RESUMO
OBJECTIVE: To determine the effects of prolonged administration of the oral NSAIDs phenylbutazone and firocoxib on concentrations of cytokines and growth factors in platelet-rich plasma (PRP) and autologous protein solution (APS). ANIMALS: 6 adult University owned horses. METHODS: Horses were randomized to receive phenylbutazone (1 g, orally, q 12 h) or firocoxib (57 mg, orally, q 24 h) for 6 days. Blood was obtained and processed for APS (Pro-Stride) and PRP (Restigen) before the administration of NSAIDs and at 7 days (1 day following cessation of NSAIDs). Horses underwent a two-week washout period, during which blood was obtained at 14 days and 21 days. The protocol was repeated with a crossover design. PRP and APS were analyzed for concentrations of platelets, leukocytes, and several cytokines (IL-1ß, IL-10, IL-6, IL-8, and tumor necrosis factor-α) and growth factors (PDGF, FGF-2, and TGF-ß1) using immunoassays. Plasma was evaluated for drug concentrations. RESULTS: No significant differences existed in concentrations of growth factors and cytokines before or after prolonged administration of NSAIDs. There were significant differences in concentrations of leukocytes and platelets in PRP compared to APS, with higher concentrations of leukocytes at the day 7 time point (T) in APS (phenylbutazone) and in concentrations of platelets in APS at T0 (firocoxib) and in APS at T7 (phenylbutazone). CLINICAL RELEVANCE: Veterinarians can recommend the administration of these oral NSAIDs prior to obtaining blood for PRP and APS provided a single-day washout period is instituted.
Assuntos
4-Butirolactona , Anti-Inflamatórios não Esteroides , Citocinas , Peptídeos e Proteínas de Sinalização Intercelular , Fenilbutazona , Plasma Rico em Plaquetas , Sulfonas , Animais , Cavalos/sangue , 4-Butirolactona/análogos & derivados , 4-Butirolactona/administração & dosagem , 4-Butirolactona/farmacologia , Citocinas/sangue , Sulfonas/administração & dosagem , Sulfonas/farmacologia , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/farmacologia , Fenilbutazona/administração & dosagem , Fenilbutazona/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Administração Oral , Masculino , Estudos Cross-Over , FemininoRESUMO
Acyl-homoserine lactones (AHLs) are quorum-sensing signaling molecules in Gram-negative bacteria and positively regulate biofilm formation in Salmonella under specific conditions. In this study, biofilm formation in Salmonella enterica was evaluated at 28 and 37 °C, under aerobic and anaerobic conditions. Additionally, the influence of the N-dodecanoyl-DL-homoserine lactone (C12-HSL) on biofilm formation and the expression of genes related to the synthesis of structural components, regulation, and quorum sensing was assessed under anaerobiosis at 28 and 37 °C. Biofilm formation was found not to be influenced by the atmospheric conditions at 28 °C. However, it was reduced at 37 °C under anaerobiosis. C12-HSL enhanced biofilm formation at 37 °C under anaerobiosis and increased the expression of the adrA and luxS genes, suggesting an increase in c-di-GMP, a second messenger that controls essential physiological functions in bacteria. These results provide new insights into the regulation of biofilm formation in Salmonella under anaerobic conditions.
Assuntos
Percepção de Quorum , Salmonella enteritidis , Percepção de Quorum/genética , Salmonella enteritidis/genética , Biofilmes , Anaerobiose , 4-Butirolactona/farmacologia , 4-Butirolactona/metabolismo , Acil-ButirolactonasRESUMO
Bacterial quorum sensing (QS) and biofilm formation are promising targets for developing new therapies to treat chronic infections. Herein, we report the stereoselective synthesis of 18 new analogs of natural cadiolides. Among the new compounds, substances 8b, 8f, 8i, 9a, 9b and 9e completely inhibited the biofilm formation of Escherichia coli RP347 in vitro. In addition, compound 8b interfered acyl-homoserine lactone (AHL) mediated QS, while 9e interrupted the QS via autoinducer-2 (AI-2). Biological assays also revealed that synthetic intermediates alkynones are potent inhibitors of AI-2 and AHL-mediated QS. These results indicate that cadiolides and alkynones are good candidates for further structural modification for a new generation of more potent antimicrobial agents.
Assuntos
4-Butirolactona/farmacologia , Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , 4-Butirolactona/síntese química , Antibacterianos/síntese química , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Testes de Sensibilidade Microbiana , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/fisiologia , EstereoisomerismoRESUMO
Neurodegenerative disorders, including Tauopathies that involve tau protein, base their pathological mechanism on forming proteinaceous aggregates, which has a deleterious effect on cells triggering an inflammatory response. Moreover, tau inhibitors can exert their mechanism of action through noncovalent and covalent interactions. Thus, Michael's addition appears as a feasible type of interaction involving an α, ß unsaturated carbonyl moiety to avoid pathological confirmation and further cytotoxicity. Moreover, we isolated three compounds from Antarctic lichens Cladonia cariosa and Himantormia lugubris: protolichesterinic acid (1), fumarprotocetraric acid (2), and lichesterinic acid (3). The maleimide cysteine labeling assay showed that compounds 1, 2, and 3 inhibit at 50 µM, but compounds 2 and 3 are statistically significant. Based on its inhibition capacity, we decided to test compound 2 further. Thus, our results suggest that compound 2 remodel soluble oligomers and diminish ß sheet content, as demonstrated through ThT experiments. Hence, we added externally treated oligomers with compound 2 to demonstrate that they are harmless in cell culture. First, the morphology of cells in the presence of aggregates does not suffer evident changes compared to the control. Additionally, the externally added aggregates do not provoke a substantial LDH release compared to the control, indicating that treated oligomers do not provoke membrane damage in cell culture compared with aggregates alone. Thus, in the present work, we demonstrated that Michael's acceptors found in lichens could serve as a scaffold to explore different mechanisms of action to turn tau aggregates into harmless species.
Assuntos
Fumaratos/farmacologia , Agregados Proteicos/efeitos dos fármacos , Proteínas tau/antagonistas & inibidores , Proteínas tau/metabolismo , 4-Butirolactona/análogos & derivados , 4-Butirolactona/farmacologia , Regiões Antárticas , Ascomicetos/metabolismo , Linhagem Celular Tumoral , Humanos , Líquens/metabolismo , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Parmeliaceae/metabolismo , Tauopatias/tratamento farmacológico , Tauopatias/metabolismoRESUMO
This study evaluated the effects of oocyte meiosis inhibitors roscovitine (ROS) and butyrolactone I (BL-I) on in vitro production of bovine embryos. Bovine oocytes were maintained in pre in vitro maturation (pre-IVM) with 25 µM ROS or 100 µM BL-I for 24 h to delay meiosis and for 24 h in in vitro maturation (IVM). Following this treatment, the nuclear maturation index was evaluated. All embryos degenerated following this procedure. In the second set of experiments, oocytes were maintained for 6 or 12 h in pre-IVM with the following three treatments: ROS (25 µM or 12.5 µM), BL-I (100 µM or 50 µM) or a combination of both drugs (6.25 µM ROS and 12.5 µM BL-I). Oocytes were cultivated for 18 or 12 h in IVM. When a meiosis-inducing agent was used during pre-IVM for 24 h, more degenerated oocytes were observed at the end of the IVM period. This effect decreased when the meiotic blocking period was reduced to 6 or 12 h. No significant differences were observed in the blastocyst production rate of oocytes in pre-IVM for 6 h with ROS, BL-I, or ROS + BL-I compared with that of the control group (P > 0.05). However, inhibition of oocytes for 12 h resulted in decreased embryo production compared with that in the controls (P < 0.05). There was no difference in the post-vitrification embryo re-expansion rate between the study groups, showing that the meiotic inhibition for 6 or 12 h did not alter the embryo cryopreservation process.
Assuntos
4-Butirolactona/análogos & derivados , Blastocisto/citologia , Embrião de Mamíferos/citologia , Fertilização in vitro/efeitos dos fármacos , Técnicas de Maturação in Vitro de Oócitos , Meiose , Oócitos/citologia , Roscovitina/farmacologia , 4-Butirolactona/farmacologia , Animais , Blastocisto/efeitos dos fármacos , Bovinos , Embrião de Mamíferos/efeitos dos fármacos , Feminino , Oócitos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologiaRESUMO
This study describes the in vitro anthelmintic activity of a hydroalcoholic extract from the fruit of Piper cubeba and its major isolated components against the eggs and larvae of gastrointestinal nematodes obtained from naturally-infected ovines. In vitro anthelmintic activity was evaluated using the egg hatch test (EHT), larval development test (LDT) and L3 migration inhibition test (LMT). The extract showed ovicidal and larvicidal activity, with an EC50 of 200⯵g/mL and 83.00⯵g/mL in the EHT and LDT, respectively. The extract inhibited 100% of larval migration at the lowest tested concentration (95⯵g/mL). The crude extract was purified using successive silica gel chromatographic columns, which revealed the lignans hinokinin, cubebin and dihydrocubebin as the major compounds that were present, which were then used in in vitro tests. Cubebin, dihydrocubebin and hinokinin showed higher activity than the crude extract, with an EC50 for ovicidal activity of 150.00⯵g/mL, 186.70⯵g/mL and 68.38⯵g/mL, respectively. In the LDT, cubebin presented an EC50 of 14.89⯵g/mL and dihydrocubebin of 30.75⯵g/mL. Hinokinin inhibited 100% the larval development at all concentrations evaluated. In the LMT, dihydrocubebin inhibited 100% the larval migration in all concentrations evaluated while cubebin and hinokinin showed EC50 values of 0.89⯵g/mL and 0.34⯵g/mL, respectively. P. cubeba extract is rich in several classes of active compounds, but here we demonstrate that the described anthelmintic activity may be related to the presence of these lignans, which are present in larger concentrations than other components of the extract. Our results demonstrate for first time the anthelmintic activity against gastrointestinal nematodes in sheep for this class of special metabolites that are present in P. cubeba fruit. However, future detailed studies are needed to evaluate the effectiveness of P. cubeba fruits extract and active lignans in in vivo tests.
Assuntos
Enteropatias Parasitárias/veterinária , Lignanas/farmacologia , Nematoides/efeitos dos fármacos , Infecções por Nematoides/veterinária , Piper/química , Extratos Vegetais/farmacologia , 4-Butirolactona/análogos & derivados , 4-Butirolactona/química , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , Animais , Benzodioxóis/química , Benzodioxóis/isolamento & purificação , Benzodioxóis/farmacologia , Cromatografia em Gel/veterinária , Dioxolanos/química , Dioxolanos/isolamento & purificação , Dioxolanos/farmacologia , Fezes/parasitologia , Frutas/química , Enteropatias Parasitárias/tratamento farmacológico , Enteropatias Parasitárias/parasitologia , Larva/efeitos dos fármacos , Larva/crescimento & desenvolvimento , Larva/fisiologia , Lignanas/química , Lignanas/isolamento & purificação , Microscopia Eletrônica de Varredura/veterinária , Nematoides/crescimento & desenvolvimento , Nematoides/fisiologia , Infecções por Nematoides/tratamento farmacológico , Infecções por Nematoides/parasitologia , Óvulo/efeitos dos fármacos , Óvulo/fisiologia , Extratos Vegetais/química , Ovinos , Doenças dos Ovinos/parasitologiaRESUMO
Mitosis has been traditionally considered a metabolically inactive phase. We have previously shown, however, that extensive alterations in lipids occur as the cells traverse mitosis, including increased de novo fatty acid (FA) and phosphatidylcholine (PtdCho) synthesis and decreased lysophospholipid content. Given the diverse structural and functional properties of these lipids, we sought to study their metabolic fate and their importance for cell cycle completion. Here we show that FA and PtdCho synthesized at the mitotic exit are destined to the nuclear envelope. Importantly, FA and PtdCho synthesis, but not the decrease in lysophospholipid content, are necessary for cell cycle completion beyond G2/M. Moreover, the presence of alternative pathways for PtdCho synthesis renders the cells less sensitive to its inhibition than to the impairment of FA synthesis. FA synthesis, thus, represents a cell cycle-related metabolic vulnerability that could be exploited for combined chemotherapy. We explored the combination of fatty acid synthase (FASN) inhibition with agents that act at different phases of the cell cycle. Our results show that the effect of FASN inhibition may be enhanced under some drug combinations.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Ácido Graxo Sintases/antagonistas & inibidores , Ácidos Graxos/biossíntese , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Lipogênese/efeitos dos fármacos , Mitose/efeitos dos fármacos , Membrana Nuclear/metabolismo , Fosfatidilcolinas/biossíntese , 4-Butirolactona/análogos & derivados , 4-Butirolactona/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Etoposídeo/farmacologia , Ácido Graxo Sintases/metabolismo , Células HeLa , Humanos , Lipogênese/fisiologia , Lisofosfolipídeos/biossíntese , Lisofosfolipídeos/química , Mitose/fisiologia , Membrana Nuclear/efeitos dos fármacos , Membrana Nuclear/enzimologiaRESUMO
BACKGROUND: From a previous screening of Brazilian biodiversity for antitrypanosomal activity, the n-hexane extract from twigs of Nectandra oppositifolia (Lauraceae) demonstrated in vitro activity against Trypanosoma cruzi. PURPOSE: To perform the isolation and chemical characterization of bioactive compounds from n-hexane extract from twigs of N. oppositifolia and evaluate their therapeutical potential as well as to elucidate their mechanism of action against T. cruzi. METHODS/STUDY DESIGN: Bioactivity-guided fractionation of the n-hexane extract from twigs of N. oppositifolia afforded three related butenolides: isolinderanolide D (1), isolinderanolide E (2) and secosubamolide A (3). These compounds were evaluated in vitro against T. cruzi (trypomastigote and amastigote forms) and against NCTC (L929) cells for mammalian cytotoxicity. Additionally, phenotypic analyzes of compounds-treated parasites were performed: alterations in the plasma membrane permeability, plasma membrane electric potential (ΔΨp), mitochondrial membrane potential (ΔΨm) and induction of ROS. RESULTS: Compounds 1-3 were effective against T. cruzi, with IC50 values of 12.9, 29.9 and 12.5 µM for trypomastigotes and 25.3, 10.1 and 12.3 µM for intracellular amastigotes. Furthermore, it was observed alteration in the mitochondrial membrane potential (ΔΨm) of parasites treated with butenolides 1-3. These compounds caused no alteration to the parasite plasma membrane, and the deregulation of the mitochondria might be an early event to cell death. In addition, in silico studies showed that all butenolides were predicted to be non-mutagenic, non-carcinogenic, non hERG blockers, with acceptable human intestinal absorption, low inhibitory promiscuity with the main five CYP isoforms, and with high metabolic stability. Otherwise, tested butenolides showed unfavorable blood-brain barrier penetration (BBB+). CONCLUSION: Our results demonstrated the anti-T. cruzi effects of compounds 1-3 isolated from N. oppositifolia and indicated that the lethal effect of these compounds in trypomastigotes of T. cruzi could be associated to the alteration in the mitochondrial membrane potential (ΔΨm).
Assuntos
4-Butirolactona/análogos & derivados , Lauraceae/química , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , 4-Butirolactona/uso terapêutico , Animais , Brasil , Membrana Celular/efeitos dos fármacos , Doença de Chagas/tratamento farmacológico , HumanosRESUMO
OBJECTIVES: Epiisopiloturine (EPI) and epiisopilosine (EPIIS) are side products in the pharmaceutical industry. The present study aimed to investigate the anti-inflammatory potential of the alkaloids EPI and EPIIS in human neutrophils and mechanical hyperalgesia in mice. METHODS: Neutrophils (5 × 106 cells/ml) incubated with EPI and EPIIS and stimulated by the addition of N-formyl-methionyl-leucyl-phenylalanine or phorbol 12-myristate-13-acetate. The release of myeloperoxidase (MPO), reactive oxygen species (ROS) production, calcium influx, gene expression of NF-κB and pro-inflammatory cytokines production were evaluated. It was also investigated the effect these alkaloids on carrageenan-induced mechanical hyperalgesia model in mice. KEY FINDINGS: We demonstrated that both EPI and EPIIS inhibited the degranulation of activated neutrophils. This effect was accompanied by the reduction in ROS, the prevention of the increase in intracellular Ca2+ and decrease in the density of cytosolic NF-κB, and inhibition of TNF-α and IL-6 production. Evaluating hypernociception in mice, EPI and EPIIS inhibited carrageenan-induced inflammatory hypernociception and reduced MPO levels. CONCLUSIONS: The results obtained suggest EPI and EPIIS not only inhibit neutrophils functions in vitro, but also exhibits anti-inflammatory properties in vivo, acting through the modulation of the activation and/or accumulation of neutrophils in the inflammatory focus. Thus, EPI and EPIIS possess promising anti-inflammatory therapeutic potential.
Assuntos
4-Butirolactona/análogos & derivados , Alcaloides/farmacologia , Anti-Inflamatórios/farmacologia , Imidazóis/farmacologia , Neutrófilos/efeitos dos fármacos , 4-Butirolactona/farmacologia , Animais , Cálcio/metabolismo , Humanos , Hiperalgesia/metabolismo , Interleucina-6/metabolismo , Masculino , Camundongos , N-Formilmetionina Leucil-Fenilalanina , NF-kappa B/metabolismo , Neutrófilos/metabolismo , Peroxidase/efeitos dos fármacos , Peroxidase/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Acetato de Tetradecanoilforbol , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Embryo mobility occurs as a result of prostaglandin production by the embryo and endometrium, promoting uterine smooth muscle contractions, which propels the embryonic vesicle through the lumen. Non-steroidal anti-inflammatory drugs (NSAIDs), as flunixin meglumine, are routinely used in equine medicine and can alter the conceptus mobility if applied in early pregnancy, which may impair maternal recognition of pregnancy. The objective of this study was to evaluate and compare the effect of flunixin meglumine (FM; 1.1â¯mg/kg IV), firocoxib (FIRO; 0.2â¯mg/kg PO), and meloxicam (ML; 0.6â¯mg/kg, IV), on the embryo mobility. Thirty mares were divided into three groups (nâ¯=â¯10 per treatment). After the pregnancy diagnosis on day 12 after ovulation, the embryo mobility was evaluated by transrectal ultrasonography every 5â¯min for 1â¯h in order to visualize the location of the embryo. In all mares, three evaluations were performed: immediately before treatment (pre-treatment), after NSAID administration and 24â¯h after treatment. In group FM, embryo mobility decreased, from 5.8⯱â¯0.3 movements/hour (m/h) to 2.3⯱â¯0.5â¯m/h (pâ¯<â¯0.05) and, after 24â¯h the values were similar to the pre-treatment evaluation (5.9⯱â¯0.2â¯m/h). Likewise, ML treatment caused a decrease of embryo movements, from 5.9⯱â¯0.3 to 1.9⯱â¯0.3â¯m/h (pâ¯<â¯0.05), 24â¯h after treatment values were 5.7⯱â¯0.4â¯m/h. Treatment with FIRO did not interfere with embryo mobility (5.7⯱â¯0.4; 5.8⯱â¯0.3 and 5.6⯱â¯0.3 embryo movements in the first, second and third evaluation, respectively). In conclusion, FIRO was the only NSAID that did not alter the embryo mobility and may be the safest NSAID for use in early pregnant mares.