ß-Caryophyllene nanoparticles design and development: Controlled drug delivery of cannabinoid CB2 agonist as a strategic tool towards neurodegeneration.

The sesquiterpene ß-caryophyllene (BCP) is a structurally singular cannabinoid and a selective agonist of the CB2 receptor, which in addition to being expressed in the CNS, is intrinsically expressed within the immune system and lacks psychoactivity. Nanoencapsulation of BCP can allow its controlled release into the CNS and intranasal administration. Thus, a protocol for nanoencapsulation of BCP was developed and optimized in order to adjust the desired bioactive content and physicochemical parameters. The formulation was assessed regarding nanoparticle size, zeta potential, morphology, pH, osmolarity, stability, and drug release kinetics in vitro. The final composition of the BCP nanoparticles presented in its organic phase (OP) Tween 20 (0.25%), BCP (0.1%), and PEG 400 (5%); and in its aqueous phase (AP) ultrapure water and poloxamer P188 (0.25%). The formulation showed to be suitable for intranasal administration, presenting pH 6.5 and osmolarity of 150 mmol/kg. The mean particle diameter was 147.2 nm, PDI 0.052, and zeta potential of -24.5. The accelerated stability test showed that nanoparticles were stable for up to 1 month, when reversible creaming effect occurred. Besides, it was noted a low rate of particle accumulation and particle size distribution remained unchanged. BCP nanoparticles were shown to be promptly released in physiological medium (up to 60 min). In this work, a formulation containing ß-caryophyllene nanoparticles suitable for physiological administration and preclinical tests was successfully developed.

Polyvinyl alcohol-based electrospun matrix as a delivery system for nanoemulsion containing chalcone against Leishmania (Leishmania) amazonensis.

Cutaneous leishmaniasis is a worldwide public health problem. Conventional therapies, in addition to the high cost, have many adverse effects and cases of parasite's resistance. Chalcones are secondary metabolites precursors in the flavonoid pathway and can be obtained naturally, but with low yield from plant raw material. Thus, the use of synthetic chalcones has been a promising strategy for the development of molecules with leishmanicidal activity. Thus, this work aimed to develop a controlled release system of two synthetic chalcone (trans-chalcones and 3'-(trifluormethyl)-chalcone) using polyvinyl alcohol nanofibers (PVA) as scaffold. The association of chalcones to the nanofibers was made by nanoemulsions (NE) thereof, i.e., a colloidal system on a nanometric scale, which allows compounds with opposite polarities to remain miscible and stable throughout their manipulation. Chalcone nanoemulsions were developed using the spontaneous emulsification technique. The NE were characterized regarding their particle size, polydispersion index (PDI), and zeta potential. The results showed NE with spherical shape, absolute values of zeta potential were higher than 30 mV and homogeneous distribution pattern (PDI < 0.3). Dynamics light scattering (DLS) analysis showed similar hydrodynamic rays, i.e., 180 nm (trans-chalcone NE) and 178 nm (NE containing 3'-(trifluormethyl)-chalcone, in addition to presenting encapsulation efficiency values close to 100 %. Subsequently, the NE were added to a polymeric solution of polyvinyl alcohol (PVA) and processed via the electrospinning technique affording a PVA matrix (15 %, w/v) nanofiber containing the chalcones NE at 1 mg.mL-1. In a follow-up experiment, the skin permeation assay of the PVA matrix-chalcone NE was performed in vitro using Franz type diffusion cells and porcine ear as biological model of study. The results showed that the treatments with the nanofibers containing the chalcone NE were retained mainly in the stratum corneum, while the NE suspensions containing chalcone were retained in the epidermis and dermis. This result is thought to be relevant, since parasites are located mainly in the dermis. Further, in vitro assay against the amastigote form of L. (L) amazonensis, showed IC50 values to trans-chalcone and 3'-(trifluormethyl)-chalcone of 24.42 ± 6.76 µg.mL-1 and 15.36 ± 4.61 µg.mL-1, respectively. In addition to improving the solubility of the compounds tested in culture medium without using organic solvents, chalcones in nano-emulsified form reduced the IC50 to 9.09 ± 1.24 µg.mL-1 (trans-chalcone) and 10.27 ± 2.27 µg.mL-1 (3'-(trifluormethyl)-chalcone) which confirmed the potential of the nanoemulsion containing chalcone for cutaneous leishmaniasis treatment.

A simplified radiosynthesis of [18 F]MK-6240 for tau PET imaging.

[18 F]MK-6240 (6-(fluoro)-3-(1H-pyrrolo[2,3-c]pyridin-1-yl)isoquinolin-5-amine) is a highly selective PET radiotracer for the in vivo imaging of neurofibrillary tangles (NFTs). [18 F]MK-6240 was synthesized in one step from its bis-Boc protected precursor N-[(tert-butoxy)carbonyl]-N-(6-nitro-3-[1H-pyrrolo[2,3-c]pyridin-1-yl]isoquinolin-5-yl) carbamate in DMSO using [18 F] fluoride with TEA HCO3 with step-wise heating up to 150°C, resulting in an isolated radiochemical yield of 9.8% ± 1.8% (n = 3) calculated from the end of bombardment (5.2% ± 1.0% calculated from the end of synthesis). This new synthetic approach eliminates the acidic deprotection of the bis-Boc 18 F-labeled intermediate, which reduces the number of operations necessary for the synthesis as well as losses, which occur during deprotection and neutralization of the crude product mixture prior to the HPLC purification. The synthesis was performed automatically with a single-use cassette on an IBA Synthera+ synthesis module. This synthesis method affords the radioligand with a reliable radiochemical yield, high radiochemical purity, and a high molar activity. [18 F]MK-6240 synthesized with this method has been regularly (n > 60) used in our ongoing human and animal PET imaging studies.

Nanotechnology: A Promising Tool Towards Wound Healing.

Nanotechnology is an exciting emerging field with multiple applications in skin regeneration. Nanofibers have gained special attention in skin regeneration based on their structural similarity to the extracellular matrix. A wide variety of polymeric nanofibers with distinct properties have been developed and tested as scaffolds for skin regeneration. Besides providing support for tissue repair, nanofibrous materials can act as delivery systems for drugs, proteins, growth factors, and other molecules. Moreover, the morphology, biodegradability, and other functionalities of nanofibrous materials can be controlled towards specific conditions of wound healing. Other nanostructured drug delivery systems, such as nanoparticles, micelles, nanoemulsions, and liposomes, have been used to improve wound healing at different stages. These nanoscale delivery systems have demonstrated several benefits for the wound healing process, including reduced cytotoxicity of drugs, administration of poorly water-soluble drugs, improved skin penetration, controlled release properties, antimicrobial activity, and protection of drugs against light, temperature, enzymes or pH degradation, as well as stimulation of fibroblast proliferation and reduced inflammation.

(-)-ß-Caryophyllene, a CB2 Receptor-Selective Phytocannabinoid, Suppresses Motor Paralysis and Neuroinflammation in a Murine Model of Multiple Sclerosis.

(-)-ß-caryophyllene (BCP), a cannabinoid receptor type 2 (CB2)-selective phytocannabinoid, has already been shown in precedent literature to exhibit both anti-inflammatory and analgesic effects in mouse models of inflammatory and neuropathic pain. Herein, we endeavored to investigate the therapeutic potential of BCP on experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS). Furthermore, we sought to demonstrate some of the mechanisms that underlie the modulation BCP exerts on autoimmune activated T cells, the pro-inflammatory scenery of the central nervous system (CNS), and demyelination. Our findings demonstrate that BCP significantly ameliorates both the clinical and pathological parameters of EAE. In addition, data hereby presented indicates that mechanisms underlying BCP immunomodulatory effect seems to be linked to its ability to inhibit microglial cells, CD4+ and CD8+ T lymphocytes, as well as protein expression of pro-inflammatory cytokines. Furthermore, it diminished axonal demyelination and modulated Th1/Treg immune balance through the activation of CB2 receptor. Altogether, our study represents significant implications for clinical research and strongly supports the effectiveness of BCP as a novel molecule to target in the development of effective therapeutic agents for MS.

Essential oil from Pterodon emarginatus seeds ameliorates experimental autoimmune encephalomyelitis by modulating Th1/Treg cell balance.

ETHNOPHARMACOLOGICAL RELEVANCE: Pterodon emarginatus Vogel is a medicinal plant commonly used in Brazilian traditional medicine as a folk therapy due to its immunosuppressive, anti-inflammatory, anti-rheumatic, healing, tonic and depurative activities. The essential oil (EO) of Pterodon emarginatus is composed of volatile aromatic terpenes and phenyl propanoids, mainly, ß-elemene and ß-caryophyllene sesquiterpenes. Here we reported the effects and some underlying mechanisms of action of EO during murine model of MS, the experimental autoimmune encephalomyelitis (EAE). MATERIALS AND METHODS: EO (50 and 100 mg/kg) was orally administered during the entire period of development of EAE (preventive treatment, day 0-25). In vitro and in vivo immunological responses were evaluated by ELISA, immunohistochemistry, immunofluorescence and flow cytometry. RESULTS: We provide evidence that EO of Pterodon emarginatus (100 mg/kg, p.o.) significantly attenuates neurological signs and also the development of EAE. Furthermore, at the same dose EO consistently inhibited Th1 cell-mediated immune response and upregulated Treg response in vitro. Moreover, the EO inhibited both microglial activation and expression of iNOS, associated with inhibition of axonal demyelization and neuronal death during the development of the disease. CONCLUSION: This is the first experimental evidence showing that oral administration of EO consistently reduces and limits the severity and development of EAE, mainly, through the modulation of Th1/Treg immune balance, and might represent a helpful new tool for control immunoinflammatory conditions, such as MS.

Spatial reference memory deficits precede motor dysfunction in an experimental autoimmune encephalomyelitis model: the role of kallikrein-kinin system.

Multiple sclerosis (MS) is a progressive T cell-mediated autoimmune demyelinating inflammatory disease of the central nervous system (CNS). Although it is recognized that cognitive deficits represent a manifestation of the disease, the underlying pathogenic mechanisms remain unknown. Here we provide evidence of spatial reference memory impairments during the pre-motor phase of experimental autoimmune encephalomyelitis (EAE) in mice. Specifically, these cognitive deficits were accompanied by down-regulation of choline acetyltransferase (ChAT) mRNA expression on day 5 and 11 post-immunization, and up-regulation of inflammatory cytokines in the hippocampus and prefrontal cortex. Moreover, a marked increase in B1R mRNA expression occurred selectively in the hippocampus, whereas protein level was up-regulated in both brain areas. Genetic deletion of kinin B1R attenuated cognitive deficits and cholinergic dysfunction, and blocked mRNA expression of both IL-17 and IFN-γ in the prefrontal cortex, lymph node and spleen of mice subjected to EAE. The discovery of kinin receptors, mainly B1R, as a target for controlling neuroinflammatory response, as well as the cognitive deficits induced by EAE may foster the therapeutic exploitation of the kallikrein-kinin system (KKS), in particular for the treatment of autoimmune disorders, such as MS, mainly during pre-symptomatic phase.