Carotid Artery Balloon Injury Research Articles

Percutaneous endoluminal arterial cryoenergy improves vascular remodelling after angioplasty

This study aimed to investigate the effect of percutaneous endoluminal arterial cryoenergy immediately after balloon angioplasty on vascular remodeling. Restenosis, the main complication after coronary artery angioplasty, is a complex phenomenon in which vascular remodeling plays a prominent role. Observations of reduced scarring in freeze-induced wounds suggest potential value for cryoenergy in the prevention of restenosis. Juvenile swine underwent a first oversized balloon injury in both carotid arteries (CA) (3 injury sites/artery) and a second injury at day 14. At that time, one CA was randomly assigned to endoluminal cryoenergy of the sequential segments (proximal, medial, distal) at -15 degrees C, -30 degrees C, and -50 degrees C for 120 sec, and the other CA was used as control. Animals were sacrificed 28 days after the second procedure. Compared with intact reference segments, angioplasty reduced both the luminal (LA) and the external elastic lamina (EEL) areas from 6.66+/-0.59 to 3.13+/-0.54 mm(2) (p<0.05) and from 8.81+/-0.81 to 6.48+/-0.52 mm(2) (p<0.05), respectively. Cryoenergy, at the temperature with maximal benefits (-50 degrees C), caused a temperature-related protection, as the LA was maintained (6.79+/-0.89 versus 7.18+/-0.78 mm(2) for reference) and the EEL area increased from 9.12+/-0.78 to 12.98+/-1.07 mm(2), p<0.05. Moreover, cryoenergy increased the density of collagen III (p<0.05) which correlated with the maintenance of the LA (r=0.8045, p<0.009). Cryoenergy prevents late luminal loss after double-injury angioplasty by improving vascular remodeling, and is an interesting new therapeutic approach for the prevention of restenosis after angioplasty. The increased synthesis of collagen III appears to be involved in this phenomenon and could be a potential method of stabilizing the vulnerable plaque.

Inhibition of neointimal hyperplasia by a specific thrombin inhibitor

Objective—Restenosis secondary to neointimal hyperplasia remains the major limiting factor after vascular interventions. Thrombin generated in high concentrations at the site of vascular injury plays a central role in thrombosis and hemostasis. Thrombin has also been implicated as a mitogen for smooth muscle cell proliferation that contributes to restenosis. This study was designed to determine the effects of a specific thrombin inhibitor on neointimal hyperplasia after balloon injury in a rat carotid artery model.Design—A total of 47 male Sprague–Dawley rats were divided into five groups. All groups underwent balloon injury of the left carotid artery. A specific thrombin inhibitor, inogatran, was given in four different regimens: low and high dose injections, short‐term infusion for 3 h, and long‐term infusion for 1 week. After 2 weeks the animals were killed and the carotid neointima/media area ratio and the luminal narrowing were calculated.Results—All treatments significantly reduced the neointimal hyperplasia. Inogatran given as a long‐term infusion for 1 week had the lowest neointima/media ratio compared with the other groups. The percentage of lumen narrowing was also significantly lower in all treatment groups compared with the control group.Conclusion—A specific direct thrombin inhibitor, inogatran, reduces neointimal hyperplasia after arterial injury in rats. A more prolonged administration of the thrombin inhibitor gave a further reduction of the neointimal hyperplasia. It seems that inhibition of thrombin activity is not only important early after injury, but also later. This could have clinical implications in the treatment of restenosis and needs to be further evaluated.

Estradiol and progestins differentially modulate leukocyte infiltration after vascular injury.

Inflammation plays an important role in the response to endoluminal vascular injury. Estrogen (17beta-estradiol, E2) inhibits neointima formation in animal models, and the progestin medroxyprogesterone acetate (MPA) blocks this effect. This study tested the hypothesis that E2 inhibits the migration of inflammatory cells, particularly granulocytes, into the rat carotid arteries after acute endoluminal injury and that MPA blocks this effect. Ovariectomized rats were randomly divided into subgroups and treated with E2, MPA, E2+MPA, or vehicle and subjected to balloon injury of the right carotid artery. After 1, 3, or 7 days, rats were euthanized, and carotid arteries (injured and control) were analyzed for inflammatory cells by flow cytometry. At 1 day, granulocytes (HIS48+ and CD45+), monocyte/macrophages (Mar1+ and CD45+), and T lymphocytes (CD3+ and CD45+) were increased 26-fold, 12-fold, and 3-fold, respectively, in injured compared with contralateral control arteries of vehicle-treated rats. Granulocytes and monocyte/macrophages decreased markedly by 3 days. E2 reduced the granulocyte and monocyte/macrophage populations of injured vessels by approximately 50% and increased T lymphocytes. MPA had no independent effect on inflammatory cells but completely blocked the effect of E2. Immunohistochemical examination verified these findings and localized inflammatory cells to the adventitial and periadventitial domains of injured vessels. E2 may limit the neointimal response to endoluminal vascular injury, at least in part, by limiting leukocyte entry from adventitial/periadventitial tissues into injured vessels early in the injury response.

The neointimal response to endovascular injury is increased in obese Zucker rats.

Restenosis after revascularization procedures is accelerated in persons with type 2 diabetes. The current study tested the hypothesis that the neointimal response to endovascular injury is enhanced in female obese Zucker (OZ) rats, a model of type 2 diabetes. Animals were randomized to receive either a standard diet (SD) or a diabetogenic diet (DD) for 6 weeks. Four weeks later, balloon injury of the right common carotid artery was induced. All rats were euthanized 2 weeks after injury. Lean Zucker (LZ) rats served as controls. At the time of death, plasma glucose was elevated in OZ rats fed a SD (208 +/- 13 mg/dl) and a DD (288 +/- 21 mg/dl) compared to corresponding LZ rats (SD: 153 +/- 8; DD: 132 +/- 7 mg/dl). The ratio of high-density lipoprotein cholesterol (HDLc) to total cholesterol (Totc), an index of atherogenicity, was reduced in OZ rats on both diets (SD: 0.77 +/- 0.06; DD: 0.80 +/- 0.09) compared to LZ controls (SD: 1.11 +/- 0.02; DD: 1.20 +/- 0.05). Histomorphometric analysis of injured arteries showed that the intima to media (I : M) ratio was significantly increased in OZ (1.37 +/- 0.07) compared to LZ (0.79 +/- 0.08) rats. Elevations in plasma glucose and triglycerides (Tg) correlated positively and decreases in HDLc negatively with an increased I : M ratio. Administration of the DD did not further enhance the I : M ratio in LZ (0.87 +/- 0.06) or OZ (1.29 +/- 0.09) rats. These results suggest that neointima formation following endoluminal injury of the carotid artery is enhanced at an early stage in the development of diabetes mellitus.

Gene transfer of the inducible nos inhibits intimal hyperplasia in a model of diabetic vascular injury

Introduction: Cardiovascular complications of diabetes contribute significantly to the morbidity of this disease. Intimal hyperplasia (IH), which limits our current therapies, is a disease process which disproportionately affects patients with diabetes. The pathophysiology of diabetic vascular disease may be, in part, secondary to decreased bioavailability of nitric oxide (NO). The purpose of these experiments was to test the hypothesis that gene transfer of inducible NO synthase (iNOS) prevents IH following balloon injury in a rodent model of diabetes. Methods: Obese Zucker rats were utilized as a model of Type II Diabetes. These rats carry a homozygous recessive gene (fa/fa) making them resistant to leptin. Their lean counterparts (fa/-) are leptin-sensitive and were used as controls. The obese rats underwent carotid artery balloon injury followed by either adenoviral iNOS or lacZ gene transfer at the time of injury (2x106 PFU per artery). The animals were sacrificed at 3, 7, and 21 days and the degree of injury assessed. Glucose, cholesterol, and insulin levels were also checked. Values are expressed as mean ± standard error and statistical analysis was performed using one-way ANOVA. Results: The obese Zucker rats had marked elevations in baseline glucose, cholesterol, and serum insulin levels compared to lean controls. IH was worse in obese rats compared to controls, with intima:media ratios (I:M) of 0.90 ± 0.14 vs 0.54 ± 0.08, respectively, at 21 days. AdlacZ treatment did not significantly affect the development of IH compared to untreated controls in both lean and obese rats. Adenoviral iNOS gene transfer significantly reduced IH by 37% in lean and 64% in obese rats, with I:M ratios of 0.34 ± 0.03 and 0.33 ± 0.07, respectively. Conclusion: Vascular injury resulted in greater IH in diabetic rodents, raising the concern that diabetes might also reduce the efficacy of therapies that have been shown to inhibit IH in normal animals. Adenoviral iNOS gene transfer effectively inhibits the formation of IH in control rats. The presence of diabetes did not reduce the efficacy of this treatment. Instead, iNOS gene transfer had a greater protective effect in obese diabetic Zucker rats. iNOS gene transfer may be a useful clinical adjunct to vascular interventions in diabetic patients.

Angiogenesis inhibitor TNP-470 (AGM-1470) suppresses vascular smooth muscle cell proliferation after balloon injury in rats

Background. Although TNP-470, a synthetic analog of fumagillin, may inhibit vascular intimal hyperplasia, the effects of TNP-470 on smooth muscle cell (SMC) proliferation have not been demonstrated in vivo. The aim of this study was to confirm the effect of TNP-470 on vascular SMC proliferation using a rat carotid artery balloon injury model. Materials and methods. Rats were treated with vehicle or with TNP-470 at low dosage (10 mg/kg), medium dosage (20 mg/kg), or high dosage (40 mg/kg). The animals received subcutaneous injections of materials three times a week from the day following balloon injury. All rats were sacrificed at 2 weeks after injury. The ratio of intimal-to-medial cross-sectional areas (I/M ratio) and the PCNA labeling index was calculated for each group. The DNA synthesis of cultured SMCs was also evaluated using [ 3H]thymidine incorporation assays. Smooth muscle cells were stimulated with basic fibroblast growth factor and TNP-470 (0.01–100 ng/ml) were added. Results. The inhibition of intimal hyperplasia increased in a dose-dependent manner. TNP-470 also decreased PCNA expression in the neointima and inhibited DNA synthesis of cultured SMCs. Conclusion. TNP-470 may be useful in the prevention of vascular intimal hyperplasia.

C-reactive protein upregulates angiotensin type 1 receptors in vascular smooth muscle.

Accumulating evidence suggests that C-reactive protein (CRP), in addition to predicting vascular disease, may actively facilitate lesion formation by inciting endothelial cell activation. Given the central importance of angiotensin type 1 receptor (AT1-R) in the pathogenesis of atherosclerosis, we examined the effects of CRP on AT1-R expression and kinetics in vascular smooth muscle (VSM) cells. In addition, the effects of CRP on VSM migration, proliferation, and reactive oxygen species (ROS) production were evaluated in the presence and absence of the angiotensin receptor blocker, losartan. Lastly, the effects of CRP (and losartan) on neointimal formation were examined in vivo in a rat carotid angioplasty model. The effects of human recombinant CRP (0 to 100 microg/mL) on AT1-R transcript, mRNA stability, and protein expression were studied in cultured human VSM cells. AT1-R binding was assessed with 125I-labeled angiotensin II (Ang II). VSM migration was assessed with wound cell migration assays, whereas VSM proliferation was determined with [3H]-incorporation and cell number. The effects of CRP (and losartan) on Ang II-induced ROS production were evaluated by 2',7'-dichlorofluorescein fluorescence. Lastly, the effects of CRP (and losartan) on neointimal formation, VSM cell migration, proliferation, and matrix formation were studied in vivo in a rat carotid artery balloon injury model. CRP markedly upregulated AT1-R mRNA and protein expression and increased AT1-R number on VSM cells. CRP promoted VSM migration and proliferation in vitro and increased ROS production. Furthermore, CRP potentiated the effects of Ang II on these processes. In the rat carotid artery angioplasty model, exposure to CRP resulted in an increase in cell migration and proliferation, collagen and elastin content, and AT1-R expression, as well as an increase in neointimal formation; these effects were attenuated by losartan. CRP, at concentrations known to predict cardiovascular events, upregulates AT1-R-mediated atherosclerotic events in vascular smooth muscle in vitro and in vivo. These data lend credence to the notion that CRP functions as a proatherosclerotic factor as well as a powerful risk marker.

Novel NAD(P)H oxidase inhibitor suppresses angioplasty-induced superoxide and neointimal hyperplasia of rat carotid artery.

Neointimal proliferation occurring after vascular or endovascular procedures is a major complication leading to end-organ or limb ischemia. In experimental models, balloon injury has been shown to induce NAD(P)H oxidase to produce vascular superoxide anion (O2*-) production, which has been implicated in cell proliferation, but a direct link is still unclear. We postulated that inhibition of arterial NAD(P)H oxidase, resulting in decreased O2*-, would lessen the neointimal hyperplasia caused by balloon injury to the common carotid artery (CCA). Sprague-Dawley rats were implanted with osmotic minipumps containing either vehicle, a cell-permeant peptide that inhibits NAD(P)H oxidase (gp91ds-tat, 10 mg/kg per day), or a scrambled peptide control (scrmb-tat). Two days after pump implantation, the left CCA was injured using an intravascular balloon embolectomy catheter (2F Fogarty). Systolic blood pressure was monitored by tail cuff. Fourteen days after injury, CCAs were harvested and analyzed by digital morphometry. Rats in both groups remained normotensive, with no significant differences in systolic blood pressure. Reactive oxygen species measurements after injury indicated a significant reduction in vascular O2*- in rats infused with gp91ds-tat, and the neointima/media area and thickness ratios were significantly lower in their arteries compared with control. On the contrary, no significant change in overall CCA diameter was observed in any group. Our data indicate that in response to balloon injury of the rat carotid artery, NAD(P)H oxidase activity contributes to neointimal hyperplasia and is involved in vascular cell proliferation and migration during restenosis.

4P-1148 Celecoxib, cyclooxygenase-2 (COX-2) inhibitor, reduces neointimal hyperplasia after balloon injury of rat carotid artery via the Akt pathway

4P-1148 Celecoxib, cyclooxygenase-2 (COX-2) inhibitor, reduces neointimal hyperplasia after balloon injury of rat carotid artery via the Akt pathway

The Effect of Farnesyl Transferase Inhibitor on the Proliferation of Vascular Smooth Muscle Cells and Neointima Formation

Background and Objectives:We tested the hypothesis that prolonged oral administration of farnesyl transferase inhibitor (FTI, LB42908a, MW=604, LG chemical, Korea) inhibits the proliferation and neointimal thickening of smooth muscle cells (SMC) in a rat carotid injury model. Materials and Methods:Cultured rat aortic vascular SMCs were exposed to sequential concentrations of FTI, and the proliferation inhibition analyzed using the MTT assay. In the rat carotid injury model, the FTI, at 3 dose levels (low-dose;10 mg/kg, bid;mid-dose;50 mg/kg, bid;high-dose;100 mg/kg, bid), or as a placebo, was administered orally, twice a day for 14 days, starting from 30 minutes before injury, until sacrifice. The histo-morphometric analysis was performed. The immunohistochemical detection of the proliferating cell nuclear antigen (PCNA) was performed for 3 days. Results:FTI inhibited the PDGF or FBS-induced cellular proliferations in a dose dependently manner. In the rat carotid artery balloon injury model, the mean neointimal area was significantly less in the mid-dose group than in the placebo and lowdose groups (control:0.35±0.04 mm, low-dose:0.23±0.04 mm and mid-dose:0.19±0.04 mm, p<0.05), and the mean ratio of the neointima to medial areas were significantly less in the mid-dose group than in the placebo and low dose group (placebo:3.02±0.34, low-dose:2.24±0.54 and mid-dose:1.47±0.31, p<0.05). The labeling index of the PCNA was significantly less in the mid-dose group than in the placebo and low-dose groups (control:71±9, low-dose:73±9, mid-dose:54±14 and high-dose:53±9, p<0.05). Conclusion:The FTI inhibits SMC proliferation in a dose dependent manner. The prolonged oral administration of FTI, for 14 days, is effective in reducing neointimal hyperplasia in the rat carotid balloon injury model. (Korean Circulation J 2003; 33 (4):311-320)

4P-1152 Green tea catechins inhibit neointima hyperplasia in a rat carotid artery balloon injury model through manipulationof matrix metalloproteinase activity

4P-1152 Green tea catechins inhibit neointima hyperplasia in a rat carotid artery balloon injury model through manipulationof matrix metalloproteinase activity

Local expression of C-type natriuretic peptide suppresses inflammation, eliminates shear stress-induced thrombosis, and prevents neointima formation through enhanced nitric oxide production in rabbit injured carotid arteries.

We previously observed that adenovirus-mediated expression of C-type natriuretic peptide (CNP) markedly inhibits neointima formation after balloon injury in rat carotid arteries, suggesting that CNP has multiple effects over its modest inhibitory effect on cellular proliferation. We hypothesized that local expression of CNP might have antithrombotic and antiinflammatory effects. Balloon-injured rabbit carotid arteries were infected with an adenovirus expressing human CNP (AdCNP), human tissue factor pathway inhibitor (AdTFPI), or bacterial beta-galactosidase (AdLacZ) or infused with saline. Seven days later, shear stress-induced thrombosis was evaluated by cyclic flow variation (CFV), reflecting recurrent cycles of thrombus formation and dislodgment. CFV was observed in all AdLacZ-infected and saline-infused arteries but not in arteries infected with AdCNP or AdTFPI even in the presence of epinephrine. Injury increased the expressions of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) and infiltration of macrophages. However, these effects were markedly reduced in AdCNP-treated arteries but not in AdTFPI-infected ones. In AdCNP-infected arteries, injury-induced expression of inducible NO synthase (iNOS) was enhanced, leading to increased NO generation. Interestingly, when the enhanced NO production was inhibited, neither inhibitory effect was observed, and suppression of neointima formation by CNP was canceled. Our study demonstrates that overexpression of CNP shows antithrombotic and antiinflammatory effects and reduces neointima formation mainly through enhanced NO production.

Systemic alpha 1A-adrenoceptor antagonist inhibits neointimal growth after balloon injury of rat carotid artery.

Previous in vitro and in vivo studies have shown that norepinephrine, acting through alpha(1A)-adrenoceptors, stimulates hypertrophy, proliferation, and migration of vascular smooth muscle cells and adventitial fibroblasts and may contribute to neointimal growth, lumen loss, and inward remodeling caused by iatrogenic wall injury and vascular disease. Our present aim was to determine whether intravenous administration of the alpha(1A)-adrenoceptor antagonist KMD-3213, at dosages without systemic hemodynamic effects, inhibits wall growth after injury. Inhibition of alpha(1A)-adrenoceptors with 12.8 and 32 microg/kg KMD-3213 had no effect on arterial pressure or renal and hindquarter resistances in anesthetized rats. A second group then received carotid balloon injury and continuous intravenous KMD-3213 at 4 and 10 microg x kg(-1) x h(-1) for 2 wk. Mean, systolic, and diastolic arterial pressures and heart rate of conscious unrestrained rats were unaffected. KMD-3213 reduced neointima growth by approximately 30 and 46% at the two doses (P < 0.01). These data support the novel hypothesis that a direct alpha(1A)-adrenoceptor-dependent trophic action of catecholamines is augmented by injury and may contribute significantly to hypertrophic vascular disease.

Regulation of syndecan-4 expression with mechanical stress during the development of angioplasty-induced intimal thickening

In this study, we postulated that acute mechanical strain of the arterial wall induces changes in syndecan-4 expression that in turn may modulate cellular events, which promote neointima formation. Correlative in vivo and in vitro studies were performed to determine whether: (1) balloon injury of the rat carotid artery induces spatially and temporally regulated changes in syndecan-4 messenger RNA (mRNA) and protein expression; (2) the application of biaxial mechanical strain to cultured vascular wall cells regulates the expression of syndecan-4 mRNA and protein and its cell surface distribution and surface shedding; and (3) shed syndecan-4 directly effects cell motility behavior. We observed that syndecan-4 mRNA and protein expression were regulated in a temporally and spatially specific manner, such that initial expression of syndecan-4 was localized to adventitial cells followed by later expression in the neointima. Notably, in vitro stimulation of isolated adventitial fibroblasts with a mechanical stretch protocol that was designed to mimic in vivo balloon injury produced a rapid increase in syndecan-4 (4.3-fold increase; P < .001) that was accompanied both by enhanced protein shedding and by the disassembly of focal adhesions. Also significant was that shed syndecan-4, induced with mechanically conditioned media, appeared to be responsible for a chemotactic response of adventitial fibroblasts ( P = .005). These results indicate that mechanical strain tightly regulates syndecan-4 expression, which may, in part, provide a mechanism for the activation of myofibroblast migration from the tunica adventitia into the neointima. (J Vasc Surg 2002;36:361-70.)

Topical application of β-radiation to reduce intimal hyperplasia after carotid artery balloon injury in rabbit: A possible application for brachytherapy in vascular surgery

Purpose: Endovascular brachytherapy for the prevention of intimal hyperplasia (IH) and restenosis after balloon/stent angioplasty has proven effective both in animal preparations and clinical trials. A variety of β-emitting isotopes and catheter-based devices have been developed for the delivery of low-dose radiation in clinical coronary and peripheral trials. No platform, however, has yet been developed for brachytherapy in concert with vascular surgical operations. The purpose of this study was to evaluate the vascular histopathologic response following balloon injury to rabbit carotid arteries with and without topically applied low-dose β-radiation. Methods: The β-emitting isotope strontium-90 (Sr-90) was conjugated onto the matrix of polypropylene (PLYP) mesh. Rabbit carotid arteries were balloon-injured with a #2 embolectomy catheter. Six carotid arteries were wrapped with nonradioactive PLYP mesh (controls) and Sr-90 (∼90 μCi) PLYP mesh in order to deliver low-dose radiation to the vessel wall from the external (adventitial) surface. Tissue was harvested at 6 weeks and processed for histologic examination. Results: There was consistent blockade of fibrocellular neointima formation with virtually no neointima present in all treated segments, compared to moderate neointima formation in controls. Medial thinning and smooth muscle cell (SMC) necrosis were also associated with topical brachytherapy. Conclusion: β-Radiation applied by an externally wrapped PLYP mesh labeled with Sr-90 markedly suppressed neointima formation in an animal vascular surgical injury model. Further studies, however, are necessary to determine a suitable isotope and dosage for clinical application.

Upregulation of Nox-based NAD(P)H oxidases in restenosis after carotid injury.

Restenosis, a frequent complication of coronary angioplasty, is associated with increased superoxide (O2*(-)) production. Although the molecular identity of the responsible oxidase is unclear, an NAD(P)H oxidase appears to be involved. In smooth muscle, p22phox and 2 homologues of gp91phox, nox1 and nox4, are expressed, whereas fibroblasts contain gp91phox. To begin investigating the possibility that these oxidase components might contribute to the increased O2*(-) that accompanies neointimal formation, we measured their expression after balloon injury of the rat carotid artery. The increase in O2*(-) production 3 to 15 days after surgery was not due to inflammatory cell infiltration but appeared to be derived from medial and neointimal smooth muscle cells and adventitial fibroblasts. Nox1 and p22phox mRNAs were increased 2.7- and 3.6-fold, respectively, at day 3 after injury and remained elevated for 15 days. gp91Phox was increased 7 to 15 days after injury, and nox4 expression was increased 2-fold, but only at day 15 after surgery. These results confirm and extend our previous in vitro data and suggest that in the vasculature, the nox-based NAD(P)H oxidases serve different functions. This dynamic regulation of oxidase components may be critical to smooth muscle phenotypic modulation in restenosis and atherosclerosis.

Neointimal hyperplasia in balloon-injured rat carotid arteries: The influence of hyperhomocysteinemia

Purpose: Hyperhomocyst(e)inemia (hH[e]) is a risk factor for atherosclerosis. Neointimal hyperplasia (NH) after vessel injury can contribute to atherosclerosis. In this study, we investigated the effects of hH(e) on NH formation after arterial balloon injury in rats. Methods: Lewis rats that were given a hH(e)-inducing (high methionine, low folate) or normal diet for 150 days underwent common carotid artery (CCA) balloon injury. Two and 4 weeks after injury, CCAs were formalin perfusion-fixed, sectioned, and stained for elastin. Neointimal index (NI, percent lumen occlusion) and neointima (N) and media (M) area were measured by using computer-interfaced microscopy. Results: Plasma homocyst(e)ine (H[e]) levels were elevated in rats given the study diet compared with rats given the normal diet at days 40 and 90 (69 ± 8 and 73 ± 9 μmol/L vs 4 ± 0.4 and 4 ± 0.6 μmol/L, P <.001). After balloon injury, the CCA NI and N/M ratio, but not the M area, were increased by hH(e) compared with normal plasma H(e) (2 weeks [n = 6,7]: NI = 7.3 ± 1.7 vs 2.9 ± 0.7, P =.002, and N/M = 0.31 ± 0.08 vs 0.08 ± 0.02, P <.001; 4 weeks [n = 4,7]: NI = 13.1 ± 2.2 vs 6.3 ± 1.3, P =.002, and N/M = 0.36 ± 0.08 vs 0.17 ± 0.03, P <.001). Conclusion: hH(e) accelerates NH in a rat CCA balloon-injury model. The effect of hH(e) on NH may contribute to increased atherosclerosis in humans with hH(e). (J Vasc Surg 2002;35:158-65.)

Nox Response to Injury

Szocs and colleagues,1 in this issue of Arteriosclerosis, Thrombosis and Vascular Biology , have added to our understanding of the role of NAD(P)H oxidase and reactive oxygen species in vascular injury. The study was undertaken by using the rat carotid artery balloon injury model, a well-characterized model of neointimal formation. In this model, vascular balloon injury leads to medial smooth muscle cell proliferation and migration across the internal elastic lamina to form the neointima.2,3 Adventitial myofibroblasts may also migrate across the media and into the neointima within the first week after balloon injury.4 The processes of proliferation and migration of smooth muscle cells and fibroblasts in vitro are critically dependent on the production of reactive oxygen species.5–8 Moreover, several reports suggest that vascular production of reactive oxygen species increases rapidly after balloon injury, and that treatment with nonspecific antioxidant regimens can inhibit experimental neointimal formation.9–14 Although the relevance of the rat balloon injury model to human post-angioplasty restenosis is questionable,15 the well-defined kinetics of neointimal formation in this model provide a unique opportunity to explore the cellular and enzymatic sources of reactive oxygen species during vascular cell proliferation and migration in response to injury in vivo. See page 21 Recent studies indicate that nonphagocytic NAD(P)H oxidases are major sources of reactive oxygen species in the vascular wall. Like the phagocytic respiratory burst NADPH oxidase, non-phagocytic NAD(P)H oxidases reduce molecular oxygen to generate superoxide, which is in turn converted to hydrogen peroxide. However, unlike the phagocytic NADPH oxidase, the NAD(P)H oxidases present in blood vessels are constitutively active, producing relatively low levels of reactive oxygen …

Intraperitoneal infusion of homocysteine increases intimal hyperplasia in balloon-injured rat carotid arteries

Hyperhomocysteinemia is a significant risk factor in atherosclerosis and thrombosis. However, its role in the development of intimal hyperplasia after arterial reconstructive procedures remains uncertain. We therefore studied the effect of homocysteine on intimal hyperplasia in a rat model of carotid artery balloon injury. Twenty-four Spraque–Dawley rats were divided into three groups: control (saline infusion), and low dose (0.14 mg/day) and high dose (0.71 mg/day) homocysteine delivered continuously via osmotic pumps implanted intraperitoneally. All animals underwent left common carotid artery balloon denudation with sacrifice after 14 days. Plasma homocysteine levels, intimal hyperplasia, and cell proliferation of rat carotid arteries were determined. In vitro rat smooth muscle cell (SMC) proliferation with homocysteine treatment was also performed. Plasma homocysteine levels at sacrifice were 1.80±0.35, 2.65±0.05 and 3.50±0.22 μM in three groups, respectively. Intimal hyperplasia developed in all balloon-injured arteries in both control and homocysteine-treated animals. The intimal area and intima/media area ratio were increased by 92% ( P<0.05) and 105% ( P<0.05), respectively, in the high dose-homocysteine-treated animals as compared to the control animals. Homocysteine (high dose) also significantly promoted the intimal cell proliferation (bromodeoxyuridine incorporation) by 2.2-fold as compared to controls. Furthermore, homocysteine treatment in the cell culture study showed a concentration-dependent increase of rat SMC proliferation. These data demonstrate that the continuous intraperitoneal administration of homocysteine significantly increases intimal hyperplasia and SMC proliferation after carotid artery balloon injury in the rat as well as in vitro SMC proliferation. This study suggests that, following arterial reconstructive procedures, elevated plasma homocysteine may increase the complications of clinical restenoses that are associated with intimal hyperplasia.

Differential Cell Cycle Progression Patterns of Infiltrating Leukocytes and Resident Cells After Balloon Injury of the Rat Carotid Artery

The heterogeneous nature of the cell populations involved in vascular repair remains a major hurdle for the assessment of the cellular events that take place in injured arteries. The present experiments were designed to estimate the proportions and cell cycle progression of infiltrating leukocytes versus resident vascular cells after balloon injury of the rat common carotid artery. After tissue disaggregation, cell suspension samples from each artery were analyzed by flow cytometry. Cells were stained with anti-CD45 or anti-alpha-smooth muscle actin antibodies to identify leukocytes and smooth muscle cells, respectively. A day after injury, a 12-fold increase in CD45+ leukocytes was found. Double labeling with CD45 and CD-3, ED-1, or granulocyte markers revealed that most infiltrating cells were monocytes and granulocytes. Approximately 14% of infiltrating leukocytes were found to enter apoptosis at day 1, and 17% entered S phase at day 3. In contrast, the highest proliferation rate of resident alpha-smooth muscle actin-positive cells was observed at day 7 (19%). The present results demonstrate that infiltrating leukocytes and resident vascular smooth muscle cells have dissimilar cell cycle profiles. Furthermore, our study demonstrates the feasibility of using flow cytometry to quantitatively determine the cell types and their relative activation state in injured arteries.