Daniela Peter

Degradation of phospholipids by oxidative stress—Exceptional significance of cardiolipin

Abstract The aim of this study was to investigate the effect of oxidative stress on mitochondrial phospholipids. In this context, this study investigated (i) the content of phosphatidylethanolamine (PE), phosphatidylcholine (PC) and cardiolipin (CL), (ii) the correlation of CL degradation with mitochondrial function and (iii) the correlation of CL degradation and CL oxidation. Oxidative stress induced by iron/ascorbate caused a dramatic decrease of these phospholipids, in which CL was the most sensitive phospholipid. Even moderate oxidative stress by hypoxia/reoxygenation caused a decrease in CL that was parallelled by a decrease in active respiration of isolated rat heart mitochondria. The relation between oxidative stress, CL degradation and CL oxidation was studied by in vitro treatment of commercially available CL with superoxide anion radicals and H2O2. The degradation of CL was mediated by H2O2 and required the presence of cytochrome c. Other peroxidases such as horse radish peroxidase and glutathione peroxidase had no effect. Cytochrome c in the presence of H2O2 caused CL oxidation. The data demonstrate that oxidative stress may cause degradation of phospholipids by oxidation, in particular CL; resulting in mitochondrial dysfunction.

Serum Concentrations of F2-Isoprostanes and 4-Hydroxynonenal in Hemodialysis Patients in Relation to Inflammation and Renal Anemia.

Background Patients with end-stage renal disease (ESRD) undergoing hemodialysis (HD) are apparently exposed to enhanced oxidative stress and to inflammation. It was the aim of this study to characterize the state of systemic oxidative stress of ESRD patients before and following HD using highly specific biomarkers, F2-isoprostanes and 4-hydroxynonenal (HNE). Furthermore the question should be answered, if there are associations between inflammation and systemic oxidative stress and/or between systemic oxidative stress and renal anemia, which is more or less typical for HD patients. Patients and methods Concentrations of F2-isoprostanes, HNE, C-reactive protein (CRP) as marker of inflammation, and hemoglobin were measured in serum samples of patients with ESRD before and after HD and of healthy control persons for comparison. Total (esterified plus free) F2-isoprostanes were quantified by highly sensitive gas chromatography/mass spectrometry technique, HNE by thin layer chromatography and HPLC/UV detection, CRP by immunoturbidimetry and hemoglobin by clinico-chemical routine assay. Results 1. HD patients showed significantly higher serum concentrations of F2-isoprostanes and HNE than healthy human control subjects. 2. Total (esterified plus free) F2-isoprostane levels before HD were not significantly different from those after HD, whereas HNE levels were significantly decreased in patients after HD. 3. F2-isoprostane concentrations in HD patients correlated with the levels of CRP, whereas HNE concentrations inversely correlated with the content of hemoglobin. Conclusion Both, F2-isoprostanes and HNE serum concentrations are useful oxidative stress parameters in ESRD patients undergoing HD. Whereas HNE strongly correlates with the severity of renal anemia, leading to left heart insufficiency, F2-isoprostanes (sum of free plus esterified) highly correlate with the degree of inflammation.

Prevention of free fatty acid-induced lipid accumulation, oxidative stress, and cell death in primary hepatocyte cultures by a Gynostemma pentaphyllum extract

Hepatocytes of a primary cell culture that are exposed to high glucose, insulin, and linoleic (LA) acid concentration respond with lipid accumulation, oxidative stress up to cell death. Such alterations are typically found in patients with non-alcoholic fatty liver disease (NAFLD). We used this cellular model to study the effect of an ethanolic Gynostemma pentaphyllum (GP) extract in NAFLD. When hepatocytes were cultured in the presence of high insulin, glucose, and LA concentration the extract completely protected the cells from cell death. In parallel, the extract prevented accumulation of triglycerides (TGs) and cholesterol as well as oxidative stress. Our data further demonstrate that GP stimulates the production of nitric oxide (NO) in hepatocytes and affects the molecular composition of the mitochondrial phospholipid cardiolipin (CL). We conclude that GP is able to protect hepatocytes from cell death, lipid accumulation, and oxidative stress caused by diabetic-like metabolism and lipotoxicity. Therefore, GP could be beneficial for patients with diabetes mellitus and NAFLD.

Oxysterols Are Increased in Plasma of End-Stage Renal Disease Patients

Background/Aims: Oxidative stress occurs in chronic renal failure patients undergoing hemodialysis (HD). The objective of our study was to measure oxidation products of cholesterols, so-called oxysterols, in the serum of HD patients in comparison to healthy control persons. Methods: In 42 HD patients, plasma oxysterols were measured before and after HD. The values were compared with those in 40 healthy controls. The following cholesterol derivatives were analyzed: dienes, 7β-OH, β-epoxy, α-epoxy, 20α-OH, α-triol, and 7-keto cholesterol. Results: In HD patients, serum levels of oxysterols are increased in comparison to controls. The highest values were measured for β-epoxy cholesterol and for 20α-OH cholesterol. During HD oxysterol concentrations increased, obviously by water removal and concentration of nondialyzable compounds. Conclusion: Due to oxidative stress which is known as a typical sign of chronic renal failure the plasma concentrations of oxysterols are also significantly increased in comparison to healthy controls. This underlines the data on accelerated lipid peroxidation in end-stage renal disease (ESRD) patients. Accumulated oxysterols which are accused of exerting atherosclerosis-stimulating effects, which can contribute to the increased cardiovascular risk of ESRD patients, could either induce atherosclerosis via signaling or chronic effects. Direct chemical reactions stimulating plaque formation can be excluded because of the low levels of oxysterols. The share of oxysterols within the total cholesterol ranges from 4 to 15‰.

Cardiolipin composition correlates with prostate cancer cell proliferation

Prostate cancer (PC) is the second most diagnosed cancer in men. It has been recognized that diet can play a crucial role in PC genesis and progression. In this context, free fatty acids are considered as modulators of cell proliferation. Recently, a relationship between the composition of the mitochondrial phospholipid cardiolipin (CL) and cell proliferation has been discussed. The aim of this study was to analyse the interrelationship between CL composition and the proliferation of prostate cells by exposing PC-3 tumour cells to different fatty acids and by analysing the CL composition in prostate tissue from PC patients after prostatectomy. Among the applied fatty acids, palmitic acid was found to stimulate proliferation of PC-3 cells, whereas oleic acid (OA) had an inhibiting effect. The lipidomic analysis of CL revealed that fatty acids supplied to PC-3 cells were incorporated into CL molecules. Further, the CL content of palmitoleic acid (C16:1) exclusively correlated with the proliferation of PC-3 cells. The CL composition significantly differed between tumour and normal prostate tissue from PC patients. In five out of six patients, the CL content of palmitoleic acid was higher in tumour prostate tissue in comparison to normal prostate tissue. Our data illustrate that the composition of CL can be easily modified by the fatty acid environment of cells. OA was most effective in decreasing the amount of palmitoleic acid within the CL molecules and deceleration of PC-3 cell proliferation. In conclusion, a diet rich in OA might be beneficial in protecting from rapid proliferation of PC cells.

Palmitate protects hepatocytes from oxidative stress and triacylglyceride accumulation by stimulation of nitric oxide synthesis in the presence of high glucose and insulin concentration

Abstract Excessive flux of free fatty acids (FFA) into the liver contributes to liver impairment in non-alcoholic fatty liver disease (NAFLD). It remains unclear how FFA contribute to impairment of hepatocytes. This study treated hepatocytes with linoleic acid and palmitate to investigate the early event triggering FFA-mediated impairment. It determined cell viability, content of nitrite/nitrate and triacylglycerides (TG), inducible nitric oxide synthase (iNOS) protein, oxidation of cardiolipin (CL) as well as formation of F2-isoprostanes in the presence of insulin and glucose. Linoleic acid caused significant decrease in cell viability. It is shown that palmitate caused induction of iNOS resulting in increased nitrite/nitrate concentration and slight increase in TG content. Linoleic acid led to a decrease in nitrite/nitrate concentration parallelled by massive TG accumulation in combination with increased oxidation of CL and increased F2-isoprostane levels. It is concluded that nitric oxide (NO) concentration regulates FFA-dependent TG accumulation and oxidative stress in rat hepatocytes.

Effects of siRNA-dependent knock-down of cardiolipin synthase and tafazzin on mitochondria and proliferation of glioma cells

The mitochondrial phospholipid cardiolipin (CL) has been implicated with mitochondrial morphology, function, and cell proliferation. Changes in CL are often paralleled by changes in the lipid environment of mitochondria that may contribute to mitochondrial function and proliferation. This study aimed to separate the effects of CL content and CL composition from cellular free fatty acid distribution on bioenergetics and proliferation in C6 glioma cells. To this end, cardiolipin synthase and the CL remodelling enzyme, tafazzin, were knocked-down by siRNA in C6 cells. After 72 h of cultivation, we analysed CL composition by means of LC/MS/MS, distribution of cellular fatty acids by means of gas chromatography, and determined oxygen consumption and proliferation. Knock-down of cardiolipin synthase affected the cellular CL content in the presence of linoleic acid (LA) in the culture medium. Knock-down of tafazzin had no consequence with respect to the pattern of cellular fatty acids but caused a decrease in cell proliferation. It significantly changed the distribution of molecular CL species, increased CL content, decreased oxygen consumption, and decreased cell proliferation when cultured in the presence of linoleic acid (LA). The addition of linoleic acid to the culture medium caused significant changes in the pattern of cellular fatty acids and the composition of molecular CL species. These data suggest that tafazzin is required for efficient bioenergetics and for proliferation of glioma cells. Supplementation of fatty acids can be a powerful tool to direct specific changes in these parameters.