Circulating Atherogenic Multiple-Modified Low-Density Lipoprotein: Pathophysiology and Clinical Applications

Low-density lipoprotein (LDL) circulating in human bloodstream is the source of lipids that accumulate in arterial intimal cells in atherosclerosis. In-vitro–modified LDL (acetylated, exposed to malondialdehyde, oxidized with transition metal ions, etc.) is atherogenic, that is, it causes accumulation of lipids in cultured cells. We have found that LDL circulating in the atherosclerosis patients’ blood is atherogenic, while LDL from healthy donors is not. Atherogenic LDL was found to be desialylated. Moreover, only the desialylated subfraction of human LDL was atherogenic. Desialylated LDL is generally denser, smaller, and more electronegative than native LDL. Consequently, these LDL types are multiply modified, and according to our observations, desialylation is probably the principal and foremost cause of lipoprotein atherogenicity. It was found that desialylated LDL of coronary atherosclerosis patients was also oxidized. Complex formation further increases LDL atherogenicity, with LDL associates, immune complexes with antibodies recognizing modified LDL and complexes with extracellular matrix components being most atherogenic. We hypothesized that a nonlipid factor might be extracted from the blood serum using a column with immobilized LDL. This treatment not only allowed revealing the nonlipid factor of blood atherogenicity but also opened the prospect for reducing atherogenicity in patients

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

Use of Natural Products for Direct Anti-Atherosclerotic Therapy

Atherosclerosis and vascular disorders, which result from atherosclerosis, represent one of the major problems in the modern medicine and public health. Atherosclerosis is characterized by structural and functional changes of large arteries. The approaches for the treatment of atherosclerosis require at least the prevention of growth of atherosclerotic lesions and reduction in the lipid core mass, which would followed by plaque stabilization. Taken together, these approaches could theoretically result in the regression of arterial lesions. Atherosclerosis develops in the arterial wall and remains asymptomatic until ischemia of distal organs is evident. Therapy of clinical manifestations of atherosclerosis is largely aimed at reducing symptoms or affecting hemodynamic response and often does not affect the cause or course of disease, namely the atherosclerotic lesion itself. Of course, anti-atherosclerotic effects of statins revealed in many prospective clinical trials may be considered; however, statins have never been recognized as the drugs indicated just for direct treatment or prevention of atherosclerosis. They are used predominately in the course of hypolipidemic therapy, and the effects of treatment are estimated by success in reaching the target level of low density lipoprotein (LDL) cholesterol, but not the regression of atherosclerotic lesion or intimamedia thickness. The last should be considered as beneficial effect, which is mainly due to pleiotropic mechanisms of action. Atherosclerosis develops over many years, so anti-atherosclerotic therapy should be a long-term or even lifelong therapy. Tachyphylaxis, long-term toxicity and cost amongst other issues may present problems for the use of conventional medications in a long-term. Drugs based on natural products can be a good alternative

The Interaction of Plasma Sialylated and Desialylated Lipoproteins with Collagen from the Intima and Media of Uninvolved and Atherosclerotic Human Aorta

We have evaluated the binding of sialylated and desialylated lipoproteins to collagen isolated from the proteoglycan and musculoelastic layers of intima and media of uninvolved human aorta and atherosclerotic lesions. Comparing various collagen preparations from the uninvolved intima-media, the binding of sialylated apoB-containing lipoproteins was best to collagen from the intimal PG-rich layer. Binding of sialylated apoB-containing lipoproteins to collagen from this layer of fatty streak and fibroatheroma was 1.4- and 3.1-fold lower, respectively, in comparison with normal intima. Desialylated VLDL versus sialylated one exhibited a greater binding (1.4- to 3.0-fold) to all the collagen preparations examined. Desialylated IDL and LDL showed a higher binding than sialylated ones when collagen from the intimal layers of fibroatheroma was used. Binding of desialylated HDL to collagen from the intimal PG-rich layer of normal tissue, initial lesion, and fatty streak was 1.2- to 2.0-fold higher compared with sialylated HDL

Low Density Lipoprotein-Containing Circulating Immune Complexes : Role in Atherosclerosis and Diagnostic Value

Peer reviewe

Role of Phagocytosis in the Pro-Inflammatory Response in LDL-Induced Foam Cell Formation; a Transcriptome Analysis

Excessive accumulation of lipid inclusions in the arterial wall cells (foam cell formation) caused by modified low-density lipoprotein (LDL) is the earliest and most noticeable manifestation of atherosclerosis. The mechanisms of foam cell formation are not fully understood and can involve altered lipid uptake, impaired lipid metabolism, or both. Recently, we have identified the top 10 master regulators that were involved in the accumulation of cholesterol in cultured macrophages induced by the incubation with modified LDL. It was found that most of the identified master regulators were related to the regulation of the inflammatory immune response, but not to lipid metabolism. A possible explanation for this unexpected result is a stimulation of the phagocytic activity of macrophages by modified LDL particle associates that have a relatively large size. In the current study, we investigated gene regulation in macrophages using transcriptome analysis to test the hypothesis that the primary event occurring upon the interaction of modified LDL and macrophages is the stimulation of phagocytosis, which subsequently triggers the pro-inflammatory immune response. We identified genes that were up- or downregulated following the exposure of cultured cells to modified LDL or latex beads (inert phagocytosis stimulators). Most of the identified master regulators were involved in the innate immune response, and some of them were encoding major pro-inflammatory proteins. The obtained results indicated that pro-inflammatory response to phagocytosis stimulation precedes the accumulation of intracellular lipids and possibly contributes to the formation of foam cells. In this way, the currently recognized hypothesis that the accumulation of lipids triggers the pro-inflammatory response was not confirmed. Comparative analysis of master regulators revealed similarities in the genetic regulation of the interaction of macrophages with naturally occurring LDL and desialylated LDL. Oxidized and desialylated LDL affected a different spectrum of genes than naturally occurring LDL. These observations suggest that desialylation is the most important modification of LDL occurring in vivo. Thus, modified LDL caused the gene regulation characteristic of the stimulation of phagocytosis. Additionally, the knock-down effect of five master regulators, such as IL15, EIF2AK3, F2RL1, TSPYL2, and ANXA1, on intracellular lipid accumulation was tested. We knocked down these genes in primary macrophages derived from human monocytes. The addition of atherogenic naturally occurring LDL caused a significant accumulation of cholesterol in the control cells. The knock-down of the EIF2AK3 and IL15 genes completely prevented cholesterol accumulation in cultured macrophages. The knock-down of the ANXA1 gene caused a further decrease in cholesterol content in cultured macrophages. At the same time, knock-down of F2RL1 and TSPYL2 did not cause an effect. The results obtained allowed us to explain in which way the inflammatory response and the accumulation of cholesterol are related confirming our hypothesis of atherogenesis development based on the following viewpoints: LDL particles undergo atherogenic modifications that, in turn, accompanied by the formation of self-associates; large LDL associates stimulate phagocytosis; as a result of phagocytosis stimulation, pro-inflammatory molecules are secreted; these molecules cause or at least contribute to the accumulation of intracellular cholesterol. Therefore, it became obvious that the primary event in this sequence is not the accumulation of cholesterol but an inflammatory response

Approach to Reduction of Blood Atherogenicity

We have earlier found that blood sera of patients with coronary heart disease (CHD) increase lipid levels in cells cultured from subendothelial intima of human aorta. We have also revealed that the ability of blood sera to raise intracellular cholesterol; that is, their atherogenicity is caused by at least modified low density lipoprotein (LDL) circulating in the blood of patients and autoantibodies to modified LDL. In the present work we have demonstrated significant impact of nonlipid factor(s) to blood atherogenicity. We have developed an approach to removal of nonlipid atherogenicity factor(s) from blood serum based on the use of immobilized LDL. This approach was used for extracorporeal perfusion of patient’s blood through the column with immobilized LDL. Pilot clinical study confirmed the efficacy of this approach for prevention of coronary atherosclerosis progression

How do macrophages sense modified low-density lipoproteins?

In atherosclerosis, serum lipoproteins undergo various chemical modifications that impair their normal function. Modification of low density lipoprotein (LDL) such as oxidation, glycation, carbamylation, glucooxidation, etc. makes LDL particles more proatherogenic. Macrophages are responsible for clearance of modified LDL to prevent cytotoxicity, tissue injury, inflammation, and metabolic disturbances. They develop an advanced sensing arsenal composed of various pattern recognition receptors (PRRs) capable of recognizing and binding foreign or altered-self targets for further inactivation and degradation. Modified LDL can be sensed and taken up by macrophages with a battery of scavenger receptors (SRs), of which SR-A1, CD36, and LOX1 play a major role. However, in atherosclerosis, lipid balance is deregulated that induces inability of macrophages to completely recycle modified LDL and leads to lipid deposition and transformation of macrophages to foam cells. SRs also mediate various pathogenic effects of modified LDL on macrophages through activation of the intracellular signaling network. Other PRRs such Toll-like receptors can also interact with modified LDL and mediate their effects independently or in cooperation with SRs

Paraoxonase and atherosclerosis-related cardiovascular diseases

In humans, three paraoxonase (PON1, PON2, and PON3) genes are clustered on chromosome 7 at a locus that spans a distance around 170 kb. These genes are highly homologous to each other and have a similar protein structural organization. PON2 is the intracellular enzyme, which is expressed in many tissues and organs, while two other members of PON gene family are produced by liver and associate with high density lipoprotein (HDL). The lactonase activity is the ancestral. Besides lactones and organic phosphates, PONs can hydrolyze and therefore detoxify oxidized low density lipoprotein and homocysteine thiolactone, i.e. two cytotoxic compounds with a strong proatherogenic action. Indeed, PONs possess numerous atheroprotective properties, which include antioxidant activity, anti-inflammatory action, preserving HDL function, stimulation of cholesterol efflux, anti-apoptosis, anti-thrombosis, and anti-adhesion. PON genetic polymorphisms contribute to susceptibility/protection from atherosclerosis-related diseases. The bright antiatherogenic activity of the PON cluster makes it a promising target for the development of new therapeutic strategies

Thrombospondins: A Role in Cardiovascular Disease

Thrombospondins (TSPs) represent extracellular matrix (ECM) proteins belonging to the TSP family that comprises five members. All TSPs have a complex multidomain structure that permits the interaction with various partners including other ECM proteins, cytokines, receptors, growth factors, etc. Among TSPs, TSP1, TSP2, and TSP4 are the most studied and functionally tested. TSP1 possesses anti-angiogenic activity and is able to activate transforming growth factor (TGF)-β, a potent profibrotic and anti-inflammatory factor. Both TSP2 and TSP4 are implicated in the control of ECM composition in hypertrophic hearts. TSP1, TSP2, and TSP4 also influence cardiac remodeling by affecting collagen production, activity of matrix metalloproteinases and TGF-β signaling, myofibroblast differentiation, cardiomyocyte apoptosis, and stretch-mediated enhancement of myocardial contraction. The development and evaluation of TSP-deficient animal models provided an option to assess the contribution of TSPs to cardiovascular pathology such as (myocardial infarction) MI, cardiac hypertrophy, heart failure, atherosclerosis, and aortic valve stenosis. Targeting of TSPs has a significant therapeutic value for treatment of cardiovascular disease. The activation of cardiac TSP signaling in stress and pressure overload may be therefore beneficial