Characterization of the De Novo Biosynthetic Enzyme of Platelet Activating Factor, DDT-Insensitive Cholinephosphotransferase, of Human Mesangial Cells

Platelet activating factor (PAF), a potent inflammatory mediator, is implicated in several proinflammatory/inflammatory diseases such as glomerulonephritis, glomerulosclerosis, atherosclerosis, cancer, allergy, and diabetes. PAF can be produced by several renal cells under appropriate stimuli and it is thought to be implicated in renal diseases. The aim of this study is the characterization of DTT-insensitive cholinephosphotransferase (PAF-CPT) of human mesangial cell (HMC), the main regulatory enzyme of PAF de novo biosynthetic pathway. Microsomal fractions of mesangial cells were isolated and enzymatic activity and kinetic parameters were determined by TLC and in vitro biological test in rabbit washed platelets. The effect of bovine serum albumin (BSA), dithiothreitol (DTT), divalent cations (Mg2+ and Ca2+), EDTA, and various chemicals on the activity of PAF-CPT of HMC was also studied. Moreover, preliminary in vitro tests have been performed with several anti-inflammatory factors such as drugs (simvastatin, IFNa, rupatadine, tinzaparin, and salicylic acid) and bioactive compounds of Mediterranean diet (resveratrol and lipids of olive oil, olive pomace, sea bass “Dicentrarchus labrax,” and gilthead sea bream “Sparus aurata”). The results indicated that the above compounds can influence PAF-CPT activity of HMC

Hydroxyl-platelet-activating factor exists in blood of healthy volunteers and periodontal patients.

Periodontal diseases are localized chronic inflammatory conditions of the gingival and underlying bone and connective tissue. Platelet-activating factor (PAF), a potent inflammatory phospholipid mediator that has been previously detected in elevated levels in inflamed gingival tissues, in gingival crevicular fluid and in saliva, is implicated in periodontal disease. Our results from previous studies showed that the biologically active phospholipid detected in gingival crevicular fluid is a hydroxyl-PAF analogue. In this study, hydroxyl-PAF analogue was detected for the first time in human blood derived from patients with chronic periodontitis as well as from periodontally healthy volunteers. The hydroxyl-PAF analogue was purified by high-performance liquid chromatography, detected by biological assays and identified by electrospray analysis. In addition, the quantitative determination of PAF and hydroxyl-PAF analogue (expressed as PAF-like activity) showed a statistically significant increase in the ratio of hydroxyl-PAF analogue levels to PAF levels in periodontal patients, suggesting that this bioactive lipid may play a role in oral inflammation

Beneficial anti‐platelet and anti‐inflammatory properties of irish apple juice and cider bioactives

peer-reviewedSeveral bioactives from fruit juices and beverages like phenolics, nucleotides and polar lipids (PL) have exhibited anti‐platelet cardio‐protective properties. However, apple juice and cider lipid bioactives have not been evaluated so far. The aim of this study was to investigate the an‐ ti‐platelet and anti‐inflammatory effects and structure activity relationships of Irish apple juice and Real Irish cider lipid bioactives against the platelet‐activating factor (PAF)‐ and adenosine di‐ phosphate (ADP)‐related thrombotic and inflammatory manifestations in human platelets. Total Lipids (TL) were extracted from low, moderate and high in tannins apple juices and from their de‐ rived‐through‐fermentation cider products, as well as from commercial apple juice and cider. These were separated into neutral lipids (NL) and PL, while all lipid extracts were further assessed for their ability to inhibit aggregation of human platelets induced by PAF and ADP. In all cases, PL exhibited the strongest anti‐platelet bioactivities and were further separated by high‐performance liquid chromatography (HPLC) analysis into PL subclasses/fractions that were also assessed for their antiplatelet potency. The PL from low in tannins apple juice exhibited the strongest an‐ tiplatelet effects against PAF and ADP, while PL from its fermented cider product were less active. Moreover, the phosphatidylcholines (PC) in apple juices and the phosphatidylethanolamines (PE) in apple ciders were the most bioactive HPLC‐derived PL subclasses against PAF‐induced platelet aggregation. Structural elucidation of the fatty acid composition by gas chromatography mass spectra (GCMS) analysis showed that PL from all samples are rich in beneficial monounsaturated fatty acids (MUFA) and omega 3 (n‐3) polyunsaturated fatty acids (PUFA), providing a possible explanation for their strong anti‐platelet properties, while the favorable low levels of their ome‐ ga‐6/omega‐3 (n‐6/n‐3) PUFA ratio, especially for the bioactive PC and PE subclasses, further support an anti‐inflammatory cardio‐protective potency for these apple products. In conclusion, Irish apple juice and Real Irish cider were found to possess bioactive PL compounds with strong antiplatelet and anti‐inflammatory properties, while fermentation seems to be an important mod‐ ulating factor on their lipid content, structures and bioactivities. However, further studies are needed to evaluate these effects

Fish polar lipids retard atherosclerosis in rabbits by down-regulating PAF biosynthesis and up-regulating PAF catabolism

<p>Abstract</p> <p>Background</p> <p>Platelet activating factor (PAF) has been proposed as a key factor and initial trigger in atherosclerosis. Recently, a modulation of PAF metabolism by bioactive food constituents has been suggested. In this study we investigated the effect of fish polar lipid consumption on PAF metabolism.</p> <p>Results</p> <p>The specific activities of four PAF metabolic enzymes; in leukocytes, platelets and plasma, and PAF concentration; either in blood cells or plasma were determined. Samples were acquired at the beginning and at the end of a previously conducted study in male New Zealand white rabbits that were fed for 45 days with atherogenic diet supplemented (group-B, n = 6) or not (group-A, n = 6) with gilthead sea bream (<it>Sparus aurata</it>) polar lipids.</p> <p>The specific activity of PAF-Acetylhydrolase (PAF-AH); a catabolic enzyme of PAF, was decreased in rabbits' platelets of both A and B groups and in rabbits' leukocytes of group A (p < 0.05). On the other hand the specific activity of Lipoprotein-associated Phospholipase A2 (Lp-PLA2); the catabolic enzyme of PAF in plasma was increased in both A and B groups in both leukocytes and platelets (p < 0.05). PAF-cholinephosphotransferase (PAF-CPT); a biosynthetic enzyme of PAF showed increased specific activity only in rabbits' leukocytes of group A (p < 0.05). Neither of the two groups showed any change in Lyso-PAF-acetyltransferase (Lyso-PAF-AT) specific activity (p > 0.05). Free and bound PAF levels increased in group A while decreased in group B (p < 0.05).</p> <p>Conclusions</p> <p>Gilthead sea bream (<it>Sparus aurata</it>) polar lipids modulate PAF metabolism upon atherosclerotic conditions in rabbits leading to lower PAF levels and activity in blood of rabbits with reduced early atherosclerotic lesions compared to control group.</p

Structurally Diverse Metal Coordination Compounds, Bearing Imidodiphosphinate and Diphosphinoamine Ligands, as Potential Inhibitors of the Platelet Activating Factor

Metal complexes bearing dichalcogenated imidodiphosphinate [R2P(E)NP(E)R2′]− ligands (E = O, S, Se, Te), which act as (E,E) chelates, exhibit a remarkable variety of three-dimensional structures. A series of such complexes, namely, square-planar [Cu{(OPPh2)(OPPh2)N-O, O}2], tetrahedral [Zn{(EPPh2)(EPPh2)N-E,E}2], E = O, S, and octahedral [Ga{(OPPh2)(OPPh2)N-O,O}3], were tested as potential inhibitors of either the platelet activating factor (PAF)- or thrombin-induced aggregation in both washed rabbit platelets and rabbit platelet rich plasma. For comparison, square-planar [Ni{(Ph2P)2N-S-CHMePh-P, P}X2], X = Cl, Br, the corresponding metal salts of all complexes and the (OPPh2)(OPPh2)NH ligand were also investigated. Ga(O,O)3 showed the highest anti-PAF activity but did not inhibit the thrombin-related pathway, whereas Zn(S,S)2, with also a significant PAF inhibitory effect, exhibited the highest thrombin-related inhibition. Zn(O,O)2 and Cu(O,O)2 inhibited moderately both PAF and thrombin, being more effective towards PAF. This work shows that the PAF-inhibitory action depends on the structure of the complexes studied, with the bulkier Ga(O,O)3 being the most efficient and selective inhibitor

Structural Elucidation of Irish Ale Bioactive Polar Lipids with Antithrombotic Properties

The structures of bioactive polar lipids (PLs) of Irish ale with potent antithrombotic and cardioprotective properties were elucidated. Ale PL was fractionated by preparative thin layer chromatography (TLC) into subclasses, and their antithrombotic effect was assessed against human platelet aggregation induced by the pro-inflammatory mediator, platelet-activating factor (PAF). The fatty acid content and the overall structures of ale PL were elucidated by liquid chromatography mass spectrometry (LC-MS). Phosphatidylcholines (PC) and molecules of the sphingomyelin (SM) family exhibited the strongest anti-PAF effects, followed by phosphatidylethanolamines (PE). PC contained higher amounts of omega-3 polyunsaturated fatty acids (n-3 PUFA) and thus the lowest n-6/n-3 ratio. Bioactive diacyl and alkyl-acyl PC and PE molecules bearing n-3 PUFA at their sn-2 position, especially docosahexaenoic acid (DHA) and &alpha;-linolenic acid (ALA) but mostly oleic acid (OA), were identified in both PC and PE subclasses. Eicosapentaenoic acid (EPA) was present only in bioactive PC molecules and not in PE, explaining the lower anti-PAF effects of PE. Bioactive sphingolipid and glycolipid molecules with reported anti-inflammatory and anti-tumour properties, such as specific ceramides and glucosylcerebrosides with sphingosine, phytosphingosine and dihydrosphingosine bases but also specific monogalactodiglycerides and SM species bearing ALA at their sn-2 position, were identified in the SM subclass, providing a rational for its strong bioactivities against the PAF pathway. Further studies are required on the health benefits of bioactive PL from beer and brewery by-products

Comment on “Optimal Nutritional Status for a Well-Functioning Immune System Is an Important Factor to Protect against Viral Infections. Nutrients 2020, 12, 1181”

The novel coronavirus (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has engulfed the world since December 2019 [...

The anti‐inflammatory and antithrombotic properties of bioactives from orange, sanguine and clementine luices and from their remaining by‐products

The anti‐oxidant properties of vitamin C and of phenolic compounds of citrus fruits are well established. However, the evaluation of the anti‐inflammatory and antithrombotic potential of both vitamin C and of the more amphiphilic and lipophilic components of citrus fruits needs further attention. In this study, the anti‐inflammatory and antithrombotic properties of vitamin C and of freshly squeezed juices and their lipid bioactives from the Navalina and Sanguine orange varieties and the Clementine variety of mandarins, as well as from their remaining by‐products, were evaluated against the inflammatory and thrombotic pathways of the platelet‐activating factor (PAF) and thrombin in platelets, as well as against PAF‐biosynthesis in leukocytes. The non‐oxidized juices of these citrus fruits and a vitamin C supplement showed stronger anti‐PAF and antithrombin effects than their oxidized versions through their general anti‐oxidant effect in platelets. The total lipids (TLs) and the HPLC‐derived fractions of phenolic compounds and of polar lipid bioactives from both juices and their peels’ by‐products showed a more specific stronger inhibitory effect against the inflammatory and thrombotic pathways of PAF and thrombin in platelets, while these bioactives strongly inhibited also the specific enzyme activities of the main biosynthetic enzymes of PAF in leukocytes. The stronger bioactivity of the dietary bioactives found in the juices of these citrus fruits against specific biochemical pathways of inflammation and thrombosis seems to act with synergy with the anti‐oxidant potential of its vitamin C content, which further supports the notion that these juices are functional foods with anti‐inflammatory and protective health benefits. In addition, the presence of these dietary bioactive phenolic compounds and polar lipid bioactives in the remaining peels’ wastes further enhance the valorization of such food industry by‐products as potential sources of anti‐inflammatory bioactives to be used as ingredients for novel functional products.</p

Study on the metabolism of platelet activating factor (PAF) in human mesangial ceels - effect of anti-inflammatory, anti-cancer and anti-retroviral factors on PAF biological activities and the enzymes of PAF metabolism

This study is a part of a general concept of the study of the implication of Platelet Activating Factor (PAF) and its metabolism in the mechanisms that result in renal diseases, cancer and AIDS. PAF metabolism was studied in human mesangial cells (HMC) and several factors that exhibit their beneficial effect in the above diseases also through their effect in PAF metabolism and its biological actions, were clarified. In particular: 1. A new enzymatic assay for the determination of the biosynthetic enzyme of PAF, PAF-CPT, was developed for the first time. Ιt was found that the activity of PAF-CPT is present in HMC and its biochemical properties and kinetic parameters in HMC were established for the first time. 2. Several drugs (simvastatin, IFNa, rupatadine, tinzaparin, and acetylsalicylic acid) and bioactive compounds of Mediterranean diet (polar lipids of olive oil, olive pomace, sea bass “Dicentrarchus labrax,” and gilthead sea bream “Sparus aurata” and resveratrol) exhibited an inhibitory effect on PAF-CPT activity. Simvastatin and polar lipids of olive pomace inhibited also the biosynthetic enzyme of PAF, Lyso-PAF-AT, while they induced the basic degradative enzyme of PAF, PAF-AH, of HMC. 3. Finally, several anti-HIV drugs, backbones and regimens of highly active antiretroviral therapy exhibited inhibitory effect against PAF-activity. Lopinavir -r and Tenofovir DF also inhibited the biosynthetic enzymes PAF-CPT and Lyso-PAF-AT, while lopinavir-r induced PAF-AH activity of HMC.Η παρούσα μελέτη εντάσσεται στο γενικότερο πλαίσιο μελέτης της συμμετοχής του Παράγοντα Ενεργοποίησης Αιμοπεταλίων (PAF) στους μηχανισμούς πρόκλησης των νεφρικών παθήσεων, του καρκίνου και του AIDS. Μελετήθηκε ο μεταβολισμός του PAF σε ανθρώπινα μεσαγγειακά κύτταρα (HMC) και καθορίσθηκαν παράγοντες που επιδρώντας στην δράση και στα επίπεδα του παραγόμενου PAF, επιδεικνύουν ευεργετικές ιδιότητες στις προαναφερθείσες σημαντικές παθολογικές καταστάσεις. Συγκεκριμένα: 1. Αναπτύχθηκε νέα μέθοδος προσδιορισμού της ενζυμικής δραστικότητας του βιοσυνθετικού ενζύμου του PAF, PAF-CPT. Σύμφωνα με αυτήν, προσδιορίστηκε η δραστικότητα της PAF-CPT και καθορίστηκαν τα βιοχημικά χαρακτηριστικά και οι κινητικές παράμετροι του σε HMC. 2. Διάφορες φαρμακευτικές ουσίες (ρουπαταδίνη, συμβαστατίνη, ακετυλοσαλικυλικό οξύ, τινζαπαρίνη και IFNα), και βιολογικώς δραστικά συστατικά τροφίμων της Μεσογειακής δίαιτας (πολικά λιποειδή από ελαιόλαδο, ελαιοπυρήνα, τσιπούρα, λαυράκι και ρεσβερατρόλη) αναστέλλουν in vitro την PAF-CPT των HMC. Η συμβαστατίνη και τα πολικά λιποειδή από ελαιοπυρήνα ανέστειλαν και την Lyso-PAF-AT (βιοσυνθετικό ένζυμο του PAF) ενώ διέγειραν το βασικό αποικοδομητικό ένζυμο του PAF, PAF-AH. 3. Τέλος, διάφορα αντι-HIV φάρμακα και συνδυασμοί τους της αντιρετροϊκής θεραπείας επιδεικνύουν μια ανασταλτική επίδραση έναντι της βιολογικής δράσης του PAF. Δύο από αυτά τα φάρμακα (Lopinavir-r και Tenofovir-DF) ανέστειλαν τις PAF-CPT και Lyso-PAF-AT, ενώ το Lopinavir -r ενεργοποιεί την PAF-AH των HMC