Tafazzin-an acyltransferase-is involved in cardiolipin (CL) remodeling. CL is associated with mitochondrial function, structure and more recently with cell proliferation. Various tafazzin isoforms exist in humans. The role of these isoforms in cardiolipin remodeling is unknown. Aim of this study was to investigate if specific isoforms like Δ5 can restore the wild type phenotype with respect to CL composition, cellular proliferation and gene expression profile. In addition, we aimed to determine the molecular mechanism by which tafazzin can modulate gene expression by applying promoter analysis and (Ingenuity Pathway Analyis) IPA to genes regulated by TAZ-deficiency. Expression of Δ5 and rat full length TAZ in C6-TAZ- cells could fully restore CL composition and-as proven for Δ5-this is naturally associated with restoration of mitochondrial respiration. A similar restoration of CL-composition could not be observed after re-expression of an enzymatically dead full-length rat TAZ (H69L; TAZMut). Re-expression of only rat full length TAZ could restore proliferation rate. Surprisingly, the Δ5 variant failed to restore wild-type proliferation. Further, as expected, re-expression of the TAZMut variant completely failed to reverse the gene expression changes, whereas re-expression of the TAZ-FL variant largely did so and the Δ5 variant to somewhat less extent. Very likely TAZ-deficiency provokes substantial long-lasting changes in cellular lipid metabolism which contribute to changes in proliferation and gene expression, and are not or only very slowly reversible.

The mitochondrial phospholipid (CL) has been linked to mitochondrial and cellular functions. It has been postulated that the composition of CL is of impact for mitochondrial energy metabolism and cell proliferation. Although a correlation between CL composition and proliferation could be demonstrated for several cell types, evidence for a causal relationship remains obscure. Here, we applied two independent approaches, i) supplementation of fatty acids and ii) knock-out of the phospholipid remodeling enzyme tafazzin, to manipulate CL composition and analyzed the response on proliferation of C6 glioma cells. Both strategies caused substantial changes in the distribution of cellular fatty acids as well as in the distribution of fatty acids incorporated in CL that were accompanied by changes of the composition of molecular CL species. These changes did not correlate with cell proliferation. However, knock-out of tafazzin caused dramatic reduction in proliferation of C6 glioma cells independent of CL composition. The mechanism of tafazzin-dependent restriction of proliferation remains unclear. Among the various fatty acids administered only palmitic acid restricted cell proliferation by induction of cell death.

There is growing evidence for the contribution of the activated coagulation factor X (FXa) in the development of chronic inflammatory lung diseases. Therefore, we aimed to investigate effects of exogenous FXa on mitochondrial and metabolic function as well as the induction of inflammatory molecules in type II alveolar epithelial cells. Effects of FXa on epithelial cells were investigated in A549 cell line. Activation of extracellular signal-regulated kinase (ERK) and induction of inflammatory molecules were examined by immunoblot and gene expression analysis. Mitochondrial function was assessed by the measurement of oxygen consumption during maximal oxidative phosphorylation and quantitative determination of cardiolipin oxidation. Apoptosis was tested using a caspase 3 antibody. Metabolic activity and lactate dehydrogenase assay were applied for the detection of cellular viability. FXa activated ERK1/2 and induced an increase in the expression of pro-inflammatory cytokines, which was prevented by an inhibitor of FXa, edoxaban, or an inhibitor of protease-activated receptor 1, vorapaxar. Exposure to FXa caused mitochondrial alteration with restricted capacity for ATP generation, which was effectively prevented by edoxaban, vorapaxar and GB83 (inhibitor of protease-activated receptor 2). Of note, exposure to FXa did not initiate apoptosis in epithelial cells. FXa-dependent pro-inflammatory state and impairment of mitochondria did not reach the level of significance in lung epithelial cells. However, these effects might limit regenerative potency of lung epithelial cells, particular under clinical circumstances where lung injury causes exposure to clotting factors.

Vitamin B6 deficiency during pregnancy translates into a severe vitamin B6 deficiency (plasma levels decreased by 97%) in new-born rats. Further, hallmarks are increased (+89%) concentrations of homocysteine, gross changes in gene methylation and expression, and metabolic alterations including lipid metabolism. This study focuses on determining the effects of vitamin B6-deficiency on cardiolipin composition and oxidative phosphorylation in liver. For this purpose, hepatic cardiolipin composition was analyzed by means of LC/MS/MS, and mitochondrial oxygen consumption was determined by using a Clark-type electrode in a rat model of vitamin B6 deficiency. Liver mitochondria from new-born rats with pre-term vitamin B6 deficiency responded with substantial alterations in cardiolipin composition that include the following changes in the amounts of cardiolipin incorporated fatty acids: increase in C16, decrease in C18, decrease in saturated fatty acid, as well as increase in amount of oxidized cardiolipin species. These changes were accompanied by significantly decreased capacity of oxidative phosphorylation. In conclusion, vitamin B6 deficiency in new born rats induces massive alterations of cardiolipin composition and function of liver mitochondria. These findings support the importance of sufficient periconceptional supply of vitamin B6 to prevent vitamin B6 deficiency. Impact statement Vitamin B6 (VitB6) is an active co-enzyme for more than 150 enzymes and is required for a great diversity of biosynthesis and metabolic reactions. There is an increased need for VitB6 during pregnancy and sufficient supply of VitB6 is crucial for the prevention of cleft palate and neural tube defects. We show that liver mitochondria from new-born rats with pre-term VitB6 deficiency respond with substantial alterations in cardiolipin (CL) composition and in the amount of oxidized CL species. These changes are associated with a decrease in the efficiency of oxidative phosphorylation. The results of this study support the significance of sufficient supply of VitB6 during pregnancy (and periconceptional) for diminishing the number of early abortions and minimizing malformation. The established link between VitB6 deficiency, CL composition, and mitochondrial respiration/energy production provides mechanistic insight as to how the VitB6 deficiency translates into the known pathophysiological and clinically relevant conditions.

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.

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.

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 F-2-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 F-2-isoprostane levels. It is concluded that nitric oxide (NO) concentration regulates FFA-dependent TG accumulation and oxidative stress in rat hepatocytes.

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.

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, F-2-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 F-2-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) F-2-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 F-2-isoprostanes and HNE than healthy human control subjects. 2. Total (esterified plus free) F-2-isoprostane levels before HD were not significantly different from those after HD, whereas HNE levels were significantly decreased in patients after HD. 3. F-2-isoprostane concentrations in HD patients correlated with the levels of CRP, whereas HNE concentrations inversely correlated with the content of hemoglobin.Conclusion: Both, F-2-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, F-2-isoprostanes (sum of free plus esterified) highly correlate with the degree of inflammation.

F2-isoprostanes diclofenac cyclo-oxygenase TO THE EDITOR UVB irradiation of the skin induces a complex cascade of acute inflammation, characterized by erythema, edema, and immunosuppression and is according to chronic exposure subsequently linked to the initiation and progression of skin cancer (Matsumura and Ananthaswamy, 2004Matsumura Y. Ananthaswamy H.N. Toxic effects of ultraviolet radiation on the skin.Toxicol Appl Pharmacol. 2004; 195: 298-308Crossref PubMed Scopus (762) Google Scholar). The role of inflammatory mediators in skin diseases is of critical importance. This includes the release of cytokines, NO, and oxidized arachidonic acid derivatives (Reilly et al., 2000Reilly D.M. Parslew R. Sharpe G.R. Powell S. Green M.R. Inflammatory mediators in normal, sensitive and diseased skin types.Acta Derm Venereol. 2000; 80: 171-174Crossref PubMed Scopus (71) Google Scholar; Rhodes et al., 2001Rhodes L.E. Belgi G. Parslew R. McLoughlin L. Clough G.F. Friedmann P.S. Ultraviolet-B-induced erythema is mediated by nitric oxide and prostaglandin E2 in combination.J Invest Dermatol. 2001; 117: 880-885Crossref PubMed Google Scholar). The so-called eicosanoid lipid mediators are formed by two main pathways: enzymatically and non-enzymatically. Inflammatory stimuli induce the liberation of arachidonic acid from membrane phospholipids and the formation of prostaglandins, which are generated by cyclo-oxygenase (COX) isoenzymes and are blocked by non-steroidal anti-inflammatory drugs. Prostaglandins are known as bioactive products with vasoregulatory properties acting in a synergistic or antagonistic manner. Whereas PGE2 mediates vasodilatation, PGF2α is responsible for vasoconstriction. The non-enzymatical pathway of arachidonic acid oxidation includes free radical mechanisms and synthesis of F2-isoprostanes (F2-IsoPs), which are mostly bound to the backbone of phospholipids (Morrow et al., 1992Morrow J.D. Awad J.A. Boss H.J. Blair I.A. Roberts L.J. Non-cyclooxygenase-derived prostanoids (F2-isoprostanes) are formed in situ in phospholipids.Proc Natl Acad Sci (USA). 1992; 89: 10721-10725Crossref PubMed Scopus (640) Google Scholar). F2-IsoPs are a novel group of prostaglandin-like compounds with potent biological activities, as evidenced by their pulmonary and renal vasoconstrictive effects (Lawson et al., 1999Lawson J.A. Rokach J. FitzGerald G.A. Isoprostanes: formation, analysis and use as indices of lipid peroxidation in vivo.J Biol Chem. 1999; 274: 24441-24444Crossref PubMed Scopus (342) Google Scholar). Several studies have indicated associations between isoprostanes and severe inflammatory conditions (Basu, 2004Basu S. Isoprostanes: novel bioactive products of lipid peroxidation.Free Radic Res. 2004; 38: 105-122Crossref PubMed Scopus (188) Google Scholar). Although F2-IsoPs are preferentially formed non-enzymatically, a COX-dependent generation can not be excluded (Klein et al., 1996Klein T. Reutter F. Schweer H. Seyberth H.W. Nüsing R.M. Generation of the isoprostane 8-epi-prostaglandin F2α in vitro and in vivo via the cyclooxygenases.J Pharmacol Exp Ther. 1996; 282: 1658-1665Google Scholar; Lawson et al., 1999Lawson J.A. Rokach J. FitzGerald G.A. Isoprostanes: formation, analysis and use as indices of lipid peroxidation in vivo.J Biol Chem. 1999; 274: 24441-24444Crossref PubMed Scopus (342) Google Scholar). Therefore, F2-IsoPs are not only biomarkers of lipid peroxidation and potential in vivo indicators of oxidant stress in various clinical conditions, but also mediators of acute inflammation (Basu, 2004Basu S. Isoprostanes: novel bioactive products of lipid peroxidation.Free Radic Res. 2004; 38: 105-122Crossref PubMed Scopus (188) Google Scholar; Montuschi et al., 2004Montuschi P. Barnes P.J. Roberts L.J. Isoprostanes: markers and mediators of oxidative stress.FASEB J. 2004; 18: 1791-1800Crossref PubMed Scopus (548) Google Scholar). Diclofenac (DCLF) belongs to the most common non-steroidal anti-inflammatory drugs used for the treatment of pain and inflammation. DCLF (2-[(2,6-dichlorophenyl)amino] phenyl-acetate) non-selectively inhibits COX-1 and COX-2, which both catalyze prostaglandin synthesis (Hickey et al., 2001Hickey E.J. Raje R.R. Reid V.E. Gross S.M. Ray S.D. Diclofenac induced in vivo nephrotoxicity may involve oxidative stress-mediated massive genomic DNA fragmentation and apoptotic cell death.Free Radic Biol Med. 2001; 31: 139-152Crossref PubMed Scopus (160) Google Scholar). It was the aim of our study to test the hypotheses that1.low-dose UVB irradiation influences the concentrations of F2-IsoPs and prostaglandins in vitro in human HaCaT keratinocytes and in vivo in the interstitial fluid of the dermis and this could be a dose-dependent action.2.application of non-toxic concentrations of DCLF can lower the levels of enzymatically and of non-enzymatically formed lipid mediators as well.3.HaCaT keratinocytes may serve as a suitable in vitro model to reflect UVB- and DCLF-induced alterations of lipid mediators in vivo. In this study, we showed early evidence that in HaCaT keratinocytes both F2-IsoPs and PGF2α can reliably be quantified by gas chromatography–mass spectrometry (negative ion chemical ionization–selected ion monitoring (Table 1a) and that their concentrations were dose-dependently enhanced with low-dose UVB irradiation between 30 and 100 mJ/cm2 (Figure 1). 8-iso-PGF2α as marker of non-enzymatic arachidonic acid oxidation exclusively reflects phospholipid-bound F2-IsoPs, because (i) it occurred in HaCaT keratinocytes completely in the esterified and not in the free form (experiments according to Wiswedel et al., 2004Wiswedel I. Hirsch D. Kropf S. Gruening M. Pfister E. Schewe T. et al.Flavanol-rich cocoa drink lowers plasma F2-isoprostane concentrations in humans.Free Radic Biol Med. 2004; 37: 411-421Crossref PubMed Scopus (136) Google Scholar) and (ii) no 8-iso-PGF2α was released and detected in the cell-free supernatant (Quist et al., 2006Quist S.R. Simmel F. Wiswedel I. Neubert R. Gollnick H. Influence of green and black tea, epigallocatechin-3-gallate and theaflavin on prostanoid synthesis in vitro and in vivo using microdialysis.Proceedings of the 13th Biennial Congress of the International Society for Free Radical Research – SFRR. Medimond, Davos, Switzerland2006: pp 293-297Google Scholar). Concerning 9α,11α-PGF2α, it can be assumed that the main part represents COX-catalyzed generation. This was derived from the predominant occurrence in the free and non-esterified form (about 77%) and by the considerable release of PGF2α in the supernatant (Quist et al., 2006Quist S.R. Simmel F. Wiswedel I. Neubert R. Gollnick H. Influence of green and black tea, epigallocatechin-3-gallate and theaflavin on prostanoid synthesis in vitro and in vivo using microdialysis.Proceedings of the 13th Biennial Congress of the International Society for Free Radical Research – SFRR. Medimond, Davos, Switzerland2006: pp 293-297Google Scholar). COX-derived PGE2 was not detectable in HaCaT cells, but was completely released in the supernatant (Table 1a). The UVB-induced enhancement of PGE2 was definitely more pronounced compared with the other mediators.Table 1aLevels of F2-isoprostanes/prostaglandins in HaCaT keratinocytes and supernatants – Influence of UVB irradiation and DCLF8-iso-PGF2α9α,11α-PGF2αPGE2(pmol/mg protein in HaCaTs)(pmol/mg protein in HaCaTs)(pmol/ml supernatant)Non-irradiated cells0.23±0.030.50±0.060.21±0.08(n=26)(n=22)(n=3)UVB (30 mJ/cm2)0.35±0.061.21±0.2862.5±18.8(n=19)(n=17)(n=3)UVB (30 mJ/cm2)0.32±0.070.95±0.03*n.d.+2.5 μm DCLF(n=6)(n=6)UVB (30 mJ/cm2)0.23±0.05**For 8-iso-PGF2a, UVB+5µm DCLF is significantly different from UVB (P<0.05). In the case of 9a,11a-PGF2a, UVB+2.5µm DCLF as well as UVB+5µm DCLF are significantly different vs UVB (P=0.010 and 0.004); Student's t-test.0.79±0.18*0.25±0.05+5.0 μm DCLF(n=6)(n=6)(n=3)DCLF, diclofenac; n.d., not determined.Values are means±SEM for HaCaT cells and±SD for supernatants. Values of non-irradiated and UVB-irradiated HaCaT keratinocytes (30 mJ/cm2) are identical with those of Figure 1.* *For 8-iso-PGF2a, UVB+5 µm DCLF is significantly different from UVB (P<0.05). In the case of 9a,11a-PGF2a, UVB+2.5 µm DCLF as well as UVB+5 µm DCLF are significantly different vs UVB (P=0.010 and 0.004); Student's t-test. Open table in a new tab DCLF, diclofenac; n.d., not determined. Values are means±SEM for HaCaT cells and±SD for supernatants. Values of non-irradiated and UVB-irradiated HaCaT keratinocytes (30 mJ/cm2) are identical with those of Figure 1. In Figure 1, it is shown that 8-iso-PGF2α concentrations were slightly and more continuously increased up to 100 mJ/cm2, whereas 9α,11α-PGF2α concentrations already reached a maximum at 50 mJ/cm2 with a more than fivefold enhancement compared with the non-irradiated control values and dropped down thereafter to about half-maximum levels. The decrease of PGF2α in HaCaTs may reflect the enhanced release in the supernatant as shown by Quist et al., 2006Quist S.R. Simmel F. Wiswedel I. Neubert R. Gollnick H. Influence of green and black tea, epigallocatechin-3-gallate and theaflavin on prostanoid synthesis in vitro and in vivo using microdialysis.Proceedings of the 13th Biennial Congress of the International Society for Free Radical Research – SFRR. Medimond, Davos, Switzerland2006: pp 293-297Google Scholar or a partial transformation to the more stable 15-keto-dihydro-PGF2α, which was not analyzed in this study. The different UVB dependence of 8-iso-PGF2α and 9α,11α-PGF2α may be an additional indication for their different origin: free radical-catalyzed or COX-derived. The results correspond to those of Pentland et al., 1990Pentland A.P. Mahoney M. Jacobs S.C. Holtzmann M.J. Enhanced prostaglandin synthesis after ultraviolet injury is mediated by endogenous histamine stimulation. A mechanism for irradiation erythema.J Clin Invest. 1990; 86: 566-574Crossref PubMed Scopus (97) Google Scholar showing fivefold increases of PGF2α and PGE2 in human skin explants following 120 mJ/cm2 UVB irradiation and those of Buckman et al., 1998Buckman S.Y. Gresham A. Hale P. Hruza G. Anast J. Masferrer J. et al.COX-2 expression is induced by UVB exposure in human skin: implication for the development of skin cancer.Carcinogenesis. 1998; 19: 723-729Crossref PubMed Scopus (491) Google Scholar, demonstrating Western blots with sixfold increases of COX-2 protein in cultured human keratinocytes 24 hours following 30 mJ/cm2 UVB exposure. In a next set of experiments, the influence of DCLF on the UVB-induced enhancement of prostanoids was investigated (Table 1a). The DCLF concentrations used in HaCaT keratinocytes were between 1 and 5 μm and a low, but effective, UVB dose of 30 mJ/cm2 was chosen. Concentrations of 1 μm DCLF or less were not effective at all and concentrations of 10 μm and higher were proven to be toxic to HaCaT cell cultures. The UVB-induced increases of 8-iso-PGF2α and of 9α,11α-PGF2α were suppressed by about 35 % with 5 μm DCLF, whereas only that of 9α,11α–PGF2α were suppressed by about 12 % with 2.5 μm DCLF (Table 1a). PGE2 concentrations were followed in the supernatant. In contrast to the mediators before, they were about 300-fold enhanced by UVB and nearly totally reduced to control levels by DCLF (Table 1a). The results obtained with HaCaT keratinocytes in vitro have been qualitatively confirmed using a microdialysis technique, which has been used in human studies for more than 10 years (Stahl et al., 2002Stahl M. Bouw R. Jackson A. Pay V. Human microdialysis.Curr Pharm Biotechnol. 2002; 3: 165-178Crossref PubMed Scopus (50) Google Scholar; Grundmann et al., 2004Grundmann J.U. Wiswedel I. Hirsch D. Gollnick H.P.M. Detection of monohydroxyeicosatetraenoic acids and F2-isoprostanes in microdialysis samples of human UV-irradiated skin by gas chromatography-mass spectrometry.Skin Pharmacol Physiol. 2004; 17: 37-41Crossref PubMed Scopus (23) Google Scholar). Six healthy female volunteers, 18–30 years old, were included in the study after giving their written, informed consent and after approval by the ethics committee of the Medical Faculty, Otto-von-Guericke-University Magdeburg, Germany. The Declarations of Helsinki Principles were followed. Microdialysis allows the "near in vivo" measurement of endogeneous eicosanoid mediators released in the interstitial space of the dermis under control and inflammatory conditions (Table 1b). Whereas UVB- and DCLF-induced alterations of 9α,11α-PGF2α concentrations were not significant, the concentrations of 8-iso-PGF2α and PGE2 were four and 2.7-fold enhanced 24 hours following UVB treatment in comparison with healthy untreated skin. DCLF, topically applied as cream or gel, immediately after UVB irradiation, prevented the generation of an erythema and reduced the levels of 8-iso-PGF2α and PGE2 remarkably (Table 1b). The anti-inflammatory effect of DCLF was more pronounced for the 5% cream than for the 1% gel. As found in HaCaT cells, DCLF did not only influence the prostaglandin, but additionally the isoprostane pathway. The so-called radical scavenging or antioxidant side effects of DCLF were also described by Aruoma and Halliwell, 1988Aruoma O.I. Halliwell B. The iron-binding and hydroxyl radical scavenging action of anti-inflammatory drugs.Xenobiotica. 1988; 18: 459-470Crossref PubMed Scopus (139) Google Scholar and Maffei Facino et al., 1993Maffei Facino R. Carini M. Aldini G. Saibene L. Macciocchi A. Antioxidant profile of nimesulide, indomethacin and diclofenac in phosphatidylcholine liposomes as membrane model.Int J Tissue React. 1993; 15: 225-234PubMed Google Scholar.Table 1bLevels of F2-isoprostanes/prostaglandins in microdialysis samples of human skin – influence of UVB irradiation and treatment with DCLF8-iso-PGF2α9α,11α- PGF2αPGE2(pmol/ml microdialysate)(pmol/ml microdialysate)(pmol/ml microdialysate)Non-irradiated skin0.25±0.110.53±0.250.62±0.14UVB1.00±0.22*P<0.05 vs non-irradiated control0.38±0.281.65±0.23*P<0.05 vs non-irradiated controlUVB+DCLF (1%)0.59±0.09*P<0.05 vs UVB without DCLF.0.62±0.350.97±0.13*P<0.05 vs UVB without DCLF.UVB+DCLF (5%)0.41±0.12*P<0.05 vs UVB without DCLF.0.55±0.300.81±0.12*P<0.05 vs UVB without DCLF.DCLF, diclofenac.Values are means±SEM of n=6 healthy volunteers.UVB irradiation (about 20–40 mJ/cm2) caused the formation of erythema of stages 3–5, which were completely suppressed by DCLF immediately applicated following UVB irradiation. Samples were taken 24 h later; after an additional hour of "baseline" microdialysis for equilibration. Microdialysate samples of about 2 hours have to be combined for triplicate analyses. Differences in the levels of prostanoids are significant for 8-iso-PGF2α and PGE2 according to Student's t-test.* P<0.05 vs non-irradiated control** P<0.05 vs UVB without DCLF. Open table in a new tab DCLF, diclofenac. Values are means±SEM of n=6 healthy volunteers. UVB irradiation (about 20–40 mJ/cm2) caused the formation of erythema of stages 3–5, which were completely suppressed by DCLF immediately applicated following UVB irradiation. Samples were taken 24 h later; after an additional hour of "baseline" microdialysis for equilibration. Microdialysate samples of about 2 hours have to be combined for triplicate analyses. Differences in the levels of prostanoids are significant for 8-iso-PGF2α and PGE2 according to Student's t-test. Prostanoid concentrations measured by gas chromatography–mass spectrometry in this study were on the same level as isoprostane concentrations reported by Karamouzis et al., 2004Karamouzis I. Christoulas K. Grekas D. Giannoulis K. Vamvakoudis E. Mandroukas K. The response of muscle interstitial F2-isoprostane (8-iso-PGF2alpha) during dynamic muscle concentrations in humans.Prostaglandins Leukot Essent Fatty Acids. 2004; 71: 87-90Abstract Full Text Full Text PDF PubMed Scopus (17) Google Scholar in dialysate samples from muscle and PGE2 levels in microdialysates of human skin reported by Fairweather et al., 2004Fairweather I. McGlone F. Reilly D. Rukwied R. Controlled dermal cell damage as human in vivo model for localised pain and inflammation.Inflamm Res. 2004; 53: 118-123Crossref PubMed Scopus (12) Google Scholar. UVB-induced increases of 8-iso-PGF2α were also shown by Schneider et al., 2006Schneider L.A. Bloch W. Kopp K. Hainzl A. Rettberg P. Wlaschek M. et al.8-isoprostane is a dose-related biomarker for photo-oxidative ultraviolet (UV) B damage in vivo: a pilot study with personal UV dosimetry.Br J Dermatol. 2006; 6: 1147-1154Crossref Scopus (27) Google Scholar in human skin biopsies and by Kuhn et al., 2006Kuhn M. Wolber R. Kolbe L. Schnorr O. Sies H. Solar-simulated radiation induces secretion of IL-6 and production of isoprostanes in human skin in vivo.Arch Dermatol Res. 2006; 297: 477-479Crossref PubMed Scopus (18) Google Scholar in suction blister fluid 24 hours post-irradiation. The authors state no conflict of interest. The experimental work was supported by grants of the Federal State of Saxony-Anhalt (FKZ: 3173A/0089 M) and from the BMBF-Forschungsverbundprojekt (FKZ 01ZZ0407). We thank Rita Adolf and Marita Lotzing for excellent technical assistance. Methods. Figure S1. Standard calibration curves for 8-iso-PGF2α and 9α,11α-PGF2α. Download .pdf (.05 MB) Help with pdf files Supplementary Information

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, 7 -OH, epoxy, -epoxy, 20 -OH, -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 -epoxy cholesterol and for 20 -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 Published online: March 7, 2006

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, 7beta-OH, beta-epoxy, alpha-epoxy, 20alpha-OH, alpha-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 beta-epoxy cholesterol and for 20alpha-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 per thousand.

Patients with end-stage renal disease (ESRD) represent a growing number of patients in the cardiac catheterization laboratories worldwide. This is a consequence of the growing absolute number of ESRD patients in developed countries, better noninvasive diagnostic tools, better catheterization facilities and last-but-not-least better education of referring physicians about the incidence and prognosis of coronary artery disease (CAD) for patients with ESRD. There is growing evidence of the positive impact of coronary revascularization on long-term outcome of these patients. ESRD patients have a high comorbidity and are therefore better candidates for the less invasive approach using percutaneous coronary intervention (PCI) rather than coronary artery bypass surgery (CABG). From the view of the interventional cardiologist, ESRD patients represent one of the most challenging patient cohort concerning technical challenges and potential risk of complication for the patient. Percutaneous coronary intervention (PCI) including debulking techniques and stent implantation is the current standard therapy for patients with symptomatic single-vessel disease (SVD) and the preferred therapy for most patients with focal, polyfocal or even diffuse multi-vessel disease (MVD). Coronary bypass surgery is reserved for a dePublished online: March 7, 2006 Priv.-Doz. Dr. med. Wolfgang Bocksch, Department of Internal Medicine-Cardiology Cardiac Catheterization Laboratories, Charité-Campus Virchow-Klinikum Universitätsmedizin Berlin, Humboldt Universität zu Berlin and Freie Universität Berlin Augustenburger Platz 1, DE–13353 Berlin (Germany) Tel. +49 30 450 553 283, Fax +49 30 450 553 961, E-Mail wolfgang.bocksch@charite.de © 2005 S. Karger AG, Basel 1420–4096/05/0286–0275$22.00/0 Accessible online at: