Healing of two microarterial anastomoses with diameter mismatch.

BACKGROUND: The use of fascial perforating vessels as recipients for microvascular composite tissue autotransplants has led to vessel diameter discrepancy becoming an increasingly common finding. Little evidence, however, is available to direct the choice of anastomotic technique where a discrepancy exists. We have been studying two methods of anastomosing arteries where a small-to-large discrepancy exists-a 45° section of the smaller vessel, and invaginating the smaller vessel inside the larger. As part of this work, this study examines intimal hyperplasia and healing of the two methods. MATERIALS AND METHODS: A previously described paired Wistar rat femoral axis model was used. Anastomoses were performed, one on each side, and specimens were harvested in groups at 24 h, 1 wk, 6 wk, and 8 mo. Inflammation, necrosis, and fibrosis in each layer of the vessel wall and intimal hyperplasia were each scored by an assessor blinded to the group and anastomotic technique. RESULTS: Significant differences in healing were found. The invagination technique induced less inflammation, and caused less endothelial and medial necrosis than the oblique cut end-to-end method. Intimal hyperplasia was most pronounced at 6 wk, but no evidence of a difference in the severity of intimal hyperplasia between the two methods was found. CONCLUSIONS: The invaginating anastomosis causes less inflammation and less vessel wall necrosis than the oblique end-to-end method in this model. This finding, alongside results from previous work, suggests that this is the better method to deal with a small-to-large microarterial diameter discrepancy in the range 1:1.5 to 1:2.5.

Nitric oxide synthases in infants and children with pulmonary hypertension and congenital heart disease.

RATIONALE: Nitric oxide is an important regulator of vascular tone in the pulmonary circulation. Surgical correction of congenital heart disease limits pulmonary hypertension to a brief period. OBJECTIVES: The study has measured expression of endothelial (eNOS), inducible (iNOS), and neuronal nitric oxide synthase (nNOS) in the lungs from biopsies of infants with pulmonary hypertension secondary to cardiac abnormalities (n = 26), compared to a control group who did not have pulmonary or cardiac disease (n = 8). METHODS: eNOS, iNOS and nNOS were identified by immunohistochemistry and quantified in specific cell types. MEASUREMENTS AND MAIN RESULTS: Significant increases of eNOS and iNOS staining were found in pulmonary vascular endothelial cells of patients with congenital heart disease compared to control infants. These changes were confined to endothelial cells and not present in other cell types. Patients who strongly expressed eNOS also had strong expression of iNOS. CONCLUSION: Upregulation of eNOS and iNOS occurs at an early stage of pulmonary hypertension, and may be a compensatory mechanism limiting the rise in pulmonary artery pressure.

Nitric oxide generation by NO donors is enhanced following balloon injury in the porcine coronary artery.

Vasospasm is a complication of cardiological procedures such as balloon angioplasty and may be related to vascular oxidant stress. Although nitric oxide donor drugs are often administered to prevent vasospasm, the response to these drugs in balloon-injured arteries has not been studied. Pig coronary arteries were balloon-injured in vitro and relaxations to nitric oxide (NO)-donating and NO-independent vasodilators studied. Generation of superoxide in response to injury was assayed using dihydroethidium. NO formation on addition of the NO donor drugs was studied using an amperometric sensor. Expression of nitrotyrosine, a peroxynitrite marker, was probed using immunocytochemistry. In vitro injury enhanced sensitivity to the NO donors SNAP and SpermineNONOate but blunted the response to isoprenaline or chromakalim. With both donors, NO formation was significantly enhanced in the presence of an injured vessel. Vascular superoxide generation was also increased throughout the vessel wall and a small increase in nitrotyrosine was detected in the injured vessel media following addition of SNAP. In conclusion, injured vessels were more sensitive to NO donors and this appears to be due to enhanced NO generation by the donor molecule. Increased formation of peroxynitrite within the injured vessel may contribute to the enhanced relaxation in injured vessels.

Adrenomedullin acts via nitric oxide and peroxynitrite to protect against myocardial ischaemia-induced arrhythmias in anaesthetized rats.

1. The overall aim of this study was to determine if adrenomedullin (AM) protects against myocardial ischaemia (MI)-induced arrhythmias via nitric oxide (NO) and peroxynitrite. 2. In sham-operated rats, the effects of in vivo administration of a bolus dose of AM (1 nmol kg-1) was assessed on arterial blood pressure (BP), ex vivo leukocyte reactive oxygen species generation and nitrotyrosine deposition (a marker for peroxynitrite formation) in the coronary endothelium. 3. In pentobarbitone-anaesthetized rats subjected to ligation of the left main coronary artery for 30 min, the effects of a bolus dose of AM (1 nmol kg-1, i.v.; n=19) or saline (n=18) given 5 min pre-occlusion were assessed on the number and incidence of cardiac arrhythmias. In a further series of experiments, some animals received infusions of the NO synthase inhibitor N(G)-nitro-L-arginine (LNNA) (0.5 mg kg-1 min-1) or the peroxynitrite scavenger N-mercaptopropionyl-glycine (MPG) (20 mg kg-1 h-1) before AM. 4. AM treatment significantly reduced mean arterial blood pressure (MABP) and increased ex vivo chemiluminescence (CL) generation from leukocytes in sham-operated animals. AM also enhanced the staining for nitrotyrosine in the endothelium of coronary arteries. 5. AM significantly reduced the number of total ventricular ectopic beats that occurred during ischaemia (from 1185+/-101 to 520+/-74; P<0.05) and the incidences of ventricular fibrillation (from 61 to 26%; P<0.05). AM also induced a significant fall in MABP prior to occlusion. AM-induced cardioprotection was abrogated in animals treated with the NO synthase inhibitor LNNA and the peroxynitrite scavenger MPG. 6. This study has shown that AM exhibits an antiarrhythmic effect through a mechanism that may involve generation of NO and peroxynitrite.

TNFalpha increases the inflammatory response to vascular balloon injury without accelerating neointimal formation.

There is now clear evidence for a contributory role of inflammatory processes to restenosis following vascular balloon injury and stent implantation. The aim of the present study was to study the effects of TNFalpha, administered locally in vivo immediately following balloon angioplasty, on the leukocyte adhesive response and extent of neointimal formation in a rabbit model of subclavian artery injury. Initial in vitro studies were performed with normal isolated artery rings to assess the vascular adhesive response to TNFalpha or IL-1beta. Pre-incubation with either cytokine prior to addition of (51)Cr-labelled leukocytes enhanced the adhesion of leukocytes to the artery in both a time- and concentration-dependent manner. Although both cytokines induced an increase in the expression of the adhesion molecules ICAM-1 and VCAM-1, only antibodies to ICAM-1 blocked the enhanced adhesion induced by the cytokines. In artery segments retrieved from rabbits that had previously undergone subclavian artery angioplasty either 24 h or 8 days previously, there was an injury-induced increase in adhesion of leukocytes assessed ex vivo. In segments obtained from rabbits that received a 15 min local infusion of TNFalpha (2 ng/min) to the injured artery immediately after the angioplasty procedure, leukocyte adhesion assessed ex vivo was further significantly enhanced. The pro-adhesive effect of TNFalpha was associated with an increased expression of both ICAM-1 and VCAM-1. However, TNFalpha administration did not alter the extent of neointimal formation observed 8 days after injury. These findings suggest that while TNFalpha may play a role following vascular injury, it does not act alone to induce neointimal formation. Thus anti-inflammatory strategies targeted at multiple cytokines may be more appropriate than targeting a single cytokine to reduce the response to vascular injury.

Endothelial nitric oxide synthase (NOS) is upregulated in rapid progressive pulmonary hypertension of the newborn.

OBJECTIVE: To provide evidence for the upregulation of endothelial nitric oxide synthase (eNOS) or inducible nitric oxide synthase (iNOS) in the assumed imbalance in the pathophysiology of rapid progressive pulmonary hypertension of the newborn (RPPHN), which is characterized by abnormal hypertrophy of the pulmonary arterioles and arteries leading to increased pulmonary vascular resistance. Furthermore, to determine the cellular source and topographic distribution of eNOS and iNOS. MATERIAL AND METHODS: Lung biopsies were taken from two term neonates with clinical and echocardiographic evidence of RPPH and of three controls. Biopsies were obtained at an early stage of the disease as well as at post mortem and examined immunohistochemically for the presence of eNOS, iNOS and nitrotyrosine. RESULTS: The endothelial cells of pulmonary arterioles stained significantly for eNOS protein in RPPHN patients. This was not the case in the control infants. There were no differences for nitrotyrosine or iNOS between RPPHN patients and controls. CONCLUSION: Rapid progressive pulmonary hypertension of the newborn leads to compensatory induction of eNOS synthesis specifically in endothelial cells of the pulmonary arterioles. This mechanism of compensation can lead to delayed presentation of RPPHN during the late neonatal period. Exogenous inhaled nitric oxide therapy does not lead to suppression of the endogenous synthesis of nitric oxide.

Correlation of changes in nitric oxide synthase, superoxide dismutase and nitrotyrosine with endothelial regeneration and neointimal hyperplasia in the balloon-injured rabbit subclavian artery.

OBJECTIVES: Alterations in nitric oxide (NO) and superoxide production within the wall of injured vessels may modulate the development and eventual extent of neointima after balloon injury. METHODS: In this study we have characterized a rabbit model of subclavian artery injury and have used immunocytochemistry to detect NO synthase (NOS) isoforms, Cu-Zn superoxide dismutase (SOD) and nitrotyrosine in the injured vessel from 2 h to 28 days after injury. RESULTS: At 48 h after injury, when cellular proliferation that will ultimately form the neointima is commencing, there was upregulation of inducible NOS, Cu-Zn SOD and nitrotyrosine. Recovery of endothelial NOS occurred at 28 days after injury, when the neointima is stabilizing and cellular proliferation has slowed down. There was no increase in neuronal NOS at any time point studied. CONCLUSIONS: NO may serve to limit the development of neointima while superoxide may attenuate the effect of NO by formation of peroxynitrite, detected as increased nitrotyrosine staining. Upregulation of Cu-Zn SOD would limit superoxide both at sites of inflammation in the vessel wall from 48 h and in the adventitia up to 28 days after injury. Very early intervention to protect NO may reduce neointimal size.

Bright-blood T(2)-weighted MRI has high diagnostic accuracy for myocardial hemorrhage in myocardial infarction: a preclinical validation study in swine.

BACKGROUND: Myocardial hemorrhage after myocardial infarction (MI) usually goes undetected. We investigated the diagnostic accuracy of bright-blood T(2)-weighted cardiac MRI for myocardial hemorrhage in experimental MI. METHODS AND RESULTS: MI was created in swine by occluding the left anterior descending (n=10) or circumflex (n=5) coronary arteries for 90 minutes followed by reperfusion for ≤3 days (n=2), 10 days (n=7), or 60 days (n=6). MRI was performed at 1.5 T, using bright-blood T(2)-prepared steady-state free-precession, T(2)* and early (1 minute) and late (10-15 minutes) gadolinium enhancement (EGE, LGE, respectively) MRI. Left ventricular sections and histology were assessed for hemorrhage by an experienced cardiac pathologist blinded to the MRI data. Hypointense regions on T(2)-weighted and contrast-enhanced MRI were independently determined by 3 cardiologists experienced in MRI who were also blinded to the pathology results. Eighty ventricular pathological sections were matched with MRI (n=68 for EGE MRI). All sections with evidence of MI (n=63, 79%) also exhibited hyperintense zones consistent with edema on T(2)-weighted MRI and infarct on LGE MRI. Myocardial hemorrhage occurred in 49 left ventricular sections (61%) and corresponded with signal voids on 48 T(2)-weighted (98%) and 26 LGE-MRI (53%). Alternatively, signal voids occurred in the absence of hemorrhage in 3 T(2)-weighted (90% specificity) and 5 LGE MRI (84% specificity). On EGE MRI, 27 of 43 cases of early microvascular obstruction corresponded with hemorrhage (63% sensitivity), whereas 5 of 25 defects occurred in the absence of hemorrhage (80% specificity). The positive and negative predictive values for pathological evidence of hemorrhage were 94% and 96% for T(2)-weighted, 84% and 55% for LGE MRI, and 85% and 56% for EGE MRI. CONCLUSIONS: Bright-blood T(2)-weighted MRI has high diagnostic accuracy for myocardial hemorrhage.