TET2 Protects Against Vascular Smooth Muscle Cell Apoptosis and Intimal Thickening in Transplant Vasculopathy.

BACKGROUND: Coronary allograft vasculopathy (CAV) is a devastating sequela of heart transplant in which arterial intimal thickening limits coronary blood flow. There are currently no targeted therapies to prevent or reduce this pathology that leads to transplant failure. Vascular smooth muscle cell (VSMC) phenotypic plasticity is critical in CAV neointima formation. TET2 (TET methylcytosine dioxygenase 2) is an important epigenetic regulator of VSMC phenotype, but the role of TET2 in the progression of CAV is unknown. METHODS: We assessed TET2 expression and activity in human CAV and renal transplant samples. We also used the sex-mismatched murine aortic graft model of graft arteriopathy (GA) in wild-type and inducible smooth muscle-specific Tet2 knockout mice; and in vitro studies in murine and human VSMCs using knockdown, overexpression, and transcriptomic approaches to assess the role of TET2 in VSMC responses to IFNγ (interferon γ), a cytokine elaborated by T cells that drives CAV progression. RESULTS: In the present study, we found that TET2 expression and activity are negatively regulated in human CAV and renal transplant samples and in the murine aortic graft model of GA. IFNγ was sufficient to repress TET2 and induce an activated VSMC phenotype in vitro. TET2 depletion mimicked the effects of IFNγ, and TET2 overexpression rescued IFNγ-induced dedifferentiation. VSMC-specific TET2 depletion in aortic grafts, and in the femoral wire restenosis model, resulted in increased VSMC apoptosis and medial thinning. In GA, this apoptosis was tightly correlated with proliferation. In vitro, TET2-deficient VSMCs undergo apoptosis more readily in response to IFNγ and expressed a signature of increased susceptibility to extrinsic apoptotic signaling. Enhancing TET2 enzymatic activity with high-dose ascorbic acid rescued the effect of GA-induced VSMC apoptosis and intimal thickening in a TET2-dependent manner. CONCLUSIONS: TET2 is repressed in CAV and GA, likely mediated by IFNγ. TET2 serves to protect VSMCs from apoptosis in the context of transplant vasculopathy or IFNγ stimulation. Promoting TET2 activity in vivo with systemic ascorbic acid reduces VSMC apoptosis and intimal thickening. These data suggest that promoting TET2 activity in CAV may be an effective strategy for limiting CAV progression.

The Seed Coat Extract of Black Soybean Decreases Nicotine-Induced Vascular Fiber Degradation by Suppressing Matrix Metalloproteinase 2 Expression.

Abdominal aortic aneurysm (AAA) is a vascular disease characterized by weakening of vascular walls and progressive dilation of the abdominal aorta. Nicotine, the main component of tobacco, is reportedly associated with the development and rupture of AAA. It is desirable to attenuate the destructive effect of nicotine on vascular walls, using dietary food components. However, effective methods for preventing AAA progression using dietary food components remain unestablished. This study focuses on proanthocyanidins, well known for their potent antioxidant activity. We speculated that proanthocyanidins can suppress nicotine-induced weakening of vascular walls. To estimate the effect of black soybean seed coat extract (BSSCE), rich in proanthocyanidins, on nicotine-induced weakening of the aortic wall, mice were divided into four groups: the control diet and distilled water group (named C), BSSCE solution diet and distilled water group (named B), control diet and 0.5 mg/mL nicotine solution group (named CN), and BSSCE solution diet and 0.5 mg/mL nicotine solution group (named BN). Nicotine-induced degradation of elastin and collagen fibers were significantly suppressed in BN group. The positive areas for matrix metalloproteinase (MMP)-2 and oxidative stress in BN group were significantly decreased compared to those in CN group. These results suggest that proanthocyanidins-rich BSSCE can prevent the weakening of the aortic wall via inhibiting MMP-2 upregulation.

The resolution of inflammation through omega-3 fatty acids in atherosclerosis, intimal hyperplasia, and vascular calcification.

Omega-3 fatty acids serve as the substrate for the formation of a group of lipid mediators that mediate the resolution of inflammation. The cardiovascular inflammatory response in atherosclerosis and vascular injury is characterized by a failure in the resolution of inflammation, resulting in a chronic inflammatory response. The proresolving lipid mediator resolvin E1 (RvE1) is formed by enzymatic conversion of the omega-3 fatty acid eicosapentaenoic acid (EPA), and signals resolution of inflammation through its receptor ChemR23. Importantly, the resolution of cardiovascular inflammation is an active, multifactorial process that involves modulation of the immune response, direct actions on the vascular wall, as well as close interactions between macrophages and vascular smooth muscle cells. Promoting anti-atherogenic signalling through the stimulation of endogenous resolution of inflammation pathways may provide a novel therapeutic strategy in cardiovascular prevention.

Fucoidan alleviates high-fat diet-induced dyslipidemia and atherosclerosis in ApoE(shl) mice deficient in apolipoprotein E expression.

Fucoidan, a sulfated polysaccharide extracted from brown seaweeds, possesses many biological activities including anti-inflammatory and antioxidant activities. We aimed to investigate the protective effects of fucoidan on dyslipidemia and atherosclerosis in apolipoprotein E-deficient mice (ApoE(shl) mice) and to elucidate its molecular targets in the liver by using a transcriptomic approach. For 12weeks, ApoE(shl) mice were fed a high-fat diet (HFD) supplemented with either 1% or 5% fucoidan. Fucoidan supplementation significantly reduced tissue weight (liver and white adipose tissue), blood lipid, total cholesterol (TC), triglyceride (TG), non-high-density lipoprotein cholesterol (non-HDL-C) and glucose levels in HFD-fed ApoE(shl) mice but increased plasma lipoprotein lipase (LPL) activity and HDL-C levels. Fucoidan also reduced hepatic steatosis levels (liver size, TC and TG levels, and lipid peroxidation) and increased white adipose tissue LPL activity. DNA microarray analysis and quantitative reverse transcription-polymerase chain reaction demonstrated differential expression of genes encoding proteins involved in lipid metabolism, energy homeostasis and insulin sensitivity, by activating Ppara and inactivating Srebf1. Fucoidan supplementation markedly reduced the thickness of the lipid-rich plaque, lipid peroxidation and foaming macrophage accumulation in the aorta in HFD-fed ApoE(shl) mice. Thus, fucoidan supplementation appears to have anti-dyslipidemic and anti-atherosclerotic effects by inducing LPL activity and inhibiting the effects of inflammation and oxidative stress in HFD-fed ApoE(shl) mice.

Combination of spices and herbal extract restores macrophage foam cell migration and abrogates the athero-inflammatory signalling cascade of atherogenesis.

The trapping of lipid-laden macrophages in the arterial intima is a critical but reversible step in atherogenesis. However, information about possible treatments for this condition is lacking. Here, we hypothesized that combining the polyphenol-rich fractions (PHC) of commonly consumed spices (Allium sativum L (Liliaceae), Zingiber officinale R (Zingiberaceae), Curcuma longa L (Zingiberaceae)) and herbs (Terminalia arjuna (R) W & A (Combretaceae) and Cyperus rotundus L (Cyperaceae)) prevents foam cell formation and atherogenesis. Using an in vitro foam cell formation assay, we found that PHC significantly inhibited lipid-laden macrophage foam cell formation compared to the depleted polyphenol fraction of PHC (F-PHC). We further observed that PHC attenuated the LDL and LPS induced CD36, p-FAK and PPAR-γ protein expression in macrophages and increased their migration. NK-κB-DNA interaction, TNF-α, ROS generation, and MMP9 and MMP2 protein expression were suppressed in PHC-treated macrophages. The anti-atherosclerotic activity of PHC was investigated in a high fat- and cholesterol-fed rabbit model. The inhibition of foam cell deposition within the aortic intima and atheroma formation confirmed the atheroprotective activity of PHC. Therefore, we conclude that the armoury of polyphenols in PHC attenuates the CD36 signalling cascade-mediated foam cell formation, enhances the migration of these cells and prevents atherogenesis.

Tongxinluo inhibits vascular inflammation and neointimal hyperplasia through blockade of the positive feedback loop between miR-155 and TNF-α.

Tongxinluo (TXL), a traditional Chinese medicine, has multiple vasoprotective effects, including anti-inflammation. MicroRNA-155 (miR-155) is involved in vascular inflammation and atherosclerosis. However, a direct relationship between TXL and miR-155 in the development of vascular inflammation and remodeling had not yet been shown. The objective of the present study was to investigate whether TXL exerts an inhibitory effect on the vascular inflammatory response and neointimal hyperplasia by regulating miR-155 expression. Using the carotid artery ligation model in mice, we have shown that TXL dose dependently inhibited neointimal formation and reduced the vascular inflammatory response by inhibiting inflammatory cytokine production and macrophage infiltration. miR-155 was induced by carotid artery ligation, and neointimal hyperplasia was strongly reduced in miR-155(−/−) mice. In contrast, miR-155 overexpression partly reversed the inhibitory effect of TXL on neointimal hyperplasia. In bone marrow-derived macrophages, miR-155 and TNF-α formed a positive feedback loop to promote the inflammatory response, which could be blocked by TXL. Furthermore, TXL increased Akt1 protein expression and phosphorylation in TNF-α-stimulated marrow-derived macrophages, and knockdown of Akt1 abrogated the TXL-induced suppression of miR-155. In conclusion, TXL inhibits the vascular inflammatory response and neointimal hyperplasia induced by carotid artery ligation in mice. Suppression of miR-155 expression mediated by Akt1 and blockade of the feedback loop between miR-155 and TNF-α are important pathways whereby TXL exerts its vasoprotective effects.

Nitric oxide and nitrite-based therapeutic opportunities in intimal hyperplasia.

Vascular intimal hyperplasia (IH) limits the long term efficacy of current surgical and percutaneous therapies for atherosclerotic disease. There are extensive changes in gene expression and cell signaling in response to vascular therapies, including changes in nitric oxide (NO) signaling. NO is well recognized for its vasoregulatory properties and has been investigated as a therapeutic treatment for its vasoprotective abilities. The circulating molecules nitrite (NO(2)(-)) and nitrate (NO(3)(-)), once thought to be stable products of NO metabolism, are now recognized as important circulating reservoirs of NO and represent a complementary source of NO in contrast to the classic L-arginine-NO-synthase pathway. Here we review the background of IH, its relationship with the NO and nitrite/nitrate pathways, and current and future therapeutic opportunities for these molecules.

Chinese red yeast rice attenuates the development of angiotensin II-induced abdominal aortic aneurysm and atherosclerosis.

OBJECTIVE: Abdominal aortic aneurysm (AAA) is a chronic vascular disease characterized by medial degradation and inflammation. No medical approaches have been validated for treating AAA, and therapeutic options are limited to regular surveillance leading to surgical intervention. This study aimed to investigate whether administration of Chinese red yeast rice (Monascus purpureus; RYR) suppressed angiotensin II (AngII)-induced AAA and atherosclerosis. METHODS AND RESULTS: Apolipoprotein E-deficient male mice fed a normal diet were administered either RYR extract (200 mg/kg/day) or vehicle by gavage for 1 week before initiating AngII infusion (1000 ng/kg/min) via subcutaneous osmotic pumps for 28 days. Red yeast rice extract administration significantly suppressed AngII-induced expansion of suprarenal diameter and area (P<.05). Furthermore, RYR extract significantly reduced atherosclerotic lesion areas in both the intima of aortic arches and cross sections of aortic roots (P<.05). These effects were associated with reductions of serum total cholesterol, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, matrix metalloproteinase (MMP) 2 and increases of serum macrophage migration inhibitory factor, but no changes in serum interleukin (IL) 1α, IL-6, monocyte chemoattractant protein 1, MMP-9 and expression of MMP-2 and MMP-9 in aortic walls. CONCLUSIONS: This study demonstrated that RYR extract administration suppressed AngII-induced AAA and atherosclerosis associated with regulating inflammation responses independent of lipid-lowering effects. Red yeast rice may have preventive potential for patients with AAA.

Cynanchum wilfordii ameliorates hypertension and endothelial dysfunction in rats fed with high fat/cholesterol diets.

Hypercholesterolemia increases the incidence of atherosclerosis and its pathologic complications. This study was performed to test the effect of an ethanol extract of Cynanchum wilfordii (ECW) on vascular dysfunction in rats fed with high fat/cholesterol diets (HFCD). Male rats were fed a HFCD consisting of 7.5% cocoa butter and 1.25% cholesterol, with or without 100, 200 mg/day/kg ECW. Rats fed with HFCD increased body weight associated with an increase in plasma low-density lipoprotein (LDL) cholesterol level. Chronic ECW treatment in HFCD-fed rats lessened LDL cholesterol and triglyceride levels as well as elevated high-density lipoprotein (HDL) cholesterol. Chronic ECW treatment recovered the HFCD-induced increase in systolic blood pressure, maintained smooth and soft intima endothelial layers by the decrease of intima-media thickness. ECW significantly recovered the diet-induced decrease in vasorelaxation to acetylcholine, high-dose ECW apparently increased vasorelaxation response to sodium nitroprusside in rats fed with HFCD. ECW clearly restored the HFCD-induced reduction in endothelial nitric oxide (NO) synthase expression and Akt expression levels in aortic tissue, leading to improve endothelial function through an increase in endothelium-derived NO production. Furthermore, treatment of ECW significantly recovered the HFCD-induced decrease in aortic cGMP levels in rats. These findings suggest that ECW ameliorates hypertension and endothelial dysfunction via improvement of NO/cGMP signaling pathway in aortic tissue of rats fed with HFCD, suggesting a vascular protective role for this herb in the treatment and prevention of atherosclerotic vascular disease.

Salviae Miltiorrhizae Radix and Puerariae Lobatae Radix herbal formula mediates anti-atherosclerosis by modulating key atherogenic events both in vascular smooth muscle cells and endothelial cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Salviae Miltiorrhizae Radix (Danshen) and Puerariae Lobatae Radix (Gegen) are principal herbs have long been used in combination for treating cardiovascular disease. AIMS OF STUDY: Danshen and Gegen in the ratio of 7:3 (DGW) have significantly reduced the carotid intimal-media thickening (IMT) in patients in our previous clinical study. In the present study, we have demonstrated the mechanisms on IMT reduction by investigating its key processes on both vascular smooth muscle cell (vSMC) and endothelial cells. MATERIALS AND METHODS: The anti-proliferative effects of DGW on platelet-derived growth factor (PDGF) induced vSMC proliferation were studied by cell proliferation, cell cycle distribution, p-ERK and cyclin D expression level. The anti-migratory effect of DGW was investigated by using transwell apparatus. For human umbilical endothelial cells (HUVEC), the inhibitory effects of DGW on TNF-alpha induced cell adhesion, cell adhesion molecules expression, MCP-1 and IL-6 production were investigated. RESULTS: DGW significantly inhibited A7r5 proliferation and exhibited G1/S cell cycle arrest by suppressing both p-ERK and cyclin D expression. Moreover, DGW showed anti-migratory effect against PDGF-induced A7r5 migration. In addition, DGW inhibited the cell adhesion as well as the expression of ICAM-1 and VCAM-1, the production of MCP-1 but not IL-6 in TNF-α stimulated HUVECs. CONCLUSIONS: Our study provided strong scientific evidence on IMT reduction in patients by modulating the key atherogenic events in both vSMC and endothelial cells.

Smooth muscle calponin: an unconventional CArG-dependent gene that antagonizes neointimal formation.

OBJECTIVE: Smooth muscle calponin (CNN1) contains multiple conserved intronic CArG elements that bind serum response factor and display enhancer activity in vitro. The objectives here were to evaluate these CArG elements for activity in transgenic mice and determine the effect of human CNN1 on injury-induced vascular remodeling. METHODS AND RESULTS: Mice carrying a lacZ reporter under control of intronic CArG elements in the human CNN1 gene failed to show smooth muscle cell (SMC)-restricted activity. However, deletion of the orthologous sequences in mice abolished endogenous Cnn1 promoter activity, suggesting their necessity for in vivo Cnn1 expression. Mice carrying a 38-kb bacterial artificial chromosome (BAC) harboring the human CNN1 gene displayed SMC- restricted expression of the corresponding CNN1 protein, as measured by immunohistochemistry and Western blotting. Extensive BAC recombineering studies revealed the absolute necessity of a single intronic CArG element for correct SMC-restricted expression of human CNN1. Overexpressing human CNN1 suppressed neointimal formation following arterial injury. Mice with an identical BAC carrying mutations in CArG elements that inhibit human CNN1 expression showed outward remodeling and neointimal formation. CONCLUSIONS: A single intronic CArG element is necessary but insufficient for proper CNN1 expression in vivo. CNN1 overexpression antagonizes arterial injury-induced neointimal formation.

EGb761, a Ginkgo biloba extract, is effective against atherosclerosis in vitro, and in a rat model of type 2 diabetes.

BACKGROUND: EGb761, a standardized Ginkgo biloba extract, has antioxidant and antiplatelet aggregation and thus might protect against atherosclerosis. However, molecular and functional properties of EGb761 and its major subcomponents have not been well characterized. We investigated the effect of EGb761 and its major subcomponents (bilobalide, kaemferol, and quercetin) on preventing atherosclerosis in vitro, and in a rat model of type 2 diabetes. METHODS AND RESULTS: EGb761 (100 and 200 mg/kg) or normal saline (control) were administered to Otsuka Long-Evans Tokushima Fatty rats, an obese insulin-resistant rat model, for 6 weeks (from 3 weeks before to 3 weeks after carotid artery injury). Immunohistochemical staining was performed to investigate cell proliferation and apoptosis in the injured arteries. Cell migration, caspase-3 activity and DNA fragmentation, monocyte adhesion, and ICAM-1/VCAM-1 levels were explored in vitro. Treatment with EGb761 dose-dependently reduced intima-media ratio, proliferation of vascular smooth muscle cells (VSMCs) and induced greater apoptosis than the controls. Proliferation and migration of VSMCs in vitro were also decreased by the treatment of EGb761. Glucose homeostasis and circulating adiponectin levels were improved, and plasma hsCRP concentrations were decreased in the treatment groups. Caspase-3 activity and DNA fragmentation increased while monocyte adhesion and ICAM-1/VCAM-1 levels decreased significantly. Among subcomponents of EGb761, kaemferol and quercetin reduced VSMC migration and increased caspase activity. CONCLUSIONS: EGb761 has a protective role in the development of atherosclerosis and is a potential therapeutic agent for preventing atherosclerosis.

Microcalcifications in early intimal lesions of atherosclerotic human coronary arteries.

Although calcium (Ca) precipitation may play a pathogenic role in atherosclerosis, information on temporal patterns of microcalcifications in human coronary arteries, their relation to expression of calcification-regulating proteins, and colocalization with iron (Fe) and zinc (Zn) is scarce. Human coronary arteries were analyzed post mortem with a proton microprobe for element concentrations and stained (immuno)histochemically for morphological and calcification-regulating proteins. Microcalcifications were occasionally observed in preatheroma type I atherosclerotic intimal lesions. Their abundance increased in type II, III, and IV lesions. Moreover, their appearance preceded increased expression of calcification-regulating proteins, such as osteocalcin and bone morphogenetic protein-2. In contrast, their presence coincided with increased expression of uncarboxylated matrix Gla protein (MGP), whereas the content of carboxylated MGP was increased in type III and IV lesions, indicating delayed posttranslational conversion of biologically inactive into active MGP. Ca/phosphorus ratios of the microcalcifications varied from 1.6 to 3.0, including amorphous Ca phosphates. Approximately 75% of microcalcifications colocalized with the accumulation of Fe and Zn. We conclude that Ca microprecipitation occurs in the early stages of atherosclerosis, inferring a pathogenic role in the sequel of events, resulting in overt atherosclerotic lesions. Microcalcifications may be caused by local events triggering the precipitation of Ca rather than by increased expression of calcification-regulating proteins. The high degree of colocalization with Fe and Zn suggests a mutual relationship between these trace elements and early deposition of Ca salts.

Factors associated with early atherosclerosis and arterial calcifications in young subjects with a benign phenotype of obesity.

We assessed (i) the association between early arterial disease and factors linked to adiposity, dietary habits, and family in a young cohort of 151 obese children and adolescents with less than or equal to one cardiovascular (CV) risk factor, (ii) whether in subjects with carotid calcifications there was an imbalance of calcium-phosphorus homeostasis. Measurement included: carotid ultrasound, oral glucose tolerance test, anthropometry, body composition, dietary history, white blood cells count, lipids, uric acid, adiponectin, insulin, C-reactive protein, plasminogen activator inhibitor 1 (PAI-1), 25-hydroxyvitamin D, parathyroid hormone (PTH), calcium and phosphorus. Obese children with carotid artery intima media thickness (cIMT) values >75° percentile (0.55 mm), compared to those with lower cIMT, were more obese, more often pubertal and had higher prevalence of family history of CV disease (CVD) (P < 0.05), higher plasma PAI-1 and uric acid (P < 0.001) and lower adiponectin (P < 0.05) and high-density lipoprotein (HDL) cholesterol levels (P < 0.05). After adjustment for sex, age, puberty, obesity, and insulin levels, only PAI-I remained significantly different between the two groups (10.9 (7.2-29.8) vs. 6.2 (4.3-10.6) ng/ml, P < 0.001). Dietary intake did not affect cIMT values. Eight percent of subjects showed nonatherosclerotic carotid calcifications with patchy pattern. These children had a worse lipid profile (P < 0.05) and higher plasma PTH levels (48.6 ± 21.5 vs 38.5 ± 16.9 pg/ml, P < 0.05) that were inversely associated with 25-hydroxyvitamin D levels (r = 0.245, P < 0.01). Present results suggest that (i) several adiposity-related factors may play a role in promoting the development of early arterial diseases in young subjects with a benign phenotype of obesity, (ii) a PTH rise resulting from a subclinical imbalance in calcium-phosphorus homeostasis may affect the biological process of vascular calcifications.

Salvianolic acid B inhibits low-density lipoprotein oxidation and neointimal hyperplasia in endothelium-denuded hypercholesterolaemic rabbits.

BACKGROUND: Atherosclerosis and restenosis are inflammatory responses involving free radicals and lipid peroxidation and may be prevented/cured by antioxidant-mediated lipid peroxidation inhibition. Salvianolic acid (Sal B), a water-soluble antioxidant obtained from a Chinese medicinal herb, is believed to have multiple preventive and therapeutic effects against human vascular diseases. In this study the in vitro and in vivo inhibitory effects of Sal B on oxidative stress were determined. RESULTS: In human aortic endothelial cells (HAECs), Sal B reduced oxidative stress, inhibited low-density lipoprotein (LDL) oxidation and reduced oxidised LDL-induced cytotoxicity. Sal B inhibited Cu(2+) -induced LDL oxidation in vitro (with a potency 16.3 times that of probucol) and attenuated HAEC-mediated LDL oxidation as well as reactive oxygen species (ROS) production. In cholesterol-fed New Zealand White rabbits (with probucol as positive control), Sal B intake reduced Cu(2+) -induced LDL oxidation, lipid deposition in the thoracic aorta, intimal thickness of the aortic arch and thoracic aorta and neointimal formation in the abdominal aorta. CONCLUSION: The data obtained in this study suggest that Sal B protects HAECs from oxidative injury-mediated cell death via inhibition of ROS production. The antioxidant activity of Sal B may help explain its efficacy in the treatment of vascular diseases.

Total Ginsenosides suppress the neointimal hyperplasia of rat carotid artery induced by balloon injury.

Ginsenosides, the active components found in Panax ginseng, have been reported to inhibit the cardiac hypertrophy in rats. This study aims to observe the potential effect of total ginsenosides (TG) on the hypertrophic vascular diseases. The model of vascular neointimal hyperplasia was established by rubbing the endothelia of the common carotid artery with a balloon in male Sprague Dawley rats. TG (15 mg/kg/day, 45 mg/kg/day), L-arginine (L-arg) 200 mg/kg/day, and NG-nitro-L-arginine-methyl ester (L-NAME) 100 mg/kg/day used with the same dose of L-arg or TG 45 mg/kg/day were given for 7 and 14 consecutive days after surgery. TG and L-arg administrations significantly ameliorated the histopathology of injured carotid artery, which was abolished or blunted by L-NAME, an NOS inhibitor; TG and L-arg could also remarkably reduce the expression of proliferating cell nuclear antigen (PCNA), a proliferation marker of vascular smooth muscle cells(VSMCs), in neointima of the injured artery wall. Further study indicated that balloon injury caused a decreased superoxide dismutase (SOD) activity and an elevated malondialdehyde (MDA) content in plasma, and reduced the cGMP level in the artery wall, which were reversed by TG. It was concluded that TG suppress the rat carotid artery neointimal hyperplasia induced by balloon injury, which may be involved in its anti-oxidative action and enhancing the inhibition effects of NO/cGMP on VSMC proliferation.

Molecular mechanism of green microalgae, Dunaliella salina, involved in attenuating balloon injury-induced neointimal formation.

The pathological mechanism of restenosis is primarily attributed to excessive proliferation of vascular smooth muscle cells (VSMC). The preventive effects of ethanol extract of Dunaliella salina (EDS) on balloon injury-induced neointimal formation were investigated. To explore its molecular mechanism in regulating cell proliferation, we first showed that EDS markedly reduced the human aortic smooth muscle cell proliferation via the inhibition of 5'-bromo-2'-deoxyuridine (BrdU) incorporation at 40 and 80 microg/ml. This was further supported by the G0/G1-phase arrest using a flow cytometric analysis. In an in vivo study, EDS at 40 and 80 microg/ml was previously administered to the Sprague-Dawley rats and found that the thickness of neointima, and the ratio of neointima:media were also reduced. EDS inhibited VSMC proliferation in a dose-dependent manner following stimulation of VSMC cultures with 15 % fetal bovine serum (FBS). Suppressed by EDS were 15 % FBS-stimulated intracellular Raf, phosphorylated extracellular signal-regulated kinases (p-Erk) involved in cell-cycle arrest and proliferating cell nuclear antigen. Phosphorylated focal adhesion kinase (p-FAK) was also suppressed by EDS. Also active caspase-9, caspase-3 and cleaved poly(ADP-ribose) polymerase (PARP) protein expression levels were increased by administration with EDS; the apoptotic pathway may play an important role in the regulatory effects of EDS on cell growth. These observations provide a mechanism of EDS in attenuating cell proliferation, thus as a potential intervention for restenosis.

c-Jun regulates shear- and injury-inducible Egr-1 expression, vein graft stenosis after autologous end-to-side transplantation in rabbits, and intimal hyperplasia in human saphenous veins.

Coronary artery bypass graft failure represents an unsolved problem in interventional cardiology and heart surgery. Late occlusion of autologous saphenous vein bypass grafts is a consequence of neointima formation underpinned by smooth muscle cell (SMC) migration and proliferation. Poor long term patency and the lack of pharmacologic agents that prevent graft failure necessitate effective alternative therapies. Our objective here was to evaluate the effect of targeted inhibition of the bZIP transcription factor c-Jun on intimal hyperplasia in human saphenous veins and vein graft stenosis after autologous end-to-side transplantation. DNAzymes targeting c-Jun attenuated intimal hyperplasia in human saphenous vein explants. Adenovirus-forced c-Jun expression stimulated SMC proliferation, proliferating cell nuclear antigen, and MMP-2 expression. c-Jun DNAzymes abrogated Adeno-c-Jun-inducible SMC growth and wound repair and reduced intimal thickening in jugular veins of New Zealand white rabbits 4 weeks after autologous end-to-side transplantation to carotid arteries. Conversely, in a DNAzyme-free setting, Adeno-c-Jun potentiated neointima formation in the veins compared with Adeno-LacZ. Inducible c-Jun expression is ERK1/2- and JNK-dependent but p38-independent. Injury- and shear-inducible c-Jun controls early growth response-1. These data demonstrate that strategies targeting c-Jun may be useful for the prevention of vein graft stenosis. Control of one important shear-responsive transcription factor by another indicates the existence of transcriptional amplification mechanisms that magnify the vascular response to cell injury or stress through inducible transcriptional networks.

Inhibition of aortic intimal hyperplasia and cell cycle protein and extracellular matrix protein expressions by BuYang HuanWu Decoction.

AIM OF THE STUDY: The inhibitive effect of BuYang HuanWu Decoction (BYHWD) and its major components on vascular intimal hyperplasia and the expressions of cell cycle protein and extracellular matrix protein. MATERIALS AND METHODS: Sprague-Dawley rats were randomly divided into sham-operated, control, alkaloid, glycoside, BYHWD and atorvastatin groups. Rat aorta intima in all groups were injured by insesion of domestic balloon catheter into the aortae except sham-operated rats. Drugs were administrated orally from the second day after vascular injury and continued for 14 days. The injured segments of aortae were collected on the sixteenth day after operation to observe the morphological changes of vascular structure and to examine the expressions of proteins in vascular cells associated with cell cycle including proliferating cell nuclear antigen(PCNA), cyclinD(1) and cyclinE, and extracellular matrix(ECM) proteins including collagen I (Col-I) and fibronectin (FN), further to discover the involved biologically active substances and the potential mechanisms. RESULTS: Alkaloid and glycosid isolated from BYHWD were more effective than BYHWD in the inhibition of intimal hyperplasia and the expressions of PCNA, cyclinD(1), cyclinE, Col-I and FN, suggesting that alkaloid and glycoside may be the main components of BYHWD responsible for the observed inhibition of excessive hyperplasia of vascular intima. CONCLUSIONS: The mechanism associated with the anti-hyperplasia activity of BYHWD and its effective components may be related to the blockage of cell cycles of VSMC, and the inhibition of the ECM protein synthesis, even the increased degradation of ECM proteins.