans and rats are characterized by the absence of external elastic lamina, as compared with systemic arteries. A type of multipolar cell resembling the interstitial cell of Cajal is present in the cerebral arteries of humans. Its function is unknown. Earlier reports of cerebral valves have been shown to represent intimal cushions near the branching points of the cerebral arteries. Intravascular bridges present in human cerebral arteries, have not been reported in rats. Finally, the presence of vascular remodeling, as found in the cerebral arterioles of hypertensive rats, remains to be established in the cerebral arterioles of human hypertensives.(ABSTRACT TRUNCATED AT 400 WORDS)

Structural and functional consequence of neonatal sympathectomy on the blood vessels of spontaneously hypertensive rats.

Neonatal sympathectomy of spontaneously hypertensive rats (SHR) and control Wistar-Kyoto rats (WKY) was performed by a combined treatment with antiserum to nerve growth factor and guanethidine during the first 4 weeks after birth. The development of hypertension was completely prevented in the treated SHR: at 28 to 30 weeks of age, systolic blood pressure of treated SHR was 139 +/- 2 mm Hg as compared with 195 +/- 8 mm Hg in untreated SHR. The extent of sympathectomy was verified by histofluorescence. Fluorescence histochemistry for catecholamine-containing nerves showed a complete absence of adrenergic nerves in the mesenteric arteries of treated rats. A supersensitivity to norepinephrine was exhibited by mesenteric arteries, anococcygeus muscle, and tail arteries from the treated SHR and WKY. In the mesenteric vascular bed, maximal response to norepinephrine was significantly reduced by sympathectomy. Sympathectomy also abolished the responses (e.g., generation of excitatory junctional potentials) of tail arteries to electrical stimulation of perivascular nerves. Morphometric measurements of three categories of mesenteric arteries showed that sympathectomy had no effect on the hypertrophic change of smooth muscle cells in the conducting vessels, but it prevented the hyperplastic changes of the muscle cells from reactive, muscular arteries and small resistance vessels. These results suggest that one of the primary roles of the overactive sympathetic nervous system in the development of hypertension in SHR is manifested through its trophic effect on the arteries of SHR. This trophic effect appears to cause a hyperplastic change in the smooth muscle cells in the reactive and resistance vessels, thereby contributing to the development of hypertension in older SHR.

Morphometric study of structural changes in the mesenteric blood vessels of spontaneously hypertensive rats.

Structural changes of three categories of mesenteric arteries (representing elastic, muscular and arteriolar vessels) from 10- to 12-week-old and 28-week-old spontaneously hypertensive rats (SHR) were studied morphometrically at the light microscope level, and the results compared with age-matched Wistar-Kyoto normotensive rats. In 10- to 12-week-old SHR, hypertrophy of the vessel wall occurred only in the muscular and arteriolar vessels. At 28 weeks, further thickening of the vessel wall occurred in the muscular and arteriolar vessels, and the superior mesenteric artery (elastic vessel) was also thickened in the SHR. There was no evidence that the wall of the relaxed hypertrophied vessels encroached upon the lumen of the vessel. The structural basis for the increase in the vessel wall thickness varied with vessel type. In the superior mesenteric artery, increase in the media at 28 weeks of age would be consistent with hypertrophy of the smooth muscle cells. In the large muscular arteries, at 10-12 weeks of age, increase in medial mass occurred with increase in the number of the smooth muscle cell layers whereas at 28 weeks further increase in media could be due to hypertrophy of the smooth muscle cells. In the small arteriolar vessels, medial enlargement was due at all ages to an increase in the number of smooth muscle layers. Our results show that in the SHR hypertrophy of the media occurs not only in the small arteriolar vessels, but also in large elastic and muscular arteries.

Endothelium-dependent relaxation factor released by perivascular adipose tissue.

Objective Recent studies have demonstrated that perivascular adipose tissue (PVAT) releases vascular relaxation factor(s), but the identity of this relaxation factor remains unknown. Here, we examined if angiotensin 1-7 [Ang-(1-7)] is one of the relaxation factors released by PVAT. Method Morphological and functional methods were used to study aorta from adult Wistar rats. Results Immunohistochemical staining showed abundant presence of Ang-(1-7) in aortic PVAT. In vessels with PVAT removed but intact endothelium (PVAT − E+), contraction induced by phenylephrine was attenuated by preincubation with Ang-(1-7). PVAT − E+ vessels precontracted with phenylephrine showed a concentration-dependent relaxation response to Ang-(1-7), and this response was abolished by the removal of endothelium. Relaxation response induced by Ang-(1-7) was also prevented by Ang-(1-7) receptor (Mas) antagonist (A779), nitric oxide synthase inhibitor, and nitric oxide scavenger. Ang-(1-7) did not cause a relaxation response in aorta precontracted with KCl, and the relaxation response to Ang-(1-7) was also blocked by calcium-dependent potassium (KCa) channel blockers. Incubation of PVAT + E+ vessels with A779 or angiotensin-converting enzyme 2 inhibitor DX600 or angiotensin-converting enzyme inhibitor enalaprilat increased the contraction induced by phenylephrine. Transfer of donor solution incubated with PVAT + E+ vessel to recipient PVAT − E+ vessel caused a relaxation response. This relaxation response was abolished when donor vessels were incubated with DX600 or enalaprilat or when recipient vessels were incubated with A779. Conclusion Ang-(1-7) released by PVAT acts on the endothelium to cause the release of nitric oxide, and nitric oxide acts as a hyperpolarizing factor through KCa channels to cause relaxation of the blood vessel.

Prevention of hypertension and vascular changes by captopril treatment.

Treatment of female spontaneously hypertensive rats (SHR) and control Wistar-Kyoto (WKY) rats with captopril was carried out by the addition of the drug in the drinking water throughout pregnancy and lactation and after weaning. At 28 weeks of age, average systolic blood pressure of treated SHR was 113±3 mm Hg, which was below that of control SHR (188±3 mm Hg) and WKY rats (124±3 mm Hg). Body weight and heart rate of the SHR were not affected by the treatment Tissue level of catecholamines was increased by captopril treatment in the superior cervical ganglia but remained unchanged in the plasma, heart, mesenteric arteries, and the adrenal glands of both SHR and WKY rats. Left ventricular weight, wall thickness, and internal diameter of the left ventricle in the SHR were reduced by the treatment Morphometric measurements of the mesenteric arteries showed that vascular alterations present in the control SHR were prevented by the treatment In the superior mesenteric artery and large mesenteric artery, smaller lumen size at maximal relaxation found in the control SHR was normalized to the level of the WKY rats. Hypertrophy of the medial wall in the superior mesenteric, large and small mesenteric arteries, and an increase in the number of smooth muscle cell layers in the large mesenteric artery of the SHR were prevented by the treatment. Perfusion study of the mesenteric vascular bed showed that reactivity of these vessels to norepinephrine was reduced, and sensitivity to norepinephrine (as determined by the effective dose that causes 50% of maximal response) was increased in the SHR by captopril treatment Sensitivity of the tail artery in response to norepinephrine was not altered by the treatment We conclude that long-term treatment with captopril of SHR before and after birth prevented the development of hypertension, structural and functional alterations of the mesenteric arteries, and cardiac hypertrophy.

Mechanisms of hydrogen-peroxide-induced biphasic response in rat mesenteric artery

In phenylephrine (PHE) (1 μM)‐precontracted superior mesenteric arteries from adult rats, low concentration of hydrogen peroxide (H2O2, 10–100 μM) caused only contraction, while high concentration of H2O2 (0.3–1 mM) caused a biphasic response: a transient contraction followed by a relaxation response. Endothelium removal did not affect the biphasic response. 7,7‐Dimethyl‐(5Z,8Z)‐eicosadienoic acid, diclofenac, furegrelate, or SQ 29548 greatly inhibited the contraction but did not affect the relaxation. 17‐Octadecynoic acid, eicosatriynoic acid, ICI 198615, SQ 22536, or ODQ did not inhibit the biphasic response. KCl at 40 mM inhibited the relaxation response to H2O2 by 98±24%. 4‐Aminopyridine (4‐AP) inhibited while tetraethylammonium chloride (TEA), charybdotoxin, or glibenclamide attenuated the relaxation response. A combination of 4‐AP, TEA and glibenclamide mimicked the effects of 40 mM KCl. Iberiotoxin, apamin, or barium chloride did not inhibit the relaxation response. H2O2 at 1 mM hyperpolarized membrane potential and reversibly augmented K+ current in smooth muscle cells of mesenteric artery. These effects of H2O2 were attenuated significantly by 4‐AP. In summary, in PHE‐precontracted rat mesenteric artery: (1) the response to H2O2 shifted qualitatively from contraction to a biphasic response as H2O2 increased to 0.3 mM or higher; (2) the relaxation response is caused by the activation of K+ channels, with voltage‐dependent K+ channels playing a primary role; and the contraction is likely to be mediated by thromboxane A2; (3) the K+ channel activation by H2O2 is independent of phospholipase A2, cyclooxygenase, lipoxygenase, cytochrome P450 monooxygenase, adenylate or guanylate cyclase.

Hydrogen peroxide induces a greater contraction in mesenteric arteries of spontaneously hypertensive rats through thromboxane A2 production

Hydrogen peroxide (H2O2) caused a transient contraction in endothelium‐intact (E+) and ‐denuded (E−) mesenteric arteries (MA) from 8 – 10‐month‐old spontaneously hypertensive rats (SHR) and normotensive Wistar‐Kyoto rats (WKY) in a concentration‐dependent manner (10−5 M to 10−3 M). The contraction to H2O2 in MA (E+ or E−) was greater in SHR than in WKY. Removal of endothelium potentiated the contraction to H2O2 in WKY but not in SHR. Tachyphylaxis to H2O2 was less prominent in SHR than in WKY. The contraction of aorta to H2O2 (5×10−4 M), expressed as a percentage of 80 mM KCl‐induced contraction, was approximately half of that found in the MA. A greater contraction was found in E+ but not E− SHR aortic rings. The contraction of MA to H2O2 (5×10−4 M) was greatly inhibited by SQ 29548 and ICI 192605 (thromboxane A2 (TXA2)/prostaglandin H2 receptor antagonists), quinacrine (a phospholipase A2 (PLA2) inhibitor), indomethacin and diclofenac (cyclooxygenase (COX) inhibitors), and furegrelate (a TXA2 synthase inhibitor). Production of thromboxane B2 induced by H2O2 (5×10−4 M) was greater in SHR MA than in WKY, and was inhibited by quinacrine, indomethacin and diclofenac, and furegrelate, but not by SQ 29584 and ICI 192605. These results suggested (1) that SHR MA exhibits a higher contraction involving an increased smooth muscle reactivity and less tachyphylaxis to H2O2 than WKY; (2) that a greater production of TXA2 through activation of PLA2‐COX‐TXA2 synthase pathway appeared to be responsible for the enhanced contraction in SHR MA. The enhanced vascular response to H2O2 may be related to hypertension in SHR.

Structural and reactivity alterations of the renal vasculature of spontaneously hypertensive rats prior to and during established hypertension.

The renal vasculatures of Wlstar Kyoto spontaneously hypertensive rats (SHR), prior to (4-5 week) and during established hypertension (21 week) and those of age-matched Wistar Kyoto normotensive rats (WKY) were morphometrically and pharmacologically studied. Under dilated conditions, the vascular resistances (RVR) of the isolated kidneys of young and adult SHR were similar to WKY. Morphometric measurements of renal vasculature indicated that the cross-sectional area of the intima and ad ventitia and its subcomponents were similar in adult SHR and WKY. With the exception of the preglomerular arterioles, all the renal arteries of adult SHR exhibited elevated cross-sectional quantities of total media, medial smooth muscle cells (SMCs), and extracellular space. Analysis of the SMCs indicated the presence of increased numbers of SMC layers and/or an increase in the SMC volume-to-surface area ratio in arteries sampled from adult SHR. Vascular contraction produced by infusing norepinephrine, BaCl2, angiotensin II, or by stimulating the renal nerves elevated the RVR to a greater degree hi adult SHR than in WKY. The sensitivity of the renal vasculature to the various contractile agents was similar in adult SHR and WKY. When compared with WKY, prehypertensive SHR also exhibited increased cross-sectional quantities of arterial media and elevated amplitudes of RVR change in response to norepinephrine and renal nerve stimulation. However, the vascular contractile sensitivity to norepinephrine was reduced. Our results indicate that renovascular wall thickening and the hypercontractile reactivity associated with such a change precedes hypertension in SHR. In prehypertensive SHR, elevations in RVR might be counterbalanced by a decreased norepinephrine sensitivity. An increase in the norepinephrine contractile sensitivity and further vascular thickening with age could elevate the RVR and establish hypertension.

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.