Leonardo Gomes Peixoto

Identification of major royal jelly proteins in the brain of the honeybee Apis mellifera.

The consumption of royal jelly (RJ) determines the differences between castes and behavioral development in the honeybee Apis mellifera. However, it is not known whether the proteins of RJ are related to these differences, or which proteins are responsible for the changes. To understand the functions of RJ proteins that are present in other tissues of the bee, in addition to hypopharyngeal gland, we used a polyclonal antibody anti-MRJP1 to investigate the presence of this protein in nervous system of honeybee. This study showed the presence of three polypeptides (p57, p70 and p128) in specific tissues of bee brain. Mushroom body, optic lobe and antennal lobe neuropils all contained proteins recognized by anti-MRJP1. Proteomic analysis showed that the three polypeptides are correlated with proteins of the MRJP family. p57 is correlated with MRJP1, p70 with MRJP3, while p128 may be an oligomeric form or a new polypeptide. Immunostaining of the brain and hypopharyngeal gland revealed differential expression of MRJPs in various brain regions and in different honeybee castes and subcastes. The identification and localization of these MRJPs contribute to the elucidation of the biological roles of this protein family.

Chronic Stress Induces a Hyporeactivity of the Autonomic Nervous System in Response to Acute Mental Stressor and Impairs Cognitive Performance in Business Executives

The present study examined the incidence of chronic stress in business executives (109 subjects: 75 male and 34 female) and its relationship with cortisol levels, cognitive performance, and autonomic nervous system (ANS) reactivity after an acute mental stressor. Blood samples were collected from the subjects to measure cortisol concentration. After the sample collection, the subjects completed the Lipp Inventory of Stress Symptoms for Adults and the Stroop Color-Word Test to evaluate stress and cognitive performance levels, respectively. Saliva samples were collected prior to, immediately after, and five minutes after the test. The results revealed that 90.1% of the stressed subjects experienced stress phases that are considered chronic stress. At rest, the subjects with chronic stress showed higher cortisol levels, and no gender differences were observed. No differences were found between the stressed and non-stressed subjects regarding salivary amylase activity prior to test. Chronic stress also impaired performance on the Stroop test, which revealed higher rates of error and longer reaction times in the incongruent stimulus task independently of gender. For the congruent stimulus task of the Stroop test, the stressed males presented a higher rate of errors than the non-stressed males and a longer reaction time than the stressed females. After the acute mental stressor, the non-stressed male group showed an increase in salivary alpha-amylase activity, which returned to the initial values five minutes after the test; this ANS reactivity was not observed in the chronically stressed male subjects. The ANS responses of the non-stressed vs stressed female groups were not different prior to or after the Stroop test. This study is the first to demonstrate a blunted reactivity of the ANS when male subjects with chronic psychological stress were subjected to an acute mental stressor, and this change could contribute to impairments in cognitive performance.

Peel of araticum fruit (Annona crassiflora Mart.) as a source of antioxidant compounds with α-amylase, α-glucosidase and glycation inhibitory activities

Annona crassiflora Mart., whose fruit is popularly known as araticum, is a member of the Annonaceae family found in the Brazilian Cerrado. Although this plant has several medicinal uses, its bioactive molecules are not fully understood. A bioguided assay was performed to identify the main bioactive compounds of A. crassiflora fruit peel from the ethanol extract fractions with antioxidant capacity and α-amylase, α-glucosidase and glycation inhibitory activities. Ethyl acetate and n-butanol fractions showed, respectively, higher antioxidant capacity (DPPH IC50 1.5±0.1 and 0.8±0.1μgmL-1, ORAC 3355±164 and 2714±79μmoltroloxeq/g, and FRAP 888±16 and 921±9μmoltroloxeq/g) and inhibitory activities against α-amylase (IC50 4.5±0.8 and 1.7±0.3μgmL-1), α-glucosidase (IC50 554.5±158.6 and 787.8±140.6μgmL-1) and glycation (IC50 14.3±3.3 and 16.0±4.2μgmL-1), and lower cytotoxicity, compared to the other fractions and crude ethanol extract. The HPLC-ESI-MS/MS analysis identified various biomolecules known as potent antioxidants, such as chlorogenic acid, (epi)catechin, procyanidins, caffeoyl-hexosides, quercetin-glucosides and kaempferol. The fruit peel of A. crassiflora, a specie from Cerrado, the Brazilian Savanna, provided a source of antioxidant compounds with properties to block carbohydrate digestive enzymes and formation of glycation products. Thus, there is potential to use the by-products of araticum in order to identify and isolate phytochemicals for application in nutraceutical supplements, food additives and pharmaceuticals products.

The Role of Metformin in Controlling Oxidative Stress in Muscle of Diabetic Rats

Metformin can act in muscle, inhibiting the complex I of the electron transport chain and decreasing mitochondrial reactive oxygen species. Our hypothesis is that the inhibition of complex I can minimize damage oxidative in muscles of hypoinsulinemic rats. The present study investigated the effects of insulin and/or metformin treatment on oxidative stress levels in the gastrocnemius muscle of diabetic rats. Rats were rendered diabetic (D) with an injection of streptozotocin and were submitted to treatment with insulin (D+I), metformin (D+M), or insulin plus metformin (D+I+M) for 7 days. The body weight, glycemic control, and insulin resistance were evaluated. Then, oxidative stress levels, glutathione antioxidant defense system, and antioxidant status were analyzed in the gastrocnemius muscle of hypoinsulinemic rats. The body weight decreased in D+M compared to ND rats. D+I and D+I+M rats decreased the glycemia and D+I+M rats increased the insulin sensitivity compared to D rats. D+I+M reduced the oxidative stress levels and the activity of catalase and superoxide dismutase in skeletal muscle when compared to D+I rats. In conclusion, our results reveal that dual therapy with metformin and insulin promotes more benefits to oxidative stress control in muscle of hypoinsulinemic rats than insulinotherapy alone.

Royal jelly decreases corticosterone levels and improves the brain antioxidant system in restraint and cold stressed rats

Restraint and cold stress induces the hypothalamic-pituitary-adrenal (HPA) axis to release corticosterone from the adrenal gland, which can worsen the antioxidant defense system in the central nervous system. Here, we investigated the corticosterone levels and the antioxidant defense system in the cerebellum and brain, as well as in its isolated regions, such as cerebral cortex, striatum and hippocampus of stressed rats supplemented with royal jelly (RJ). Wistar rats were supplemented with RJ for 14days and the stress induction started on the 7th day. Stressed rats increased corticosterone levels, glycemia and lipid peroxidation in the brain and cerebellum, cerebral cortex and hippocampus besides reduced glutathione defense system in the brain and striatum. Rats supplemented with RJ decreased corticosterone, maintained glycemia and decreased lipid peroxidation in the brain, cerebellum, as well as striatum and hippocampus, besides improved glutathione defense system in cerebral cortex and striatum. This study suggests an anti-stress and neuroprotective effect of RJ under stress conditions.

Myosins and DYNLL1/LC8 in the honey bee (Apis mellifera L.) brain.

Honey bees have brain structures with specialized and developed systems of communication that account for memory, learning capacity and behavioral organization with a set of genes homologous to vertebrate genes. Many microtubule- and actin-based molecular motors are involved in axonal/dendritic transport. Myosin-Va is present in the honey bee Apis mellifera nervous system of the larvae and adult castes and subcastes. DYNLL1/LC8 and myosin-IIb, -VI and -IXb have also been detected in the adult brain. SNARE proteins, such as CaMKII, clathrin, syntaxin, SNAP25, munc18, synaptophysin and synaptotagmin, are also expressed in the honey bee brain. Honey bee myosin-Va displayed ATP-dependent solubility and was associated with DYNLL1/LC8 and SNARE proteins in the membrane vesicle-enriched fraction. Myosin-Va expression was also decreased after the intracerebral injection of melittin and NMDA. The immunolocalization of myosin-Va and -IV, DYNLL1/LC8, and synaptophysin in mushroom bodies, and optical and antennal lobes was compared with the brain morphology based on Neo-Timm histochemistry and revealed a distinct and punctate distribution. This result suggested that the pattern of localization is associated with neuron function. Therefore, our data indicated that the roles of myosins, DYNLL1/LC8, and SNARE proteins in the nervous and visual systems of honey bees should be further studied under different developmental, caste and behavioral conditions.

The Effects of Isoflavone Supplementation Plus Combined Exercise on Lipid Levels, and Inflammatory and Oxidative Stress Markers in Postmenopausal Women

This study tested the effect of isoflavone supplementation in addition to combined exercise training on plasma lipid levels, inflammatory markers and oxidative stress in postmenopausal women. Thirty-two healthy and non-obese postmenopausal women without hormone therapy were randomly assigned to exercise + placebo (PLA; n = 15) or exercise + isoflavone supplementation (ISO; n = 17) groups. They performed 30 sessions of combined exercises (aerobic plus resistance) over ten weeks and consumed 100 mg of isoflavone supplementation or placebo. Blood samples were collected after an overnight fast to analyze the lipid profile, interleukin-6 (IL-6), interleukin-8 (IL-8), superoxide dismutase (SOD), total antioxidant capacity (FRAP), and thiobarbituric acid reactive substances (TBARS), before and after ten weeks of the intervention. There were no differences in the changes (pre vs. post) between groups for any of the inflammatory markers, oxidative stress markers or lipid profile variables. However, interleukin-8 was different between pre- and post-tests (p < 0.001) in both groups (Δ = 7.61 and 5.61 pg/mL) as were cholesterol levels (p < 0.05), with no interaction between groups. The combination of isoflavone supplementation and exercise training did not alter oxidative stress markers in postmenopausal women, but exercise training alone may increase IL-8 and decrease total cholesterol levels.

Antioxidant and anti-glycation capacities of some medicinal plants and their potential inhibitory against digestive enzymes related to type 2 diabetes mellitus

ETHNOPHARMACOLOGICAL RELEVANCE Plants preparations are used by traditional medicine in the treatment of various diseases, such as type-2 diabetes mellitus. Some medicinal plants are capable of controlling the complications of this metabolic disease at different levels, for example, providing antioxidant compounds that act against oxidative stress and protein glycation and others which are capable of inhibiting the catalysis of digestive enzymes and thus contribute to the reduction of hyperglycemia and hyperlipidemia. Our objective was to investigate the antioxidant and anti-glycation activities of some medicinal plants and their potential inhibitory against α-amylase, α-glucosidase and pancreatic lipase activities. MATERIAL AND METHODS Based on the ethnobotanical researches carried out by academic studies conducted at the Federal University of Uberlandia, ten plants traditionally used in the treatment of type-2 diabetes mellitus were selected. Ethanol (EtOH) and hexane (Hex) extracts of specific parts of these plants were used in enzymatic assays to evaluate their inhibitory potential against α-amylase, α-glucosidase and lipase, as well as their antioxidant (DPPH, ORAC and FRAP) and anti-glycation (BSA/fructose model) capacities. RESULTS The results indicate that EtOH extract of four of the ten analyzed plants exhibited more than 70% of antioxidant and anti-glycation capacities, and α-amylase and lipase inhibitory activities; no extract was able to inhibit more than 40% the α-glucosidase activity. The EtOH extracts of Bauhinia forficata and Syzygium. cumini inhibited α-amylase (IC50 8.17 ± 2.24 and 401.8 ± 14.7 μg/mL, respectively), whereas EtOH extracts of B. forficata, Chamomilla recutita and Echinodorus grandiflorus inhibited lipase (IC50 59.6 ± 10.8, 264.2 ± 87.2 and 115.8 ± 57.1 μg/mL, respectively). In addition, EtOH extracts of B. forficata, S. cumini, C. recutita and E. grandiflorus showed, respectively, higher antioxidant capacity (DPPH IC50 0.7 ± 0.1, 2.5 ± 0.2, 1.3 ± 0.2 and 35.3 ± 9.0 μg/mL) and anti-glycation activity (IC50 22.7 ± 4.4, 246.2 ± 81.7, 18.5 ± 2.8 and 339.0 ± 91.0 μg/mL). CONCLUSIONS EtOH extracts of four of the ten species popularly cited for treatment of type 2 diabetes mellitus have shown promising antioxidant and anti-glycation properties, as well as the ability to inhibit the digestive enzymes α-amylase and lipase. Thus, our results open new possibilities for further studies in order to evaluate the antidiabetic potential of these medicinal plants.

Effect of saliva collection methods and oral hygiene on salivary biomarkers

Abstract The aim of this study was to evaluate the influence of unstimulated and stimulated saliva collection methods, as well as tooth brushing, on the secretion rate of salivary total protein, nitrite, total antioxidant capacity and alpha-amylase. Saliva of 14 healthy individuals were collected with stimulation using Salivette®, Parafilm® and chewing gum and without stimulation from spit with and without fluid accumulation, before and after oral hygiene. Total protein, nitrite, total antioxidant capacity and alpha-amylase concentration (sAA) were evaluated. The collection of saliva stimulated with Parafilm® and chewing gum increased the salivary flow (1.5 ± 0.4 and 3.4 ± 0.7 mL/min, respectively) and the secretion rate of salivary total protein (1.0 ± 0.2 and 2.3 ± 0.5 mg/min, respectively). Also, chewing gum increases the salivary nitrite secretion (213 ± 58 nmol/min) and total antioxidant capacity (410 ± 47 nmol trolox eq/min). Interestingly, the unstimulated method without saliva accumulation prior to collection resulted in low sAA levels (23,531 ± 7979 pixel density). Furthermore, oral hygiene decreased salivary flow (1.3 ± 0.5 to 1.0 ± 0.4 mL/min), reduced the secretion rate of total protein (1.0 ± 0.5 to 0.6 ± 0.2 mg/min, p < .05) and increased sAA (13,159 ± 7114 to 20,075 ± 25,656 pixel density, p < .05). The type of stimulation can activate autonomous receptors responsible for the secretion and composition of saliva. Therefore, the evaluation of saliva collection methods and oral hygiene on salivary biomarkers is important for understanding and standardizing variations in salivary composition to strengthen the use of saliva as a diagnostic fluid.

Adaptogenic potential of royal jelly in liver of rats exposed to chronic stress

Restraint and cold stress increase both corticosterone and glycemia, which lead to oxidative damages in hepatic tissue. This study assessed the effect of royal jelly (RJ) supplementation on the corticosterone level, glycemia, plasma enzymes and hepatic antioxidant system in restraint and cold stressed rats. Wistar rats were allocated into no-stress, stress, no-stress supplemented with RJ and stress supplemented with RJ groups. Initially, RJ (200mg/Kg) was administered for fourteen days and stressed groups were submitted to chronic stress from the seventh day. The results showed that RJ supplementation decreases corticosterone levels and improves glycemia control after stress induction. RJ supplementation also decreased the body weight, AST, ALP and GGT. Moreover, RJ improved total antioxidant capacity, SOD activity and reduced GSH, GR and lipoperoxidation in the liver. Thus, RJ supplementation reestablished the corticosterone levels and the hepatic antioxidant system in stressed rats, indicating an adaptogenic and hepatoprotective potential of RJ.