Nanodispersions of beta-carotene: effects on antioxidant enzymes and cytotoxic properties

Beta-carotene is a carotenoid precursor of vitamin A, known for its biological activities. Due to its high hydrophobicity, nanonization processes, i.e. the transformation into nanoparticles, can improve its water affinity, and therefore the activity in aqueous systems. The objective of this study was to produce beta-carotene nanoparticles by the solid dispersion method and to evaluate their effects on the activity of glutathione-S-transferase and acetylcholinesterase enzymes using Drosophila melanogaster (DM) homogenate, the superoxide dismutase- and catalase-like activities under in vitro conditions, and their cytotoxic properties against tumor and non-tumor cells. The formed nanometric beta-carotene particles resulted in stable colloids, readily dispersed in water, able to modulate acetylcholinesterase (AChE) activity, and presenting high potential to control the cholinergic system. Beta-carotene nanoparticles, at concentrations much lower than the pure pristine beta-carotene, presented in vitro mimetic activity to superoxide dismutase and altered glutathione-S-transferase activity in DM tissue. The content of hydrogen peroxide was neither affected by the nanoparticles (in aqueous solution) nor by pristine beta-carotene (in DMSO). In the cytotoxic assays, beta-carotene nanoparticles dispersed in water showed activity against four different tumor cell lines. Overall, beta-carotene nanoparticles presented significant bioactivity in aqueous medium surpassing their high hydrophobicity constraint.The authors thank CNPq, CAPES and Fundação Araucária for the support. The authors are also grateful to the Foundation for Science and Technology (FCT, Portugal) for financial support to CIMO (strategic project UID/AGR/00690/2013) and R. Calhelha contract, and to the project POCI-01-0145- FEDER-006984 – Associate Laboratory LSRE-LCM funded by the FEDER through COMPETE2020 – Programa Operacional Competitividade e Internacionalização (POCI) – and by national funds through FCT. This work was also funded by the European Structural and Investment Funds (FEEI) through the Regional Operational Program North 2020, within the scope of Project NORTE-01-0145-FEDER-023289: DeCodE and Project Mobilizador ValorNatural®.info:eu-repo/semantics/publishedVersio

Anxiolytic-like effects of acute and chronic treatment with Achillea millefolium L. extract

AbstractEthnopharmacological relevanceAchillea millefolium L. (Asteraceae), known as yarrow (“mil folhas”), has been used as folk medicine for gastrointestinal disorders, inflammation, anxiety, and insomnia.AimTo evaluate the potential anxiolytic-like effect of hydroalcoholic extract of Achillea millefolium L. in animal models.MethodsThe present study evaluated the effects of the hydroalcoholic extract from the aerial parts of Achillea millefolium L. in mice subjected to the elevated plus-maze, marble-burying, and open-field tests. Additionally, the GABAA/benzodiazepine (BDZ) mediation of the effects of Achillea millefolium was evaluated by pretreatment with the noncompetitive GABAA receptor antagonist picrotoxin and the BDZ antagonist flumazenil and by [3H]-flunitrazepam binding to the BDZ site on the GABAA receptor.ResultsAchillea millefolium exerted anxiolytic-like effects in the elevated plus-maze and marble-burying test after acute and chronic (25 days) administration at doses that did not alter locomotor activity. This behavioral profile was similar to diazepam. The effects of Achillea millefolium in the elevated plus-maze were not altered by picrotoxin pretreatment but were partially blocked by flumazenil. Furthermore, Achillea millefolium did not induce any changes in [3H]-flunitrazepam binding.ConclusionThe results indicate that the orally administered hydroalcoholic extract of Achillea millefolium L. exerted anxiolytic-like effects that likely were not mediated by GABAA/BDZ neurotransmission and did not present tolerance after short-term, repeated administration

The involvement of the TRPA1 receptor in a mouse model of sympathetically maintained neuropathic pain

AbstractSympathetic fibres maintain some forms of neuropathic pain, but the underlying mechanisms are poorly understood. Therefore, this study investigated the possible involvement of transient receptor potential ankyrin 1 (TRPA1) and the role of the sympathetic nervous system (involved in sympathetically maintained neuropathic pain) in a model of neuropathic pain induced by sciatic nerve chronic constriction injury (CCI) in mice. Systemic injection of the selective TRPA1 antagonist HC-030031 reversed the mechanical and cold allodynia that was induced by sciatic nerve chronic constriction injury (CCI). Nerve injury also sensitised mice to nociception, which was induced by the intraplantar injection of a low dose of the TRPA1 agonist allyl isothiocyanate without changing TRPA1 immunoreactivity in the injected paw. Furthermore, chemical sympathectomy produced by guanethidine largely prevented CCI-induced mechanical and cold allodynia. CCI also induced a norepinephrine-triggered nociception that was inhibited by an α-adrenoceptor antagonist, norepinephrine transporter block and monoamine oxidase inhibition. Finally, the peripheral injection of HC-030031 also largely reduced CCI-induced norepinephrine nociception and mechanical or cold allodynia. Taken together, the present findings reveal a critical role of TRPA1 in mechanical and cold hypersensitivity and norepinephrine hypersensitivity following nerve injury. Finally, our results suggest that TRPA1 antagonism may be useful to treat patients who present sympathetically maintained neuropathic pain

Mechanisms Involved in the Nociception Triggered by the Venom of the Armed Spider <i>Phoneutria nigriventer</i>

<div><p>Background</p><p> The frequency of accidental spider bites in Brazil is growing, and poisoning due to bites from the spider genus <i>Phoneutria nigriventer</i> is the second most frequent source of such accidents. Intense local pain is the major symptom reported after bites of <i>P. nigriventer</i>, although the mechanisms involved are still poorly understood. Therefore, the aim of this study was to identify the mechanisms involved in nociception triggered by the venom of <i>Phoneutria nigriventer</i> (PNV).</p><p>Methodology/Principal Findings</p><p> Twenty microliters of PNV or PBS was injected into the mouse paw (intraplantar, i.pl.). The time spent licking the injected paw was considered indicative of the level of nociception. I.pl. injection of PNV produced spontaneous nociception, which was reduced by arachnid antivenin (ArAv), local anaesthetics, opioids, acetaminophen and dipyrone, but not indomethacin. Boiling or dialysing the venom reduced the nociception induced by the venom. PNV-induced nociception is not dependent on glutamate or histamine receptors or on mast cell degranulation, but it is mediated by the stimulation of sensory fibres that contain serotonin 4 (5-HT₄) and vanilloid receptors (TRPV1). We detected a kallikrein-like kinin-generating enzyme activity in tissue treated with PNV, which also contributes to nociception. Inhibition of enzymatic activity or administration of a receptor antagonist for kinin B₂ was able to inhibit the nociception induced by PNV. PNV nociception was also reduced by the blockade of tetrodotoxin-sensitive Na⁺ channels, acid-sensitive ion channels (ASIC) and TRPV1 receptors.</p><p>Conclusion/Significance</p><p>Results suggest that both low- and high-molecular-weight toxins of PNV produce spontaneous nociception through direct or indirect action of kinin B₂, TRPV1, 5-HT₄ or ASIC receptors and voltage-dependent sodium channels present in sensory neurons but not in mast cells. Understanding the mechanisms involved in nociception caused by PNV are of interest not only for better treating poisoning by <i>P. nigriventer</i> but also appreciating the diversity of targets triggered by PNV toxins.</p></div

Role of tissue kallikrein and kinins in PNV-induced nociception.

<p>The effect of i.pl. treatment with the kallikrein inhibitors aprotinin (10 µg/paw, A) and SBTI (3 µg/paw, B) or the B₂ receptor antagonist HOE140 (3 nmol/paw, D) on PNV-induced (3 µg/paw, i.pl.) nociception in mice. Each column represents the mean± S.E.M of 5–6 mice (A, B and C). The asterisks denote the significance levels. *P<0.05, **P<0.01 compared with the PNV group (A, B, C: Student's t-test).</p

Effect of boiling or dialysing venom in the PNV-induced nociception.

<p>The nociceptive effect produced by i.pl. injection of boiled (A) or dialysed (B) PNV (3 µg/paw) in mice. Each column represents the mean± S.E.M of 6 mice. The asterisks denote the significance levels. **P<0.01, ***P<0.001 compared with the PBS group. ^#P<0.05, ^##P<0.01 compared with the non-boiled or non-dialysed PNV group (one-way ANOVA followed by the Student-Newman-Keuls test).</p

Involvement of TRPV1, ASIC, cyclooxygenase or sodium channels in PNV-induced nociception.

<p>The effect of i.pl. treatment with the acid-sensitive ion channel (ASIC) blocker amiloride (100 nmol/paw, A), the cyclooxygenase inhibitor indomethacin (30 µmol/paw, B), the sodium channel blocker tetrodotoxin (TTX) (20 pmol/paw, C) or the selective TRPV1 antagonist SB366791 (1 nmol/paw, D) on PNV-induced (3 µg/paw, i.pl.) nociception in mice. E) The specific binding of [³H]-resiniferatoxin to spinal cord membranes in the presence or absence of capsaicin (10 µM) or PNV (1.5–150 µg/ml). Each column represents the mean± S.E.M of 5–6 mice (A–D) of 3 experiments carried out in duplicate (E). The asterisks denote the significance levels. *P<0.05, ***P<0.001, compared with the vehicle group (A–D: Student's t-test, E: one-way ANOVA followed by the Student-Newman-Keuls test).</p