Aronia melanocarpa fruit extract exhibits anti-inflammatory activity in human aortic endothelial cells.

BACKGROUND: Altered expression of cell adhesion molecules (CAMs) has been implicated in a variety of chronic inflammatory conditions, including atherosclerosis. Regulation of adhesion molecule expression by specific redox-sensitive mechanisms has been reported. Additionally, it has been observed that the extract of Aronia melanocarpa (A. Melanocarpa) fruits, rich in polyphenols, exhibits potent anti-oxidant properties and displays cardioprotective activity. METHODS AND RESULTS: Human aortic endothelial cells (HAECs) were pretreated with various concentrations (primarily 50 µg/mL) of Aronia Melanocarpa fruit extract prior to treatment with TNFα (10 ng/mL) for various periods of time. The surface protein and mRNA expression of ICAM-1 and VCAM-1 were determined using flow cytometry and real-time RT-PCR, respectively. Adhesion of peripheral blood mononuclear leucocytes (PBMLs) to TNFα-treated HAECs was evaluated by an adhesion assay. Activation of NF-κB was evaluated by measuring NF-κB p65 phosphorylation using flow cytometry. ROS production was determined by reduction in fluorescent 2',7'-dichlorofluorescein diacetate (DCFH-DA). Tested A. Melanocarpa extract significantly inhibited the expression of ICAM-1 and VCAM-1, attenuated the phosphorylation of NF-κB p65 and decreased intracellular ROS production in TNFα-treated HAECs. CONCLUSION: We conclude that A. Melanocarpa fruit extract exhibits anti-inflammatory effects in HAECs by inhibiting the expression of endothelial CAMs, activation of NF-κB and production of ROS.

Trans fatty acids induce a proinflammatory response in endothelial cells through ROS-dependent nuclear factor-κB activation.

UNLABELLED: It has been shown that increased intake of trans fatty acids (TFAs) is associated with a higher risk of cardiovascular disease. In this study, we have investigated the effects of linoelaidic (LA) and elaidic (EA) acids on the proinflammatory response in endothelial cells, a key step in vascular disease. Human aortic endothelial cells (HAECs) were treated with different concentrations (100 µmol/l in most experiments) of LA or EA for different periods of time. The surface protein and mRNA expression of ICAM-1 and VCAM-1 were determined by flow cytometry and real time RT-PCR, respectively. Adhesion of leukocytes to TFA-treated HAECs was evaluated by an adhesion assay. Activation of nuclear factor-κB (NF-κB) was evaluated by measuring NF-κB p65 phosphorylation using flow cytometry. ROS production was determined by the reduction of fluorescent 2',7'-dichlorofluorescein diacetate (DCFH-DA). LA treatment significantly increased protein and mRNA levels of ICAM-1 and VCAM-1, leukocyte adhesion to HAECs, phosphorylation of NF-κB and ROS generation. Similar effects were achieved for cells incubated with EA. Experiments with HAECs pretreated with pyrrolidine dithiocarbamate, an inhibitor of NF-κB, revealed that both LA and EA-mediated induction of ICAM-1 and VCAM-1 is mainly regulated by NF-κB. The ROS production induced by both of the studied acids was inhibited in the presence of diphenyleneiodonium (DPI), a NADPH oxidase inhibitor, suggesting ROS production through the activation of NADPH oxidase. Furthermore, LA or EA-induced ICAM-1 and VCAM-1 expression, activation of NF-κB and adhesion of leukocytes to HAECs were abolished in the presence of DPI. CONCLUSION: TFAs present in our diet have a direct proinflammatory effect, which promotes leukocyte adhesion to the endothelium through ROS-dependent NF-κB activation.

Propionate reduces the cytokine-induced VCAM-1 and ICAM-1 expression by inhibiting nuclear factor-kappa B (NF-kappaB) activation.

Adhesion and migration of leukocytes into the surrounding tissue are crucial steps in inflammation, immunity and atherogenesis. Expression of cell adhesion molecules by endothelial cells plays a role in these processes. Propionate is a naturally occurring short chain fatty acid produced by bacterial fermentation of dietary fibre. High intake of dietary fibre has been associated with an improved bowel function and with a reduced risk of cardiovascular disease. However, the molecular mechanisms responsible for these effects remain unknown. In this study, the effects of propionate on the expression of endothelial leukocyte adhesion molecules by cytokine-stimulated human umbilical vein endothelial cells (HUVEC) were investigated. Pretreatment of HUVEC with propionate significantly inhibited the tumor necrosis factor alpha (TNF-alpha)-induced expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1) in a time- and dose-dependent manner. At 10 mM, propionate also inhibited the interleukin-1 (IL- 1)-mediated VCAM-1 and ICAM-1 expression, with the latter effect being more pronounced, as well as decreased the TNF-alpha-induced VCAM-1 and ICAM-1 mRNA expression in a similar manner. The decrease in VCAM-1 and ICAM-1 expression was associated with a reduction of adherence of monocytes and lymphocytes to the cytokine-stimulated HUVEC. In addition, propionate significantly inhibited the TNF-alpha-induced activation of nuclear factor-kappa B (NF-kappaB) and significantly increased the expression of peroxisome proliferator-activated receptor alpha (PPARalpha) in HUVEC. These results demonstrate that propionate may have antiinflammatory and possibly antiatherogenic properties. Our findings warrant further investigation into the therapeutic effects of propionate on a number of pathological events nvolving leukocyte recruitment.

Trans fatty acids induce apoptosis in human endothelial cells.

The present study was designed to investigate the hypothesis that trans fatty acids can induce apoptosis of human umbilical vein endothelial cells (HUVEC). To test this hypothesis apoptosis was measured in HUVEC treated with 0.1, 1.0 or 5.0 mM trans elaidic acid (t-18:1) or linoelaidic acid (t,t-18:2) for 24 hours. For the detection of apoptosis, TdT-mediated dUTP nick end labelling assay (TUNEL), cell binding of annexin V and propidium iodide uptake were measured. Active Caspase-3 and cleaved PARP (poly-ADP-ribose polymerase) were also measured in the cell lysate. Moreover, cellular ability to produce ROS (reactive oxygen species) was measured by DCF fluorescence Both acids studied induce both early (annexin-positive cells) and late stages of apoptosis (cells stained by propidium iodide) in a dose-dependent manner. Also the appearance of TUNEL-positive cells was induced by both trans fatty acids tested, in a dose dependent manner. Both trans acids induce apoptosis through their effect on Caspase-3 activity and on intracellular ROS production. It is worth emphasising that linoelaidic acid proved to be a more potent inducer of apoptosis and ROS production in endothelial cells than elaidic acid. The present studies suggest that trans fatty acids may play a role in damaging and death of vascular endothelial cells in atherosclerosis.

Simvastatin modulates TNFalpha-induced adhesion molecules expression in human endothelial cells.

Adhesion and transendothelial migration of leukocytes into the vascular wall is a crucial step in atherogenesis. Expression of cell adhesion molecules by endothelial cells plays a leading role in this process. We investigated the effect of simvastatin, an inhibitor of HMG-CoA reductase administered to reduce plasma levels of LDL-cholesterol, on the expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular cell adhesion molecule-1 (ICAM-1) by human umbilical vein endothelial cells (HUVEC) stimulated with tumor necrosis factor alpha (TNFalpha). We found the expression to be significantly inhibited by the drug in a time and concentration-dependent manner and to a greater extent in the case of VCAM-1 as compared with ICAM-1. In TNFalpha-stimulated HUVEC, simvastatin decreased VCAM-1 and ICAM-1 mRNA levels, inhibited TNFalpha-induced activation of nuclear factor kappaB (NF-kappaB) and enhanced expression of peroxisome proliferator-activated receptor alpha (PPARalpha). These effects were associated with reduction of adherence of monocytes and lymphocytes to HUVEC. The present findings suggest that the benefits of statins in vascular disease may include the inhibition of expression of VCAM-1 and ICAM-1 through effects on NF-kappaB.

A pleiotropic antiatherogenic action of ibuprofen.

Ibuprofen is a cyclooxygenase (COX-1 and COX-2) inhibitor known to reduce the production of prostaglandins that play prominent role in inflammation. Other properties of the drug, aside from its anti-inflammatory effects, have been recently studied. In this paper we shall discuss several properties of ibuprofen that making the drug interesting for treatment of conditions associated with atherosclerosis. Ibuprofen exerts pleiotropic effects such as inhibition of adhesion and transendothelial migration of leukocytes, suppressing intracellular production of reactive oxygen species and oxidative modification of LDL. Interestingly, ibuprofen increased HDL cholesterol levels and reduced the level of triglicerides. Ibuprofen can also modulate efficiency of fibrynolisis by inhibiting production of plasminogen activator inhibitor (PAI-1). This properties of ibuprofen may be due to changing the activity of transcription factors. Ibuprofen inhibits the activation of NF-kB and activates PPARa and PPARg.

Selective inhibition by probucol of vascular cell adhesion molecule-1 (VCAM-1) expression in human vascular endothelial cells.

An early event in atherogenesis is the adhesion of monocytes to endothelium via adhesion molecules, such as VCAM-1 and intracellular adhesion molecule-1 (ICAM-1). It has been suggested that VCAM-1 plays a very important role in the recruitment of monocytes in atherosclerosis. Probucol is a potent inhibitor of atherosclerosis in animal models. However, the mechanism of its antiatherogenic effect is poorly understood. The aim of our study was to evaluate whether probucol can influence the expression of endothelial cell adhesion molecules and endothelial adhesiveness. The study was performed on cultured human umbilical vein endothelial cells (HUVEC). HUVEC were pretreated with probucol (50 microM) at different time periods before stimulation with TNFalpha (100 U ml(-1)) or IL-1beta (100 U ml(-1)). The protein expression of VCAM-1 and ICAM-1 was measured by flow cytometry. VCAM-1 mRNA expression was measured by reverse transcription polymerase chain reaction (RT PCR). Probucol time dependently reduced agonist-induced VCAM-1 ( approximately 45%, 48 h) surface protein and mRNA expression ( approximately 40%, 48 h) in HUVEC, but not ICAM-1 surface protein expression. Decreased VCAM-1 expression was associated with reduction ( approximately 40%) of adherence between cytokine-stimulated HUVEC and peripheral blood mononuclear leukocytes (PBMC). Our results suggest that the antiatherogenic effect of probucol may, in part, be due to a downregulation of VCAM-1 expression.

Ibuprofen inhibits adhesiveness of monocytes to endothelium and reduces cellular oxidative stress in smokers and non-smokers.

BACKGROUND: Cigarette smoking is a major risk factor in atherosclerosis and a useful model from which to study chronic inflammation. We compared monocyte function, lipid profiles and inflammatory markers in smokers and non-smokers, before and after oral ibuprofen intake. The adhesion of freshly isolated monocytes to native and tumour necrosis factor alpha (TNFalpha) stimulated human umbilical vein endothelial cells (HUVEC), as well as superoxide anion (O2-) levels and hydrogen peroxide (H2O2) production in resting and phorbol myristate acetate (PMA) stimulated monocytes were determined. MATERIALS AND METHODS: A group of nine smokers without any other coronary risk factor was compared with an age-matched group of 9 non-smokers. Tests were performed before and after a two-week course of oral ibuprofen (600 mg day-1). RESULTS: In smokers before ibuprofen, monocyte adhesion to native and TNFalpha-stimulated HUVEC was increased (P < 0001 and P < 0.01, respectively), and so were O2- levels in native and PMA-stimulated monocytes (P < 0.01 and P < 0.001, respectively). Ibuprofen reduced the adhesion of monocytes to native and stimulated HUVEC (P < 0.001) and O2- generation by resting and PMA-stimulated cells (P < 0.01) in both groups. H2O2 production by resting and PMA-stimulated monocytes was reduced in smokers and non-smokers (P < 0.01). Interestingly, ibuprofen increased HDL cholesterol levels in smokers (P < 0.01) and non-smokers (P < 0.001), and reduced the level of triglycerides in smokers (P < 0.05). CONCLUSION: Oral administration of ibuprofen reduced the adhesion of monocytes to HUVEC, suppressed oxidative stress and increased HDL cholesterol levels in smokers and non-smokers.

Selective inhibition by alpha-tocopherol of vascular cell adhesion molecule-1 expression in human vascular endothelial cells.

An early event in atherogenesis is the adhesion of monocytes to endothelium via adhesion molecules such as VCAM-1 and intracellular adhesion molecule-1 (ICAM-1). It has been suggested that VCAM-1 plays a very important role in recruitment of monocytes in atherosclerosis. Several studies suggest that vitamin E has antiatherosclerotic properties. However, the mechanism of its antiatherogenic effect awaits elucidation. The aim of our study was to evaluate whether alpha-tocopherol can influence expression of endothelial cell adhesion molecules and endothelial adhesiveness. The study was performed on cultured human umbilical vein endothelial cells (HUVEC). HUVEC were pretreated with alpha-tocopherol (50 micromol/l) in different times before stimulation with TNFalpha (100 U/ml) or IL-1beta (100 U/ml). Protein expression of VCAM-1 and ICAM-1 was measured by flow cytometry. mRNA expression of VCAM-1 was measured by reverse transcription polymerase chain reaction (RT-PCR). alpha-Tocopherol time dependently reduced agonist-induced VCAM-1 in both surface protein (about 40%, 48 h) and mRNA (about 35%, 48 h) expression in HUVEC but not ICAM-1 surface protein expression. Inhibitory effect of alpha-tocopherol was dependent on culture condition of HUVEC. Decreased VCAM-1 expression was associated with reduction (about 40%) of adherence between cytokine-stimulated HUVEC and peripheral blood mononuclear leukocytes (PBMC). Our results suggest that the antiatherogenic effect of alpha-tocopherol may in part be due to a downregulation of VCAM-1 expression.

Ibuprofen protects low density lipoproteins against oxidative modification.

Oxidative modification of LDL by vascular cells has been proposed as the mechanism by which LDL become atherogenic. The effect of ibuprofen on LDL modification by copper ions, monocytes and endothelial cells was studied by measuring lipid peroxidation products. Ibuprofen inhibited LDL oxidation in a dose-dependent manner over a concentration range of 0.1 to 2.0 mM. Ibuprofen (2 mM, 100 microg/ml LDL) reduced the amount of lipid peroxides formed during 2 and 6 h incubation in the presence of copper ions by 52 and 28%, respectively. Weak free radical scavenging activity of ibuprofen was observed in the DPPH test. The protective effect of ibuprofen was more marked when oxidation was induced by monocytes or endothelial cells. Ibuprofen (1 mM, 100 microg/ml LDL) reduced the amount of lipid peroxides generated in LDL during monocyte-mediated oxidation by 40%. HUVEC-mediated oxidation of LDL in the absence and presence of Cu2+ was reduced by 32 and 39%, respectively. More lipid peroxides appeared when endothelial cells were stimulated by IL-1beta or TNFalpha and the inhibitory effect of ibuprofen in this case was more pronounced. Ibuprofen (1 mM, 100 microg/ml LDL) reduced the amount of lipid peroxides formed during incubation of LDL with IL-1beta-stimulated HUVEC by 43%. The figures in the absence and presence of Cu2+ for HUVEC stimulated with TNFalpha were 56 and 59%, respectively. To assess the possibility that ibuprofen acts by lowering the production rate of reactive oxygen species, the intracellular concentration of H2O2 was measured. Ibuprofen (1 mM) reduced intracellular production of hydrogen peroxide in PMA-stimulated mononuclear cells by 69%. When HUVEC were stimulated by IL-1beta or TNFalpha the reduction was 62% and 66%, respectively.

[Atherosclerosis as an inflammatory process. Potential role of adhesion molecules in development of atherosclerosis]. / Miazdzyca jako proces zapalny. Potencjalna rola molekul adhezyjnych w procesie powstawania miazdzycy.

Adherence of monocytes and their subsequent migration into intima appears to be important early event in the formation of foam cell rich atherosclerotic lesion, both in humans and experimental animals. There is increasing evidence that vascular endothelial cells play an active role in these process. In this paper, current knowledge about adhesion molecules presented on the surface of the endothelial cells has been summarized and their role in adhesion and migration or leucocyte both in inflammation and atherosclerosis was characterized. Current concept of pathogenesis of atherosclerosis has been presented, emphasizing the similarities of the mechanism involved to those in inflammation.

[Role of adhesion molecules in interactions of endothelium with leukocytes in inflammatory processes]. / Rola czastek adhezyjnych w interakcjach sródblonka z leukocytami w procesie zapalnym.

This paper reviews recent work regarding the molecules that mediate leukocyte-endothelial cell adhesion, describes the underlying principles of leukocyte migration, and discusses a model of the sequence of events that allows a leukocyte to attach to endothelium and migrate into tissue. Pathophysiological importance of the adhesion molecules in the development of different states of inflammation has been also presented.