DAXX, ATRX, and MSI in PanNET and Their Metastases: Correlation with Histopathological Data and Prognosis.

INTRODUCTION: Studies on pancreatic neuroendocrine tumors (PanNETs) regarding loss of ATRX, DAXX, or frequency of microsatellite instability (MSI) show inconclusive results. So far, data on corresponding metastaseshave not been published. METHODS: We performed immunohistochemistry (IHC) of ATRX, DAXX, MSH2, MSH6, MLH1, and PMS2 on 74 PanNETs and 19 metastases. ATRX- and DAXX-negative PanNETs were further sequenced for mutations. We used polymerase chain reaction for MSI on cases with IHC loss of MSH2, MSH6, MLH1, and PMS2. RESULTS: Immunohistochemical loss of DAXX and ATRX was observed in 8/74 (11%) and 6/74 (8%) PanNETs. Loss of DAXX immunoreactivity was statistically associated with higher tumor grade and showed a tendency toward a decreased overall survival. Sequencing of DAXX- (7/11 [64%]) and ATRX-negative (5/11 [45%]) PanNETs revealed a mutation in 6/7 (86%) and 2/5 (40%). The specificity of immunohistochemical loss of DAXX and ATRX for mutation was 80% and 67%, respectively. The expression status of DAXX compared to primary tumor differs in 2/12 (17%) lymph node metastases. We further identified 3/74 (4%) tumors as MSI, associated with a poor prognosis. DISCUSSION/CONCLUSION: Our study supports the hypothesis that a loss of DAXX immunoreactivity can identify a more aggressive subtype of PanNET with high confidence, while ATRX loss is a weaker indicator. Our results also strengthen the role of DAXX immunolabeling as a prognostic marker. We could show that ATRX might be less suitable as a surrogate for sequencing. Our results indicate that IHC of DAXX and ATRX may identify PanNET subtypes as targets for more aggressive therapy.

Transformation Products of Organic Contaminants and Residues-Overview of Current Simulation Methods.

The formation of transformation products (TPs) from contaminants and residues is becoming an increasing focus of scientific community. All organic compounds can form different TPs, thus demonstrating the complexity and interdisciplinarity of this topic. The properties of TPs could stand in relation to the unchanged substance or be more harmful and persistent. To get important information about the generated TPs, methods are needed to simulate natural and manmade transformation processes. Current tools are based on metabolism studies, photochemical methods, electrochemical methods, and Fenton's reagent. Finally, most transformation processes are based on redox reactions. This review aims to compare these methods for structurally different compounds. The groups of pesticides, pharmaceuticals, brominated flame retardants, and mycotoxins were selected as important residues/contaminants relating to their worldwide occurrence and impact to health, food, and environmental safety issues. Thus, there is an increasing need for investigation of transformation processes and identification of TPs by fast and reliable methods.

Synthesis and Structural Identification of a Biaryl Ether-Linked Zearalenone Dimer.

A new dimer of the food-relevant mycotoxin zearalenone was isolated after electrochemical and chemical oxidation. The structure was determined as a 16-O-15'-biaryl ether-linked dimer based on spectroscopic analyses (¹H- and 13C-NMR, COSY, HMBC, and HSQCAD) and high-resolution mass spectrometry analysis (Q-TOF).

Electrochemical simulation of biotransformation reactions of citrinin and dihydroergocristine compared to UV irradiation and Fenton-like reaction.

Mycotoxins occur widely in foodstuffs and cause a variety of mold-related health risks to humans and animals. Elucidation of the metabolic fate of mycotoxins and the growing number of newly discovered mycotoxins have enhanced the demand for fast and reliable simulation methods. The viability of electrochemistry coupled with mass spectrometry (EC/ESI-MS), Fenton-like oxidation, and UV irradiation for the simulation of oxidative phase I metabolism of the mycotoxins citrinin (CIT) and dihydroergocristine (DHEC) was investigated. The specific reaction products are compared with metabolites produced by human and rat liver microsomes in vitro. Depending on the applied potential between 0 and 2000 mV vs. Pd/H2 by using a flow-through cell, CIT and DHEC are oxidized to various products. Besides dehydrogenation and dealkylation reactions, several hydroxylated DHEC and CIT species are produced by EC and Fenton-like reaction, separated and analyzed by LC-MS/MS and ESI-HRMS. Compared to reaction products from performed microsomal incubations, several mono- and dihydroxylated DHEC species were found to be similar to the reaction products of EC, Fenton-like reaction, and UV-induced oxidation. Consequentially, nonmicrosomal efficient and economic simulation techniques can be useful in early-stage metabolic studies, even if one-to-one simulation is not always feasible.

CYP-13A12 of the nematode Caenorhabditis elegans is a PUFA-epoxygenase involved in behavioural response to reoxygenation.

A specific behavioural response of Caenorhabditis elegans, the rapid increase of locomotion in response to anoxia/reoxygenation called the O2-ON response, has been used to model key aspects of ischaemia/reperfusion injury. A genetic suppressor screen demonstrated a direct causal role of CYP (cytochrome P450)-13A12 in this response and suggested that CYP-eicosanoids, which in mammals influence the contractility of cardiomyocytes and vascular smooth muscle cells, might function in C. elegans as specific regulators of the body muscle cell activity. In the present study we show that co-expression of CYP-13A12 with the NADPH-CYP-reductase EMB-8 in insect cells resulted in the reconstitution of an active microsomal mono-oxygenase system that metabolized EPA (eicosapentaenoic acid) and also AA (arachidonic acid) to specific sets of regioisomeric epoxy and hydroxy derivatives. The main products included 17,18-EEQ (17,18-epoxyeicosatetraenoic acid) from EPA and 14,15-EET (14,15-epoxyeicosatrienoic acid) from AA. Locomotion assays showed that the defective O2-ON response of C20-PUFA (polyunsaturated fatty acid)-deficient, Δ-12 and Δ-6 fatty acid desaturase mutants (fat-2 and fat-3 respectively) can be restored by feeding the nematodes AA or EPA, but not ETYA (eicosatetraynoic acid), a non-metabolizable AA analogue. Short-term incubation with 17,18-EEQ was sufficient to rescue the impaired locomotion of the fat-3 strain. The endogenous level of free 17,18-EEQ declined during anoxia and was rapidly restored in response to reoxygenation. On the basis of these results, we suggest that CYP-dependent eicosanoids such as 17,18-EEQ function as signalling molecules in the regulation of the O2-ON response in C. elegans. Remarkably, the exogenously administered 17,18-EEQ increased the locomotion activity under normoxic conditions and was effective not only with C20-PUFA-deficient mutants, but to a lesser extent also with wild-type worms.

ADP-Ribosylation Factor-Interacting Protein 2 Acts as a Novel Regulator of Mitophagy and Autophagy in Podocytes in Diabetic Nephropathy.

(1) Background: Differentiated podocytes are particularly vulnerable to oxidative stress and cellular waste products. The disease-related loss of postmitotic podocytes is a direct indicator of renal disease progression and aging. Podocytes use highly specific regulated networks of autophagy and endocytosis that counteract the increasing number of damaged protein aggregates and help maintain cellular homeostasis. Here, we demonstrate that ARFIP2 is a regulator of autophagy and mitophagy in podocytes both in vitro and in vivo. (2) Methods: In a recent molecular regulatory network analysis of mouse glomeruli, we identified ADP-ribosylation factor-interacting protein 2 (Arfip2), a cytoskeletal regulator and cofactor of ATG9-mediated autophagosome formation, to be differentially expressed with age. We generated an Arfip2-deficient immortalized podocyte cell line using the CRISPR/Cas technique to investigate the significance of Arfip2 for renal homeostasis in vitro. For the in vivo analyses of Arfip2 deficiency, we used a mouse model of Streptozotozin-induced type I diabetes and investigated physiological data and (patho)histological (ultra)structural modifications. (3) Results: ARFIP2 deficiency in immortalized human podocytes impedes autophagy. Beyond this, ARFIP2 deficiency in human podocytes interferes with ATG9A trafficking and the PINK1-Parkin pathway, leading to the compromised fission of mitochondria and short-term increase in mitochondrial respiration and induction of mitophagy. In diabetic mice, Arfip2 deficiency deteriorates autophagy and leads to foot process effacement, histopathological changes, and early albuminuria. (4) Conclusions: In summary, we show that ARFIP2 is a novel regulator of autophagy and mitochondrial homeostasis in podocytes by facilitating ATG9A trafficking during PINK1/Parkin-regulated mitophagy.

Root uptake and metabolization of Alternaria toxins by winter wheat plants using a hydroponic system.

Fungi of the genus Alternaria are ubiquitous in the environment. Their mycotoxins can leach out of contaminated plants or crop debris into the soil entering the plant via the roots. We aim to evaluate the importance of this entry pathway and its contribution to the overall content of Alternaria toxins (ATs) in wheat plants to better understand the soil-plant-phytopathogen system. A hydroponic cultivation system was established and wheat plants were cultivated for up to two weeks under optimal climate conditions. One half of the plants was treated with a nutrient solution spiked with alternariol (AOH), alternariol monomethyl ether (AME), and tenuazonic acid (TeA), whereas the other half of the plants was cultivated without mycotoxins. Plants were harvested after 1 and 2 weeks and analyzed using a QuEChERS-based extraction and an in-house validated LC-MS/MS method for quantification of the ATs in roots, crowns, and leaves separately. ATs were taken up by the roots and transported throughout the plant up to the leaves after 1 as well as 2 weeks of cultivation with the roots showing the highest ATs levels followed by the crowns and the leaves. In addition, numerous AOH and AME conjugates like glucosides, malonyl glucosides, sulfates, and di/trihexosides were detected in different plant compartments and identified by high-resolution mass spectrometry. This is the first study demonstrating the uptake of ATs in vivo using a hydroponic system and whole wheat plants examining both the distribution of ATs within the plant compartments and the modification of ATs by the wheat plants.

Negative polaron and triplet exciton diffusion in organometallic "molecular wires".

The dynamics of negative polaron and triplet exciton transport within a series of monodisperse platinum (Pt) acetylide oligomers is reported. The oligomers consist of Pt-acetylide repeats, [PtL(2)-C≡C-Ph-C≡C-](n) (where L = PBu(3) and Ph = 1,4-phenylene, n = 2, 3, 6, and 10), capped with naphthalene diimide (NDI) end groups. The Pt-acetylide segments are electro- and photoactive, and they serve as conduits for transport of electrons (negative polaron) and triplet excitons. The NDI end groups are relatively strong acceptors, serving as traps for the carriers. Negative polaron transport is studied by using pulse radiolysis/transient absorption at the Brookhaven National Laboratory Laser-Electron Accelerator Facility (LEAF). Electrons are rapidly attached to the oligomers, with some fraction initially residing upon the Pt-acetylide chains. The dynamics of transport are resolved by monitoring the spectral changes associated with transfer of electrons from the chain to the NDI end group. Triplet exciton transport is studied by femtosecond-picosecond transient absorption spectroscopy. Near-UV excitation leads to rapid production of triplet excitons localized on the Pt-acetylide chains. The excitons transport to the chain ends, where they are annihilated by charge separation with the NDI end group. The dynamics of triplet transport are resolved by transient absorption spectroscopy, taking advantage of the changes in spectra associated with decay of the triplet exciton and rise of the charge-separated state. The results indicate that negative polarons and excitons are transported rapidly, on average moving distances of ~3 nm in less than 200 ps. Analysis of the dynamics suggests diffusive transport by a site-to-site hopping mechanism with hopping times of ~27 ps for triplets and <10 ps for electrons.

Standard reference values of the upper body posture in healthy middle-aged female adults in Germany.

In order to classify and analyze the parameters of upper body posture, a baseline in form of standard values is demanded. To this date, standard values have only been published for healthy young women. Data for female adults between 51 and 60 years are lacking. 101 symptom-free female volunteers aged 51-60 (55.16 ± 2.89) years. The mean height of the volunteers was 1.66 ± 0.62 m, with a mean body weight of 69.3 ± 11.88 kg and an average BMI of 25.02 ± 4.55 kg/m2. By means of video raster stereography, a 3D-scan of the upper back surface was measured in a habitual standing position. The confidence interval, tolerance range and ICCs were calculated for all parameters. The habitual standing position is almost symmetrical in the frontal plane the most prominent deviation being a slightly more ventral position of the left shoulder blade in comparison to the right. The upper body (spine position) is inclined ventrally with a minor tilt to the left. In the sagittal plane, the kyphosis angle of the thoracic spine is greater than the lordosis angle of the lumbar spine. The pelvis is virtually evenly balanced with deviations from an ideal position falling under the measurement error margin of 1 mm/1°. There were also BMI influenced postural variations in the sagittal plane and shoulder distance. The ICCs are calculated from three repeated measurements and all parameters can be classified as "almost perfect". Deflections from an ideally symmetric spinal alignment in women aged 51-60 years are small-scaled, with a minimal frontal-left inclination and accentuated sigmoidal shape of the spine. Postural parameters presented in this survey allow for comparisons with other studies as well as the evaluation of clinical diagnostics and applications.

Fatal attraction of Caenorhabditis elegans to predatory fungi through 6-methyl-salicylic acid.

Salicylic acid is a phenolic phytohormone which controls plant growth and development. A methyl ester (MSA) derivative thereof is volatile and involved in plant-insect or plant-plant communication. Here we show that the nematode-trapping fungus Duddingtonia flagrans uses a methyl-salicylic acid isomer, 6-MSA as morphogen for spatiotemporal control of trap formation and as chemoattractant to lure Caenorhabditis elegans into fungal colonies. 6-MSA is the product of a polyketide synthase and an intermediate in the biosynthesis of arthrosporols. The polyketide synthase (ArtA), produces 6-MSA in hyphal tips, and is uncoupled from other enzymes required for the conversion of 6-MSA to arthrosporols, which are produced in older hyphae. 6-MSA and arthrosporols both block trap formation. The presence of nematodes inhibits 6-MSA and arthrosporol biosyntheses and thereby enables trap formation. 6-MSA and arthrosporols are thus morphogens with some functions similar to quorum-sensing molecules. We show that 6-MSA is important in interkingdom communication between fungi and nematodes.

Heme-Iron-Induced Production of 4-Hydroxynonenal in Intestinal Lumen May Have Extra-Intestinal Consequences through Protein-Adduct Formation.

Some epidemiological studies show that heme iron consumption, in red meat, is associated to the development of several chronic diseases, including cancers and cardio-metabolic diseases. As heme iron intestinal absorption is finely regulated, we hypothesized that heme iron may act indirectly, through the peroxidation of dietary lipids, in food or in the intestinal lumen during digestion. This heme-iron-induced lipid peroxidation provokes the generation of toxic lipid oxidation products that could be absorbed, such as 4-hydroxynonenal (HNE). In a first experiment, heme iron given to rats by oral gavage together with the linoleic-acid-rich safflower oil induced the formation of HNE in the intestinal lumen. The HNE major urinary metabolite was elevated in the urine of the treated rats, indicating that this compound has been absorbed. In a second experiment, we showed that stable isotope-labeled HNE given orally to rats was able to reach non-intestinal tissues as a bioactive form and to make protein-adducts in heart, liver and skeletal muscle tissues. The presence of HNE-protein adducts in those tissues suggests a putative biological role of diet-originating HNE in extra-intestinal organs. This finding could have major consequences on the onset/development of chronic diseases associated with red meat over-consumption, and more largely to peroxidation-prone food consumption.

Complexes of the Mycotoxins Citrinin and Ochratoxin A with Aluminum Ions and their Spectroscopic Properties.

The sensitive detection of the mycotoxin citrinin (CIT) utilizing its fluorescence requires approaches to enhance the emission. In this respect, we studied the complexation of CIT and ochratoxin A (OTA) with Al3+ in methanol using absorption and fluorescence spectroscopy. In this context, an isocratic high performance liquid chromatography (HPLC) method using a polymer column and a fluorescence detector was also developed that enables the separation of the metal ion complexes from the free ligands and non-complexed Al3+. CIT and OTA showed distinct changes in their absorption and fluorescence properties upon Al3+-coordination, and the fluorescence of CIT was considerably enhanced. Analysis of the photometrically assessed titration of CIT and OTA with Al3+ using the Job plot method revealed 1:2 and 1:1 stoichiometries for the Al3+ complexes of CIT (Al:CIT) and OTA (Al:OTA), respectively. In the case of CIT, only one ß-diketone moiety participates in Al3+ coordination. These findings can be elegantly exploited for signal amplification and provide the base to reduce the limit of detection for CIT quantification by about an order of magnitude, as revealed by HPLC measurements using a fluorescence detector.

Toxicity Assay for Citrinin, Zearalenone and Zearalenone-14-Sulfate Using the Nematode Caenorhabditis elegans as Model Organism.

To keep pace with the rising number of detected mycotoxins, there is a growing need for fast and reliable toxicity tests to assess potential threats to food safety. Toxicity tests with the bacterial-feeding nematode Caenorhabditis elegans as the model organism are well established. In this study the C. elegans wildtype strain N2 (var. Bristol) was used to investigate the toxic effects of the food-relevant mycotoxins citrinin (CIT) and zearalenone-14-sulfate (ZEA-14-S) and zearalenone (ZEA) on different life cycle parameters including reproduction, thermal and oxidative stress resistance and lifespan. The metabolization of the mycotoxins by the nematodes in vivo was investigated using HPLC-MS/MS. ZEA was metabolized in vivo to the reduced isomers α-zearalenol (α-ZEL) and β-ZEL. ZEA-14-S was reduced to α-/β-ZEL-14-sulfate and CIT was metabolized to mono-hydroxylated CIT. All mycotoxins tested led to a significant decrease in the number of nematode offspring produced. ZEA and CIT displayed negative effects on stress tolerance levels and for CIT an additional shortening of the mean lifespan was observed. In the case of ZEA-14-S, however, the mean lifespan was prolonged. The presented study shows the applicability of C. elegans for toxicity testing of emerging food mycotoxins for the purpose of assigning potential health threats.

Biosynthesis and Characterization of Zearalenone-14-Sulfate, Zearalenone-14-Glucoside and Zearalenone-16-Glucoside Using Common Fungal Strains.

Zearalenone (ZEN) and its phase II sulfate and glucoside metabolites have been detected in food and feed commodities. After consumption, the conjugates can be hydrolyzed by the human intestinal microbiota leading to liberation of ZEN that implies an underestimation of the true ZEN exposure. To include ZEN conjugates in routine analysis, reliable standards are needed, which are currently not available. Thus, the aim of the present study was to develop a facilitated biosynthesis of ZEN-14-sulfate, ZEN-14-glucoside and ZEN-16-glucoside. A metabolite screening was conducted by adding ZEN to liquid fungi cultures of known ZEN conjugating Aspergillus and Rhizopus strains. Cultivation conditions and ZEN incubation time were varied. All media samples were analyzed for metabolite formation by HPLC-MS/MS. In addition, a consecutive biosynthesis was developed by using Fusarium graminearum for ZEN biosynthesis with subsequent conjugation of the toxin by utilizing Aspergillus and Rhizopus species. ZEN-14-sulfate (yield: 49%) is exclusively formed by Aspergillus oryzae. ZEN-14-glucoside (yield: 67%) and ZEN-16-glucoside (yield: 39%) are formed by Rhizopus oryzae and Rhizopusoligosporus, respectively. Purities of ≥73% ZEN-14-sulfate, ≥82% ZEN-14-glucoside and ≥50% ZEN-16-glucoside were obtained by ¹H-NMR. In total, under optimized cultivation conditions, fungi can be easily utilized for a targeted and regioselective synthesis of ZEN conjugates.

Radical ion states of platinum acetylide oligomers.

The ion radicals of two series of platinum acetylide oligomers have been subjected to study by electrochemical and pulse radiolysis/transient absorption methods. One series of oligomers, Ptn, has the general structure Ph-C[triple bond]C-[Pt(PBu3)2-C[triple bond]C-(1,4-Ph)-C[triple bond]C-]n-Pt(PBu3)2-C[triple bond]C-Ph (where x=0-4, Ph=phenyl and 1,4-Ph=1,4-phenylene). The second series of oligomers, Pt4Tn, contain a thiophene oligomer core, -C[triple bond]C-(2,5-Th)n-C[triple bond]C- (where n=1-3 and 2,5-Th=2,5-thienylene), capped on both ends with -Pt(PBu3)2-C[triple bond]C-(1,4-Ph)-C[triple bond]C-Pt(PBu3)2-C[triple bond]C-Ph segments. Electrochemical studies reveal that all of the oligomers feature reversible or quasi-reversible one-electron oxidation at potentials less than 1 V versus SCE. These oxidations are assigned to the formation of radical cations on the platinum acetylide chains. For the longer oligomers multiple, reversible one-electron waves are observed at potentials less than 1 V, indicating that multiple positive polarons can be produced on the oligomers. Pulse-radiolysis/transient absorption spectroscopy has been used to study the spectra and dynamics of the cation and anion radical states of the oligomers in dichloroethane and tetrahydrofuran solutions, respectively. All of the ion radicals exhibit two allowed absorption bands: one in the visible region and the second in the near-infrared region. The ion radical spectra shift with oligomer length, suggesting that the polarons are delocalized to some extent on the platinum acetylide chains. Analysis of the electrochemical and pulse radiolysis data combined with the density functional theory calculations on model ion radicals provides insight into the electronic structure of the positive and negative ion radical states of the oligomers. A key conclusion of the work is that the polaron states are concentrated on relatively short oligomer segments.

Multiple reasons for an inefficient STAT1 response upon IL-6-type cytokine stimulation.

IL-6-type cytokines play an important role during inflammation and the immune response. In addition, they are involved in haematopoiesis, liver and neuronal regeneration, embryonic development and fertility. We found that IL-6-type cytokine stimulation of cell lines and primary human macrophages results in a different distribution of the DNA-binding competent STAT dimer species in the cytosol and nucleus as demonstrated by electrophoretic mobility shift assays. In the absence of detergent, STAT3/STAT3, STAT1/STAT3 were the predominant species in the cytoplasm while STAT3/STAT3 was predominant in the nucleus. However, in detergent containing total cellular lysates and nuclear fractions prepared with detergent containing buffers, the STAT1/STAT1 homodimer was as prominent or even more prominent than STAT3/STAT3 and STAT1/STAT3. We were interested in the cause of this discrepancy since STAT1-regulated genes have not been described to be expressed upon IL-6-type cytokine stimulation. In addition to the more transient STAT1 activation, IL-6-type cytokines such as IL-6 and OSM lead to a much less efficient STAT1 activation compared to the potent STAT1 activators IFNgamma and IFNalpha. Studies with STAT1-deficient cells revealed that STAT1 activation does not seem to be an important competitive process to STAT3 activation arguing again for a very inefficient STAT1 activation upon IL-6-type cytokine stimulation. We also describe that pY-STAT3 is much more efficiently shuttled into the nucleus than pY-STAT1.

"Twin peaks": searching for 4-hydroxynonenal urinary metabolites after oral administration in rats.

4-Hydroxynonenal (HNE) is a cytotoxic and genotoxic lipid oxidation secondary product which is formed endogenously upon peroxidation of cellular n-6 fatty acids. However, it can also be formed in food or during digestion, upon peroxidation of dietary lipids. Several studies have evidenced that we are exposed through food to significant concentrations of HNE that could pose a toxicological concern. It is then of importance to known how HNE is metabolized after oral administration. Although its metabolism has been studied after intravenous administration in order to mimick endogenous formation, its in vivo fate after oral administration had never been studied. In order to identify and quantify urinary HNE metabolites after oral administration in rats, radioactive and stable isotopes of HNE were used and urine was analyzed by radio-chromatography (radio-HPLC) and chromatography coupled with High Resolution Mass Spectrometry (HPLC-HRMS). Radioactivity distribution revealed that 48% of the administered radioactivity was excreted into urine and 15% into feces after 24h, while 3% were measured in intestinal contents and 2% in major organs, mostly in the liver. Urinary radio-HPLC profiles revealed 22 major peaks accounting for 88% of the urinary radioactivity. For identification purpose, HNE and its stable isotope [1,2-(13)C]-HNE were given at equimolar dose to be able to univocally identify HNE metabolites by tracking twin peaks on HPLC-HRMS spectra. The major peak was identified as 9-hydroxy-nonenoic acid (27% of the urinary radioactivity) followed by classical HNE mercapturic acid derivatives (the mercapturic acid conjugate of di-hydroxynonane (DHN-MA), the mercapturic acid conjugate of 4-hydroxynonenoic acid (HNA-MA) in its opened and lactone form) and by metabolites that are oxidized in the terminal position. New urinary metabolites as thiomethyl and glucuronide conjugates were also evidenced. Some analyses were also performed on feces and gastro-intestinal contents, revealing the presence of tritiated water that could originate from beta-oxidation reactions.

Epoxides Derived from Dietary Dihomo-Gamma-Linolenic Acid Induce Germ Cell Death in C. elegans.

Dietary fats are not created equally, slight differences in structure lead to crucial differences in function. Muticellular organisms use polyunsaturated fatty acid as substrates to produce potent signaling molecules crucial for many physiological processes, including reproduction. Here we explored the mechanism responsible for germ cell loss induced by dietary supplementation of dihomo-gamma-linolenic acid (DGLA, 20:3n-6) in the roundworm Caenorhabditis elegans. In this study we found that C. elegans CYP-33E2 activity produces a range of epoxy and hydroxy metabolites from dietary DGLA. Knockdown of cyp-33E2 suppressed the DGLA-induced sterility phenotype. Additionally, direct exposure of two specific DGLA-derived epoxy products, 8,9- and 14,15-epoxyeicosadienoic acids, produced germ cell abnormalities in the C. elegans gonad. We propose that sterility is mediated by the production of toxic DGLA-derived epoxides that trigger germ cell destruction. These studies are the first to establish a biological activity for a CYP-produced metabolite of DGLA.

Antiatherogenic and antitumoral properties of Opuntia cladodes: inhibition of low density lipoprotein oxidation by vascular cells, and protection against the cytotoxicity of lipid oxidation product 4-hydroxynonenal in a colorectal cancer cellular model.

Opuntia species have been used for thousands of years as a folk medicine in the treatment of diseases. However, the components and protective mechanisms are still unclear. We make the hypothesis that Opuntia species may protect the development of oxidative stress-associated diseases, such as atherosclerosis or colon cancer, via their antioxidant properties. We investigated the protective effect of Opuntia cladode powder against the oxidation of low-density lipoprotein (LDL) evoked by vascular endothelial cells, an important risk factor for atherosclerosis development, and the toxicity of 4-hydroxynonenal (a major lipid peroxidation product) on normal (Apc +/+) and preneoplastic (Apc min/+) immortalized epithelial colon cells. Various Opuntia species classified according to their degree of domestication, from the wildest (Opuntia streptacantha, Opuntia hyptiacantha, Opuntia megacantha), medium (Opuntia albicarpa), to the most domesticated (Opuntia ficus-indica) were tested. Cladode powders prepared from these Opuntia species significantly inhibited LDL oxidation induced by incubation with murine endothelial cells and the subsequent foam cell formation of RAW 264.7 murine macrophages and cytotoxicity on murine endothelial cells. Moreover, Opuntia cladode powder blocked the promotion of colon cancer development on an in vitro model of colonocytes. It may be noted that the phenolic acid and flavonoids content, the antioxidant capacity, and the protective effect were relatively similar in all the cladode powders from wild (O. streptacantha) and domesticated Opuntia. Altogether, these data confirm the therapeutic potential of Opuntia cladodes in diseases associated with oxidative stress.