[Metabolic surgery and remission of type 2 diabetes]. / Metabolische Chirurgie und Remission des Diabetes mellitus Typ 2.

A new therapeutic field is developing from bariatric surgery. Gastrointestinal surgery does not only seem to be a very efficacious method for weight reduction but may also have the potential to reduce obesity-related metabolic disorders. Even if there is still a lack of prospective randomized trials evaluating the correct indications for metabolic surgery in patients with type 2 diabetes mellitus, there is the legitimate expectation that the surgical approach can be successful. This article provides an overview of the current state of bariatric surgery and gives a surgical perspective on the treatment of type 2 diabetes mellitus.

Fluvastatin/fenofibrate vs. simvastatin/ezetimibe in patients with metabolic syndrome: different effects on LDL-profiles.

BACKGROUND: Patients with metabolic syndrome (MS) and type 2 diabetes (T2DM) show increased risk for coronary artery disease. Lipoprotein metabolism is characterized by elevated triglycerides (TG), low high-density lipoprotein cholesterol (HDL-C) and predominance of atherogenic small, dense low-density lipoprotein (sdLDL), while low-density lipoprotein (LDL) cholesterol is only slightly elevated. METHODS: Multicentre, randomized, open-label cross-over study investigating the effect of combination of fluvastatin/fenofibrate (80/200 mg) (F&F) on LDL-subfractions compared with combination of simvastatin/ezetimibe (20/10 mg) (S&E) in patients with MS/T2DM. RESULTS: Seventy-five patients were randomized, 69 completed the study and LDL-subfractions of 56 patients were analysed. Thirty-eight out of 56 patients (68%) showed a profile dominated by sdLDL. In these, TG and total cholesterol (TC) were elevated compared with non-sdLDL patients. In all patients, reduction of TC and LDL cholesterol (LDL-C) by S&E was stronger than by F&F. The increase of HDL-C was stronger with S&E in the non-sdLDL group, whereas in the sdLDL group, there was no difference between treatments. In non-sdLDL patients, there was no effect on TG or LDL-radius. However, in the sdLDL group, F&F was more effective in reducing TG and increased LDL radius, whereas S&E reduced LDL radius even further. CONCLUSIONS: S&E is more efficient in reducing TC and LDL-C. This is also true for HDL-C increase in non-sdLDL patients. However, in patients with sdLDL, F&F was more efficient in reducing TG and increasing LDL radius.

Epicatechin selectively prevents nitration but not oxidation reactions of peroxynitrite.

The flavanol (-)-epicatechin has been found to protect against damage inflicted by peroxynitrite, an inflammatory intermediate. Here, epicatechin was tested in systems of increasing complexity. The compound efficiently protected against nitration of protein tyrosine residues by peroxynitrite (IC(50) approximately 0.02 mol epicatechin/mol peroxynitrite). However, at epicatechin concentrations completely preventing nitration of tyrosine by peroxynitrite, protection against the oxidative inactivation of glyceraldehyde-3-phosphate dehydrogenase or soybean lipoxygenase-1 was marginal (IC(50) > 1 mol epicatechin/mol peroxynitrite), approximately two orders of magnitude less. Likewise, epicatechin was relatively ineffective against oxidation of thiols in cell lysates, and against the oxidation of 2',7'-dichlorodihydrofluorescein in cultured cells. The activation of the kinases Akt/protein kinase B, ERK1/2 and p38-MAPK by peroxynitrite in murine aorta endothelial cells was not altered by epicatechin, suggesting that activation of these kinases is due to processes other than tyrosine nitration.

Evidence for the presence of phospholipid hydroperoxide glutathione peroxidase in human platelets: implications for its involvement in the regulatory network of the 12-lipoxygenase pathway of arachidonic acid metabolism.

The 12-lipoxygenase pathway of arachidonic acid metabolism in platelets and other cells is bifurcated into a reduction route yielding 12-hydroxyeicosatetraenoic acid (12-HETE) and an isomerization route forming hepoxilins. Here we show for the first time the presence of phospholipid hydroperoxide glutathione peroxidase (PHGPx) protein and its activity in platelets. The ratio of the activity of PHGPx to that of cytosolic glutathione peroxidase (GPx-1) was consistently found to be approx. 1:60 in platelets and UT7 megakaryoblasts. Moreover, short-lived PHGPx mRNA was detected in megakaryocytes but not in platelets. Carboxymethylation of selenium-containing glutathione peroxidases by iodoacetate, which results in the inactivation of PHGPx and GPx-1 without inhibition of 12-lipoxygenase, markedly altered the pattern of arachidonic acid metabolism in human platelets. Whereas the formation of 12-HETE was inhibited by 80%, a concomitant accumulation of 12-hydroperoxyeicosatetraenoic acid (12-HpETE) by two orders of magnitude as well as the formation of hepoxilins A(3) and B(3) were observed. The formation of hepoxilins also occurred when 12-HpETE was added to untreated platelets. In selenium-deficient UT7 cells, which were devoid of GPx-1 but not of PHGPx, the reduction of 12-HPETE was retained, albeit with a lower rate than in control cells containing GPx-1. We therefore believe that both GPx-1 and PHGPx are involved in the regulatory network of the 12-lipoxygenase pathway in platelets and other mammalian cells. Moreover, the diminution of hydroperoxide tone in platelets incubated with arachidonic acid leads primarily to the formation of 12-HETE, whereas the increase in hydroperoxide tone (a situation found under oxidative stress or selenium deficiency or on incubation with 12-HPETE) partly diverts the 12-lipoxygenase pathway from the reduction route to the isomerization route, thus resulting in the formation of hepoxilins.

Polyphenols of cocoa: inhibition of mammalian 15-lipoxygenase.

Some cocoas and chocolates are rich in (-)-epicatechin and its related oligomers, the procyanidins. Fractions of these compounds, isolated from the seeds of Theobroma cacao, caused dose-dependent inhibition of isolated rabbit 15-lipoxygenase-1 with the larger oligomers being more active; the decamer fraction revealed an IC50 of 0.8 microM. Among the monomeric flavanols, epigallocatechin gallate (IC50 = 4 microM) and epicatechin gallate (5 microM) were more potent than (-)-epicatechin (IC50 = 60 microM). (-)-Epicatechin and procyanidin nonamer also inhibited the formation of 15-hydroxy-eicosatetraenoic acid from arachidonic acid in rabbit smooth muscle cells transfected with human 15-lipoxygenase-1. In contrast, inhibition of the lipoxygenase pathway in J774A.1 cells transfected with porcine leukocyte-type 12-lipoxygenase (another representative of the 12/15-lipoxygenase family) was only observed upon sonication of the cells, suggesting a membrane barrier for flavanols in these cells. Moreover, epicatechin (IC50 approx. 15 microM) and the procyanidin decamer inhibited recombinant human platelet 12-lipoxygenase. These observations suggest general lipoxygenase-inhibitory potency of flavanols and procyanidins that may contribute to their putative beneficial effects on the cardiovascular system in man. Thus, they may provide a plausible explanation for recent literature reports indicating that procyanidins decrease the leukotriene/prostacyclin ratio in humans and human aortic endothelial cells.

Phospholipase A(2)s and lipid peroxidation.

Lipid peroxidation of membrane phospholipids can proceed both enzymatically via the mammalian 15-lipoxygenase-1 or the NADPH-cytochrome P-450 reductase system and non-enzymatically. In some cells, such as reticulocytes, this process is biologically programmed, whereas in the majority of biological systems lipid peroxidation is a deleterious process that has to be repaired via a deacylation-reacylation cycle of phospholipid metabolism. Several reports in the literature pinpoint a stimulation by lipid peroxidation of the activity of secretory phospholipase A(2)s (mainly pancreatic and snake venom enzymes) which was originally interpreted as a repair function. However, recent experiments from our laboratory have demonstrated that in mixtures of lipoxygenated and native phospholipids the former are not preferably cleaved by either secretory or cytosolic phospholipase A(2)s. We propose that the platelet activating factor (PAF) acetylhydrolases of type II, which cleave preferentially peroxidised or lipoxygenated phospholipids, are competent for the phospholipid repair, irrespective of their role in PAF metabolism. A corresponding role of Ca(2+)-independent phospholipase A(2), which has been proposed to be involved in phospholipid remodelling in biomembranes, has not been addressed so far. Direct and indirect 15-lipoxygenation of phospholipids in biomembranes modulates cell signalling by several ways. The stimulation of phospholipase A(2)-mediated arachidonic acid release may constitute an alternative route of the arachidonic acid cascade. Thus, 15-lipoxygenase-mediated oxygenation of membrane phospholipids and its interaction with phospholipase A(2)s may play a crucial role in the pathogenesis of diseases, such as bronchial asthma and atherosclerosis.

Mapping the distribution of 3-hydroxylipins in the Mucorales using immunofluorescence microscopy.

The distribution of endogenous 3-hydroxylipins (3-OH oxylipins) in representatives of the Mucorales was mapped using immunofluorescence microscopy. Strains of each of the following genera were examined: Absidia, Actinomucor, Cunninghamella, Mortierella (subgenus Micromucor), Mortierella (subgenus Mortierella), Mucor and Rhizomucor. Immunofluorescence microscopy was carried out using an antibody that was raised against 3R-hydroxy-5Z,8Z, 11 Z, 14Z-eicosatetraenoic acid (3R-HETE), which cross-reacts with other 3-OH oxylipins. Subsequently, the occurrence and distribution of the antibody on the various reproductive stages of each fungus was noted. In Absidia, Actinomucor, Mortierella (subgenus Micromucor), Mucor and Rhizomucor, 3-OH oxylipins were found to be associated with the columellae and/or wall of the sporangium. In the representative of Cunninghamella, the 3-OH oxylipins were associated with the single-spored sporangiola. No 3-OH oxylipins were detected in the strains representing Mortierella (subgenus Mortierella).

Arachidonic acid stimulates cell growth and forms a novel oxygenated metabolite in Candida albicans.

Infection of human tissues by Candida albicans has been reported to cause the release of arachidonic acid (AA), eicosanoids and other proinflammatory mediators from host cells. Therefore, we investigated the interaction of this pathogen with AA. AA stimulated cell growth at micromolar concentrations when used as a sole carbon source. Moreover, it selectively inhibited the antimycin A-resistant alternative oxidase. [1-(14)C]AA was completely metabolised by C. albicans. Only one-seventh of the radioactivity metabolised was found in CO(2), whereas two-thirds occurred in carbohydrates suggesting a predominant role of the glyoxalate shunt of citrate cycle. About 1% of radioactivity was found in polar lipids including eicosanoids. A novel AA metabolite, which revealed immunoreactivity with an antibody against 3(R)-hydroxy-oxylipins, was identified as 3, 18-dihydroxy-5,8,11,14-eicosatetraenoic acid. Using immunofluorescence microscopy, endogenous 3(R)-hydroxy-oxylipins were found in hyphae but not in yeast cells. Such compounds have recently been shown to be connected with the sexual stage of the life cycle of Dipodascopsis uninucleata. Together, we propose that infection-mediated release of AA from host cells may modulate cell growth, morphogenesis and invasiveness of C. albicans by several modes. A better understanding of its role is thus promising for novel approaches towards the treatment of human mycoses.

Biological actions of the free acid of hepoxilin A3 on human neutrophils.

In earlier reports and reviews, it was suggested that unlike its methyl ester, the free acid form of the 12-lipoxygenase-derived eicosanoid hepoxilin A3 (HXA3) does not enter neutrophils and other cells. Therefore, in the past, most studies on the biological activities of HXA3 on human neutrophils were conducted with its methyl ester. Here, we present evidence that free HXA3 is biologically active towards human neutrophils at submicromolar concentrations, which may occur under certain circumstances in vivo. Thus, HXA3 caused chemotaxis at concentrations as low as 30-40 nM, an effect which was attenuated at higher concentrations of this eicosanoid. Its chemotactic potency proved to be comparable to that of leukotriene B4, but higher than that of the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP), and greatly exceeded that of the other 12-lipoxygenase metabolite, 12(S)-hydroxy-5,8,10,14-eicosatetraenoic acid, which was inactive at comparable concentrations. The chemotactic activity of HXA3 was not abolished by serum albumin, but it was suppressed by pertussis toxin. Unlike fMLP, at this concentration range HXA3 did not cause respiratory burst or aggregation of the neutrophils or activation of protein kinase C. These observations suggest a remarkably selective and specific receptor-mediated process. At concentrations higher than 1 microM, HXA3 gives rise to an instantaneous release of calcium from intracellular stores which causes, however, only a slight, if any, liberation of arachidonic acid. On the other hand, pretreatment of the neutrophils with submicromolar concentrations of HXA3 significantly blunts the liberation of arachidonic acid caused by fMLP.

Metabolic suppression of platelet-type 12-lipoxygenase in human uterine cervix with invasive carcinoma.

Several types of lipoxygenases with various functions occur in mammalian cells. Although the presence of 12-lipoxygenase activity has been reported in uterine tissues, neither its type nor its biological functions have yet been established. Moreover, the putative role of uterine 12-lipoxygenase in cervical cancer has not been addressed before. Homogenates of uterine tissues from women without cancer and from patients with invasive cervical carcinoma were incubated with (1-(14)C)-arachidonic acid under various conditions and the labelled reaction products were analyzed both by thin-layer chromatography and by high-pressure liquid chromatography. 12-Lipoxygenase protein was estimated by Western blot using anti-serum against recombinant human platelet-type 12-lipoxygenase. Highest concentrations and activities of 12-lipoxygenase were found in the exocervix. The formation of 12S-hydroxy-5Z,8Z,10E, 14Z-eicosatetraenoic acid (12-HETE) was stimulated by micromolar concentrations of 13S-hydroperoxy-9Z,11E-octadecadienoic acid, suggesting metabolic control of the 12-lipoxygenase activity via the hydroperoxide tone. Immunohistochemical investigation revealed that the enzyme is mainly located in the squamous epithelium, and is of platelet-type. Significantly lower values for the 12-HETE formation were found in samples from patients with invasive cervical carcinoma, whereas the amount of immunochemically detectable 12-lipoxygenase protein was unaltered. At the same time the expression levels of the bcl-2 gene were enhanced. Thus, it is concluded that during carcinogenesis the hydroperoxide-reducing capacity of the uterine cervix tissue is enhanced, possibly mediated by bcl-2 protein, and in turn metabolically suppresses the 12-lipoxygenase activity. Furthermore, the data suggest an anti-carcinogenic action of 12-lipoxygenase in human cervix, in contrast to its reported pro-carcinogenic action in breast cancer.

15-Lipoxygenation of phospholipids may precede the sn-2 cleavage by phospholipases A2: reaction specificities of secretory and cytosolic phospholipases A2 towards native and 15-lipoxygenated arachidonoyl phospholipids.

Reticulocyte-type 15-lipoxygenase is known to dioxygenate phospholipids without preceding action of phospholipases A2 (PLA2). Therefore we studied the reaction of the secretory PLA2s (sPLA2) from pancreas and snake venom, and of the human cytosolic PLA2 (cPLA2) with 1-palmitoyl-2-arachidonoyl phosphatidylcholine (PAPC) and their 15-lipoxygenated species (PAPC-OOH and PAPC-OH) either alone or as equimolar mixtures. These PLA2s cleaved PAPC-O(O)H with higher (sPLA2) or similar rates (cPLA2) as compared with native PAPC. In mixtures, however, PAPC proved to be the preferred, albeit not exclusive substrate for all three PLA2s. Thus, partial 15-lipoxygenation of phospholipids may also trigger liberation of arachidonic acid.

Biological dynamics and distribution of 3-hydroxy fatty acids in the yeast Dipodascopsis uninucleata as investigated by immunofluorescence microscopy. Evidence for a putative regulatory role in the sexual reproductive cycle.

Dipodascopsis uninucleata has been recently shown to produce 3-hydroxy polyenoic fatty acids from several exogenous polyenoic fatty acids. In order to examine whether endogenous 3-hydroxy fatty acids (3-OH-FA) may be implicated in the developmental biology of this yeast, we mapped by immunofluorescence microscopy their occurrence in fixed cells with or without cell walls using an antibody raised against 3R-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid (3R-HETE), the biotransformation product from arachidonic acid (AA). This antibody turned out to cross-react with other 3-OH-FA. 3-OH-FA were detected in situ in gametangia, asci, as well as between released ascospores, and proved to be associated with the sexual reproductive stage of the life cycle of the yeast. Acetylsalicylic acid (1 mM), which is known to suppress the formation of 3-OH-FA from exogenous polyenoic fatty acids, inhibited the occurrence of immunoreactive material as well as the sexual phase of the life cycle suggesting a prominent regulatory role of 3-OH-FA for the latter.

Oxylipin formation in fungi: biotransformation of arachidonic acid to 3-hydroxy-5,8-tetradecadienoic acid by Mucor genevensis.

The soil fungus Mucor genevensis was shown to convert exogenous arachidonic acid to the oxylipin 3-hydroxy-5Z,8Z-tetradecadienoic acid (3-HTDE) as determined by gas chromatography/mass spectrometry. This metabolite was only found in the aqueous supernatant together with free linoleic acid, but not in the final fungal biomass. In contrast, the corresponding primary arachidonic acid metabolite (3R)-hydroxy-(5Z,8Z,11Z,14Z)-eicosatetraenoic acid (3-HETE), which has been earlier shown to be produced by the yeast Dipodascopsis uninucleata, could not be detected. These observations may be plausibly explained by a retroconversion by M. genevensis of arachidonic acid to linoleic acid before the latter is metabolised to 3-HTDE.

Production of 3R-hydroxy-polyenoic fatty acids by the yeast Dipodascopsis uninucleata.

Various fatty acids were fed to the yeast Dipodascopsis uninucleata UOFS Y 128, and the extracted samples were analyzed for the accumulation of 3-hydroxy metabolites with the help of electron impact gas chromatography-mass spectrometry. Fatty acids containing of 5Z,8Z-diene system (5Z,8Z,11Z-eicosatrienoic, 5Z,8Z,11Z,14Z-eicosatetraenoic, and 5Z,8Z,11Z,14Z,17Z-eicosapentaenoic acids) yielded the corresponding 3-hydroxy-all-Z-eicosapolyenoic acids. Moreover, linoleic acid (9Z,12Z-octadecadienoic acid) and 11Z,14Z,17Z-eicosatrienoic acid were converted to the 3-hydorxylated metabolites of shorter chain length, e,g., 3-hydroxy-5Z,8Z-tetradecadienoic acid and 3-hydroxy-5Z,8Z,11Z-tetradecatrienoic acid, respectively. In contrast, no accumulation of a 3-hydroxy metabolite was observed with oleic acid (9Z-octadecenoic acid), linolelaidic acid (9E,12E-octadecadienoic acid), gamma-linolenic acid (6Z,9Z,12Z-octadecatrienoic acid), and eicosanoic acid as substrate. These findings pinpoint that the 3-hydroxylation of a fatty acid in Dipodascopsis uninucleata requires a 5Z,8Z-diene system either directly or following initial incomplete beta-oxidation. Following analysis of the enantiomer composition, the arachidonic acid metabolite was identified as 3R-hydroxy-5Z,8Z,11Z,14Z-eicosatetraenoic acid, which rules out a normal beta-oxidation as biosynthetic route to this new class of oxylipins.

[Biotransformation of the lipoxygenase inhibitor 2-hydroxy-5-methyl-laurophenone-oxime (FLM 5011)]. / Untersuchungen zur Biotransformation des Lipoxygenasehemmers 2-Hydroxy-5-methyl-laurophenon-oxim (FLM 5011).

2-Hydroxy-5-methyl-laurophenone-oxime (FLM 5011, 1) is an inhibitor of the lipoxygenase with antiinflammatory and antiallergic actions. The studies on the biotransformation using in vivo investigations and in vitro test systems resulted in finding of at least eight metabolites. Four of these compounds have been detected and identified in urine and faeces after p.o. administration in male Wistar rats. By means of cultures of hepatocytes, lymphocytes and myeloma cells additional metabolites were found and the main pathways of metabolism could be suggested. Furthermore it was possible to confirm the sequence of the metabolic reactions. First of all, 1 is hydroxylated in the omega-position of the lauryl side chain by the cytochrome P-450 system. The further oxidation to the carboxylated compound is followed by the stepwise degradation of the side chain by beta-oxidation similarly to the pathways of fatty acid metabolism. Simultaneously the oxime group is converted to the keto group. The metabolites and 1 partly occur as sulfate or glucuronide conjugates. Additionally all compounds produced by beta-oxidation are conjugated with other partners, probably amino acids. By omega-oxidation, compounds with higher inhibitory potency on the lipoxygenase than the parent compound are formed. These results suggest that the activity of 1 is partly caused by the initial metabolites.

The suppression of 5-lipoxygenation of arachidonic acid in human polymorphonuclear leucocytes by the 15-lipoxygenase product (15S)-hydroxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid: structure-activity relationship and mechanism of action.

(15S)-Hydroxy-(5Z,8Z,11Z,13E)-eicosatetraenoic acid (15-HETE) suppresses in ionophore-A23187-stimulated human polymorphonuclear leucocytes (PMN) the conversion of exogenous arachidonic acid into leukotriene B(4) (LTB4) and (5S)-hydroxy-(6E,8Z,11Z,14Z)-eicosatetraenoic acid (5-HETE). However, contrary to earlier suggestions, 15-HETE is not a genuine 5-lipoxygenase inhibitor under these conditions, but rather suppresses the 5-lipoxygenation of arachidonic acid by switching-over of substrate utilization, as judged from a sizeable formation of labelled (5S,15S)-dihydroxy-(6E,8Z,11Z,13E)-eicosatetr aen oic acid (5,15-diHETE) from 15-[1(-14)C]HETE. Identical results were obtained with human recombinant 5-lipoxygenase. In PMN the formation of 5,15-diHETE is strongly stimulated by either hydroperoxypolyenoic fatty acids or arachidonic acid, suggesting a crucial role of the hydroperoxide tone of the cell. A comparison of a selection of hydroxypolyenoic fatty acids with respect to their capability of suppressing 5-lipoxygenation of arachidonic acid revealed that 15-mono-hydroxyeicosanoids throughout exhibit the highest inhibitory potencies, whereas the other HETEs, 5,15-diHETE as well as octadecanoids, are modest or poor inhibitors. The R and S enantiomers of 15-HETE do not differ from each other, excluding a receptor-like binding of the 15-hydroxy group.