Haralabos C. Karantonis

Antithrombotic and Antiatherosclerotic Properties of Olive Oil and Olive Pomace Polar Extracts in Rabbits

Olive oil polar lipid (OOPL) extract has been reported to inhibit atherosclerosis development on rabbits. Olive pomace polar lipid (PPL) extract inhibits PAF activity in vitro and the most potent antagonist has been identified as a glycerylether-sn-2-acetyl glycolipid with common structural characteristics with the respective potent antagonist of OOPL. The aim of this study was to investigate the effect of PPL on early atherosclerosis development on rabbits and to compare it with the antiatherosclerotic effect of OOPL. OOPL and PPL inhibition potency, towards both PAF action and PAF binding, was tested in vitro on washed rabbit platelets. Consequently, rabbits were divided into three groups (A, B, and C). All groups were fed atherogenic diet for 22 days. Atherogenic diets in groups B and C were enriched with OOPL and PPL, respectively. At the end of the experimental time, rabbits were euthanized and aortic samples were examined histopathologically. OOPL and PPL inhibited PAF-induced aggregation, as well as specific PAF binding, with PPL being more potent. Free and bound PAF levels and PAF-AH activity were significantly elevated at the end of the experimental time. Plasma total cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides levels were also found increased. Groups B and C exhibited significantly increased values of EC50 compared to group A. Histopathological examination revealed that the development of early atherosclerosis lesions in groups B and C were significantly inhibited compared to group A. Significant differences were noted in the early atherosclerosis lesions between groups B and C, thus indicating that PPL exhibit its anti-atherosclerotic activity by blocking PAF receptor. Specific PAF antagonists with similar in vitro and in vivo bioactivity to those that have been previously reported in OOPL exist in PPL.

One-step separation system for the main phospholipids, glycolipids, and phenolics by normal phase HPLC. Application to polar lipid extracts from olive and sunflower oils

A new normal phase high performance liquid chromatographic system has been developed that allows efficient separation of several polar lipids, such as phospholipids, glycolipids, and phenolics. By this method, hesperidin, naringin, cerebrosides, phosphatidylcholine, sphingomyelin, platelet activating factor, lysophosphatidylcholine, digalactosyldiglycerides, and phosphatidyl-ethanolamine are separated within 60 min. A gradient elution with acetonitrile/methanol, methanol, and water was performed with a normal phase aminopropyl-modified silica gel HPLC column. The eluted lipids were monitored by UV detection. In addition, in this study, a successful application of the new method is represented for the polar lipids of olive oil and sunflower, which are also identified with chemical determinations.

Platelet-activating factor involvement in thioacetamide-induced experimental liver fibrosis and cirrhosis.

Platelet-activating factor (PAF) is a potent lipid inflammatory mediator acting on cells through its specific receptor. Plasma PAF-acetylhydrolase (PAF-AH) is the main enzyme that inactivates PAF in blood, participating in its homeostasis. The objective of this study was to investigate the involvement of PAF in the liver fibrotic process using an experimental animal model. Liver fibrosis was induced in adult male Wistar rats by administration of thioacetamide (TAA) in drinking water (300xa0mg/l) for three months. The animals were sacrificed at time 0 (control group) and after 1, 2, and 3xa0months. PAF levels in liver and blood and PAF-AH activity in plasma were determined. Liver histopathological examination was also performed. TAA administration resulted in progressively increased liver fibrosis, leading finally to the formation of cirrhotic nodules in the liver. Throughout the experiment PAF levels in liver tissue remained stable. “Total” (“free” plus “bound”) PAF levels in blood decreased, reaching statistically significant differences in the first and third months compared with the control group (Pxa0<xa00.05). “Free” PAF levels in blood were higher at one month (Pxa0<xa00.05) and decreased gradually thereafter. In all treated groups, “bound” PAF levels in blood decreased whereas plasma PAF-AH activity increased (Pxa0<xa00.05) compared with the control group. Our data indicated alterations of PAF levels in blood and PAF-AH activity during fibrosis induction, implicating participation of PAF in the liver fibrotic process.

Isolation and structural elucidation of biologically active phospholipids from Scytonema julianum (cyanobacteria).

The role of platelet-activating factor (PAF) as a mediator appeared in rather primitive organisms like protozoans and was maintained in more evolved organisms. No reports exist for the presence of PAF or PAF analogues - or even compounds that exhibit PAF-like activity - in cyanobacteria, even though they belong to a a group of organisms at a low evolutionary level where the content of alkylacyl forms of ether lipids is expected to be high. In addition, cyanobacteria serve as a rich source of novel bioactive metabolites. In the present study the total lipids of a strain of Scytonema julianum, a filamentous cyanobacterium isolated from a Greek cave, were separated into neutral lipids and phospholipids, the latter being further fractionated by HPLC. Each phospholipid fraction was tested in vitro for its ability to inhibit PAF-, arachidonic acid- and ADP-induced washed-rabbit-platelet aggregation and/or to cause platelet aggregation. Two types of phospholipids causing platelet aggregation were detected and shown to be an acetylsphingomyelin and an acylacetylglycerol phosphoacetylated glycolipid. The existence of the sphingomyelin analogues is very important, since ceramides, cerebrosides and related lipids are intracellular second messengers. The identification of the phosphoglycoglycerolipid demonstrates a new type of lipid in cyanobacteria, namely one that exhibits a biological activity very similar to that of PAF. Its presence reinforces the concept that PAF is a member of a large family of lipid mediators, apparently having different physiological roles in prokaryotic and eukaryotic organisms. In addition, Scytonema julianum contains a phosphatidylcholine (C(16:0)/(18:2)), even though bacteria in general seldom contain choline-containing phosphoacylglycerols.

Separation of polar lipids from soybean oil and cotton seed oil by one-step HPLC system. Biological activity of isolated lipids

Many epidemiological studies suggest that vegetable oils present a protective effect against atherosclerosis, in a preventive level. Although the atherogenic mechanism is not clear, a number of reports indicate that platelet activating factor (PAF) plays a critical role in atherogenesis. In this study, total lipids of two vegetable oils were separated into polar and neutral lipids. Total lipids, total polar lipids, and total neutral lipids were tested in vitro for their biological activity against washed rabbit platelets, that is their ability to inhibit platelet aggregation induced by PAF, or to cause platelet aggregation. Total polar lipids, which are more potent than total neutral lipids, were further fractionated by HPLC and each fraction was tested for its biological activity. The experimental data give an answer for the preventive protective effect of vegetable oils against atherogenesis.