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Objectives To circumvent cisplatin (CDDP) toxic effects and improve the antitumoural effect, our research group developed long-circulating and pH-sensitive liposomes containing CDDP (SpHL-CDDP). This study aimed to evaluate whether SpHL-CDDP is associated with intestinal protection under in-vitro conditions in the presence of host–microbiota, compared with free CDDP. Methods The cytotoxicity of CDDP and SpHL-CDDP were evaluated by colorimetric MTT and sulforhodamine B (SRB) assays. Epithelial proliferation was assessed by using an in-vitro wounding model in the presence of host–microbiota with intestinal epithelial cell line 6 (IEC-6) monolayers. Cytokines were determined by ELISA. Key findings Reduced cytotoxicity of SpHL-CDDP in IEC-6 cells (minimum of 1.3-fold according to the IC50 values) was observed when compared with CDDP. The presence of microbiota or CDDP reduced the wound healing. The association of microbiota and SpHL-CDDP improved the wound healing and cell number in IEC-6 cells when compared with control. These beneficial results can be associated with increased IL-6 and IL-10 levels induced by SpHL-CDDP which were affected by the presence of microbiota. Conclusions These results indicate that the presence of microbiota associated with SpHL-CDDP provided less intestinal cellular damages compared with CDDP and constitutes a promising candidate for clinical use.

Lippia alba (Mill.) N.E.Br. ex Britton & P. Wilson, Verbenaceae, is considered a great source of a bioactive volatile oil. Due to the wide range of known chemotypes, its chemical analysis is very important. Among the several activities of this volatile oil, a potential larvicidal action against Culicidae species is highlighted. However, the low water miscibility of volatile oils limits their application in aqueous media. Oil in water nano-emulsions are in the spotlight of novelty to solve this main problem. Thus, the aim of the present study was to obtain this nanostructured system with L. alba volatile oil (citral chemotype) and evaluate its larvicidal activity against Aedes aegypti and Culex quinquefasciatus larvae. The major compounds were geranial (30.02%) and neral (25.26%). Low mean droplet size (117.0 ± 1.0 nm) and low polydispersity index (0.231 ± 0.004) were observed and no major changes were observed after seven days of storage. LC50 values against C. quinquefasciatus and A. aegypti third-instar larvae were respectively 38.22 and 31.02 ppm, while LC90 values were, respectively, 59.42 and 47.19 ppm. The present study makes use of a low energy, solvent-free and ecofriendly method with reduced costs. Thus, this paper contributes significantly to phyto-nanobiotechnology of larvicidal agents, opening perspectives for the utilization of L. alba volatile oil in integrated practices of vector control. Keywords: Colloidal system, Droplet size distribution, Miniemulsion, Larvicide

In the last few decades, utilization of medicinal plants by the pharmaceutical industry has led to the identification of many new bioactive compounds. The genus Pterodon, native of the Brazilian Flora, is known for the therapeutic properties attributed to its species, which are widely used in popular medicine for their anti-inflammatory, anti-rheumatic, tonic, and depurative properties. The intrinsic low water solubility of the plant derivatives from the genus, including diterpenes with vouacapane skeletons that are partially associated with the pharmacological activities, impairs the bioavailability of these bioactive compounds. Recent studies have aimed to encapsulate Pterodon products to improve their water solubility, achieve stability, increase their efficacy, and allow clinical applications. The purpose of this paper is to review recent research on the use of nanotechnology for the development of new products from plant derivatives of the Pterodon genus in different types of micro- and nanocarriers. Therapeutic properties of their different products are also presented. Finally, an update about the current and future applications of encapsulated formulations is provided.

Aedes aegypti (Diptera: Culicidae) is the main vector of some neglected diseases, including dengue. It is very important to develop formulations that increase effectiveness of vector control with low toxicity. Quercetin is a plant-derived flavonoid that modulates the development of some insects. The low water solubility of quercetin impairs the development of water-dispersible commercial products. To circumvent this problem, the preparation of nanoformulations is considered promising. Thus, this study aimed to evaluate the effect of bulk and quercetin nanosuspension against A. aegypti larvae and also to investigate their ecotoxicity. Quercetin nanosuspension was produced by a solvent displacement method followed by solvent evaporation and was maintained in two different temperatures (4 and 25 °C). Its size distribution and zeta potential were monitored along 30 days. The influence of quercetin nanosuspension and bulk-quercetin was investigated at various concentrations against A. aegypti and the green algae Chlorella vulgaris. The quercetin nanosuspension presented higher stability at 4 °C and negative zeta potential values. Quercetin nanosuspension and bulk-quercetin adversely affected the larvae development, especially at the highest concentrations. Larvae mortality was between 44% and 100% (48 h) for quercetin nanosuspension at 100 and 500 ppm, respectively. The bulk-quercetin induced around 50% mortality regardless the concentration used at this same time-period. Absence of emerging mosquitoes from water was observed on the survival larvae of all the treated groups. Quercetin nanosuspension was less toxic than bulk-quercetin against C. vulgaris, especially at higher concentrations. These data indicate that quercetin nanosuspension may represent a potential larvicide for A. aegypti control, once they induced larvae death and inhibited the survival ones to emerge from water. In addition, it did not demonstrated ecotoxicity against a non-target organism, highlighting its better properties, when compared to the bulk-quercetin. Keywords: Culicidae, Ecotoxicity, Larvicide, Nanocarrier, Flavonoid

The plant species Libidibia ferrea (Mart. ex Tul.) LP Queiroz var. ferrea basionym of Caesalpinia ferrea (Mart. ex Tul.) is used in various regions of Brazil in folk medicine in the treatment of several health problems, especially in acute and chronic inflammatory processes. Most of the preparations employed are alcoholic. Therefore, this study aimed to evaluate the acute toxicity of the hydroethanolic extract of fruits of Libidibia ferrea (EHEFLf) in zebrafish, emphasizing the possible changes in the organic-cellular level of the gills, liver, kidneys, and intestine and on embryos. The result obtained by LC-M/MS from EHEFLf indicated a high concentration of possible polyhydroxylated substances. EHEFLf, at a dose of 2 g/kg orally, produced non-significant alterations of the analyzed organs. However, for embryos, the treatment with different concentrations demonstrated heart toxicity that was concentration-dependent. There is no evidence of a correlation of the observed effects with the phytochemical composition, and considering the species of animal used, it can be suggested that the oral use of L. ferrea hydroethanolic extract has an acceptable degree of safety for use as an oral medicinal product. and embryo results have shown significant affinity to the heart, however, it is perceived to be related to the concentrations used.

In this work, we developed new scripts to calculate the lacunarity exponent and Moran’s index of Atomic Force Microscopy (AFM) images. The lacunarity exponent was estimated by combining the Otsu binarization and gliding-box algorithm, and Moran index was introduced to evaluate the surfaces’ spatial autocorrelation. Developed scripts were first validated using numerical simulation of self-similar fractal and self-affine isotropic surfaces. Then, we successfully synthesized chitosan films with different glycerol concentrations and used the lacunarity and Moran’s index for a thorough characterization. The validation of the proposed scripts using simulated Sierpinski Carpets and 3D artificial surfaces showed promising potential for analyzing AFM images. Finally, the methodology application to AFM images of chitosan films suggested that lacunarity analysis and Moran index determination could complement thin films’ quality processing control.

Cisplatin (CDDP) is a chemotherapeutic agent widely used in several anticancer protocols for instance head and neck, testicle, ovarian, lung and peritoneal carcinomatosis. According to the literature, the use of CDDP is associated with several side effects; among them, we highlighted the mucositis. CDDP, when administered by IP, promoted significant intestinal epithelium alterations in an experimental model. Our research group has proposed that the incorporation of CDDP into long-circulating and pH-sensitive liposomes (SpHL-CDDP) could help to overcome some side effects induced by this drug. Thus, we evaluated signs of intestinal toxicity 24 h and 72 h after the administration of a single i.p dose of free CDDP or SpHL-CDDP to healthy Swiss mice. Twenty-four hours after administration of free CDDP, the mice showed signs of intestinal toxicity, principally weight loss, increased intestinal permeability associated with a decrease in expression of tight junctions, and histological damage with the presence of inflammatory infiltrates and activation of ERK1/2 and NF-κB. These changes persisted after 72 h. While signs of intestinal toxicity were also observed 24 h after administration of SpHL-CDDP, after 72 h body weight and intestinal permeability of mice in this group were similar to those of mice in the control group. In comparison with the free CDDP treatment group, 72 h after treatment mice in the SpHL-CDDP group showed better histological parameters, lower levels of inflammatory infiltrate with increased IL-10 and IgA levels, and less activation of caspase-3, ERK1/2 and NF-κB. These differences could account for the recovery of the intestinal epithelium observed in mice treated with SpHL-CDDP but not in mice treated with free CDDP. In conclusion, here we show that encapsulation of CDDP in SpHL lessens intestinal damage and that, as such, SpHL-CDDP is a promising candidate for clinical use.

The present work shows the development and evaluation of the veterinary antibiotic cloxacillin benzathine (CLOXB) loaded into poly-e-caprolactone (PCL) nanocapsules (NC), as a potential new treatment strategy to manage bovine intramammary infections, such as mastitis. Staphylococcus aureus-induced mastitis is often a recurrent disease due to the persistence of bacteria within infected cells. CLOXB-PCL NC were prepared by interfacial deposition of preformed biodegradable polymer followed by solvent displacement method. The mean diameter of NC varied from 241 to 428 nm and from 326 to 375 nm, when determined by dynamic light scattering and by atomic force microscopy, respectively. The zeta potential of NC was negative and varied from −28 to −51 mV. In vitro release studies from the NC were performed in two media under sink conditions: PBS with 1% polyethylene glycol or milk. A reversed-phase HPLC method was developed to determine the NC entrapment efficiency and kinetics of CLOXB release from the NC. Free CLOXB dissolution occurred very fast in both media, while drug release from the NC was slower and incomplete (below 50%) after 9 h. CLOXB release kinetics from polymeric NC was fitted with the Korsmeyer-Peppas model indicating that CLOXB release is governed by diffusion following Fick's law. The fluorescence confocal microscopy images of macrophage-like J774A.1 cells reveal NC uptake and internalization in vitro. In addition, antimicrobial effect of the intramammary administration of CLOXB-PCL NC in cows with mastitis resulted in no clinical signs of toxicity and allowed complete pathogen elimination after treatment. The in vivo results obtained in this work suggest that CLOXB-PCL NC could be a promising formulation for the treatment of intramammary infections in cattle, considering their physicochemical properties, release profiles and effects on bovine mastitis control.

Cutaneous leishmaniasis (CL) is a neglected tropical skin disease caused by the protozoan genus Leishmania. The treatment is restricted to a handful number of drugs that exhibit toxic effects, limited efficacy, and drug resistance. Additionally, developing an effective topical treatment is still an enormous unmet medical challenge. Natural oils, e.g. the oleoresin from P. emarginatus fruits (SO), contain various bioactive molecules, especially terpenoid compounds such as diterpenes and sesquiterpenes. However, its use in topical formulations can be impaired due to the natural barrier of the skin for low water solubility compounds. Nanoemulsions (NE) are drug delivery systems able to increase penetration of lipophilic compounds throughout the skin, improving their topical effect. In this context, we propose the use of SO-containing NE (SO-NE) for CL treatment. The SO-NE was produced by a low energy method and presented suitable physicochemical characteristic: average diameter and polydispersity index lower than 180 nm and 0.2, respectively. Leishmania (Leishmania) amazonensis-infected BALB/c mice were given topical doses of SO or SO-NE. The topical use of a combination of SO-NE and intraperitoneal meglumine antimoniate reduced lesion size by 41 % and tissue regeneration was proven by histopathological analyses. In addition, a reduction in the parasitic load and decreased in the level of IFN-γ in the lesion may be associated, as well as a lower level of the cytokine IL-10 may be associated with a less intense inflammatory process. The present study suggests that SO-NE in combination meglumine antimoniate represents a promising alternative for the topical treatment of CL caused by L. (L.) amazonensis.

The intestinal epithelium is composed of specialized epithelial cells that form a physical and biochemical barrier to commensal and pathogenic microorganisms. However, dysregulation of the epithelial barrier function can lead to increased intestinal permeability and bacterial translocation across the intestinal mucosa, which contributes to local and systemic immune activation. The increase in these parameters is associated with inflammatory bowel disease, physical exercise under heat stress, intestinal obstruction, ischemia, and mucositis, among other conditions. Lately, there has been growing interest in immunomodulatory nutrients and probiotics that can regulate host immune and inflammatory responses and possibly restore the intestinal barrier. Immunomodulators such as amino acids (glutamine, arginine, tryptophan, and citrulline), fatty acids (short-chain and omega-3 fatty acids and conjugated linoleic acids), and probiotics (Bifidobacterium, Saccharomyces, and Lactobacillus) have been reported in the literature. Here, we review the critical roles of immunomodulatory nutrients in supporting gut barrier integrity and function.

Background Candida spp is an etiologic agent of fungal infections in hospitals and resistance to treatment with antifungals has been extensively reported. Thus, it is very important to develop formulations that increase effectiveness with low toxicity. In this sense, nanocarriers have been investigated, once they modify drug biodistribution profile. Thus, this study aimed to evaluate the biodistribution of free and encapsulated 99mTc-fluconazole into nanocapsules (NCs) in an experimental immunosuppressed murine model of Candida albicans infection. Methods Fluconazole was radiolabeled with technetium-99 metastable (99mTc) and encapsulated into conventional (99mTc-Fluconazole-PLA-POLOX) and surface-modified (99mTc-Fluconazole-PLA-PEG) NCs by the interfacial deposition of the preformed biodegradable polymer [poly (D,L-lactic acid) (PLA) and PLA–PEG (polyethyleneglycol)] followed by solvent evaporation. The size distribution and zeta potential of the NCs preparations were determined in a Zetasizer by photon correlation spectroscopy and laser Doppler anemometry, respectively. Free and encapsulated 99mTc-fluconazole were administered intravenously in immunosuppressed mice bearing a local infection induced by Candida Albicans inoculation in the right thigh muscle. At pre-established time intervals, tissues and organs of interest were removed and radioactivity was measured in an automatic gamma radiation counter. Results The NCs diameter was between 200 and 400 nm with negative zeta potential values. Free 99mTc-fluconazole was more rapidly eliminated by the renal system compared to the encapsulated drug in NCs, which remained longer in blood circulation. The uptake of conventional NCs by mononuclear phagocyte system organs was higher than the one demonstrated by the surface-modified NCs. Both NCs remained longer in the infectious focus when compared to free 99mTc-fluconazole, but the results did not show a significant difference between NC formulations. Conclusion These data indicate that these NCs might represent a therapeutic alternative for the treatment of candidiasis, once they remain more time in the infectious focus, allowing high retention of 99mTc-fluconazole at this site.

The association between gluten and body weight is inconsistent. Previously, we showed that a gluten-free diet reduces weight gain without changing food intake in mice fed high-fat diets. In the present study, we investigated the effects of gluten intake on fat metabolism, thermogenesis and energy expenditure in mice fed a standard or high-fat diet.Mice were fed four different experimental diets during 8 weeks: a control-standard diet (CD), a CD added with 4.5% of wheat gluten (CD-G), a high-fat diet (HFD) and a HFD added with 4.5% of wheat gluten (HFD-G). After 8 weeks, the mice received (99m)Tc-radiolabeled gluten orally to study gluten absorption and biodistribution or they underwent indirect calorimetry. After killing, subcutaneous and brown adipose tissues (SAT and BAT) were collected to assess thermogenesis-related protein expression. Lipid metabolism was studied in adipocyte cultures from the four groups.Despite having had the same energy intake, CD-G and HFD-G mice exhibited increased body weight and fat deposits compared with their respective controls. (99m)Tc-GLU or its peptides were detected in the blood, liver and visceral adipose tissue, suggesting that gluten can even reach extraintestinal organs. Uncoupling protein-1 expression was reduced in the BAT of HFD-G and in the SAT of CD-G and HFD-G mice. Indirect calorimetry showed lower oxygen volume consumption in CD-G and HFD-G groups compared with their controls. In HFD mice, daily energy expenditure was reduced with gluten intake. Gluten also reduced adiponectin, peroxisome proliferator-activated receptor (PPAR)-α and PPARγ and hormone-sensitive lipase in cultures of isolated adipocytes from HFD mice, whereas in the CD-G group, gluten intake increased interleukin-6 expression and tended to increase that of tumor necrosis factor.Wheat gluten promotes weight gain in animals on both HFD and CD, partly by reducing the thermogenic capacity of adipose tissues.

Baccharis reticularia DC. is a plant species from the Asteraceae family that is endemic to Brazil. Despite the great importance of Baccharis genus, no study has been carried out regarding either the phytochemical composition of B. reticularia or the evaluation of its larvicidal potential. Considering the intrinsic immiscibility of essential oils, this study shows larvicidal nanoemulsions containing the B. reticularia phytochemically characterized essential oil and its main constituent against Aedes aegypti. The major compound found was d-limonene (25.7%). The essential oil inhibited the acetylcholinesterase, one of the main targets of insecticides. The required hydrophile-lipophile balance of both nanoemulsions was 15.0. The mean droplet sizes were around 90.0 nm, and no major alterations were observed after 24 h of preparation for both formulations. After 48 h of treatment, the estimated LC50 values were 118.94 μg mL−1 and 81.19 μg mL−1 for B. reticularia essential oil and d-limonene nanoemulsions, respectively. Morphological alterations evidenced by scanning electron micrography were observed on the larvae treated with the d-limonene nanoemulsion. This paper demonstrated a simple and ecofriendly method for obtaining B. reticularia essential oil and d-limonene aqueous nanoemulsions by a non-heating and solvent-free method, as promising alternatives for Aedes aegypti control.

Purpose Pancreatic cancer is the fourth most common cause of cancer-related death in the USA. This is mainly because of the chemoresistance of this type of tumor; thus, the development of novel therapeutic modalities is needed. Methods Long-circulating and pH-sensitive liposomes containing cisplatin (SpHL-CDDP) were administered systemically into pancreatic tumor-bearing mice for a period of 14 days. The antitumor efficacy and toxicity of this new treatment method on the basis of cisplatin-loaded liposomes was compared with the classical free-CDDP method. Tc-HYNIC-βAla-bombesin(7-14) tumor uptake and histopathologic findings were used to monitor and compare the two treatment modalities. Results The antitumor activity of SpHL-CDDP treatment was shown by (a) decrease in tumor volume, (b) development of tumor necrotic areas, and (c) decrease in Tc-HYNIC-βAla-bombesin(7-14) tumor uptake. Toxicity was evaluated by the development of inflammation and necrotic areas in the kidneys, liver, spleen, and intestine: toxic effects were greater with free-CDDP than SpHL-CDDP. Conclusion SpHL-CDDP showed significant antitumor activity in pancreatic cancer-bearing mice, with lower toxicity in comparison with free-CDDP.

In our previous results, we showed that wheat gluten (WG) intake increases weight gain and adiposity through reduction in oxidation of fatty acids and increase in lipid stores. However, it is not c...

Nanocapsules (NCs) of poly-e-caprolactone (PCL NC) and poly-e-caprolactone coated with chitosan (CS-PCL NC) loaded with technetium-99m methoxyisobutylisonitrile (99mTc-MIBI) were employed as a model to understand the role of these nanocarriers in the veterinary area. 99mTc-MIBI-NCs and 99mTc-MIBI were administrated by intramammary via in rats. The level of radioactivity in the blood was lower when NCs were employed in all times investigated. The results obtained suggest that NCs could be an alternative method for treatment of intramammary diseases in animals, once the data showed high retention in this tissue, increasing the drug residence time on the target.

In this paper, comparative molecular studies between authentic Saccharomyces cerevisiae strains, related species, and the strain described as Saccharomyces boulardii were performed. The response of a S. boulardii strain and a S. cerevisiae strain (W303) to different stress conditions was also evaluated. The results obtained in this study show that S. boulardii is genetically very close or nearly identical to S. cerevisiae. Metabolically and physiologically, however, it shows a very different behavior, particularly in relation to growth yield and resistance to temperature and acidic stresses, which are important characteristics for a microorganism to be used as a probiotic.Key words: Saccharomyces boulardii, probiotic, stress response, rDNA.

Pancreatic adenocarcinoma is important in oncology because of its high mortality rate. Deaths may be avoided if an early diagnosis could be achieved. Several types of tumors overexpress gastrin-releasing peptide receptors (GRPr), including pancreatic cancer cells. Thus, a radiolabeled peptide derivative of gastrin-releasing peptide (GRP) may be useful as a specific imaging probe. The purpose of the present study was to evaluate the feasibility of using (99m)Tc-HYNIC-βAla-Bombesin(7-14)as an imaging probe for Capan-1 pancreatic adenocarcinoma. Xenographic pancreatic tumor was developed in nude mice and characterized by histopathological analysis. Biodistribution studies and scintigraphic images were carried out in tumor-bearing nude mice. The two methods showed higher uptake by pancreatic tumor when compared to muscle (used as control), and the tumor-to-muscle ratio indicated that (99m)Tc-HYNIC-βAla-Bombesin (7-14)uptake was four-fold higher in tumor cells than in other tissues. Scintigraphic images also showed a clear signal at the tumor site. The present data indicate that (99m)Tc-HYNIC-βAla-Bombesin (7-14) may be useful for the detection of pancreatic adenocarcinoma.

Current diagnostic methods and imaging techniques are not able to differentiate septic and aseptic inflammation. Thus, reliable methods are sought to provide this distinction and scintigraphic imaging is an interesting option, since it is based on physiological changes. In this context, radiolabeled antimicrobial peptides have been investigated as they accumulate in infectious sites instead of aseptic inflammation. The peptide LyeTx I, from the venom of Lycosa erythrognatha, has potent antimicrobial activity. Therefore, this study aimed to synthesize LyeTx I derivatives with the chelating compound HYNIC, to evaluate their antimicrobial activity and to radiolabel them with 99mTc. Two LyeTx I derivatives, HYNIC-LyeTx I (N-terminal modification) and LyeTx I-K-HYNIC (C-terminal modification), were synthesized by Fmoc strategy and purified by RP-HPLC. The purified products were assessed by RP-HPLC and MALDI-ToF-MS analysis. Microbiological assays were performed against S. aureus (ATCC® 6538) and E. coli (ATCC® 10536) in liquid medium to calculate the MIC. The radiolabeling procedure of LyeTx I-K-HYNIC with 99mTc was performed in the presence of co-ligands (tricine and EDDA) and reducing agent (SnCl2 . 2H2O), and standardized taking into account the amount of peptide, reducing agent, pH and heating. Radiochemical purity analysis was performed by thin-layer chromatography on silica gel strips and the radiolabeled compound was assessed by RP-HPLC and radioactivity measurement of the collected fractions. Data were analyzed by ANOVA, followed by Tukey test (p-values

CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior A clindamicina, antimicrobiano pertencente ao grupo farmacológico das lincosamidas, além de sua atividade antimicrobiana, exibe atividades em modelos de dor aguda e crônica e inflamação, indicando que os efeitos imunomoduladores podem contribuir para um potencial farmacológico mais amplo. No presente estudo, investigamos os efeitos da clindamicina e de um derivado acetilado da clindamicina (DAC) em modelos experimentais de inflamação articular aguda e em um ensaio microbiológico. A inflamação das articulações foi induzida por injeção intra-articular (i.a.) de zimosano (50 μg) ou cristais de urato monossódico (UMS; 100 μg) em camundongos. Clindamicina ou DAC foram administrados por via intraperitoneal (i.p.) 1 h antes da injeção de zimosano ou UMS. Além disso, foi avaliado o efeito induzido pelaclindamicina administrada 1 h após a injeção de zimosano. Foram avaliados o limiar de retirada da pata, o diâmetro da articulação, o recrutamento de neutrófilos, a atividade da mieloperoxidase, a produção de citocinas como o fator de necrose tumoral-α (TNF-α), interleucina (IL)-1β, IL-6, CXCL-1 e a fosforilação de IκBα. Um ensaio in vitro foi usado para avaliar os efeitos induzidos pela clindamicina ou DACsobre a produção de TNF-α por macrófagos murinos RAW264.7 estimulados comzimosano. Um ensaio microbiológico in vitro foi usado para avaliar a atividade da clindamicina e DAC. A clindamicina, mas não o DAC, exibiu atividade contra as cepas de Staphylococcus aureus e Salmonella Typhimurium ATCC®. A administração de clindamicina aumentou o limiar nociceptivo e reduziu o edema articular induzido por zimosano. A atividade da clindamicina (400 mg/kg) no modelo de alodínia mecânica induzida por zimosano foi atenuada pela administração prévia de naltrexona, mas não pela glibenclamida ou AM251. A administração de clindamicina (400 mg/kg) ou DAC (436 mg/kg) aumentou o limiar nociceptivo e reduziu o edema articular, o recrutamento de neutrófilos, a atividade da mieloperoxidase, a produção de TNF-α, IL-1β, IL-6, CXCL-1 e a fosforilação de IκBα. A clindamicina e o DAC inibiram a produção in vitro de TNF-α por macrófagos RAW264.7. A injeção i.a de UMS reduziu o limiar nociceptivo e induziu edema articular por 24 h, efeitos que foram reduzidos pela administração i.p. de clindamicina (400 mg/kg). Em resumo, fornecemos uma demonstração adicional dos efeitos imunomoduladores da clindamicina, cujo mecanismo foi associado à redução da produção de citocinas, do recrutamento de neutrófilos e da atividade do NF-κB. Tais efeitos foram estendidos ao DAC, o qual apresentou atividade antimicrobiana reduzida, indicando que os derivados de clindamicina devem ser investigados como candidatos a fármacos que poderiam ser úteis no tratamento de condições inflamatórias e dolorosas. Clindamycin, a lincosamide antimicrobial, in addition to its antimicrobial activity, exhibits activities in models of acute and chronic pain and inflammation, indicating that immunomodulatory effects may contribute to a broader pharmacological potential. In the present study, we investigated the effects of clindamycin and a clindamycin acetylated derivative (CAD) in experimental models of acute joint inflammation and in a microbiological assay. Joint inflammation was induced by intra-articular injection (i.a.) of zymosan (50 μg) or monosodium urate (MSU; 100 μg) in mice. Clindamycin or CAD were administered intraperitoneally (i.p.) 1 h before injection of zymosan or MSU. In addition, the effect of clindamycin administered 1 h after zymosan injection was evaluated. The paw withdrawal threshold, joint diameter, neutrophil recruitment, myeloperoxidase activity, cytokine production such as tumor necrosis factor-α (TNFα), interleukin (IL)-1β, IL-6 and CXCL-1, and the phosphorylation of IκBα were evaluated. An in vitro assay was used to evaluate the effects induced by clindamycin or ACD on TNF-α production by zymosan-stimulated RAW264.7 cells. An in vitro microbiological assay was used to evaluate the activity of clindamycin and CAD. Clindamycin, but not CAD, exhibited activity against Staphylococcus aureus and Salmonella Typhimurium ATCC® strains. Clindamycin administration increased the nociceptive threshold and reduced zymosan-induced joint edema. The activity of clindamycin (400 mg/kg) in the zymosan-induced mechanical allodynia model was attenuated by prior administration of naltrexone, but not by glibenclamide or AM251. Administration of clindamycin (400 mg/kg) or CAD (436 mg/kg) increased the nociceptive threshold and reduced joint edema, neutrophil recruitment, myeloperoxidase activity, TNF-α, IL-1β, IL-6 production, CXCL-1 and the phosphorylation of IκBα. Clindamycin and CAD inhibited the in vitro production of TNFα by RAW264.7 macrophages. The i.a injection of MSU reduced the nociceptive threshold and induced joint edema for 24 h, effects that were reduced by the i.p. administration of clindamycin (400 mg/kg). In summary, we provide additional demonstration of the immunomodulatory effects of clindamycin. These effects may be associated with reduced cytokine production, neutrophil recruitment and NF-κB activity. Such effects were extended to CAD, which exhibited reduced antimicrobial activity. These results indicate that clindamycin derivatives should be investigated as drug candidates that could be useful in the treatment of inflammatory and painful conditions.

A administração continuada de cisplatina (CDDP), quimioterápico usado no tratamento de vários tipos de tumores sólidos, é limitada em função de reações adversas, como nefrotoxicidade, hepatotoxicidade, neurotoxicidade, mielotoxicidade e ototoxicidade. Na tentativa de contornar esses inconvenientes e melhorar o índice terapêutico da CDDP, modificações farmacotécnicas têm sido propostas, como a encapsulação da CDDP em sistemas de liberação de fármacos, tais como os lipossomas. Nesse sentido, foi proposto o desenvolvimento de lipossomas termossensíveis de cisplatina não funcionalizado (LTS-CDDP), funcionalizado com ácido hialurônico (AH) por interação eletrostática (LTS-CDDP-AH-1) e funcionalizado com AH por ligação covalente (LTS-CDDP-AH-2), no intuito de proporcionar um direcionamento ativo para a região tumoral. Inicialmente foi realizado um estudo de caracterização das formulações desenvolvidas, por meio da associação das técnicas espalhamento dinâmico da luz (DLS), microcalorimetria e espalhamento de raio-X a baixo ângulo (SAXS). A taxa de encapsulação de CDDP foi avaliada por cromatografia líquida de alta eficiência por um método de derivatização com dietilditiocarbamato. Em seguida, foi avaliada a citotoxicidade frente as células tumorais MDA-MB-231, bem como a toxicidade in vivo em camundongos Swiss sadios. Os resultados de caracterização mostraram diâmetro médio menor que 200 nm e índice de polidispersão (PDI) sugestivos de formulações homogêneas (< 0,2), indicando a ausência de agregação de vesículas mesmo após funcionalização com AH. A eficiência do revestimento na superfície do lipossoma foi atribuída a valores de potencial zeta próximos da neutralidade. Os dados de DLS mostraram uma redução significativa no diâmetro médio e contagem de partículas (Kcps) das formulações avaliadas a 40oC, e o revestimento com AH não alterou a temperatura de transição de fase das formulações. A encapsulação de CDDP foi de 9,3 ± 0,7% para LTS-CDDP-AH-2, 16,1 ± 1,9% para LTS-CDDP-AH-1 e 13,7 ± 0,9% para LTS-CDDP. O perfil obtido por SAXS para todas as formulações estudadas foi característico de uma organização lamelar independente da temperatura avaliada e apresentou dilatação da bicamada, causada por desorganização na estrutura dos lipídeos, confirmando a alteração conformacional devido ao aquecimento. No entanto, o perfil de liberação in vitro sugeriu uma possível adsorção do fármaco na bicamada fosfolipídica ocasionando uma taxa de difusão de CDDP antes de atingir a Tc (42oC), para LTS-CDDP e LTS-CDDP-AH-2. Para o LTS-CDDP-AH-1, a presença do polímero pode estar modulando essa difusão, gerando um perfil de liberação mais controlado e lento. Os resultados biológicos mostraram que LTS-CDDP apresentou maior citotoxicidade quando comparado com CDDP livre, além de levar ao aumento da ocorrência de senescência. O uso de hipertermia nos grupos tratados com LTS-CDDP e LTS-CDDP-AH-1 aumentou a ocorrência de apoptose em relação aos grupos sem hipertermia. No estudo in vivo, foi observado maior toxicidade da formulação catiônica para células saudáveis, sendo observado a ocorrência de nefrotoxicidade nos animais tratados com LTS-CDDP. Enquanto que, com o uso do ligante AH, os efeitos adversos conhecidos relacionados ao uso de CDDP foram eliminados, uma vez que os animais tratados com essa formulação não apresentaram alterações significativas nos exames hematológicos e bioquímicos, bem como nas análises histológicas. Sendo assim, a formulação de CDDP proposta, termossensível e contendo AH por interação eletrostática, apresenta características adequadas para tratamento do câncer por via intravenosa, em associação com hipertermia moderada, podendo ser capaz de reduzir a toxicidade relacionada a CDDP, representando uma alternativa potencial para veiculação desse fármaco. The continuous administration of cisplatin (CDDP), a chemotherapeutic agent used to treat various types of solid tumors, is limited due to adverse reactions, such as nephrotoxicity, hepatotoxicity, neurotoxicity, myelotoxicity, and ototoxicity. In an attempt to circumvent these inconveniences and improve the therapeutic index of CDDP, pharmacotechnical modifications have been proposed, such as the encapsulation of CDDP in drug delivery systems, like liposomes. In this sense, it was proposed to develop a thermosensitive liposome of nonfunctionalized cisplatin (TSL-CDDP), functionalized with hyaluronic acid (HA) by electrostatic interaction (TSL-CDDP-HA-1) and functionalized with HA by covalent bond (TSL-CDDP-SA-HA-2), in order to provide an active targeting for the tumor region. Initially, a detailed study was carried out to characterize the formulations developed, through the association of dynamic light scattering (DLS), microcalorimetry, and low angle X-ray scattering (SAXS) techniques. The CDDP encapsulation rate was evaluated by high performance liquid chromatography using a diethyldithiocarbamate derivatization method. Then, cytotoxicity against MDA-MB-231 tumor cells was evaluated, as well as in vivo toxicity in healthy Swiss mice. The characterization results showed an average diameter less than 200 nm and a polydispersity index (PDI) suggestive of homogeneous formulations (< 0.2), indicating the absence of vesicle aggregation even after functionalization with HA. The efficiency of the HA-coating of liposome was attributed to values of zeta potential close to neutrality. The DLS data showed a significant reduction in the mean diameter and Kcps (Kilo counts per second) of the formulations evaluated at 40oC, and the coating with HA did not alter the phase transition temperature of the formulations. CDDP encapsulation was 9.3 ± 0.7% for TSL-CDDP-SA-HA, 16.1 ± 1.9% for TSL-CDDP-HA and 13.7 ± 0.9% for TSL -CDDP. The profile obtained by SAXS for all the formulations studied was characteristic of a lamellar organization independent of the evaluated temperature and presented dilation of the bilayer, caused by disorganization in the lipid structure, confirming the conformational alteration due to heating. However, the in vitro release profile suggested possible drug adsorption in the phospholipid bilayer causing a diffusion rate of CDDP before reaching Tc (42oC), for TSLCDDP and TSL-CDDP-SA-HA. For TSL-CDDP-HA, the presence of the polymer may be modulating this diffusion, generating a more controlled and slower release profile. The biological results showed that TSL-CDDP have greater cytotoxicity when compared to free CDDP, in addition to leading to an increase in the occurrence of senescence. The use of hyperthermia in the groups treated with LTS-CDDP and LTS-CDDP-HA increased the occurrence of apoptosis compared to groups without hyperthermia. In the in vivo study, greater toxicity of the cationic formulation was observed for healthy cells, with the occurrence of nephrotoxicity in animals treated with LTS-CDDP. While, with the use of the HA ligand, the known adverse effects related to the use of CDDP were eliminated, since the animals treated with this formulation did not show significant changes in hematological and biochemical tests, as well as in histological analyzes. Thus, the proposed formulation of CDDP, thermosensitive and containing HA by electrostatic interaction, has adequate characteristics for cancer treatment intravenously, in association with moderate hyperthermia, and may be able to reduce CDDPrelated toxicity, representing a potential alternative for the delivery of this drug.