Faculty Bibliography

The endothelium is not merely a barrier but it plays a key role in the maintenance of vascular homeostasis. A dysfunctional endothelium is an early marker of the development of atherosclerotic changes and can also contribute to cardiovascular events. Vascular reactivity tests represent the most widely used methods in the clinical assessment of endothelial function and in the last two decades, several methodologies were developed to study it non invasively in the peripheral macrocirculation (conduit arteries) and microcirculation (resistance arteries and arterioles). This review will centre on the most relevant available non-invasive techniques in the research on endothelial function, their advantages and limitations. Flow mediated dilation (FMD) of the brachial artery by ultrasounds is the most widely used vascular test to assess endothelium-dependent vasodilation. Other approaches include measurement of microcirculatory reactive hyperaemia by forearm venous pletysmography or digital pulse amplitude tonometry, response to beta2 agonist by applanation tonometry or digital photoplethysmography and several test by skin laser doppler. It appears that FMD is the most reproducible test when an appropriate and accurate methodology is applied. Recently, post-ischemic vasodilation in the cavernous arteries was also suggested to study endothelial function in patients with erectile dysfunction. Systemic markers proposed as measures of NO biology, inflammatory cytokines, adhesion molecules, or markers of endothelial damage and repair have only a very limited role as a result of biological and assay availability and variability, these factors currently have a limited role in the assessment of individual patients. The optimal methodology for investigating the multifaceted aspects of endothelial dysfunction is still under debate. Therefore, no available test to assess endothelial function has sufficient sensitivity and specificity to be used yet in clinical practice. Only the growing concordant results from different reproducible and reliable non-invasive methods exploring endothelial function with different stimuli will support and strengthen experimental findings, thus providing conclusive answers in this area of research.

We report a case of decompensated porto-pulmonary hypertension closely associated with the development of intra-portocaval shunt thrombosis. A woman with Laennec's cirrhosis was hospitalized because of severe dyspnea and edema. She underwent surgical portocaval anastomosis ten years ago. Imaging studies showed massive intra-shunt thrombosis, portal hypertension, ascites, pleuro-pericardial effusions and enlargement of right cardiac cavities. Cardiac catheterization allowed to rule out coronary and left-sided heart abnormalities and led to the diagnosis of pre-capillary pulmonary hypertension. Antithrombotic treatment with low molecular weight heparin was instituted. The management also included ACE inhibitors, spironolactone, low-salt diet and lactulose. The patient was discharged and three months later we observed the disappearance of edema, ascites and pleuro-pericardial effusions, a marked body weight reduction and improved dyspnea and liver function tests. A possible link between the development of intra-shunt thrombosis and clinical decompensation in our patient was hypothesized. In fact, it has been demonstrated that the increased portal pressure, caused by occlusion of portosystemic shunt, reduces renal plasma flow and increases systemic endothelin-1 concentration. In our patient the disappearance of edematous state and improved dyspnea observed after recanalization of the shunt strongly support this hypothesis.

BACKGROUND:Essential hypertension is characterized by endothelial dysfunction, arterial stiffness, and increased oxidative stress. We evaluated the effect of short-term combined treatment with the antioxidants vitamins C and E on endothelial function, arterial stiffness, and oxidative stress in untreated essential hypertensive patients. METHODS:A randomized, double-blind, placebo-controlled, crossover study design was used to assign 30 male essential hypertensive patients to either vitamin C (1 g) and vitamin E (400 IU) or placebo for 8 weeks. Endothelium-dependent response was assessed as flow-mediated dilation (FMD) of the brachial artery. Arterial stiffness was assessed as central pulse wave velocity (PWV) and augmentation index (AIx). Plasma markers of oxidative stress and antioxidant status were measured. RESULTS:After vitamin supplementation, FMD was significantly improved. Central PWV was significantly reduced, while AIx tended to decrease. Plasma vitamin levels and antioxidant capacity increased significantly. Levels of oxidative stress decreased. Changes in central PWV were related to changes in levels of oxidative stress. CONCLUSIONS:Combined treatment with vitamins C and E has beneficial effects on endothelium-dependent vasodilation and arterial stiffness in untreated, essential hypertensive patients. This effect is associated with changes in plasma markers of oxidative stress.

We evaluated the possible role of NO in modulating tissue plasminogen activator (t-PA) release in the forearm microcirculation of normotensive subjects and hypertensive patients. Essential hypertensive patients are characterized by endothelial dysfunction because of a reduced NO availability and also show an impaired t-PA release. In healthy volunteers and essential hypertensive patients, we studied local t-PA release and forearm blood flow changes (strain-gauge plethysmography) induced by intrabrachial administration of acetylcholine (0.45 and 1.5 microg/100 mL/min) and of sodium nitroprusside (0.5 and 1.0 microg/100 mL/min), an endothelium-dependent and -independent agonist, respectively. Acetylcholine was also repeated in the presence of intra-arterial infusion of the NO synthase inhibitor N(G)-monomethyl-l-arginine (100 microg/100 mL/min). In normotensive subjects, vasodilation to acetylcholine was blunted by N(G)-monomethyl-l-arginine. In these subjects, acetylcholine infusion induced a significant, dose-dependent increase in net forearm t-PA release. N(G)-monomethyl-l-arginine significantly reduced basal t-PA release, as well as acetylcholine-induced t-PA release. In essential hypertensive patients, vasodilation to acetylcholine was reduced as compared with controls and resistant to N(G)-monomethyl-l-arginine. In contrast to what was observed in healthy control subjects, in hypertensive patients, acetylcholine had no effect on t-PA release. Similarly, N(G)-monomethyl-l-arginine failed to modify either the tonic or the agonist-induced t-PA release. Both tonic and agonist-induced release of NO are directly involved in t-PA release by endothelial cells. Essential hypertension, characterized by a reduction in tonic and stimulated NO availability, is also associated with impaired capacity of t-PA release, suggesting a major role of impaired NO availability in worsening both vasodilation and t-PA release.

Angiotensin II induces endothelial dysfunction by reducing NO availability and increasing reactive oxygen species. We assessed whether cyclooxygenase (COX)-1 or COX-2 participate in the angiotensin II-induced endothelial dysfunction in murine mesenteric small arteries and examined the role of reduced nicotinamide-adenine dinucleotide phosphate-dependent reactive oxygen species production. Mice received angiotensin II (600 ng/kg per minute, SC), saline (controls), angiotensin II + apocynin (reduced nicotinamide-adenine dinucleotide phosphate oxidase inhibitor, 2.5 mg/day), or apocynin alone for 2 weeks. Endothelial function of mesenteric arteries was assessed by pressurized myograph. In controls, acetylcholine-induced relaxation was inhibited by NG-monomethyl-L-arginine and unaffected by DFU (COX-2 inhibitor), SC-560 (COX-1 inhibitor), or ascorbic acid. In angiotensin II-infused animals, the attenuated response to acetylcholine was less sensitive to NG-monomethyl-L-arginine, unaffected by DFU, and enhanced by SC-560 and, similarly, by SQ-29548, a thromboxane-prostanoid receptor antagonist. Moreover, response to acetylcholine was unchanged by ozagrel, a thromboxane synthase inhibitor, and normalized by ascorbic acid. Apocynin prevented the angiotensin II-induced vascular dysfunctions. In angiotensin II-infused mice, RT-PCR analysis showed a significant COX-2 downregulation, whereas COX-1 expression was upregulated. These changes were unaffected by apocynin. Modulation of COX isoform by angiotensin II was also documented by immunohistochemistry. In small mesenteric vessels, the reduced NO availability and oxidant excess, which characterize endothelial dysfunction secondary to angiotensin II, are associated with a reduced COX-2 and an increased COX-1 function and expression. Angiotensin II causes an oxidative stress-independent COX-1 overexpression, whereas angiotensin II-mediated oxidant excess production stimulates COX-1 activity to produce a contracting prostanoid endowed with agonist activity on thromboxane-prostanoid receptors.

DESIGN AND PARTICIPANTS/METHODS:A double-blind, crossover, randomized study was designed to evaluate the effect of 3-month treatment with a lower versus a higher antihypertensive dosage of ramipril (5 or 10 mg/day) on nitric oxide (NO)-dependent vasodilation in 46 untreated patients with essential hypertension. Radial artery flow-mediated dilation (FMD), before and after the intra-arterial infusion of NG-monomethyl-L-arginine (L-NMMA), to block NO synthase, and the response to sublingual glyceril trinitrate (GTN, 25 microg) were measured at baseline and after the two treatment periods as a change in artery diameter (computerized system from ultrasound scans). Plasma angiotensin II and oxidative stress markers were also assessed. RESULTS:FMD was significantly (P < 0.01) lower in hypertensive patients (4.6 +/- 1.8%) than in normotensive subjects (7.1 +/- 2.6%), whereas the response to GTN was similar. L-NMMA significantly (P < 0.001) inhibited FMD in normotensive but not in hypertensive subjects. Mean 24-h ambulatory blood pressure, plasma angiotensin II and oxidative stress marker levels were similarly reduced at the end of the two treatment periods. Both dosages of ramipril significantly (P < 0.001) increased FMD (5 mg: 5.9 +/- 2.1%; 10 mg: 6.3 +/- 2.4%) without modifying the response to GTN. However, compared with baseline (11 +/- 19%), the inhibiting effect of L-NMMA on FMD (NO-dependent FMD) was significantly (P < 0.01) greater with ramipril 10 mg (49 +/- 12%) than 5 mg per day (38 +/- 15%). The improvement in FMD and NO-dependent FMD was not related to changes in plasma levels of angiotensin II or markers of oxidative stress. CONCLUSION/CONCLUSIONS:Treatment with ramipril at a higher dosage induced a greater improvement in NO-dependent vasodilation compared with the lower antihypertensive dosage in hypertensive patients.

The evaluation of the intima media thickness (IMT) of the common carotid artery (CCA) with B-mode ultrasonography represents an important index of cardiovascular risk. The IMT is defined as the distance between the leading edge of the lumen-intima interface and the leading edge of the media-adventitia interface. In order to evaluate the IMT, it is necessary to locate such edges. In this paper we developed an automatic real-time system to evaluate the IMT based on the first order absolute moment (FOAM), which is used as an edge detector, and on a pattern recognition approach. The IMT measurements were compared with manual measurements. We used regression analysis and Bland-Altman analysis to compare the results.