[Zhuyu Pills promote polarization of macrophages toward M2 phenotype to prevent atherosclerosis via PPARγ/NF-κB signaling pathway].

This article aims to investigate the effect of Zhuyu Pills on atherosclerosis and decipher the underlying mechanism. The mouse model of atherosclerosis was induced by a high-fat diet, and the total modeling period was 12 weeks. A total of 47 ApoE~(-/-) mice successfully modeled were randomized into 5 groups, including 10 in the model group, 9 in each of low-, medium-, and high-dose(130.54, 261.08 and 522.16 mg·kg~(-1)·d~(-1), respectively) Zhuyu Pills groups, and 10 in the atorvastatin calcium(10.40 mg·kg~(-1)·d~(-1)) group. In addition, 10 C57BL/6J mice were included as the normal group. The mice in the normal group and model group were administrated with an equal volume of sterile distilled water, and those in other groups with corresponding agents by gavage once a day for 12 weeks. At the end of drug intervention, the levels of total cholesterol(TC), triglyceride(TG), high-density lipoprotein cholesterol(HDL-C), and low-density lipoprotein cholesterol(LDL-C) were measured by the biochemical method. Hematoxylin-eosin(HE) staining was employed to observe the plaque distribution in the aortic region. The serum levels of pro-inflammatory cytokines tumor necrosis factor-α(TNF-α) and interleukin(IL)-6 in M1 macrophages and anti-inflammatory cytokines IL-13 and IL-4 in M2 macrophages were determined by enzyme-linked immunosorbent assay(ELISA). The expression levels of inducible nitric oxide synthase(iNOS) and arginase-1(Arg-1) were examined by immunofluorescence. Real-time fluorescence quantitative polymerase chain reaction(real-time PCR) was employed to measure the mRNA levels of peroxisome proliferator-activated receptor γ(PPARγ), nuclear factor-κB(NF-κB), Arg-1, and iNOS in the aorta. Western blot was employed to determine the protein levels of PPARγ and NF-κB in the aorta. The results showed that compared with the normal group, the modeling elevated the TC, TG, and LDL-C levels, lowered the HDL-C level, caused large area thickening of the aortic intima, elevated the TNF-α and IL-6 levels, lowered the IL-4 and IL-13 levels, down-regulated the mRNA and protein levels of PPARγ and Arg-1, and up-regulated the mRNA and protein levels of iNOS and NF-κB in the aorta(P<0.01). Compared with the model group, low-, medium-, and high-dose Zhuyu Pills and atorvastatin calcium lowered the TC, TG, and LDL-C levels, elevated the HDL-C level, reduced the plaque area in a concentration-dependent manner, lowered the TNF-α and IL-6 levels, elevated the IL-4 and IL-13 levels, up-regulated the mRNA and protein levels of PPARγ and Arg-1, and down-regulated the mRNA and protein levels of NF-κB and iNOS in the aorta(P<0.05 or P<0.01). In conclusion, Zhuyu Pills may play an anti-atherosclerosis role by regulating PPARγ/NF-κB signaling pathway, inhibiting the polarization of macrophages toward the M1 phenotype, promoting the polarization of macrophages toward the M2 phenotype, and improving the inflammatory microenvironment of macrophages.

Long-term dietary nitrate supplementation slows the progression of established atherosclerosis in ApoE-/- mice fed a high fat diet.

BACKGROUND AND AIMS: Atherosclerosis is associated with a reduction in the bioavailability and/or bioactivity of endogenous nitric oxide (NO). Dietary nitrate has been proposed as an alternate source when endogenous NO production is reduced. Our previous study demonstrated a protective effect of dietary nitrate on the development of atherosclerosis in the apoE-/- mouse model. However most patients do not present clinically until well after the disease is established. The aims of this study were to determine whether chronic dietary nitrate supplementation can prevent or reverse the progression of atherosclerosis after disease is already established, as well as to explore the underlying mechanism of these cardiovascular protective effects. METHODS: 60 apoE-/- mice were given a high fat diet (HFD) for 12 weeks to allow for the development of atherosclerosis. The mice were then randomized to (i) control group (HFD + 1 mmol/kg/day NaCl), (ii) moderate-dose group (HFD +1 mmol/kg/day NaNO3), or (iii) high-dose group (HFD + 10 mmol/kg/day NaNO3) (20/group) for a further 12 weeks. A group of apoE-/- mice (n = 20) consumed a normal laboratory chow diet for 24 weeks and were included as a reference group. RESULTS: Long-term supplementation with high dose nitrate resulted in ~ 50% reduction in plaque lesion area. Collagen expression and smooth muscle accumulation were increased, and lipid deposition and macrophage accumulation were reduced within atherosclerotic plaques of mice supplemented with high dose nitrate. These changes were associated with an increase in nitrite reductase as well as activation of the endogenous eNOS-NO pathway. CONCLUSION: Long-term high dose nitrate significantly attenuated the progression of established atherosclerosis in the apoE-/- mice fed a HFD. This appears to be mediated in part through a XOR-dependent reduction of nitrate to NO, as well as enhanced eNOS activation via increased Akt and eNOS phosphorylation.

S-1-Propenylcysteine promotes IL-10-induced M2c macrophage polarization through prolonged activation of IL-10R/STAT3 signaling.

Atherosclerosis is a chronic inflammatory disease that may lead to the development of serious cardiovascular diseases. Aged garlic extract (AGE) has been reported to ameliorate atherosclerosis, although its mode of action remains unclear. We found that AGE increased the mRNA or protein levels of arginase1 (Arg1), interleukin-10 (IL-10), CD206 and hypoxia-inducible factor 2α (HIF2α) and decreased that of CD68, HIF1α and inducible nitric oxide synthase in the aorta and spleen of apolipoprotein E knockout mice. We also found that S-1-propenylcysteine (S1PC), a characteristic sulfur compound in AGE, increased the level of IL-10-induced Arg1 mRNA and the extent of M2c-like macrophage polarization in vitro. In addition, S1PC increased the population of M2c-like macrophages, resulting in suppressed the population of M1-like macrophages and decreased lipopolysaccharide-induced production of pro-inflammatory cytokines. These effects were accompanied by prolonged phosphorylation of the IL-10 receptor α (IL-10Rα) and signal transducer and activator of transcription 3 (STAT3) that inhibited the interaction between IL-10Rα and Src homology-2-containing inositol 5'-phosphatase 1 (SHIP1). In addition, administration of S1PC elevated the M2c/M1 macrophage ratio in senescence-accelerated mice. These findings suggest that S1PC may help improve atherosclerosis due to its anti-inflammatory effect to promote IL-10-induced M2c macrophage polarization.

Floralozone protects endothelial function in atherosclerosis by ameliorating NHE1.

Endothelial dysfunction is the pathological basis of atherosclerosis. Incomplete understanding of endothelial dysfunction etiology has impeded drug development for this devastating disease despite the currently available therapies. Floralozone, an aroma flavor, specifically exists in rabbit ear grass. Recently, floralozone has been demonstrated to inhibit atherosclerosis, but the underlying mechanisms are undefined. The present study was undertaken to explore whether floralozone pharmacologically targets endothelial dysfunction and therefore exerts therapeutic effects on atherosclerosis. The Na+/H+ exchanger 1 (NHE1), a channel protein, plays a vital role in atherosclerosis. Whether NHE1 is involved in the therapeutic effects of floralozone on endothelial dysfunction has yet to be further answered. By performing oil red staining and hematoxylin-eosin staining, vascular functional study, and oxidative stress monitoring, we found that floralozone not only reduced the size of carotid atherosclerotic plaque but also prevented endothelial dysfunction in atherosclerotic rats. NHE1 expression was upregulated in the inner membrane of carotid arteries and H2O2-induced primary rat aortic endothelial cells. Inspiringly, floralozone prevented the upregulation of NHE1 in vivo and in vitro. Notably, the administration of NHE1 activator LiCl significantly weakened the protective effect of floralozone on endothelial dysfunction in vivo and in vitro. Our study demonstrated that floralozone exerted its protective effect on endothelial dysfunction in atherosclerosis by ameliorating NHE1. NHE1 maybe a drug target for the treatment of atherosclerosis, and floralozone may be an effective drug to meet the urgent needs of atherosclerosis patients by dampening NHE1.

The benzoate plant metabolite ethyl gallate prevents cellular- and vascular-lipid accumulation in experimental models of atherosclerosis.

Ethyl gallate (EG) is a well-known constituent of medicinal plants, but its effects on atherosclerosis development are not clear. In the present study, the anti-atherosclerosis effects of EG and the underlying mechanisms were explored using macrophage cultures, zebrafish and apolipoprotein (apo) E deficient mice. Treatment of macrophages with EG (20 µM) enhanced cellular cholesterol efflux to HDL, and reduced net lipid accumulation in response to oxidized LDL. Secretion of monocyte chemotactic protein-1 (MCP-1) and interleukin-6 (IL-6) from activated macrophages was also blunted by EG. Fluorescence imaging techniques revealed EG feeding of zebrafish reduced vascular lipid accumulation and inflammatory responses in vivo. Similar results were obtained in apoE-/- mice 6.5 months of age, where plaque lesions and monocyte infiltration into the artery wall were reduced by 70% and 42%, respectively, after just 6 weeks of injections with EG (20 mg/kg). HDL-cholesterol increased 2-fold, serum cholesterol efflux capacity increased by ∼30%, and the levels of MCP-1 and IL-6 were reduced with EG treatment of mice. These results suggest EG impedes early atherosclerosis development by reducing the lipid and macrophage-content of plaque. Underlying mechanisms appeared to involve HDL cholesterol efflux mechanisms and suppression of pro-inflammatory cytokine secretion.

Digestive n-6 Lipid Oxidation, a Key Trigger of Vascular Dysfunction and Atherosclerosis in the Western Diet: Protective Effects of Apple Polyphenols.

SCOPE: A main risk factor of atherosclerosis is a Western diet (WD) rich in n-6 polyunsaturated fatty acids (PUFAs) sensitive to oxidation. Their oxidation can be initiated by heme iron of red meat leading to the formation of 4-hydroxy-2-nonenal (4-HNE), a cytotoxic aldehyde. An increased 4-HNE production is implicated in endothelial dysfunction and atherosclerosis. By contrast, a diet rich in proanthocyanidins reduces oxidative stress and arterial diseases. This study evaluates the effects of a WD on vascular integrity in ApolipoproteinE (ApoE-/- ) mice and the protective capacity of apple extract and puree rich in antioxidant proanthocyanidins. METHODS AND RESULTS: ApoE-/- mice are fed during 12 weeks with a WD with or without n-6 PUFAs. Moreover, two WD + n-6 PUFAs groups are supplemented with apple puree or phenolic extract. An increase in digestive 4-HNE production associated with a rise in plasmatic 4-HNE and oxidized LDL concentrations is reported. Oxidizable n-6 PUFAs consumption is associated with a worsened endothelial dysfunction and atherosclerosis. Interestingly, supplementations with apple polyphenol extract or puree prevented these impairments while reducing oxidative stress. CONCLUSION: n-6 lipid oxidation during digestion may be a key factor of vascular impairments. Nevertheless, an antioxidant strategy can limit 4-HNE formation during digestion and thus durably protect vascular function.

Anti-atherogenic properties of Kgengwe (Citrullus lanatus) seed powder in low-density lipoprotein receptor knockout mice are mediated through beneficial alterations in inflammatory pathways.

Kgengwe fruits are commonly consumed in sub-Saharan countries. Recent reports indicated low coronary artery disease rates in those regions. To investigate anti-atherogenic properties and potential mechanisms of action of Kgengwe seed powder (KSP), male low-density lipoprotein receptor knockout (LDL-r-KO) mice were fed with an atherogenic diet supplemented with (treated, n = 10) or without (controls, n = 10) 10% (w/w) KSP for 20 weeks. Proximate analysis revealed that KSP contained 38% fibre and 15% lipids. KSP supplementation was not associated with significant changes in body weight gain rate, food intake, and plasma lipid levels. However, the average atherosclerotic lesion size in the aortic roots in the KSP-treated group was 58% smaller than that in the control group (0.26 vs 0.11 mm2, p < 0.05). This strong anti-atherogenic effect was associated with significant increases in the average plasma levels of certain cytokines such as IL-10 (6 vs 13 pg/mL, p < 0.05), GM-CSF (0.1 vs 0.2 pg/mL, p < 0.05), and EPO (7 vs 16 pg/mL, p < 0.05) along with reductions in the average levels of plasma MCP-1 (19 vs 14 pg/mL, p < 0.05) and MIP-2 (28 vs 13 pg/mL, p < 0.05). Except for relatively high levels of saturated fatty acids, KSP possesses balanced nutrient compositions with strong anti-atherogenic properties, which may be mediated through alterations in inflammatory pathways. Additional studies warrant confirmation and mechanism(s) of action of such effects. Novelty: Kgengwe seeds prevent atherogenesis in LDL-r-KO mice. Kgengwe seeds increase circulating levels of IL-10 and EPO. No reduction in plasma total cholesterol levels.

Dingxin Recipe IV attenuates atherosclerosis by regulating lipid metabolism through LXR-α/SREBP1 pathway and modulating the gut microbiota in ApoE-/- mice fed with HFD.

ETHNOPHARMACOLOGICAL RELEVANCE: Dingxin Recipe (DXR) is a traditional Chinese medicine formula that has been reported to be effective and safe treatment for cardiovascular diseases, such as arrhythmias, coronary heart disease. Dingxin Recipe IV (DXR IV) was further improved from the DXR according to the traditional use. However, the mechanism of DXR IV in atherosclerosis is unclear. AIM OF THE STUDY: This study aimed to illustrate whether DXR IV improve atherosclerosis through modulating the lipid metabolism and gut microbiota in atherosclerosis mice. MATERIALS AND METHODS: 40 male ApoE-/- mice were fed on HFD for 12 weeks and were then treated with DXR IV (1.8, 0.9, or 0.45 g/kg/d) for another 12 weeks. The decroation of DXR IV contains four traditional Chinese medicines: the dried rhizome of Coptis chinensis Franch. (15.09%), the root of Salvia miltiorrhiza Bunge (28.30%), the seed of Ziziphus jujuba Mill. (37.74%) and the fruiting body of Ganoderma lucidum (Leyss.ex Fr.) Karst. (18.87%). 8 male c57BL/6 mice fed a normal diet served as control group. The atherosclerotic plaque was quantified by oil-red O staining and masson trichrome staining. Mice feces were collected. The gut micobiota were detected by 16S rRNA gene sequencing and fecal metabolites were analyzed by 1H NMR spectroscopy. The effect of DXR IV on blood lipids (TG, TC, LDL-C, HDL-C) was investigated. The lipid metabolism related genes were determined by RT-qPCR and western blotting respectively. RESULTS: DXR IV exerted the anti-atherosclerosis effect by inhibiting the excessive cholesterol deposition in aorta and regulating the level of TG, TC, LDL-C and HDL-C. The composition of gut microbiota was changed. Interestingly, the relative abundance of Muribaculaceae and Ruminococcaceae increased after DXR IV administration, whereas the abundance of Erysipelotrichaceae decreased, which have been beneficial to lipid metabolism. Nine potential metabolic biomarkers, including acetate, butyrate, propionate, alanine, succinate, valerate, xylose, choline, glutamate, were identified, which were related to fatty acid metabolism. Further, the pathway of fatty acid was detected by the RT-qPCR and western blotting. Compared with model group, the level of LXR-α and SREBP1 decreased significantly in DXR IV group while LXR-ß, SREBP2 showed no statistical significance. It indicated that DXR IV modulated lipid metabolism by LXR-α/SREBP1 but not LXRß and SREBP2. CONCLUSIONS: DXR IV exhibits potential anti-atherosclerosis effect, which is closely related to lipid metabolism and the gut microbiota. This study may provide novel insights into the mechanism of DXR IV on atherosclerosis and a basis for promising clinical usage.

Assessment of Cardiovascular Safety of Anti-Osteoporosis Drugs.

The incidence of osteoporosis and cardiovascular disease increases with age, and there are potentially shared mechanistic associations between the two conditions. It is therefore highly relevant to understand the cardiovascular implications of osteoporosis medications. These are presented in this narrative review. Calcium supplementation could theoretically cause atheroma formation via calcium deposition, and in one study was found to be associated with myocardial infarction, but this has not been replicated. Vitamin D supplementation has been extensively investigated for cardiac benefit, but no consistent effect has been found. Despite findings in the early 21st century that menopausal hormone therapy was associated with coronary artery disease and venous thromboembolism (VTE), this therapy is now thought to be potentially safe (from a cardiac perspective) if started within the first 10 years of the menopause. Selective estrogen receptor modulators (SERMs) are associated with increased risk of VTE and may be related to fatal strokes (a subset of total strokes). Bisphosphonates could theoretically provide protection against atheroma. However, data from randomised trials and observational studies have neither robustly supported this nor consistently demonstrated the potential association with atrial fibrillation. Denosumab does not appear to be associated with cardiovascular disease and, although parathyroid hormone analogues are associated with palpitations and dizziness, no association with a defined cardiovascular pathology has been demonstrated. Finally, romosozumab has been shown to have a possible cardiovascular signal, and therefore post-market surveillance of this therapy will be vital.

The Impact of Dietary Supplementation of Whole Foods and Polyphenols on Atherosclerosis.

The purpose of this review is to highlight current research on the benefits of supplementation with foods with a diverse polyphenol composition, including fruits, vegetables, nuts, grains, oils, spices, and teas in blunting atherosclerosis. We searched PubMed for publications utilizing whole food or polyphenols prepared from whole foods in Apolipoprotein E (ApoE) or Low-Density Lipoprotein Receptor (LDLR) knockout mice, and identified 73 studies in which plaque was measured. The majority of the studies reported a reduction in plaque. Nine interventions showed no effect, while three using Agaricus blazei mushroom, HYJA-ri-4 rice variety, and safrole-2', 3'-oxide (SFO) increased plaque. The mechanisms by which atherosclerosis was reduced include improved lipid profile, antioxidant status, and cholesterol clearance, and reduced inflammation. Importantly, not all dietary interventions that reduce plaque showed an improvement in lipid profile. Additionally, we found that, out of 73 studies, only 9 used female mice and only 6 compared both sexes. Only one study compared the two models (LDLR vs. ApoE), showing that the treatment worked in one but not the other. Not all supplementations work in both male and female animals, suggesting that increasing the variety of foods with different polyphenol compositions may be more effective in mitigating atherosclerosis.

High-throughput profiling reveals perturbation of endoplasmic reticulum stress-related genes in atherosclerosis induced by high-cholesterol diet and the protective role of vitamin E.

Formation of atherosclerotic plaques, called atherogenesis, is a complex process affected by genetic and environmental factors. It was proposed that endoplasmic reticulum (ER) stress is an important factor in the pathogenesis of atherosclerosis and that vitamin E affects atherosclerotic plaque formation via its antioxidant properties. Here, we investigated ER stress-related molecular mechanisms in high-cholesterol diet (HCD, 2%)-induced atherosclerosis model and the role of vitamin E supplementation in it, beyond its antioxidant properties. The consequences of HCD and vitamin E supplementation were examined by determining protein levels of ER stress markers in aortic tissues. As vitamin E supplementation acts on several unfolded protein response (UPR) factors, it decreased ER stress induced by HCD. To elucidate the associated pathways, gene expression profiling was performed, revealing differentially expressed genes enriched in ER stress-related pathways such as the proteasome and the apoptosis pathways. We further assessed the proteasomal activity impaired by HCD in the aorta and showed that vitamin E reversed it to that of control animals. Overall, the study characterized the effects of HCD and vitamin E on ER stress-related gene expression, revealing the role of proteolytic systems during atherogenesis.

Rapamycin-Loaded Biomimetic Nanoparticles Reverse Vascular Inflammation.

RATIONALE: Through localized delivery of rapamycin via a biomimetic drug delivery system, it is possible to reduce vascular inflammation and thus the progression of vascular disease. OBJECTIVE: Use biomimetic nanoparticles to deliver rapamycin to the vessel wall to reduce inflammation in an in vivo model of atherosclerosis after a short dosing schedule. METHODS AND RESULTS: Biomimetic nanoparticles (leukosomes) were synthesized using membrane proteins purified from activated J774 macrophages. Rapamycin-loaded nanoparticles were characterized using dynamic light scattering and were found to have a diameter of 108±2.3 nm, a surface charge of -15.4±14.4 mV, and a polydispersity index of 0.11 +/ 0.2. For in vivo studies, ApoE-/- mice were fed a high-fat diet for 12 weeks. Mice were injected with either PBS, free rapamycin (5 mg/kg), or rapamycin-loaded leukosomes (Leuko-Rapa; 5 mg/kg) once daily for 7 days. In mice treated with Leuko-Rapa, flow cytometry of disaggregated aortic tissue revealed fewer proliferating macrophages in the aorta (15.6±9.79 %) compared with untreated mice (30.2±13.34 %) and rapamycin alone (26.8±9.87 %). Decreased macrophage proliferation correlated with decreased levels of MCP (monocyte chemoattractant protein)-1 and IL (interleukin)-b1 in mice treated with Leuko-Rapa. Furthermore, Leuko-Rapa-treated mice also displayed significantly decreased MMP (matrix metalloproteinases) activity in the aorta (mean difference 2554±363.9, P=9.95122×10-6). No significant changes in metabolic or inflammation markers observed in liver metabolic assays. Histological analysis showed improvements in lung morphology, with no alterations in heart, spleen, lung, or liver in Leuko-Rapa-treated mice. CONCLUSIONS: We showed that our biomimetic nanoparticles showed a decrease in proliferating macrophage population that was accompanied by the reduction of key proinflammatory cytokines and changes in plaque morphology. This proof-of-concept showed that our platform was capable of suppressing macrophage proliferation within the aorta after a short dosing schedule (7 days) and with a favorable toxicity profile. This treatment could be a promising intervention for the acute stabilization of late-stage plaques.

Central cardiovascular calcifications: supplementation with Pycnogenol® and Centellicum®: variations over 12 months.

BACKGROUND: This 'concept' registry study evaluated the efficacy of Pycnogenol® and the combination Pycnogenol® and Centella Asiatica (Centellicum®) in controlling over 12 months the increasing number of arterial/cardiac calcifications in subjects with asymptomatic atherosclerosis. METHODS: The study included 3 groups of 30 males with asymptomatic coronary calcifications. Group one was followed with standard management (SM); group 2 used SM and Pycnogenol® (150 mg/day); group 3 used the combination Pycnogenol® (150 mg/day) + Centellicum® (450 mg/day). All subjects took cardioaspirin (Bayer, 100 mg/day). RESULTS: No dropouts, no clinical events were observed in 12 months. The 3 groups had comparable demographic and medical characteristics at baseline. No tolerability problems and no side effects from supplementation were reported. After 12 months, oxidative stress was significantly decreased (P<0.05) in both groups taking Pycnogenol®. The evaluation of the number of calcifications >1 mm indicated a trend in controls using SM towards a progressive increase in calcifications. At 12 months the decrease in the number of calcifications with the combined supplements (Pycnogenol® and Centellicum®) (group 3) was -9.952% and thus significantly better that in the other two groups (P<0.05). Pycnogenol® alone was more effective than SM alone in controlling the variation in calcifications (P<0.05). Considering a 34.88% increase in SM subjects, the total absolute difference between SM (34.8%) and the decrease observed in group 3 (-9.95%) was 44.75% (P<0.02). This indicates that supplementation with the combined supplements blocks the increase in calcified areas and, possibly, in time may decrease the number of calcified spots. CONCLUSIONS: This study shows that there is a significant activity of the complex Pycnogenol®+ Centellicum® in reducing the progressive diffusion of central cardiovascular calcifications-associated with advanced plaques - in a relatively short period of time. Longer studies - focusing also on events - may better evaluate the efficacy of these standardized supplements combination on the evolution of atherosclerosis.

Delayed progression of atherosclerosis and cardiovascular events in asymptomatic patients with atherosclerotic plaques: 3-year prevention with the supplementation with Pycnogenol®+Centellicum®.

BACKGROUND: The aim of this study was the evaluation of the progression of atherosclerosis and the occurrence of cardiovascular events in asymptomatic patients with atherosclerotic plaques (Class IV and V) and arterial wall atherosclerotic lesions and intima-media thickening (IMT). METHODS: Progression of atherosclerotic lesions, oxidative stress and IMT were measured in a 3-year concept, pilot registry study. All subjects were followed with standard management (SM) - including diet and exercise - to control cardiovascular risk factors.The target measurements were: the rate of progression of the atherosclerotic lesions (the passage of subjects from one atherosclerotic class to the next class); the occurrence of "hard" cardiovascular events (i.e. myocardial infarction or strokes; angina was not considered a "hard" event). The study included 3 groups: 1) SM): 2) subjects using cardioaspirin (100 mg/day) and SM; 3) subjects following SM, taking cardioaspirin and supplemented with Pycnogenol® (150 mg/day)+Centellicum® (450 mg/day). RESULTS: The groups were comparable for age and baseline evaluations. 54 subjects completed the 3 year study with standard management only, 74 with aspirin and 56 with aspirin and Pycnogenol®+Centellicum®. The BMI of all subjects was <26. No side effects and no tolerability problems were observed with the supplements. Progression was defined by the passage of the atherosclerotic lesions from one class to the next more advanced class. Progression in the supplement group was observed in 5.3% of the subjects in comparison with a progression >20% in the other groups (P<0.05). In comparison with the SM group and the cardioaspirin group the rate of 'hard' cardiovascular events, requiring hospital admissions were <4% with the combined supplement in comparison with a value >12% in the other two groups (22.22% event rate in the SM group). The reduction produced by the aspirin only was significantly lower (P<0.05) in comparison with supplemented patients. Antiplatelet management appears to reduce a significant number of events (P<0.05) without a real effect on progression of atherosclerotic lesions. The additional parameters of carotid IMT and oxidative stress were also lower (P<0.05) with the supplements. CONCLUSIONS: In conclusion, this study indicates that the combined supplementation with Pycnogenol®+Centelicum® appears to control both the progression of atherosclerosis and the occurrence of cardiovascular events in this 3 year study. Larger studies, in a wider population with more complex and less standardized conditions may be needed.

Pycnogenol®+Centellicum®, post-stent evaluation: prevention of neointima and plaque re-growth.

BACKGROUND: The aim of this study was to evaluate the regrowth and progression of within-stent neointima after stenting as a model of accelerated atherosclerosis and the potential effects of the combination Pycnogenol® and Centellicum® in 12 months' follow-up. METHODS: Progression was defined as the passage from one arterial risk class to next, more advanced risk class in 12 months of follow-up. Each class corresponds to a different risk of cardiovascular events and progression. Three management groups were formed, treated with either standard management (SM), Pycnogenol® 150 mg/day, or a combination of Pycnogenol® 150 mg/day and Centellicum® 450 mg/day. RESULTS: No side effects or tolerability problems were observed. 82 subjects with stented arteries in class 2 were evaluated for the passage into class 3 over 12 months. This group included 82 subjects; there were no dropouts. The management subgroups were comparable at baseline. At 12 months 66.7% of subjects in the SM subgroup progressed to class 3, versus 10.7% in the Pycnogenol® group; progression was seen in 6.7% (P<0.05) of subjects supplemented with the combination. In the second section of the registry study (78 subjects with stented arteries in class 3) we evaluated the percentage of patients passing into class 4. At 12 months 53.6% of subjects using the SM progressed versus 26.9% in the subgroup using Pycnogenol® (P<0.05) and 11.5% in the Pycnogenol®+Centellicum® group (P<0.05). Across all 160 subjects in the three management groups, progression of the stented artery at 12 months was seen in 59.6% of subjects in the SM group versus 18.5% (P<0.05) in the group managed with Pycnogenol® only. The Pycnogenol®+Centellicum® combination further decreased progression down to 8.9% (P<0.05). Oxidative stress was significantly reduced (P<0.05) in the two supplement groups. CONCLUSIONS: In conclusion, the combination Pycnogenol®+Centellicum® appears to reduce the rate of progression of the neointima after stenting.

An omega-3 fatty acid plasma index ≥4% prevents progression of coronary artery plaque in patients with coronary artery disease on statin treatment.

BACKGROUND AND AIMS: Higher blood levels of the omega-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been associated with fewer cardiovascular events and lower mortality in prospective studies. Our aim was to determine a target level of EPA and DHA to prevent progression of coronary artery plaque. METHODS: 218 subjects with stable coronary artery disease on statins were randomized to high-dose EPA and DHA (3.36 g daily) or no omega-3 for 30 months. Coronary plaque volume was measured by coronary computed tomographic angiography. Plasma phospholipid levels of EPA, DHA and total fatty acids were measured by gas chromatography mass spectrometry. The omega-3 fatty acid index was calculated as EPA+DHA/total fatty acid. RESULTS: Mean (SD) age was 62.9 (7.8) years; mean (SD) LDL-C level 78.6 (27.3) mg/dL and median triglyceride level 122 mg/dL. Subjects assigned to EPA and DHA had increased plasma EPA and DHA levels variably from 1.85% to 13.02%. Plasma omega-3 fatty acid index ≥4% prevented progression of fibrous, noncalcified, calcified and total plaque in nondiabetic subjects whereas those in the lowest quartile (<3.43%) had significant progression of fibrous, calcified and total plaque. No difference was observed in diabetic subjects. CONCLUSIONS: EPA and DHA added to statins prevented coronary plaque progression in nondiabetic subjects with mean LDL-C <80 mg/dL, when an omega-3 index ≥4% was achieved. Low omega-3 index <3.43% identified nondiabetic subjects at risk of coronary plaque progression despite statin therapy. These findings highlight the importance of measuring plasma levels of omega-3 fatty acids early and at trial conclusion. Targeting an omega-3 index ≥4% maximizes cardiovascular benefit.

Zhixiong Capsule (ZXC), a traditional Chinese patent medicine, prevents atherosclerotic plaque formation in rabbit carotid artery and the related mechanism investigation based on network pharmacology and biological research.

BACKGROUND AND AIMS: Chinese patent medicine Zhixiong Capsule (ZXC) has been used in clinical treatment against blood stasis-induced dizziness and headache for many years in China. HYPOTHESIS/PURPOSE: Recent clinical observations demonstrated a good efficacy of ZXC against atherosclerotic plaque formation in carotid arteries. The aims of this study were to verify the plaque-preventing efficacy of ZXC in animals and to investigate the underlying mechanisms. STUDY DESIGN/METHODS: ZXC (185 mg/kg and 370 mg/kg) was administrated to rabbits which received collar implantation accompanied with high fat diet administration (12 days). The blood-dissolved components of ZXC were identified by an UPLC-QTOF-MS method. The key components and targets of ZXC were then predicted based on network pharmacology analysis and biological investigations. RESULTS: Compared with vehicle control group, ZXC administration (185 mg/kg) significantly prevented plaque formation and attenuated intima thickening in the collar-implanted carotid arteries, markedly decreased blood lipid level, and increased plasma IL-4 level in rabbits. A total of 23 blood-dissolved components were identified. Four ingredients (namely, kaempferol, daidzein, puerarin, miltirone) along with leech, and three targets (namely, JUN, FOS and TP53) were recognized to play important roles for ZXC bioactivity. CONCLUSION: It could be concluded that ZXC could be applied to prevent atherosclerotic plaque formation and intimal thickening in carotid arteries at the current clinical dose.

Gut Colonization with Methanogenic Archaea Lowers Plasma Trimethylamine N-oxide Concentrations in Apolipoprotein e-/- Mice.

A mechanistic link between trimethylamine N-oxide (TMAO) and atherogenesis has been reported. TMAO is generated enzymatically in the liver by the oxidation of trimethylamine (TMA), which is produced from dietary choline, carnitine and betaine by gut bacteria. It is known that certain members of methanogenic archaea (MA) could use methylated amines such as trimethylamine as growth substrates in culture. Therefore, we investigated the efficacy of gut colonization with MA on lowering plasma TMAO concentrations. Initially, we screened for the colonization potential and TMAO lowering efficacy of five MA species in C57BL/6 mice fed with high choline/TMA supplemented diet, and found out that all five species could colonize and lover plasma TMAO levels, although with different efficacies. The top performing MA, Methanobrevibacter smithii, Methanosarcina mazei, and Methanomicrococcus blatticola, were transplanted into Apoe-/- mice fed with high choline/TMA supplemented diet. Similar to C57BL/6 mice, following initial provision of the MA, there was progressive attrition of MA within fecal microbial communities post-transplantation during the initial 3 weeks of the study. In general, plasma TMAO concentrations decreased significantly in proportion to the level of MA colonization. In a subsequent experiment, use of antibiotics and repeated transplantation of Apoe-/- mice with M. smithii, led to high engraftment levels during the 9 weeks of the study, resulting in a sustained and significantly lower average plasma TMAO concentrations (18.2 ± 19.6 µM) compared to that in mock-transplanted control mice (120.8 ± 13.0 µM, p < 0.001). Compared to control Apoe-/- mice, M. smithii-colonized mice also had a 44% decrease in aortic plaque area (8,570 µm [95% CI 19587-151821] vs. 15,369 µm [95% CI [70058-237321], p = 0.34), and 52% reduction in the fat content in the atherosclerotic plaques (14,283 µm [95% CI 4,957-23,608] vs. 29,870 µm [95% CI 18,074-41,666], p = 0.10), although these differences did not reach significance. Gut colonization with M. smithii leads to a significant reduction in plasma TMAO levels, with a tendency for attenuation of atherosclerosis burden in Apoe-/- mice. The anti-atherogenic potential of MA should be further tested in adequately powered experiments.