Challenges in the Development of Drug Delivery Systems Based on Small Extracellular Vesicles for Therapy of Brain Diseases.

Small extracellular vesicles (sEVs) have ∼30-200 nm diameter size and may act as carriers of different cargoes, depending on the cell of origin or on the physiological/pathological condition. As endogenous nanovesicles, sEVs are important in intercellular communication and have many of the desirable features of an ideal drug delivery system. sEVs are naturally biocompatible, with superior targeting capability, safety profile, nanometric size, and can be loaded with both lipophilic and hydrophilic agents. Because of their biochemical and physical properties, sEVs are considered a promising strategy over other delivery vehicles in the central nervous system (CNS) since they freely cross the blood-brain barrier and they can be directed to specific nerve cells, potentiating a more precise targeting of their cargo. In addition, sEVs remain stable in the peripheral circulation, making them attractive nanocarrier systems to promote neuroregeneration. This review focuses on the recent progress in methods for manufacturing, isolating, and engineering sEVs that can be used as a therapeutic strategy to overcome neurodegeneration associated with pathologies of the CNS, with particular emphasis on Alzheimer's, Parkinson's, and amyotrophic lateral sclerosis diseases, as well as on brain tumors.

Overexpression of miR-124 in Motor Neurons Plays a Key Role in ALS Pathological Processes.

miRNA(miR)-124 is an important regulator of neurogenesis, but its upregulation in SOD1G93A motor neurons (mSOD1 MNs) was shown to associate with neurodegeneration and microglia activation. We used pre-miR-124 in wild-type (WT) MNs and anti-miR-124 in mSOD1 MNs to characterize the miR-124 pathological role. miR-124 overexpression in WT MNs produced a miRNA profile like that of mSOD1 MNs (high miR-125b; low miR-146a and miR-21), and similarly led to early apoptosis. Alterations in mSOD1 MNs were abrogated with anti-miR-124 and changes in their miRNAs mostly recapitulated by their secretome. Normalization of miR-124 levels in mSOD1 MNs prevented the dysregulation of neurite network, mitochondria dynamics, axonal transport, and synaptic signaling. Same alterations were observed in WT MNs after pre-miR-124 transfection. Secretome from mSOD1 MNs triggered spinal microglia activation, which was unno-ticed with that from anti-miR-124-modulated cells. Secretome from such modulated MNs, when added to SC organotypic cultures from mSOD1 mice in the early symptomatic stage, also coun-teracted the pathology associated to GFAP decrease, PSD-95 and CX3CL1-CX3CR1 signaling im-pairment, neuro-immune homeostatic imbalance, and enhanced miR-124 expression levels. Data suggest that miR-124 is implicated in MN degeneration and paracrine-mediated pathogenicity. We propose miR-124 as a new therapeutic target and a promising ALS biomarker in patient sub-populations.

Anti-inflammatory Effect and Toxicology Analysis of Oral Delivery Quercetin Nanosized Emulsion in Rats.

PURPOSE: This study evaluates the advantage of the quercetin encapsulation in nanosized emulsion (QU-NE) administered orally in rats in order to demonstrate its anti-oedematous and antioxidant effects as well as its toxicity. METHODS: The nanocarriers were prepared using the hot solvent diffusion with the phase inversion temperature methods. The nanocarriers physicochemical properties were then investigated. The anti-edematous activity was tested using paw edema in rats. In addition, NF-kB expression in subcutaneous tissue of the paws was accessed by immunohistochemistry while the lipid peroxidation was analyzed in the liver by malondialdehyde reaction with thiobarbituric acid. Hematological, renal and hepatic toxicity as well as the genetic damage were also evaluated. RESULTS: The results demonstrated that QU-NE exhibited pronounced anti-oedematous property comparable to drug diclofenac. This effect was associated with NF-κB pathway inhibition. The lipid peroxidation was also only reduced in rats treated with QU-NE. Besides this, no genetic damage, hematological, renal or hepatic toxicities were observed after administration of QU-NE. CONCLUSIONS: These results suggest that quercetin nanosized emulsion exhibits anti-oedematous and antioxidant properties and does not demonstrate toxic effects. This indicates that it has a potential application in the treatment of inflammatory diseases.

Curcumin-Loaded Chitosan-Coated Nanoparticles as a New Approach for the Local Treatment of Oral Cavity Cancer.

Mucoadhesive nanoparticles loaded with curcumin were developed as a new approach to deliver curcumin for the local treatment of oral cancer. PCL nanoparticles coated with chitosan displaying different molar masses were prepared by using the nanoprecipitation technique. The mucoadhesive properties of nanoparticle suspensions were demonstrated by their strong ability to interact with the glycoprotein mucin through electrostatic interactions. Similar permeation profiles of curcumin loaded in uncoated and chitosan-coated nanoparticles across porcine esophageal mucosa were verified. Curcumin concentrations retained in the mucosa suggest the possibility of a local effect of the drug. In vitro studies demonstrated that free curcumin.and curcumin loaded into nanoparticles coated with chitosan caused significant reduction of SCC-9 human oral cancer cell viability in a concentration and time-dependent manner. However, no significant cell death was observed after 24 h of treatment with unloaded nanoparticles coated with chitosan. In addition, curcumin-loaded nanoparticles showed reduced cytotoxicity, when compared with the free drug. Therefore, chitosan-coated PCL nanoparticles may be considered a promising strategy to deliver curcumin directly into the oral cavity for the treatment of oral cancer.

Orally Administered Chitosan-Coated Polycaprolactone Nanoparticles Containing Curcumin Attenuate Metastatic Melanoma in the Lungs.

The study was aimed to evaluate the effect of orally administered chitosan-coated nanoparticles containing curcumin on metastatic melanoma. Chitosan-coated nanoparticles containing curcumin were prepared, and their antimetastatic activity was investigated both in vitro and in vivo. Curcumin decreased cell viability and induced apoptosis of B16F10 melanoma cells. We observed that curcumin significantly decreased the expression of metalloproteinases, which are known to be associated with migration and proliferation of cancer cells. Importantly, treatment with chitosan-coated nanoparticles containing curcumin decreased pulmonary tumor formation in a murine model of experimental metastasis. Histological analyses confirmed the macroscopic results in which lungs of mice treated with curcumin-loaded chitosan-coated polycaprolactone nanoparticles had only a few small nodules and most of them were free of melanoma. Our findings indicate that nanoparticles coated with the mucoadhesive polymer chitosan containing curcumin may be a promising approach and/or intervention for the treatment of malignant melanoma.

Nanoparticles Made From Xyloglucan-Block-Polycaprolactone Copolymers: Safety Assessment for Drug Delivery.

Xyloglucan-block-polycaprolactone (XGO-PCL) copolymer nanoparticles have been proposed as nanocarriers for drug delivery. However, the possible harmful effects of exposure to nanoparticles still remain a concern. Therefore, the aim of this study is to evaluate the potential toxicity of XGO-PCL nanoparticles using in vitro and in vivo assays. Cytotoxicity and genotoxicity studies were conducted on MRC-5 human fetal lung fibroblast cells upon exposure to XGO-PCL nanoparticles. No significant reduction in the cell viability and no DNA damage were observed at the different concentrations tested. Erythrocyte toxicity was assessed by the incubation of nanoparticles with human blood. XGO-PCL nanoparticles induced a hemolytic ratio of less than 1%, indicating good blood compatibility. Finally, the subacute toxicity of XGO-PCL nanoparticles (10 mg/kg/day) was evaluated in BALB/c mice when administered orally or intraperitoneally for 14 days. Results of the in vivo toxicity study showed no clinical signs of toxicity, mortality, weight loss, or hematological and biochemical alterations after treatment with nanoparticles. Also, microscopic analysis of the major organs revealed no histopathological abnormalities, corroborating the previous results. Thus, it can be concluded that XGO-PCL nanoparticles induced no effect indicative of toxicity, indicating their potential use as drug delivery systems.

In vitro and in vivo anti-glioma activity of a chalcone-quinoxaline hybrid.

Chalcones are important compounds that exhibit multiple biological activities, including anti-inflammatory, antimitotic and antibacterial properties. In the present study, we have analyzed the potential anti-cancer activity of a chalcone named N9 (a hybrid chalcone-quinoxaline compound) using in vitro and in vivo experimental glioma models. Here, we report N9-induced inhibition of cell proliferation and also N9-induced cell death in a concentration-dependent manner in U87-MG glioma cells. These effects of N9 appear to be associated with its ability to inhibit the expression of cell cycle-associated proteins, and also the augmentation in the expression of the p21 (p21/Cip1) protein, a cyclin-dependent kinase inhibitor. Additionally, N9 also potentiates the production of the pro-apoptotic markers Bax and p53 via inhibition of MDM2. Moreover, our results show that N9 also significantly enhanced apoptosis of U87-MG cells with disruption of mitochondrial membrane potential, generation of ROS and caspase-9 activation. In vivo experiments carried out in a murine xenograft model of U87-MG revealed that N9 produced a significant reduction of tumors volume when compared to vehicle treated mice. Collectively, data demonstrate that N9 possess in vitro and in vivo anti-cancer activity, an effect that seems to involve the induction of p53 and p21 proteins, as well as, the activation of mitochondrial apoptosis pathway associated with the inhibition of protein MDM2. Overall, this study suggests N9 is affecting a variety of intracellular pathways related to tumor apoptosis. Perhaps N9 or derivate molecules could represent new potential drugs for cancer therapeutics.

A study of the relative importance of the peroxiredoxin-, catalase-, and glutathione-dependent systems in neural peroxide metabolism.

Cells are endowed with several overlapping peroxide-degrading systems whose relative importance is a matter of debate. In this study, three different sources of neural cells (rat hippocampal slices, rat C6 glioma cells, and mouse N2a neuroblastoma cells) were used as models to understand the relative contributions of individual peroxide-degrading systems. After a pretreatment (30 min) with specific inhibitors, each system was challenged with either H2O2 or cumene hydroperoxide (CuOOH), both at 100 µM. Hippocampal slices, C6 cells, and N2a cells showed a decrease in the H2O2 decomposition rate (23-28%) by a pretreatment with the catalase inhibitor aminotriazole. The inhibition of glutathione reductase (GR) by BCNU (1,3-bis(2-chloroethyl)-1-nitrosourea) significantly decreased H2O2 and CuOOH decomposition rates (31-77%). Inhibition of catalase was not as effective as BCNU at decreasing cell viability (MTT assay) and cell permeability or at increasing DNA damage (comet test). Impairing the thioredoxin (Trx)-dependent peroxiredoxin (Prx) recycling by thioredoxin reductase (TrxR) inhibition with auranofin neither potentiated peroxide toxicity nor decreased the peroxide-decomposition rate. The results indicate that neural peroxidatic systems depending on Trx/TrxR for recycling are not as important as those depending on GSH/GR. Dimer formation, which leads to Prx2 inactivation, was observed in hippocampal slices and N2a cells treated with H2O2, but not in C6 cells. However, Prx-SO3 formation, another form of Prx inactivation, was observed in all neural cell types tested, indicating that redox-mediated signaling pathways can be modulated in neural cells. These differences in Prx2 dimerization suggest specific redox regulation mechanisms in glia-derived (C6) compared to neuron-derived (N2a) cells and hippocampal slices.

La barrera hematoencefálica y la administración de medicamentos en el sistema nervioso central / The blood-brain barrier and drug delivery in the central nervous system

Objetivo. Ofrecer una visión actualizada de las dificultades debido a las barreras y estrategias usadas para permitir la liberación de medicamentos en el sistema nervioso central. Introducción. La dificultad en el tratamiento de muchas enfermedades del sistema nervioso central, mediante el uso de fármacos por vía intravenosa, se debe a la presencia de barreras que impiden la liberación de éstos: la barrera hematoencefálica, la barrera sangre-líquido cefalorraquídeo y la barrera sangre-aracnoides. Desarrollo. La barrera hematoencefálica es la principal barrera para el transporte de medicamentos en el cerebro, que actúa también como una barrera inmunológica y metabólica. Las células endoteliales de la barrera hematoencefálica están conectadas a un complejo de uniones mediante la interacción de proteínas transmembranales que sobresalen del interior hacia el exterior, formando una conexión entre las células endoteliales. El transporte de sustancias al cerebro depende de los mecanismos de transporte presentes en la barrera y la difusión de estos compuestos depende también de las características fisicoquímicas de la molécula. Algunas enfermedades alteran la permeabilidad de la barrera hematoencefálica y, por lo tanto, el paso de los medicamentos. Se han investigado diferentes estrategias como métodos para la administración de medicamentos. Conclusiones. La obtención de un conocimiento más profundo de los mecanismos de transporte a través de la barrera hematoencefálica y los cambios en la neuropatología proporcionarían una información importante sobre la etiología de algunas enfermedades y conduciría a mejores estrategias terapéuticas (AU)

Aim. To provide an updated view of the difficulties due to barriers and strategies used to allow the release of drugs in the entral nervous system. Introduction. The difficulty for the treatment of many diseases of the central nervous system, through the use of intravenous drugs, is due to the presence of barriers that prevent the release of the same: the blood-brain barrier, bloodcerebrospinal fluid barrier and the blood-arachnoid barrier. Development. The blood-brain barrier is the main barrier for the transport of drugs in the brain that also acts as a immunologic and metabolic barrier. The endothelial cells of the blood-brain barrier are connected to a junction complex through the interaction of transmembrane proteins that protrude from de inside to the outside, forming a connection between the endothelial cells. The transport of substances to the brain depends on the mechanisms of transport present in the barrier and the diffusion of these compounds also depends on the physicochemical characteristics of the molecule. Some diseases alter the permeability of the blood-brain barrier and thus the passage of drugs. Strategies such as the use of methods for drug delivery in the brain have been investigated. Conclusions. Further details regarding the mechanisms of transport across the blood-brain barrier and the changes in neuropathology would provide important information about the etiology of diseases and lead to better therapeutic strategies (AU)

[The blood-brain barrier and drug delivery in the central nervous system]. / La barrera hematoencefalica y la administracion de medicamentos en el sistema nervioso central.

AIM: To provide an updated view of the difficulties due to barriers and strategies used to allow the release of drugs in the central nervous system. INTRODUCTION: The difficulty for the treatment of many diseases of the central nervous system, through the use of intra-venous drugs, is due to the presence of barriers that prevent the release of the same: the blood-brain barrier, blood-cerebro-spinal fluid barrier and the blood-arachnoid barrier. DEVELOPMENT: The blood-brain barrier is the main barrier for the transport of drugs in the brain that also acts as a immunologic and metabolic barrier. The endothelial cells of the blood-brain barrier are connected to a junction complex through the interaction of transmembrane proteins that protrude from de inside to the outside, forming a connection between the endothelial cells. The transport of substances to the brain depends on the mechanisms of transport present in the barrier and the diffusion of these compounds also depends on the physicochemical characteristics of the molecule. Some diseases alter the permeability of the blood-brain barrier and thus the passage of drugs. Strategies such as the use of methods for drug delivery in the brain have been investigated. CONCLUSIONS: Further details regarding the mechanisms of transport across the blood-brain barrier and the changes in neuropathology would provide important information about the etiology of diseases and lead to better therapeutic strategies.

A homeopatia no SUS na perspectiva de estudantes da área da saúde / Homeopathy in the single health system from the perspective of students in the health professions gecioni Loch-Neckel

A homeopatia foi reconhecida como especialidade médica em 1980. Após a criação do SUS, alguns estados e municípios brasileiros começaram a oferecer atendimento homeopático aos usuários dos serviços públicos de saúde. Em 2006, foi editada a Portaria 971, que assegura o acesso à homeopatia a estes usuários, entre outras práticas integrativas e complementares. Diante disto, investigamos as percepções de estudantes dos cursos de Farmácia, Medicina e Odontologia sobre a homeopatia e sua prática no SUS, tendo por fundamento a teoria das representações sociais. Estas representações foram construídas com os seguintes parâmetros: a homeopatia como terapêutica; a relação entre a homeopatia, o SUS e o princípio da integralidade. Os estudantes souberam correlacionar o atendimento integral ao sujeito à compreensão de suas necessidades a partir de sua realidade. Constatou-se desconhecimento sobre os pressupostos teóricos da homeopatia e o não reconhecimento da incorporação da homeopatia no SUS, para a maioria dos estudantes. Pode ser sugerido que a inserção dos futuros profissionais no contexto das atividades em saúde e o acesso a outras racionalidades permitiriam uma escolha mais lúcida de suas práticas profissionais.

In Brazil, homeopathy was officially recognized as a medical specialty by the National Medical Board in 1980. Since the creation of the Unified National Health System (SUS), some Brazilian States and municipalities have begun to offer homeopathic care to users of public health services. In 2006, the government issued Ruling 971, which guarantees access to homeopathic care for these users, in addition to other integrative and complementary therapies. Based on the above, we studied the perceptions of undergraduate students of Pharmacy, Medicine, and Dentistry concerning homeopathy and its practice in the National Health System, based on the theory of social representations. These representations were constructed with the following parameters: homeopathy as therapy; and the relationship between homeopathy, the National Health System, and the principle of comprehensiveness. Students were able to correlate comprehensive patient care with understanding patients' needs based on their reality. Most students lacked knowledge on the theoretical premises of homeopathy and were unaware that it had been incorporated into the National Health System. The findings suggest that the inclusion of future professionals in the context of health activities and access to other rationales would allow a clearer choice of their professional practices.

Desafios para a ação interdisciplinar na atenção básica: implicações relativas à composição das equipes de saúde da família / Challenges to an interdisciplinary action in basic care: implications related to composition of family health teams

O Programa Saúde da Família (PSF) surge como uma nova estratégia de atenção à saúde e de reorientação do modelo de assistência. Partindo desses pressupostos, o presente artigo tem como objetivo refletir sobre a relação entre integralidade na atenção básica e a composição das equipes de saúde da família, na perspectiva dos integrantes da equipe mínima do PSF, caracterizando as possibilidades de atuação e contribuições de outros profissionais de saúde no PSF. Participaram da pesquisa profissionais com curso superior, membros de equipes de PSF de um município no sul do Brasil. A qualidade da experiência profissional ou pessoal acerca da atuação desses profissionais contribuiu para o conhecimento sobre suas possibilidades de intervenção. A investigação permitiu também analisar como a estratégia de saúde da família tem atingido os integrantes que constituem as equipes mínimas no contexto local. Além disso, evidenciou de que maneira a integralidade e a interdisciplinaridade têm sido entendidas pelos que compõem tais equipes, e as relações entre a composição das equipes e as (im)possibilidades de concretizá-la.

The Family Health Program emerges as a new strategy of health care as well as a reorientation of the assistance model. Based on these presuppositions, this article reflects on the relationship between integrality in basic care and the composition of family health teams, in the view of the Family Health Program minimum team members, characterizing the possible fields of action and the contributions of other health professionals in the Family Health Program. Undergraduates and members of a Family Health Program team from a certain city in the south of Brazil participated in this research. The quality of personal or professional expertise of these professionals contributed to a better understanding of their intervention possibilities. The investigation also allowed the analysis of how the family health strategy has been reaching the members who constitute the minimum teams at local context. In addition to that, the research highlighted how the integrality and interdisciplinarity have been understood by the teams members, and the relationships between team arrangement and the (im)possibilities of action.

[Challenges to an interdisciplinary action in basic care: implications related to composition of family health teams]. / Desafios para a ação interdisciplinar na atenção básica: implicações relativas à composição das equipes de saúde da família.

The Family Health Program emerges as a new strategy of health care as well as a reorientation of the assistance model. Based on these presuppositions, this article reflects on the relationship between integrality in basic care and the composition of family health teams, in the view of the Family Health Program minimum team members, characterizing the possible fields of action and the contributions of other health professionals in the Family Health Program. Undergraduates and members of a Family Health Program team from a certain city in the south of Brazil participated in this research. The quality of personal or professional expertise of these professionals contributed to a better understanding of their intervention possibilities. The investigation also allowed the analysis of how the family health strategy has been reaching the members who constitute the minimum teams at local context. In addition to that, the research highlighted how the integrality and interdisciplinarity have been understood by the teams members, and the relationships between team arrangement and the (im)possibilities of action.

Pharmacist contributions for basic care from the perspective of professionals of familial health care teams / Contribuições farmacêutica para a atenção básica a partir da perspectiva dos profissionais das equipes de saúde familiar

This study aimed to investigate the social representations of professionals included in the team of Family Health Strategy (physicians, nurses and dentists) respecting the action possibilities and contributions of the pharmacist for the basic care, and based on social psychology and, particularly, on the theory of social representations. The epistemological basis of the research is qualitative, and the data were collected by means of individual semi-structured interviews, which were submitted to analysis of categorical thematic content. Apparently, the majority of professionals already inserted in the team know and recognize the importance of professional pharmacists in the basic care, as well as their potential contribution to this topic. The representations were constructed according to the following parameters: a) the study object and the intervention area, b) the individual practice of every professional and c) his/her action in specific cases. The quality of the professional or personal experience concerning the action of these professionals has contributed for the knowledge about the possibilities of pharmacists' intervention in basic care.

Este estudo teve por objetivo investigar as representações sociais dos profissionais incluídos na equipe de Estratégia em Saúde da Família (médico, enfermeiro e odontólogo), sobre as possibilidades de atuação e as contribuições do farmacêutico na atenção básica, tendo por fundamento a psicologia social e, particularmente, a teoria das representações sociais. A base epistemológica da pesquisa é qualitativa, sendo os dados coletados por meio de entrevistas individuais semi-estruturadas e analisados por meio de análise de conteúdo categorial temático. Constatou-se que a maioria dos profissionais já inseridos na equipe conhece e reconhece a importância do profissional farmacêutico na atenção básica e as suas possibilidades de contribuição. As representações foram construídas a partir dos seguintes parâmetros: a) o objeto de estudo e de intervenção da área; b) as práticas de cada um dos profissionais e c) a sua atuação em casos específicos. A qualidade da experiência profissional ou pessoal acerca da atuação desses profissionais contribuiu para o conhecimento sobre as possibilidades de intervenção do farmacêutico na atenção básica.

Stealth and non-stealth nanocapsules containing camptothecin: in-vitro and in-vivo activity on B16-F10 melanoma.

Camptothecin (CPT) is an alkaloid that displays considerable antitumour activity, but clinical use has been limited by its poor water solubility and the instability of the lactone moiety (active form) in physiological media. We have therefore formulated the drug into nanocarrier systems in an attempt to improve its therapeutic properties. This study evaluates the effect of intraperitoneally administered stealth and non-stealth nanocapsules containing CPT on lung metastatic spread in mice inoculated with B16-F10 melanoma cells, and on the cytotoxic activity against B16-F10 melanoma cells in-vitro. Poly (D,L-lactide) PLA (non-stealth) and methoxy polyethylene glycol-(D,L-lactide) (PLA-PEG) (stealth) nanocapsules (49 and 66.6 kDa) were prepared by interfacial deposition of preformed polymer. CPT, as free drug or as drug-loaded nanocapsules, was administrated at a dose of 0.5 mg kg(-1) at 3-day intervals for 17 days. Free drug and CPT-loaded nanocapsules reduced the number of metastatic nodules by 45.09-91.76% (P < 0.05 vs positive control). However, only CPT-loaded PLA-PEG 49 kD nanocapsules significantly decreased the number of lung metastases when compared with free drug (P < 0.05). The administration of CPT-loaded nanocapsules and free drug did not result in neutropenia at the administered dose. The improved effectiveness of pegylated nanocapsules was attributed to protection of the drug by nanoencapsulation and to reduced uptake of particles by macrophages located in the lymph nodes. This assumption was supported by the in-vitro study, in which both PLA and 49 kDa PLA-PEG nanocapsules containing CPT were more cytotoxic than the free drug against B16-F10 melanoma cells.