N5 Groups Research Articles

Nannochloropsis oculata enhances immune response, transcription of stress, and cytokine genes in Nile tilapia subjected to air exposure stress

The beneficial role of dietary Nannochloropsis oculata (N. oculata) on health parameters of Nile tilapia (Oreochromis niloticus) under air exposure stress were investigated. Fish were fed for 4 weeks in three groups: the control group, N5 (5% of N. oculata), and N10 (10% of N. oculata). The fish in the supplemented groups were then exposed to the air stress experiment for 1 h (h) and designed as N5 + AE and N10 + AE, respectively. The results revealed that serum cortisol and glucose were elevated only in the N10 + AE group, whereas C-reactive protein (CRP) and Immunoglobulin-M (IgM) decreased and increased; respectively, in the supplemented-only groups compared to the control. Intricate changes in stress and cytokine gene expression patterns were evident. In the N10 group, the highest transcription levels of hepatic and intestinal heat shock protein 70/ HSP70 were noted, whereas lower and higher transcription levels of hepatic and intestinal HSP70 were evident in N10 + AE and N5 + AE groups, respectively. The hepatic glutathione peroxidase/ GPx, intestinal glutathione S-transferase/ GST, and GPx were induced in the N5 group but declined significantly upon air exposure in the same group. The intestinal transforming growth factor β1/ TGFβ1 and the IL-10 expression level were significantly induced in the N10 group compared to the N5 group. Interestingly, the N5 + AE group exhibited a significant upregulation of TGF-β1 and IL-10 expression levels compared to N5 and N10 + AE groups. Dietary supplementation of N. oculata maintained the liver and intestinal histomorphology and mitigated the air-exposure impacts on the induced tissue histopathological changes. Overall, both levels of N. oculata dietary supplementation in this study showed advantages in different scenarios regarding mitigation of air-exposure stress; nonetheless, these findings support the vital role of N. oculata as feed additives on fish health and in mitigating physical stressors, like air exposure stress.

56 Nobiletin affects gene expression profiles of the ERK1/2 pathway in bovine embryos produced invitro

During embryo development the embryonic genome activation (EGA) is one of the most important events and in bovine embryos it occurs at the 8- to 16-cell stage. Invitro embryo production increases the levels of reactive oxygen species (ROS), which leads to the low quality of the produced blastocysts, possibly by affecting EGA. Nobiletin is an antioxidant that affects cell cycle regulation (Huang et al. 2016 Evid. Based. Complement. Alternat. Med. 2016, 2918796, https://doi.org/10.1155/2016/2918796). Therefore, we aimed to evaluate the effect of nobiletin supplementation, in two key periods of early embryo development, on blastocyst yield and expression of selected genes of the ERK1/2 pathway and oxidative stress on produced embryos. Invitro zygotes were cultured in synthetic oviductal fluid (SOF) with 5% fetal calf serum (control, C); C with 5 or 10µM nobiletin (MedChemExpress) (N5, N10); or C with 0.03% dimethyl sulfoxide (CDMSO; vehicle for nobiletin dilution) during the minor (21–54h post-insemination (hpi): 2- to 8-cell; MNEGA; 12 replicates) or major (54–96 hpi: 8- to 16-cell; MJEGA; 10 replicates) phase of EGA. The speed of development was considered and embryos that reached ≥8 cells at 54 hpi from MNEGA phase and ≥16 cells at 96 hpi from MJEGA phase, were selected and further cultured in control medium until Day 7. Embryos at ≥8 cell (MNEGA), ≥16 cell (MJEGA) stage, and Day 7 blastocysts from both periods were snap-frozen in liquid N2 for gene expression analysis (3 pools of 10 embryos/treatment). The expression of genes related to ERK1/2 pathway (H3–3B, H3–3A, NFE2L2) and oxidative stress (GPX1) were measured by quantitative PCR; H2AFZ and ACTB were used as housekeeping genes. Statistical analysis was assessed by one-way ANOVA. At 54 hpi, irrespective of nobiletin supplementation, no differences were found in the proportion of embryos that reached the 8-cell stage between groups in both phases (≈60%). At 96 hpi, nobiletin during MJEGA showed a higher proportion of embryos reaching the 16-cell stage than control groups (≈70% vs. ≈60%, respectively; P<0.001). Blastocyst yield for MNEGA and MJEGA was higher (P<0.001) for N5 (40.0±0.8% and 46.7±0.8%) and N10 (41.0±0.9% and 54.5±1.1%) compared with C (32.0±0.6% and 38.4±1.1%) and CDMSO (31.2±0.4% and 35.8±1.0%) groups, while N10 was higher (P<0.05) compared to N5 group in MJEGA. The expression of H3–3B and H3–3A were higher (P<0.05) in 8-cell embryos from N5 and N10 groups during MNEGA; while in 16-cell embryos, H3–3B and NFE2L2 were higher (P<0.05) only in the N10 group compared with both controls during MJEGA. GPX1 was upregulated in nobiletin-supplemented groups from both phases (8- and 16-cell embryos and blastocysts) compared with controls (P<0.05). In conclusion, nobiletin supplementation during minor or major EGA has a positive effect in pre-implantation embryo development and modifies the transcription of cell cycle and oxidative stress genes in early embryos. These benefits can be attributed to its bioactivity and indicate that it might be a tool to overcome EGA and ROS disorders in bovine invitro-produced embryos.This research was funded by MINECO-Spain AGL2015-70140-R, PID2019-111641RB-I00, RTI2018-093548-B-I00; SENESCYT-Ecuador; FAPESP-Brazil 2017/20339-3, CNPq-Brazil 304276/2018-9.

Influence of protein malnutrition on cutaneous wound healing in rats

ABSTRACT Objective Investigate histological changes related to inflammatory response and collagen expression during wound healing in rats with protein malnutrition. Methods Twenty male Wistar rats underwent cutaneous surgery and were divided into two experimental groups: Malnourished (8% casein diet); Nourished (17% casein diet). Animals were euthanized after 5th and 10th days, descriptive and quantitative analyses were performed on sections stained with hematoxylin-eosin and sirius red, respectively. Statistical analysis of data using nonparametric Fisher's exact test with p<0.05 was carried out. Results At five days, increased fibroblast proliferation (p<0.01) and collagen expression (p<0.05) was observed in N5 group. After ten days, N10 and MN10 animals showed higher amount of granulation tissue and edema/inflammatory infiltrate independent of nutritional status (p>0.05), only N10 group showed fibroblast proliferation (p<0.01) and increased collagen expression (p<0.01). Conclusion Protein malnutrition seems not to influence inflammatory phase of healing, whereas it negatively effects fibroblast proliferation and collagen synthesis.

Structural and Kinetic Evidence for an Extended Hydrogen-bonding Network in Catalysis of Methyl Group Transfer: ROLE OF AN ACTIVE SITE ASPARAGINE RESIDUE IN ACTIVATION OF METHYL TRANSFER BY METHYLTRANSFERASES

The methyltetrahydrofolate (CH(3)-H(4)folate) corrinoid-iron-sulfur protein (CFeSP) methyltransferase (MeTr) catalyzes transfer of the methyl group of CH(3)-H(4)folate to cob(I)amide. This key step in anaerobic CO and CO(2) fixation is similar to the first half-reaction in the mechanisms of other cobalamin-dependent methyltransferases. Methyl transfer requires electrophilic activation of the methyl group of CH(3)-H(4)folate, which includes proton transfer to the N5 group of the pterin ring and poises the methyl group for reaction with the Co(I) nucleophile. The structure of the binary CH(3)-H(4)folate/MeTr complex (revealed here) lacks any obvious proton donor near the N5 group. Instead, an Asn residue and water molecules are found within H-bonding distance of N5. Structural and kinetic experiments described here are consistent with the involvement of an extended H-bonding network in proton transfer to N5 of the folate that includes an Asn (Asn-199 in MeTr), a conserved Asp (Asp-160), and a water molecule. This situation is reminiscent of purine nucleoside phosphorylase, which involves protonation of the purine N7 in the transition state and is accomplished by an extended H-bond network that includes water molecules, a Glu residue, and an Asn residue (Kicska, G. A., Tyler, P. C., Evans, G. B., Furneaux, R. H., Shi, W., Fedorov, A., Lewandowicz, A., Cahill, S. M., Almo, S. C., and Schramm, V. L. (2002) Biochemistry 41, 14489-14498). In MeTr, the Asn residue swings from a distant position to within H-bonding distance of the N5 atom upon CH(3)-H(4)folate binding. An N199A variant exhibits only approximately 20-fold weakened affinity for CH(3)-H(4)folate but a much more marked 20,000-40,000-fold effect on catalysis, suggesting that Asn-199 plays an important role in stabilizing a transition state or high energy intermediate for methyl transfer.

Effects of Epidural Neostigmine on Postoperative Analgesia and Stress Responses after Gastrectomy

Background: Intrathecal or epidural neostigmine has a postoperative analgesic effect. In this study we evaluated the effects of epidural neostigmine on postoperative analgesia and stress responses after gastrectomy. Methods: Eighty, ASA physical status 1 or 2, patients scheduled for gastrectomy were randomly assigned to one (n = 20) of four groups. Patients received of 0.25% bupivacaine with either saline (N0 group), neostigmine /kg (N5 group), neostigmine /kg (N10 group), neostigmine /kg (N15 group) epidurally 30 minutes before operation. All patients received identical general anesthesia. Patient-controlled epidural analgesia (PCEA) was provided by bupivacaine (0.05%) and fentanyl (/ml for postoperative analgesia. Results: The N10 and N15 groups showed reduced 24-h PCEA consumptions compared with the N0 and N5 groups (P /kg in bupivacaine produced a postoperative analgesic effect, but did not suppress stress responses after gastrectomy.

Pentazoles: proton and carbon-13 NMR spectra of some 1-arylpentazoles: kinetics and mechanism of degradation of the arylpentazole system

The proton and carbon-13 NMR spectra of a range of 1-(p-substituted-phenyl)pentazoles are reported. The influence of the –N5 group on the aryl shifts is compared with that of the –NO2 and –N3 groups. Direct kinetic measurements of the degradation of 1-arylpentazoles in CD3OD–CD2Cl2(1:1, v/v) gave a Hammett p value of + 1.25. For 1-(p-chlorophenyl)pentazole values of ΔE‡, 88.6 kJ mol–1, ΔH‡ 86.3 kJ mol–1 and ΔS‡+ 19.9 J mol–1 K–1 are obtained at –10–0 °C. These data suggest a polar two-step mechanism with rate-determining cleavage of the 1–2 bond giving an unstable azido–azo (pentazene) intermediate, the nitrogen analogue of the azido–azomethine species of tetrazole ring-opening.

Protein Insufficiency Aggravates the Enhanced Lipid Peroxidation and Reduced Activities of Antioxidative Enzymes in Rats Fed Diets High in Polyunsaturated Fat

To investigate the effects of dietary protein and polyunsaturated fat levels on tissue lipid peroxidation and antioxidative enzymes, Long-Evans male weanling rats were fed either an 8% lactalbumin diet containing 2% (L2), 5% (L5), 10% (L10), 15% (L15) or 20% (L20) soybean oil or a 20% lactalbumin diet containing 5% (N5) or 20% (N20) soybean oil for 8 weeks. The tissue thiobarbituric acid-reactive substances (TBARS) concentrations of the L2 group were similar to those of the N5 group except in plasma in which they were higher. The L5 group generally showed tissue TBARS concentrations comparable to the N20 group. Gradually increasing the dietary soybean oil level in the low protein diet further increased the tissue TBARS concentrations. The L20 group had significantly higher TBARS in RBC, liver, heart, kidney and muscle than the N20 group. The low protein-fed groups had lower activities of glutathione peroxidase (EC 1.11.1.9), superoxide dismutase (EC 1.15.1.1) and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) in liver and catalase (EC 1.11.1.6) in RBC than the N5 group. Compared with the N5 group, the N20 group also showed higher TBARS concentrations and lower activities of certain antioxidative enzymes in some tissues. The antioxidative enzyme activities decreased more drastically with the increasing dietary soybean oil level in the low protein-fed groups than in those fed a normal level of protein. Supplementation of 150 mg/kg of all-rac-α-tocopheryl acetate to the L15 diet slightly decreased the TBARS in plasma, heart and liver and restored the depressed activities of RBC superoxide dismutase and catalase. The results indicated that insufficiency of dietary protein aggravates the enhanced production of TBARS and the reduced activities of antioxidant enzyme in rats fed a high soybean oil diet.

A Double Blind Controlled Trial of N‐(3′, 4′‐Dimethoxycinnamoyl) Anthranilic Acid on Children with Bronchial Asthma

A double blind controlled trial of N-(3',4'-dimethoxycinnamoyl) anthranilic acid (N-5') was carried out for treating children with bronchial asthma to determine its efficacy and safety. The results showed that the drug was significantly more effective in the N10 and N5 groups than in the N0 group. Adverse reactions appeared only in one patient in the N10 group who complained of slight anorexia while another patient in the N0 group reported moderate diarrhoea. Moreover there was no indication of abnormality in further laboratory investigations. It can be therefore concluded that this drug is useful for the treatment of children with bronchial asthma.