Lisa Pasterk

Anti-Psoriatic Therapy Recovers High-Density Lipoprotein Composition and Function

Psoriasis is a chronic inflammatory disorder associated with increased cardiovascular mortality. Psoriasis affects high-density lipoprotein (HDL) composition, generating dysfunctional HDL particles. However, data regarding the impact of anti-psoriatic therapy on HDL composition and function are not available. HDL was isolated from 15 psoriatic patients at baseline and after effective topical and/or systemic anti-psoriatic therapy and from 15 age- and sex-matched healthy controls. HDL from psoriatic patients showed a significantly impaired capability to mobilize cholesterol from macrophages (6.4 vs. 8.0% [(3)H]cholesterol efflux, P<0.001), low paraoxonase (217 vs. 350 μM(-1) minute(-1) mg(-1) protein, P=0.011) and increased Lp-PLA2 activities (19.9 vs. 12.1 nM(-1) minute(-1) mg(-1) protein, P=0.028). Of particular interest, the anti-psoriatic therapy significantly improved serum lecithin-cholesterol acyltransferase activity and decreased total serum lipolytic activity but did not affect serum levels of HDL-cholesterol. Most importantly, these changes were associated with a significantly improved HDL-cholesterol efflux capability. Our results provide evidence that effective anti-psoriatic therapy recovers HDL composition and function, independent of serum HDL-cholesterol levels, and support to the emerging concept that HDL function may be a better marker of cardiovascular risk than HDL-cholesterol levels.

Liver disease alters high-density lipoprotein composition, metabolism and function

High-density lipoproteins (HDL) are important endogenous inhibitors of inflammatory responses. Functional impairment of HDL might contribute to the excess mortality experienced by patients with liver disease, but the effect of cirrhosis on HDL metabolism and function remain elusive. To get an integrated measure of HDL quantity and quality, we assessed several metrics of HDL function using apolipoprotein (apo) B-depleted sera from patients with compensated cirrhosis, patients with acutely decompensated cirrhosis and healthy controls. We observed that sera of cirrhotic patients showed reduced levels of HDL-cholesterol and profoundly suppressed activities of several enzymes involved in HDL maturation and metabolism. Native gel electrophoresis analyses revealed that cirrhotic serum HDL shifts towards the larger HDL2 subclass. Proteomic assessment of isolated HDL identified several proteins, including apoA-I, apoC-III, apoE, paraoxonase 1 and acute phase serum amyloid A to be significantly altered in cirrhotic patients. With regard to function, these alterations in levels, composition and structure of HDL were strongly associated with metrics of function of apoB-depleted sera, including cholesterol efflux capability, paraoxonase activity, the ability to inhibit monocyte production of cytokines and endothelial regenerative activities. Of particular interest, cholesterol efflux capacity appeared to be strongly associated with liver disease mortality. Our findings may be clinically relevant and improve our ability to monitor cirrhotic patients at high risk.

Neutrophil effector responses are suppressed by secretory phospholipase A2 modified HDL

Secretory phospholipase A2 (sPLA2) generates bioactive lysophospholipids implicated in acute and chronic inflammation, but the pathophysiologic role of sPLA2 is poorly understood. Given that high-density lipoprotein (HDL) is the major substrate for sPLA2 in plasma, we investigated the effects of sPLA2-mediated modification of HDL (sPLA2-HDL) on neutrophil function, an essential arm of the innate immune response and atherosclerosis. Treatment of neutrophils with sPLA2-HDL rapidly prevented agonist-induced neutrophil activation, including shape change, neutrophil extracellular trap formation, CD11b activation, adhesion under flow and migration of neutrophils. The cholesterol-mobilizing activity of sPLA2-HDL was markedly increased when compared to native HDL, promoting a significant reduction of cholesterol-rich signaling microdomains integral to cellular signaling pathways. Moreover, sPLA2-HDL effectively suppressed agonist-induced rise in intracellular Ca²⁺ levels. Native HDL showed no significant effects and removing lysophospholipids from sPLA2-HDL abolished all anti-inflammatory activities. Overall, our studies suggest that the increased cholesterol-mobilizing activity of sPLA2-HDL and suppression of rise in intracellular Ca²⁺ levels are likely mechanism that counteracts agonist-induced activation of neutrophils. These counterintuitive findings imply that neutrophil trafficking and effector responses are altered by sPLA2-HDL during inflammatory conditions.

Mode of renal replacement therapy determines endotoxemia and neutrophil dysfunction in chronic kidney disease

Bacterial infection and sepsis are common complications of chronic kidney disease (CKD). A vicious cycle of increased gut permeability, endotoxemia, inadequate activation of the innate immune system and resulting innate immune dysfunction is hypothesized. We assessed endotoxemia, neutrophil function and its relation to oxidative stress, inflammation and gut permeability in patients with CKD grade 3–5 without renal replacement therapy (CKD group, n = 57), patients with CKD stage 5 undergoing haemodialysis (HD, n = 32) or peritoneal dialysis (PD, n = 28) and patients after kidney transplantation (KT, n = 67) in a cross-sectional observational study. In HD patients, endotoxin serum levels were elevated and neutrophil phagocytic capacity was decreased compared to all other groups. Patients on HD had a significantly higher mortality, due to infections during follow up, compared to PD (p = 0.022). Oxidative stress, neutrophil energy charge, systemic inflammation and gut permeability could not completely explain these differences. Our findings suggest that dialysis modality and not renal function per se determine the development of neutrophil dysfunction and endotoxemia in CKD-patients. HD patients are particularly prone to neutrophil dysfunction and endotoxemia whereas neutrophil function seems to improve after KT. Multi-target approaches are therefore warranted to improve neutrophil function and potentially reduce the rate of infections with patients undergoing haemodialysis.

Oxidized plasma albumin promotes platelet-endothelial crosstalk and endothelial tissue factor expression

Plasma advanced oxidation protein products (AOPPs), a class of pro-inflammatory pathogenic mediators, accumulate in subjects with chronic kidney disease. Whether AOPPs contribute to coagulation abnormalities, which are frequently seen in uremic patients, is unknown. Here we report that AOPPs activate platelets via a CD36-mediated signaling pathway. Activation of signaling pathways by AOPP-platelet interaction resulted in the expression of several platelet activation markers and rapidly induced the expression of CD40 ligand, triggering platelet adhesion to endothelial cells and promoting endothelial tissue factor expression. AOPPs and serum tissue factor levels were considerably increased in end stage renal disease patients on hemodialysis and a significant correlation of AOPPs and serum tissue factor was found. Interestingly, serum levels of AOPPs and tissue factor were substantially lower in stable kidney transplant patients when compared with hemodialysis patients. Given that CD36 is known to transduce the effects of oxidized lipids into platelet hyperactivity, our findings reveal previously unknown pro-thrombotic activities of oxidized plasma albumin via a CD36 dependent pathway.

The EP3 Agonist Sulprostone Enhances Platelet Adhesion But Not Thrombus Formation Under Flow Conditions

Platelets express the EP2, EP3 and EP4 receptors. Prostaglandin (PG) E2 has a biphasic effect on platelets. Low concentrations of PGE2 enhance platelet aggregation through the activation of the EP3 receptors, while at high concentrations it attenuates aggregation via the EP4 receptor. Consequently, EP3 receptor inhibition was shown to inhibit artherothrombosis, but had no influence on bleeding time in vivo. In this study, we investigated the role of the EP3 receptor in adhesion and thrombus formation under flow conditions in vitro. The EP3 agonist sulprostone caused an increase in the adhesion of washed platelets to fibrinogen as well as to collagen under low shear stress, an effect that was blocked by the EP3 antagonist L-798106. In contrast, when whole blood was perfused over collagen-coated surfaces, sulprostone did not enhance binding and thrombus formation of platelets on collagen; at high concentrations it even attenuated this response. We conclude that in more physiological models of thrombus formation, the role for EP3 receptors is limited, indirectly suggesting that the primary action of PGE2 in haemostasis might be an inhibitory one.

Laropiprant attenuates EP3 and TP prostanoid receptor-mediated thrombus formation

The use of the lipid lowering agent niacin is hampered by a frequent flush response which is largely mediated by prostaglandin (PG) D2. Therefore, concomitant administration of the D-type prostanoid (DP) receptor antagonist laropiprant has been proposed to be a useful approach in preventing niacin-induced flush. However, antagonizing PGD2, which is a potent inhibitor of platelet aggregation, might pose the risk of atherothrombotic events in cardiovascular disease. In fact, we found that in vitro treatment of platelets with laropiprant prevented the inhibitory effects of PGD2 on platelet function, i.e. platelet aggregation, Ca flux, P-selectin expression, activation of glycoprotein IIb/IIIa and thrombus formation. In contrast, laropiprant did not prevent the inhibitory effects of acetylsalicylic acid or niacin on thrombus formation. At higher concentrations, laropiprant by itself attenuated platelet activation induced by thromboxane (TP) and E-type prostanoid (EP)3 receptor stimulation, as demonstrated in assays of platelet aggregation, Ca flux, P-selectin expression, and activation of glycoprotein IIb/IIIa. Inhibition of platelet function exerted by EP4 or I-type prostanoid (IP) receptors was not affected by laropiprant. These in vitro data suggest that niacin/laropiprant for the treatment of dyslipidemias might have a beneficial profile with respect to platelet function and thrombotic events in vascular disease. Citation: Philipose S, Konya V, Lazarevic M, Pasterk LM, Marsche G, et al. (2012) Laropiprant Attenuates EP3 and TP Prostanoid Receptor-Mediated Thrombus Formation. PLoS ONE 7(8): e40222. doi:10.1371/journal.pone.0040222 Editor: Christian Schulz, Heart Center Munich, Germany Received April 17, 2012; Accepted June 2, 2012; Published August 1, 2012 Copyright: 2012 Philipose et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Dr. Philipose was funded by the PhD Program in Molecular Medicine of the Medical University of Graz. This study was supported by the Jubilaumsfonds of the Austrian National Bank (OeNB, grants 13487 and 14263) and the Austrian Science Fund (FWF; grants P22521-B18, P19473-B05, P21004-B02 and P22976-B18. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: akos.heinemann@medunigraz.at

Secretory phospholipase A2 modified HDL rapidly and potently suppresses platelet activation

Levels of secretory phospholipases A2 (sPLA2) highly increase under acute and chronic inflammatory conditions. sPLA2 is mainly associated with high-density lipoproteins (HDL) and generates bioactive lysophospholipids implicated in acute and chronic inflammatory processes. Unexpectedly, pharmacological inhibition of sPLA2 in patients with acute coronary syndrome was associated with an increased risk of myocardial infarction and stroke. Given that platelets are key players in thrombosis and inflammation, we hypothesized that sPLA2-induced hydrolysis of HDL-associated phospholipids (sPLA2-HDL) generates modified HDL particles that affect platelet function. We observed that sPLA2-HDL potently and rapidly inhibited platelet aggregation induced by several agonists, P-selectin expression, GPIIb/IIIa activation and superoxide production, whereas native HDL showed little effects. sPLA2-HDL suppressed the agonist-induced rise of intracellular Ca2+ levels and phosphorylation of Akt and ERK1/2, which trigger key steps in promoting platelet activation. Importantly, sPLA2 in the absence of HDL showed no effects, whereas enrichment of HDL with lysophosphatidylcholines containing saturated fatty acids (the main sPLA2 products) mimicked sPLA2-HDL activities. Our findings suggest that sPLA2 generates lysophosphatidylcholine-enriched HDL particles that modulate platelet function under inflammatory conditions.

The EP1/EP3 receptor agonist 17-pt-PGE2 acts as an EP4 receptor agonist on endothelial barrier function and in a model of LPS-induced pulmonary inflammation.

Endothelial dysfunction is a hallmark of inflammatory conditions. We recently demonstrated that prostaglandin (PG)E2 enhances the resistance of pulmonary endothelium in vitro and counteracts lipopolysaccharide (LPS)-induced pulmonary inflammation in vivo via EP4 receptors. The aim of this study was to investigate the role of the EP1/EP3 receptor agonist 17-phenyl-trinor-(pt)-PGE2 on acute lung inflammation in a mouse model. In LPS-induced pulmonary inflammation in mice, 17-pt-PGE2 reduced neutrophil infiltration and inhibited vascular leakage. These effects were unaltered by an EP1 antagonist, but reversed by EP4 receptor antagonists. 17-pt-PGE2 increased the resistance of pulmonary microvascular endothelial cells and prevented thrombin-induced disruption of endothelial junctions. Again, these effects were not mediated via EP1 or EP3 but through activation of the EP4 receptor, as demonstrated by the lack of effect of more selective EP1 and EP3 receptor agonists, prevention of these effects by EP4 antagonists and EP4 receptor knock-down by siRNA. In contrast, the aggregation enhancing effect of 17-pt-PGE2 in human platelets was mediated via EP3 receptors. Our results demonstrate that 17-pt-PGE2 enhances the endothelial barrier in vitro on pulmonary microvascular endothelial cells, and accordingly ameliorates the recruitment of neutrophils, via EP4 receptors in vivo. This suggests a beneficial effect of 17-pt-PGE2 on pulmonary inflammatory diseases.