Breakers of advanced glycation end products restore large artery properties in experimental diabetes

Glucose and other reducing sugars react with proteins by a nonenzymatic, posttranslational modification process called nonenzymatic glycation. The formation of advanced glycation end products (AGEs) on connective tissue and matrix components accounts largely for the increase in collagen crosslinking that accompanies normal aging and which occurs at an accelerated rate in diabetes, leading to an increase in arterial stiffness. A new class of AGE crosslink “breakers” reacts with and cleaves these covalent, AGE-derived protein crosslinks. Treatment of rats with streptozotocin-induced diabetes with the AGE-breaker ALT-711 for 1–3 weeks reversed the diabetes-induced increase of large artery stiffness as measured by systemic arterial compliance, aortic impedance, and carotid artery compliance and distensibility. These findings will have considerable implications for the treatment of patients with diabetes-related complications and aging.

An epidemiological needs assessment of carotid endarterectomy in an English health region. Is the need being met?

Objective: To compare the level of provision of carotid endarterectomy (an intervention of proved efficacy for prevention of stroke in patients with symptomatic high grade carotid artery stenosis) with estimates of need.

Targeting Gβγ signaling in arterial vascular smooth muscle proliferation: A novel strategy to limit restenosis

Restenosis continues to be a major problem limiting the effectiveness of revascularization procedures. To date, the roles of heterotrimeric G proteins in the triggering of pathological vascular smooth muscle (VSM) cell proliferation have not been elucidated. βγ subunits of heterotrimeric G proteins (Gβγ) are known to activate mitogen-activated protein (MAP) kinases after stimulation of certain G protein-coupled receptors; however, their relevance in VSM mitogenesis in vitro or in vivo is not known. Using adenoviral-mediated transfer of a transgene encoding a peptide inhibitor of Gβγ signaling (βARKct), we evaluated the role of Gβγ in MAP kinase activation and proliferation in response to several mitogens, including serum, in cultured rat VSM cells. Our results include the striking finding that serum-induced proliferation of VSM cells in vitro is mediated largely via Gβγ. Furthermore, we studied the effects of in vivo adenoviral-mediated βARKct gene transfer on VSM intimal hyperplasia in a rat carotid artery restenosis model. Our in vivo results demonstrated that the presence of the βARKct in injured rat carotid arteries significantly reduced VSM intimal hyperplasia by 70%. Thus, Gβγ plays a critical role in physiological VSM proliferation, and targeted Gβγ inhibition represents a novel approach for the treatment of pathological conditions such as restenosis.

Separation of the arterial wall from blood contact using hydrogel barriers reduces intimal thickening after balloon injury in the rat: The roles of medial and luminal factors in arterial healing

The objective of this study was to clarify the relative roles of medial versus luminal factors in the induction of thickening of the arterial intima after balloon angioplasty injury. Platelet-derived growth factor (PDGF) and thrombin, both associated with thrombosis, and basic fibroblast growth factor (bFGF), stored in the arterial wall, have been implicated in this process. To unequivocally isolate the media from luminally derived factors, we used a 20-μm thick hydrogel barrier that adhered firmly to the arterial wall to block thrombus deposition after balloon-induced injury of the carotid artery of the rat. Thrombosis, bFGF mobilization, medial repopulation, and intimal thickening were measured. Blockade of postinjury arterial contact with blood prevented thrombosis and dramatically inhibited both intimal thickening and endogenous bFGF mobilization. By blocking blood contact on the two time scales of thrombosis and of intimal thickening, and by using local protein release to probe, by reconstitution, the individual roles of PDGF-BB and thrombin, we were able to conclude that a luminally derived factor other than PDGF or thrombin is required for the initiation of cellular events leading to intimal thickening after balloon injury in the rat. We further conclude that a luminally derived factor is required for mobilization of medial bFGF.

Conditional and targeted overexpression of vascular chymase causes hypertension in transgenic mice

We cloned a rat vascular chymase (RVCH) from smooth muscle cells (SMCs) that converts angiotensin I to II and is up-regulated in SMC from spontaneously hypertensive vs. normotensive rats. To determine whether increased activity of RVCH is sufficient to cause hypertension, transgenic mice were generated with targeted conditional expression of RVCH to SMC, with the use of the tetracycline-controlled transactivator (tTA). We confirmed conditional expression of RVCH by mRNA, protein, and chymase activity in the absence, but not in the presence, of dietary doxycycline. The systolic blood pressure (mmHg), measured by carotid artery cannulation at 10–12 weeks of age, was higher in tTA+/RVCH+ mice than in nonbinary transgenic littermates (136 ± 4 vs. 109 ± 3) (P < 0.05), as were the diastolic and mean pressures. Hypertension was completely reversed by doxycycline, suggesting a causal link with chymase expression. Medial thickening of mesenteric arteries from tTA+/RVCH+ mice vs. littermates (0.82 ± 0.1 vs. 0.42 ± 0.02) (P < 0.05) was associated with increased SMC proliferation, as judged by positive immunoreactivity, with the use of an antibody to the proliferating cell nuclear antigen. These structural changes were prevented by doxycycline. Perfusion myography of mesenteric arteries from tTA+/RVCH+ mice also revealed increased vasoconstriction in response to phenylephrine and impaired metacholine-induced vasodilatation when compared with littermate controls or with the doxycyline-treated group. Our studies suggest that up-regulation of this vascular chymase is sufficient to cause a hypertensive arteriopathy, and that RVCH may be a candidate gene and a therapeutic target in patients with high blood pressure.

Capgras syndrome: a clinical manifestation of watershed cerebral infarct complicating the use of extracorporeal membrane oxygenation

Ischaemic cerebral accidents are frequent following extracorporeal membrane oxygenation (ECMO), especially after fixing the reinjection cannula in the right primitive carotid artery, which leads to an interruption in downstream flow. We describe a rare and unusual symptom of cerebral ischaemic accident that is known as Capgras syndrome. This feature is interesting because it may be documented by computed tomography (CT) scan and particular electroencephalography signals. It appears that our observation represents the first documented case of Capgras syndrome complicating ECMO. This incident emphasizes the potential hazards associated with right common artery ligature for venoarterial extracorporeal membrane oxygenation (VAECMO). In addition, it shows that this psychiatric symptom (that has been interpreted psychodynamically for many years) can have an organic basis, which should be studied.

The angiotensin II type 2 (AT2) receptor antagonizes the growth effects of the AT1 receptor: gain-of-function study using gene transfer.

The type 1 angiotensin II (AT1) receptor is well characterized but the type 2 (AT2) receptor remains an enigma. We tested the hypothesis that the AT2 receptor can modulate the growth of vascular smooth muscle cells by transfecting an AT2 receptor expression vector into the balloon-injured rat carotid artery and observed that overexpression of the AT2 receptor attenuated neointimal formation. In cultured smooth muscle cells, AT2 receptor transfection reduced proliferation and inhibited mitogen-activated protein kinase activity. Furthermore, we demonstrated that the AT2 receptor mediated the developmentally regulated decrease in aortic DNA synthesis at the latter stages of gestation. These results suggest that the AT2 receptor exerts an antiproliferative effect, counteracting the growth action of AT1 receptor.

Vector-mediated delivery of 125I-labeled beta-amyloid peptide A beta 1-40 through the blood-brain barrier and binding to Alzheimer disease amyloid of the A beta 1-40/vector complex.

The brain amyloid of Alzheimer disease (AD) may potentially be imaged in patients with AD by using neuroimaging technology and a radiolabeled form of the 40-residue beta-amyloid peptide A beta 1-40 that is enabled to undergo transport through the brain capillary endothelial wall, which makes up the blood-brain barrier (BBB) in vivo. Transport of 125I-labeled A beta 1-40 (125I-A beta 1-40) through the BBB was found to be negligible by experiments with both an intravenous injection technique and an internal carotid artery perfusion method in anesthetized rats. In addition, 125I-A beta 1-40 was rapidly metabolized after either intravenous injection or internal carotid artery perfusion. BBB transport was increased and peripheral metabolism was decreased by conjugation of monobiotinylated 125I-A beta 1-40 to a vector-mediated drug delivery system, which consisted of a conjugate of streptavidin (SA) and the OX26 monoclonal antibody to the rat transferrin receptor, which undergoes receptor-mediated transcytosis through the BBB. The brain uptake, expressed as percent of injected dose delivered per gram of brain, of the 125I,bio-A beta 1-40/SA-OX26 conjugate was 0.15 +/- 0.01, a level that is 2-fold greater than the brain uptake of morphine. The binding of the 125I,bio-A beta 1-40/SA-OX26 conjugate to the amyloid of AD brain was demonstrated by both film and emulsion autoradiography performed on frozen sections of AD brain. Binding of the 125I,bio-A beta 1-40/SA-OX26 conjugate to the amyloid of AD brain was completely inhibited by high concentrations of unlabeled A beta 1-40. In conclusion, these studies show that BBB transport and access to amyloid within brain may be achieved by conjugation of A beta 1-40 to a vector-mediated BBB drug delivery system.

Potential roles of osteopontin and αVβ3 integrin in the development of coronary artery restenosis after angioplasty

Angioplasty procedures are increasingly used to reestablish blood flow in blocked atherosclerotic coronary arteries. A serious complication of these procedures is reocclusion (restenosis), which occurs in 30–50% of patients. Migration of coronary artery smooth muscle cells (CASMCs) to the site of injury caused by angioplasty and subsequent proliferation are suggested mechanisms of reocclusion. Using both cultured human CASMCs and coronary atherectomy tissues, we studied the roles of osteopontin (OPN) and one of its receptors, αvβ3 integrin, in the pathogenesis of coronary restenosis. We also measured the plasma levels of OPN before and after angioplasty and determined the effect of exogenous OPN on CASMC migration, extracellular matrix invasion, and proliferation. We found that cultured CASMCs during log phase of growth and smooth muscle cell layer of the coronary atherosclerotic tissues of patients express both OPN mRNA and protein at a significantly elevated level compared with controls. Interestingly, whereas the baseline plasma OPN levels in control samples were virtually undetectable, those in patient plasma were remarkably high. We also found that interaction of OPN with αvβ3 integrin, expressed on CASMCs, causes migration, extracellular matrix invasion, and proliferation. These effects were abolished when OPN or αvβ3 integrin gene expression in CASMCs was inhibited by specific antisense S-oligonucleotide treatment or OPN-αvβ3 interaction was blocked by treatment of CASMCs with antibodies against OPN or αvβ3 integrin. Our results demonstrate that OPN and αvβ3 integrin play critical roles in regulating cellular functions deemed essential for restenosis. In addition, these results raise the possibility that transient inhibition of OPN gene expression or blocking of OPN-αvβ3 interaction may provide a therapeutic approach to preventing restenosis.