Veronika A. Myasoedova

Anti-Atherosclerotic Therapy Based on Botanicals

Natural products including botanicals for both therapy of clinical manifestations of atherosclerosis and reduction of atherosclerosis risk factors are topics of recent patents. Only a few recent patents are relevant to the direct antiatherosclerotic therapy leading to regression of atherosclerotic lesions. Earlier, using a cellular model we have developed and patented several anti-atherosclerotic drugs. The AMAR (Atherosclerosis Monitoring and Atherogenicity Reduction) study was designed to estimate the effect of two-year treatment with time-released garlic-based drug Allicor on the progression of carotid atherosclerosis in 196 asymptomatic men aged 40-74 in double-blinded placebo-controlled randomized clinical study. The primary outcome was the rate of atherosclerosis progression, measured by high-resolution B-mode ultrasonography as the increase in carotid intima-media thickness (IMT) of the far wall of common carotid arteries. The mean rate of IMT changes in Allicor-treated group (-0.022±0.007 mm per year) was significantly different (P = 0.002) from the placebo group in which there was a moderate progression of 0.015±0.008 mm at the overall mean baseline IMT of 0.931±0.009 mm. A significant correlation was found between the changes in blood serum atherogenicity (the ability of serum to induce cholesterol accumulation in cultured cells) during the study and the changes in intima-media thickness of common carotid arteries (r = 0.144, P = 0.045). Thus, the results of AMAR study demonstrate that long-term treatment with Allicor has a direct anti-atherosclerotic effect on carotid atherosclerosis and this effect is likely to be due to serum atherogenicity inhibition. The beneficial effects of other botanicals including Inflaminat (calendula, elder and violet), phytoestrogen- rich Karinat (garlic powder, extract of grape seeds, green tea leafs, hop cones, β-carotene, α-tocopherol and ascorbic acid) on atherosclerosis have also been revealed in clinical studies which enforces a view that botanicals might represent promising drugs for anti-atherosclerotic therapy.

Peroxisome proliferator-activated receptor (PPAR) gamma in cardiovascular disorders and cardiovascular surgery

Peroxisome proliferation-activated receptor gamma (PPARγ) is a nuclear receptor regulating transcription of several genes involved mainly in fatty acid and energy metabolism. PPARγ agonists are used as insulin sensitizers for treatment of diabetes. However, according to the results of recent studies, their clinical application can be broadened. Activation of PPARγ has a wide spectrum of biological functions, regulating metabolism, reducing inflammation, influencing the balance of immune cells, inhibiting apoptosis and oxidative stress, and improving endothelial function. These effects appear to be beneficial not only in diabetes and atherosclerosis, but also in a number of other conditions, including cardiovascular surgical interventions. In this review we discuss the role of PPARγ in various conditions associated with cardiovascular risk, including diabetes mellitus, atherosclerosis, and hypertension, and will focus on current applications of PPARγ activators and their therapeutic use. We will also give an overview of the potential use of PPARγ agonists in cardiovascular surgical intervention.

Antiatherogenic effect of grape flavonoids in an ex vivo model

The effects of grape phytoestrogens on cholesterol accumulation were studied in primary culture of human blood monocytes incubated with blood serum from postmenopausal women obtained before and 2, 4, and 6 h after single intake of plant components of grapes. Phytoestrogens from grape seeds, pressed out grapes, and fermented grape ridges prevent cholesterol accumulation in cells and can be regarded as prospective components for the development of natural preparations for the prevention of atherosclerosis in postmenopausal women.

CD68/macrosialin: not just a histochemical marker

CD68 is a heavily glycosylated glycoprotein that is highly expressed in macrophages and other mononuclear phagocytes. Traditionally, CD68 is exploited as a valuable cytochemical marker to immunostain monocyte/macrophages in the histochemical analysis of inflamed tissues, tumor tissues, and other immunohistopathological applications. CD68 alone or in combination with other cell markers of tumor-associated macrophages showed a good predictive value as a prognostic marker of survival in cancer patients. Lowression of CD68 was found in the lymphoid cells, non-hematopoietic cells (fibroblasts, endothelial cells, etc), and tumor cells. Cell-specific CD68 expression and differentiated expression levels are determined by the complex interplay between transcription factors, regulatory transcriptional elements, and epigenetic factors. Human CD68 and its mouse ortholog macrosialin belong to the family of LAMP proteins located in the lysosomal membrane and share many structural similarities such as the presence of the LAMP-like domain. Except for a second LAMP-like domain present in LAMPs, CD68/microsialin has a highly glycosylated mucin-like domain involved in ligand binding. CD68 has been shown to bind oxLDL, phosphatidylserine, apoptotic cells and serve as a receptor for malaria sporozoite in liver infection. CD68 is mainly located in the endosomal/lysosomal compartment but can rapidly shuttle to the cell surface. However, the role of CD68 as a scavenger receptor remains to be confirmed. It seems that CD68 is not involved in binding bacterial/viral pathogens, innate, inflammatory or humoral immune responses, although it may potentially be involved in antigen processing/presentation. CD68 could be functionally important in osteoclasts since its deletion leads to reduced bone resorption capacity. The role of CD68 in atherosclerosis is contradictory.

Mutation C3256T of Mitochondrial Genome in White Blood Cells: Novel Genetic Marker of Atherosclerosis and Coronary Heart Disease

This study was undertaken to examine the association between the level of heteroplasmy for the mutation C3256T in human white blood cells and the extent of carotid atherosclerosis, as well as the presence of coronary heart disease (CHD), the major clinical manifestation of atherosclerosis. Totally, 191 participants (84 men, 107 women) aged 65.0 years (SD 9.4) were recruited in the study; 45 (24%) of them had CHD. High-resolution B-mode ultrasonography of carotids was used to estimate the extent of carotid atherosclerosis by measuring of the carotid intima-media thickness (cIMT). DNA samples were obtained from whole venous blood, and then PCR and pyrosequencing were carried out. On the basis of pyrosequencing data, the levels of C3256T heteroplasmy in DNA samples were calculated. The presence of the mutant allele was detected in all study participants; the level of C3256T heteroplasmy in white blood cells ranged from 5% to 74%. The highly significant relationship between C3256T heteroplasmy level and predisposition to atherosclerosis was revealed. In individuals with low predisposition to atherosclerosis the mean level of C3256T heteroplasmy was 16.8%, as compared to 23.8% in moderately predisposed subjects, and further to 25.2% and 28.3% in significantly and highly predisposed subjects, respectively. The level of C3256T heteroplasmy of mitochondrial genome in human white blood cells is a biomarker of mitochondrial dysfunction and risk factor for atherosclerosis; therefore, it can be used as an informative marker of genetic susceptibility to atherosclerosis, coronary heart disease and myocardial infarction.

Mechanisms of foam cell formation in atherosclerosis

Low-density lipoprotein (LDL) and cholesterol homeostasis in the peripheral blood is maintained by specialized cells, such as macrophages. Macrophages express a variety of scavenger receptors (SR) that interact with lipoproteins, including SR-A1, CD36, and lectin-like oxLDL receptor-1 (LOX-1). These cells also have several cholesterol transporters, including ATP-binding cassette transporter ABCA1, ABCG1, and SR-BI, that are involved in reverse cholesterol transport. Lipids internalized by phagocytosis are transported to late endosomes/lysosomes, where lysosomal acid lipase (LAL) digests cholesteryl esters releasing free cholesterol. Free cholesterol in turn is processed by acetyl-CoA acetyltransferase (ACAT1), an enzyme that transforms cholesterol to cholesteryl esters. The endoplasmic reticulum serves as a depot for maintaining newly synthesized cholesteryl esters that can be processed by neutral cholesterol ester hydrolase (NCEH), which generates free cholesterol that can exit via cholesterol transporters. In atherosclerosis, pro-inflammatory stimuli upregulate expression of scavenger receptors, especially LOX-1, and downregulate expression of cholesterol transporters. ACAT1 is also increased, while NCEH expression is reduced. This results in deposition of free and esterified cholesterol in macrophages and generation of foam cells. Moreover, other cell types, such as endothelial (ECs) and vascular smooth muscle cells (VSMCs), can also become foam cells. In this review, we discuss known pathways of foam cell formation in atherosclerosis.

The impact of interferon-regulatory factors to macrophage differentiation and polarization into M1 and M2

The mononuclear phagocytes control the body homeostasis through the involvement in resolving tissue injury and further wound healing. Indeed, local tissue microenvironmental changes can significantly influence the functional behavior of monocytes and macrophages. Such microenvironmental changes for example occur in an atherosclerotic plaque during all progression stages. In response to exogenous stimuli, macrophages show a great phenotypic plasticity and heterogeneity. Exposure of monocytes to inflammatory or anti-inflammatory conditions also induces predominant differentiation to proinflammatory (M1) or anti-inflammatory (M2) macrophage subsets and phenotype switch between macrophage subsets. The phenotype transition is accompanied with great changes in the macrophage transcriptome and regulatory networks. Interferon-regulatory factors (IRFs) play a key role in hematopoietic development of monocytes, their differentiation to macrophages, and regulating macrophage maturation, phenotypic polarization, phenotypic switch, and function. Of 9 IRFs, at least 3 (IRF-1, IRF-5, and IRF-8) are involved in the commitment of proinflammatory M1 whereas IRF-3 and IRF-4 control M2 polarization. The role of IRF-2 is context-dependent. The IRF impact on macrophage phenotype plasticity and heterogeneity is complex and involves activating and repressive function in triggering transcription of target genes.

Small Dense Low-Density Lipoprotein as Biomarker for Atherosclerotic Diseases

Low-density lipoprotein (LDL) plays a key role in the development and progression of atherosclerosis and cardiovascular disease. LDL consists of several subclasses of particles with different sizes and densities, including large buoyant (lb) and intermediate and small dense (sd) LDLs. It has been well documented that sdLDL has a greater atherogenic potential than that of other LDL subfractions and that sdLDL cholesterol (sdLDL-C) proportion is a better marker for prediction of cardiovascular disease than that of total LDL-C. Circulating sdLDL readily undergoes multiple atherogenic modifications in blood plasma, such as desialylation, glycation, and oxidation, that further increase its atherogenicity. Modified sdLDL is a potent inductor of inflammatory processes associated with cardiovascular disease. Several laboratory methods have been developed for separation of LDL subclasses, and the results obtained by different methods can not be directly compared in most cases. Recently, the development of homogeneous assays facilitated the LDL subfraction analysis making possible large clinical studies evaluating the significance of sdLDL in the development of cardiovascular disease. Further studies are needed to establish guidelines for sdLDL evaluation and correction in clinical practice.

Pluronic block copolymers inhibit low density lipoprotein self-association

Little is known about exogenous inhibitors of low-density lipoprotein (LDL) aggregation. The search for nontoxic and bioavailable inhibitors of LDL aggregation is of interest, especially considering that the suppression of the aggregation of LDL might represent a therapeutic approach. We hypothesized that amphiphilic copolymers of propylene oxide and ethylene oxide, the so-called Pluronic block copolymers, can be used to influence the aggregation of LDL. In this work we used Pluronic® P85, L61 and F68. A comparative study of the effects of Pluronic block copolymers with various hydrophilic–lipophilic properties on the aggregation process of LDL showed that Pluronic copolymers with strong hydrophobic properties (P85 and L61) at concentrations close to or greater than the respective critical concentration of micelle formation inhibited the aggregation process of LDL; however, the “hydrophilic” Pluronic F68 had no effect on the aggregation of LDL at any concentration. Thus, the study demonstrated for the first time that Pluronic® block copolymers inhibit LDL self-association. The possibility of modulating the aggregation of LDL by various Pluronic copolymers can be regarded as a prerequisite in the creation of new types of anti-atherosclerotic drugs.

Anti-Atherosclerotic Effects of a Phytoestrogen-Rich Herbal Preparation in Postmenopausal Women

The risk of cardiovascular disease and atherosclerosis progression is significantly increased after menopause, probably due to the decrease of estrogen levels. The use of hormone replacement therapy (HRT) for prevention of cardiovascular disease in older postmenopausal failed to meet expectations. Phytoestrogens may induce some improvements in climacteric symptoms, but their effect on the progression of atherosclerosis remains unclear. The reduction of cholesterol accumulation at the cellular level should lead to inhibition of the atherosclerotic process in the arterial wall. The inhibition of intracellular lipid deposition with isoflavonoids was suggested as the effective way for the prevention of plaque formation in the arterial wall. The aim of this double-blind, placebo-controlled clinical study was to investigate the effect of an isoflavonoid-rich herbal preparation on atherosclerosis progression in postmenopausal women free of overt cardiovascular disease. One hundred fifty-seven healthy postmenopausal women (age 65 ± 6) were randomized to a 500 mg isoflavonoid-rich herbal preparation containing tannins from grape seeds, green tea leaves, hop cone powder, and garlic powder, or placebo. Conventional cardiovascular risk factors and intima-media thickness of common carotid arteries (cIMT) were evaluated at the baseline and after 12 months of treatment. After 12-months follow-up, total cholesterol decreased by 6.3% in isoflavonoid-rich herbal preparation recipients (p = 0.011) and by 5.2% in placebo recipients (p = 0.020); low density lipoprotein (LDL) cholesterol decreased by 7.6% in isoflavonoid-rich herbal preparation recipients (p = 0.040) and by 5.2% in placebo recipients (non-significant, NS); high density lipoprotein (HDL) cholesterol decreased by 3.4% in isoflavonoid-rich herbal preparation recipients (NS) and by 4.5% in placebo recipients (p = 0.038); triglycerides decreased by 6.0% in isoflavonoid-rich herbal preparation recipients (NS) and by 7.1% in placebo recipients (NS). The differences between lipid changes in the isoflavonoid-rich herbal preparation and placebo recipients did not reach statistical significance (p > 0.05). Nevertheless, the mean cIMT progression was significantly lower in isoflavonoid-rich herbal preparation recipients as compared to the placebo group (6 μm, or <1%, versus 100 μm, or 13%; p < 0.001 for the difference). The growth of existing atherosclerotic plaques in isoflavonoid-rich herbal preparation recipients was inhibited by 1.5-fold (27% versus 41% in the placebo group). The obtained results demonstrate that the use of isoflavonoid-rich herbal preparation in postmenopausal women may suppress the formation of new atherosclerotic lesions and reduce the progression of existing ones, thus promising new drug for anti-atherosclerotic therapy. Nevertheless, further studies are required to confirm these findings.