Lack of Enantioselectivity in the SULT1A3‐catalyzed Sulfoconjugation of Normetanephrine Enantiomers: An In Vitro and Computational Study

(1R)‐Normetanephrine is the natural stereoisomeric substrate for sulfotransferase 1A3 (SULT1A3)‐catalyzed sulfonation. Nothing appears known on the enantioselectivity of the reaction despite its potential significance in the metabolism of adrenergic amines and in clinical biochemistry. We confronted the kinetic parameters of the sulfoconjugation of synthetic (1R)‐normetanephrine and (1S)‐normetanephrine by recombinant human SULT1A3 to a docking model of each normetanephrine enantiomer with SULT1A3 and the 3′‐phosphoadenosine‐5′‐phosphosulfate cofactor on the basis of molecular modeling and molecular dynamics simulations of the stability of the complexes. The K_M, V_max, and k_cat values for the sulfonation of (1R)‐normetanephrine, (1S)‐normetanephrine, and racemic normetanephrine were similar. In silico models were consistent with these findings as they showed that the binding modes of the two enantiomers were almost identical. In conclusion, SULT1A3 is not substrate‐enantioselective toward normetanephrine, an unexpected finding explainable by a mutual adaptability between the ligands and SULT1A3 through an “induced‐fit model” in the catalytic pocket. Chirality, 25:28‐34, 2012.© 2012 Wiley Periodicals, Inc.     

Hypoglycemic and antioxidant effects of phenolic extracts and purified hydroxytyrosol from olive mill waste in vitro and in rats

This study aimed to evaluate the effect of phenolic extract and purified hydroxytyrosol (HT) from olive mill waste (OMW) on oxidative stress and hyperglycemia in alloxan-induced diabetic rats. The OMW biophenols were extracted using ethyl acetate. The obtained extract was fractionated by solid phase extraction (SPE) experimentation to generate two fractions: (F1) and (F2). HPLC-UV and HPLC-MS analysis showed that (F1) was made of known OMW monomeric phenolics mainly hydroxytyrosol (HT) while (F2) contained oligomeric and polymeric phenols such as verbascosid and ligstrosid. (HT) was purified from (F1) using silica gel-column chromatography and silica gel-TLC techniques. In incubated pancreas, supplementation of OMW fractions enhanced insulin secretion. The administration of OMW extract fractions (F1) and (F2) as well as purified (HT) in diabetic rats caused a decrease in glucose level in plasma and an increase in renal superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) activities in liver and kidney. Furthermore, a protective action against hepatic and renal toxicity in diabetic rats was clearly observed. Furthermore, a significant decrease in hepatic and renal indices toxicity was observed, i.e. alkalines phosphatases (ALP), aspartate and lactate transaminases (AST and ALT) activities and the thiobarbituric acid-reactive substances (TBARs), total and direct bilirubin, creatinine and urea levels. In addition, (F1), (F2) and especially (HT) decreased triglycerides (TG), total-cholesterol (T-Ch) and higher HDL-cholesterol (HDL-Ch) in serum. These beneficial effects of OMW biophenols were confirmed by histological findings in hepatic, renal and pancreatic tissues of diabetic rats. This study demonstrates for the first time that OMW polyphenols and especially (HT) are efficient in inhibiting hyperglycemia and oxidative stress induced by diabetes and suggests that administration of HT may be helpful in the prevention of diabetic complications associated with oxidative stress.     

Importance de la Mérione de Shaw Meriones shawii au sein des composantes trophiques de la Chouette effraie Tyto alba en milieux steppiques de l’Algérie

The study of the diet of the Barn Owl in two steppic regions (M'Sila and Djelfa) located in the Algerian highlands is based on the analysis of the pellets of rejections collected in six stations. The analysis of 706 pellets resulting from the various stations made it possible to count 1380 individuals, represented by seven classes, 12 orders, 32 families, and 76 species of preys. The mammals are consumed with variable abundance rates between 59.1 % and 90.0 % whose predominance is assigned to the rodents (relative abundance: AR > 58 %). The latter constitute the most advantageous preys in biomass (61.4 ≤ B % ≤ 99.2). The most consumed prey is Meriones shawii, with variable rates between 31.9 % and 76.6 %. Generally, Tyto alba presents a diversified diet in the majority of the stations (0.69 ≤ E ≤ 0.76), except the station of Ain El-Hadjel (E = 0.35), with a low diversity and dominance of M. shawii (AR = 76.6 %).     

Tumor necrosis factor-alpha down-regulates human Cu/Zn superoxide dismutase 1 promoter via JNK/AP-1 signaling pathway

Overexpression of Cu/Zn superoxide dismutase 1 (SOD1) in monocytes blocks reactive oxygen species-induced inhibition of cell growth and apoptosis and renders cells resistant to the toxic effect of tumor necrosis factor (TNF)-alpha, suggesting that TNF-alpha represses the SOD1 gene in these cells. We herein show that TNF-alpha decreases SOD1 mRNA, protein, and promoter activity in U937 cells. Electrophoretic mobility-shift assays (EMSA) show that TNF-alpha decreased binding of three different complexes. Ectopic Sp1 overexpression markedly increased SOD1-basal promoter activity and partially antagonized the TNF-alpha inhibitory effect. In contrast, ectopic c-Jun overexpression mimics TNF-alpha inhibitory effects and antagonizes Sp1 stimulatory effects. In agreement with these findings, EMSA shows a TNF-alpha-induced increase in AP-1 and a decrease in Sp1 DNA binding. Disruption of the C/EBP site decreases, whereas mutation in the Sp1/Egr-1 site completely abolishes DNA-binding and promoter activity. A JNK inhibitor antagonized the negative effects of TNF-alpha on SOD1 promoter activity, suggesting that JNK signaling through c-Jun protein activation is critical for the TNF-alpha-dependent SOD1 repression. A greater understanding of the mechanisms of TNF-alpha-induced SOD1 repression could facilitate the design and development of novel therapeutic drugs for inflammatory conditions.     

Asymptomatic human CD4+ cytotoxic T-cell epitopes identified from herpes simplex virus glycoprotein B

The identification of “asymptomatic” (i.e., protective) epitopes recognized by T cells from herpes simplex virus (HSV)-seropositive healthy individuals is a prerequisite for an effective vaccine. Using the PepScan epitope mapping strategy, a library of 179 potential peptide epitopes (15-mers overlapping by 10 amino acids) was identified from HSV type 1 (HSV-1) glycoprotein B (gB), an antigen that induces protective immunity in both animal models and humans. Eighteen groups (G1 to G18) of 10 adjacent peptides each were first screened for T-cell antigenicity in 38 HSV-1-seropositive but HSV-2-seronegative individuals. Individual peptides within the two immunodominant groups (i.e., G4 and G14) were further screened with T cells from HLA-DR-genotyped and clinically defined symptomatic (n = 10) and asymptomatic (n = 10) HSV-1-seropositive healthy individuals. Peptides gB_161-175 and gB_166-180 within G4 and gB_661-675 within G14 recalled the strongest HLA-DR-dependent CD4⁺ T-cell proliferation and gamma interferon production. gB_166-180, gB_661-675, and gB_666-680 elicited ex vivo CD4⁺ cytotoxic T cells (CTLs) that lysed autologous HSV-1- and vaccinia virus (expressing gB)-infected lymphoblastoid cell lines. Interestingly, gB_166-180 and gB_666-680 peptide epitopes were strongly recognized by CD4⁺ T cells from 10 of 10 asymptomatic patients but not by CD4⁺ T cells from 10 of 10 symptomatic patients (P < 0.0001; analysis of variance posttest). Inversely, CD4⁺ T cells from symptomatic patients preferentially recognized gB_661-675 (P < 0.0001). Thus, we identified three previously unrecognized CD4⁺ CTL peptide epitopes in HSV-1 gB. Among these, gB_166-180 and gB_666-680 appear to be “asymptomatic” peptide epitopes and therefore should be considered in the design of future herpes vaccines.     

Density functional conformational study of 2-O-sulfated 3,6 anhydro-α-D-galactose and of neo-κ- and ι-carrabiose molecules in gas phase and water

We examined the conformational preferences of the 2-O-sulfated-3,6-α-D-anhydrogalactose (compound I) and two 1,3 linked disaccharides constituting-κ or ι-carrageenans using density functional and ab initio methods in gas phase and aqueous solution. Systematic modifications of two torsion angles leading to 324 and 144 starting geometries for the compound I and each disaccharide were used to generate adiabatic maps using B3LYP/6-31G(d). The lower energy conformers were then fully optimized using B3LYP, B3PW91 and MP2 with several basis sets. Overall, we discuss the impact of full relaxation on the energy and structure of the dominant conformations, present the performance comparison with previous molecular mechanics calculations if available, and determine whether our results are impacted, when polarization and diffuse functions are added to the 6-31G(d) basis set, or when the MP2 level of theory is used.     

Evaluation des dégâts par les vers blancs (Coleoptera : Scarabaeoidea) dans les parcelles de régéneration du chêne-liège (Quercus suber L.) en forêt de la Mamora (Maroc) et recherche de médiateurs chimiques pour une lutte biologique

Résumé

Au cours des dernières décennies, les essais de régénération du chêne-liège en forêt de la Mamora se sont heurtés à des attaques massives de vers blancs sur les racines des jeunes plants, avec un taux de réussite des plantations ne dépassant guère 12% dans la majorité des parcelles de régénération. La biologie de Sphodroxia maroccana Ley (Coleoptera : Melolonthidae), le ravageur principal, a été en partie élucidée, avec encore des lacunes concernant la période exacte ?émergence des mâles par rapport aux femelles, la longévité des imagos et la sex-ratio. La sècheresse esti vale est parmi les autres causes de dépérissement des jeunes plants. Lors de la première année suivant la plantation dans des parcelles expérimentales, la mortalité cumulée due aux attaques larvaires et à la sècheresse a varié entre 41% et 68% selon les blocs considérés dans les parcelles. La mortalité liée aux attaques des larves de S. maroccana était comprise entre 24 et 43%, avec une distribution en taches des dégâts, plus ou moins importantes selon la densité initiale des plants. L’isolement par micro-extraction en phase solide des effluves femelles de S. maroccana a permis ?identifier le résorcinol (1,3-benzènediol) comme composé phéromonal présumé. La fonction de cette molécule comme phéromone reste toutefois à démontrer.     

Association between adherens junctions and tight junctions via Rap1 promotes barrier protective effects of oxidized phospholipids

Previous studies showed that cyclopenthenone-containing products resulting from oxidation of a natural phospholipid, 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (OxPAPC) exhibit potent barrier-protective effects in the in vitro and in vivo models of lung endothelial cell (EC) barrier dysfunction, and these effects are associated with enhancement of peripheral actin cytoskeleton, cell-cell and cell-substrate contacts driven by activation of Rac and Cdc42 GTPases. Rap1 GTPase is another member of small GTPase family involved in control of cell-cell interactions; however, its involvement in EC barrier-protective effects by OxPAPC remains unknown. This study examined a role of Rap1 in regulation of OxPAPC-induced interactions in adherens junctions (AJ) and tight junctions (TJ) as a novel mechanism of EC barrier preservation in vitro and in vivo. Immunofluorescence analysis, subcellular fractionation, and co-immunoprecipitation assays indicate that OxPAPC promoted accumulation of AJ proteins: VE-cadherin, p120-catenin, and β-catenin; and TJ proteins: ZO-1, occludin, and JAM-A in the cell membrane, and induced novel cross-interactions between AJ and TJ protein complexes, that were dependent on OxPAPC-induced Rap1 activation. Inhibition of Rap1 function suppressed OxPAPC-mediated pulmonary EC barrier enhancement and AJ and TJ interactions in vitro, as well as inhibited protective effects of OxPAPC against ventilator-induced lung injury in vivo. These results show for the first time a role of Rap1-mediated association between adherens junctions and tight junction complexes in the OxPAPC-induced pulmonary vascular EC barrier protection.     

Environmental Effects on Male Reproductive Success and Parental Care in the Florida Flagfish Jordanella floridae

Models of optimal parental care predict that parental investment should depend on offspring value or the effect parental care has on offspring benefits. Few studies have examined the effect of external factors that influence offspring survival and the cost of care. In this study on the Florida flagfish (Jordanella floridae), a species with male parental care, we examined whether environmentally induced changes in care result from changes in egg requirements or in parental costs. We manipulated salinity and temperature, as these factors are known to affect the metabolic rate in both eggs and parents. We predicted that if the change in care behavior is determined by costs to the male then it should be paralleled by changes in non‐egg‐directed behavior. Conversely, if egg‐directed behavior changes independently of other behavior it would suggest that behavior is determined primarily by egg requirements. In addition we examined patterns of mating success under the assumption that if male care is affected by environmental factors then female preferences may change accordingly. Males decreased egg‐directed behavior (fanning and cleaning of eggs) at high salinity. Non‐egg‐directed behavior was unaffected by salinity. Temperature had no effects on behavior. Thus, we conclude that changes in egg demands are primarily responsible for the observed results. Successful males were bigger and more aggressive. This suggests that male dominance was an important determinant of male mating success. Unsuccessful males showed significantly more variation in number of red stripes with respect to salinity than successful males. Unsuccessful males may be less able to regulate color expression under varying environmental conditions, in which case color may be an indicator of male quality. We replicated the experiment early and late in the season. Males did not change their effort in care over the season. However, care (fanning) in the absence of eggs increased towards the end of the season. Since pre‐mating fanning was positively correlated to a male’s eventual mating success we conclude that males increased their effort to attract mates late in the season.