RESUMO
Upregulation of hydrogen sulfide (H2S) biosynthesis, at least in part related to the upregulation of cystathionine ß-synthetase (CBS) in cancer cells, serves as a tumor-promoting factor and has emerged as a possible molecular target for antitumor drug development. To facilitate future clinical translation, we have synthesized a variety of novel CBS-targeting, esterase-cleavable prodrugs based on the structure of the prototypical CBS inhibitor aminooxyacetic acid (AOAA). The pharmacological properties of these compounds were evaluated in cell-free assays with recombinant human CBS protein, the human colon cancer cell line HCT116, and in vivo using various tumor-bearing mice models. The prodrug YD0251 (the isopropyl ester derivative of AOAA) was selected for detailed characterization. YD0251 exhibits improved antiproliferative efficacy in cell culture models when compared to AOAA. It is up to 18 times more potent than AOAA at suppressing HCT116 tumor growth in vivo and is effective when administered to tumor-bearing mice either via subcutaneous injection or oral gavage. Patient-derived xenografts (PDTXs) with higher levels of CBS protein grew significantly larger than tumors with lower levels, and YD0251 treatment inhibited the growth of PDTXs with elevated CBS, whereas it had no significant effect on PDTXs with low CBS protein levels. The toxicity of YD0251 was assessed in mice subjected to subchronic administration of supratherapeutic doses the inhibitor; no significant alteration in circulating markers of organ injury or histopathological alterations were noted, up to 60 mg/kg/day × 5 days. In preparation to a future theranostic concept (to match CBS inhibitor therapy to high-CBS expressors), we identified a potential plasma marker of CBS-expressing tumors. Colon cancer cells produced significant levels of lanthionine, a rare metabolic intermediate of CBS-mediated H2S biosynthesis; forced expression of CBS into non-transformed epithelial cells increased lanthionine biogenesis in vitro and in vivo (measured in the urine of tumor-bearing mice). These current results may be useful to facilitate the translation of a CBS inhibition-based antitumor concept into the clinical space.
Assuntos
Ácido Amino-Oxiacético/farmacologia , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Cistationina beta-Sintase/antagonistas & inibidores , Pró-Fármacos/farmacologia , Animais , Células HCT116 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos NusRESUMO
Camel Urine (CU) is composed of components that have antitumor properties and other therapeutic benefits. Regardless of short-term preliminary CU genotoxicity is reported, comprehensive genotoxic studies are limited. In this study, sensitive in vitro and in vivo genotoxic bioassays such as mitotic index (MI), chromosomal aberrations (CA), micronucleated polychromatic erythrocytes (MPE), and analysis of primary spermatocytes were employed. The adventitious roots of Allium cepa L. and mice (Mus musculus), as an experimental mammalian system, were employed to assess the MI and CA of CU induced by sodium nitrate and cyclophosphamide respectively. In contrast, other clastogenic assays were studied in mice (Mus musculus). Twenty-eight days of four repeated doses (2.5, 5, 25, and 50 mL/kg BW) of CU were tested and compared with three doses (10, 25, and 50 mg/kg BW) cyclophosphamide as a positive control and deionized water as the negative control. The results proved that cytological examination of CU was cytotoxic since a decrease in mitotic activity (16.8-1.1) was observed, since the significant reduction in cell proliferation in A. cepa L. and also in mice bone marrow cells. On the other hand, CU did not induce a clastogenic effect since no significant stickiness, fragment, multinucleoli were observed compared to the control group. Additionally, the data showed that CU decreased the CA when mice had received cyclophosphamide (25 mg BW) followed by CU doses. CU was found to be cytotoxic but no clastogenic effect. Furthermore, it possesses anticlastogenic properties. The observed results suggest that CU in whole or the metabolites present in CU could be a potent drug target. Further research is warranted to study the complete metabolites profiling and to study the molecular mechanisms.
Assuntos
Mutagênicos/toxicidade , Urina , Animais , Antineoplásicos/farmacologia , Células da Medula Óssea/efeitos dos fármacos , Camelus , Ciclofosfamida/farmacologia , Sistemas de Liberação de Medicamentos , CamundongosRESUMO
OBJECTIVE: Activation of the constitutive nuclear and mitochondrial enzyme poly (ADP-ribose) polymerase (PARP) has been implicated in the pathogenesis of cell dysfunction, inflammation, and organ failure in various forms of critical illness. The objective of our study was to evaluate the efficacy and safety of the clinically approved PARP inhibitor olaparib in an experimental model of pancreatitis in vivo and in a pancreatic cell line subjected to oxidative stress in vitro. The preclinical studies were complemented with analysis of clinical samples to detect PARP activation in pancreatitis. METHODS: Mice were subjected to cerulein-induced pancreatitis; circulating mediators and circulating organ injury markers; pancreatic myeloperoxidase and malondialdehyde levels were measured and histology of the pancreas was assessed. In human pancreatic duct epithelial cells (HPDE) subjected to oxidative stress, PARP activation was measured by PAR Western blotting and cell viability and DNA integrity were quantified. In clinical samples, PARP activation was assessed by PAR (the enzymatic product of PARP) immunohistochemistry. RESULTS: In male mice subjected to pancreatitis, olaparib (3âmg/kg i.p.) improved pancreatic function: it reduced pancreatic myeloperoxidase and malondialdehyde levels, attenuated the plasma amylase levels, and improved the histological picture of the pancreas. It also attenuated the plasma levels of pro-inflammatory mediators (TNF-α, IL-1ß, IL-2, IL-4, IL-6, IL-12, IP-10, KC) but not MCP-1, RANTES, or the anti-inflammatory cytokine IL-10. Finally, it prevented the slight, but significant increase in plasma blood urea nitrogen level, suggesting improved renal function. The protective effect of olaparib was also confirmed in female mice. In HPDE cells subjected to oxidative stress olaparib (1âµM) inhibited PARP activity, protected against the loss of cell viability, and prevented the loss of cellular NAD levels. Olaparib, at 1µM to 30âµM did not have any adverse effects on DNA integrity. In human pancreatic samples from patients who died of pancreatitis, increased accumulation of PAR was demonstrated. CONCLUSION: Olaparib improves organ function and tempers the hyperinflammatory response in pancreatitis. It also protects against pancreatic cell injury in vitro without adversely affecting DNA integrity. Repurposing and eventual clinical introduction of this clinically approved PARP inhibitor may be warranted for the experimental therapy of pancreatitis.
Assuntos
Pancreatite/tratamento farmacológico , Pancreatite/patologia , Ftalazinas/uso terapêutico , Piperazinas/uso terapêutico , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Animais , Técnicas de Cultura de Células , Linhagem Celular , Ceruletídeo , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Oxidativo , Ductos Pancreáticos/efeitos dos fármacos , Ductos Pancreáticos/patologia , Pancreatite/etiologiaRESUMO
Poly(ADP-ribose) polymerase (PARP) is involved in the pathogenesis of cell dysfunction, inflammation and organ failure during septic shock. The goal of the current study was to investigate the efficacy and safety of the clinically approved PARP inhibitor olaparib in experimental models of oxidative stress in vitro and in sepsis in vivo. In mice subjected to cecal ligation and puncture (CLP) organ injury markers, circulating and splenic immune cell distributions, circulating mediators, DNA integrity and survival was measured. In U937 cells subjected to oxidative stress, cellular bioenergetics, viability and DNA integrity were measured. Olaparib was used to inhibit PARP. The results show that in adult male mice subjected to CLP, olaparib (1-10 mg/kg i.p.) improved multiorgan dysfunction. Olaparib treatment reduced the degree of bacterial CFUs. Olaparib attenuated the increases in the levels of several circulating mediators in the plasma. In the spleen, the number of CD4+ and CD8+ lymphocytes were reduced in response to CLP; this reduction was inhibited by olaparib treatment. Treg but not Th17 lymphocytes increased in response to CLP; these cell populations were reduced in sepsis when the animals received olaparib. The Th17/Treg ratio was lower in CLP-olaparib group than in the CLP control group. Analysis of miRNA expression identified a multitude of changes in spleen and circulating white blood cell miRNA levels after CLP; olaparib treatment selectively modulated these responses. Olaparib extended the survival rate of mice subjected to CLP. In contrast to males, in female mice olaparib did not have significant protective effects in CLP. In aged mice olaparib exerted beneficial effects that were less pronounced than the effects obtained in young adult males. In in vitro experiments in U937 cells subjected to oxidative stress, olaparib (1-100 µM) inhibited PARP activity, protected against the loss of cell viability, preserved NAD+ levels and improved cellular bioenergetics. In none of the in vivo or in vitro experiments did we observe any adverse effects of olaparib on nuclear or mitochondrial DNA integrity. In conclusion, olaparib improves organ function and extends survival in septic shock. Repurposing and eventual clinical introduction of this clinically approved PARP inhibitor may be warranted for the experimental therapy of septic shock.
Assuntos
Anti-Inflamatórios/uso terapêutico , Ftalazinas/uso terapêutico , Piperazinas/uso terapêutico , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Sepse/tratamento farmacológico , Animais , Anti-Inflamatórios/farmacologia , Ceco , Citocinas/sangue , DNA/efeitos dos fármacos , Reposicionamento de Medicamentos , Feminino , Humanos , Ligadura , Fígado/efeitos dos fármacos , Fígado/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Contagem de Linfócitos , Masculino , Camundongos Endogâmicos C57BL , Estresse Oxidativo/efeitos dos fármacos , Ftalazinas/farmacologia , Piperazinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Punções , Sepse/sangue , Sepse/imunologia , Sepse/patologia , Baço/efeitos dos fármacos , Baço/imunologia , Baço/patologia , Células U937RESUMO
This report identifies mitochondrial DNA (mtDNA) as a target and active mediator that links low-level oxidative stress to inflammatory response in pulmonary epithelial cells. Extrusion of mtDNA into the bronchoalveolar lavage fluid occurs as an early event in mice subjected to cigarette smoke injury, concomitantly with the depletion of mtDNA in the lung tissue. In cultured lung epithelial cells, prolonged, low-level oxidative stress damages the mtDNA, without any detectable damage to the nuclear DNA. In turn, cellular depletion of the mtDNA occurs, together with a transient remodeling of cellular bioenergetics and morphology - all without any detectable impairment in overall cell viability. Damaged mtDNA first enters the cytoplasm, where it binds to Z-DNA binding protein 1 (ZBP1) and triggers inflammation via the TANK-binding kinase 1 /interferon regulatory factor 3 signaling pathway. Fragments of the mtDNA are subsequently released into the extracellular space via exosomes. MtDNA-containing exosomes are capable of inducing an inflammatory response in naïve (non-oxidatively stressed) epithelial cells. In vivo, administration of isolated mtDNA into the in lungs of naïve mice induces the production of pro-inflammatory mediators, without histopathologic evidence of tissue injury. We propose that mtDNA-specific damage, and subsequent activation of the ZBP1 pathway, is a mechanism that links prolonged, low-level oxidative stress to autocrine and paracrine inflammation during the early stages of inflammatory lung disease.
Assuntos
Dano ao DNA/genética , DNA Mitocondrial/genética , Células Epiteliais/metabolismo , Glicoproteínas/genética , Inflamação/genética , Mitocôndrias/genética , Estresse Oxidativo/genética , Animais , Linhagem Celular , Proteínas de Ligação a DNA/genética , Pulmão/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Oxirredução , Proteínas Serina-Treonina Quinases/genética , Proteínas de Ligação a RNARESUMO
BACKGROUND AND PURPOSE: The PARP inhibitor olaparib has recently been approved for human use for the therapy of cancer. Considering the role of PARP in critical illness, we tested the effect of olaparib in a murine model of burn injury, in order to begin exploring the feasibility of repurposing olaparib for the therapy of burn patients. EXPERIMENTAL APPROACH: Mice were subjected to scald burn injury and randomized into vehicle or olaparib (10 mg·kg-1 ·day-1 i.p.) groups. Outcome variables included indices of organ injury, clinical chemistry parameters, plasma levels of inflammatory mediators (at 24 h, 7 and 21 days) and burn wound size (at 21 days). KEY RESULTS: Olaparib reduced myeloperoxidase levels in heart and lung homogenates and reduced malondialdehyde levels in all tissues 24 h post-burn. Olaparib also reduced circulating alkaline aminotransferase, amylase and blood urea nitrogen and creatinine levels, indicative of protection against hepatic, pancreatic and renal dysfunction. Pro-inflammatory mediator (TNF-α, IL-1ß, IFN-γ, GCSF, GM-CSF, eotaxin, KC, MIP-1-α and IL-3, 6 and 12) levels as well as the levels of several mediators that are generally considered anti-inflammatory (IL-4, 10 and 13) were reduced by olaparib. Plasma troponin-I levels (an indicator of skeletal muscle damage) was also attenuated by olaparib. Finally, olaparib stimulated wound healing. CONCLUSIONS AND IMPLICATIONS: The clinically approved PARP inhibitor olaparib improves organ function, suppresses inflammatory responses and accelerates wound healing in murine burn injury. The data raise the potential utility of olaparib for severe burn injury. LINKED ARTICLES: This article is part of a themed section on Inventing New Therapies Without Reinventing the Wheel: The Power of Drug Repurposing. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.2/issuetoc.
Assuntos
Queimaduras/tratamento farmacológico , Inflamação/tratamento farmacológico , Ftalazinas/uso terapêutico , Piperazinas/uso terapêutico , Cicatrização/efeitos dos fármacos , Animais , Queimaduras/sangue , Modelos Animais de Doenças , Mediadores da Inflamação/sangue , Pulmão/metabolismo , Masculino , Malondialdeído/metabolismo , Camundongos , Miocárdio/metabolismo , Peroxidase/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/uso terapêutico , Troponina T/sangueRESUMO
Considering the role of H2S in critical illness, the aim of this study was to compare the outcome of burn in wild-type mice and in mice deficient in CSE, one of the principal mammalian H2S-generating enzymes. Animals were subjected to scald burn. Outcome variables included indices of organ injury, clinical chemistry parameters and plasma levels of inflammatory mediators. Plasma levels of H2S significantly increased in response to burn in wild-type mice, but remained unchanged in CSE-/- mice. Expression of the three H2S-producing enzymes (CSE, CBS and 3-MST) in the lung and liver, and the capacity of tissue homogenates to produce H2S, however, was not affected by burn. In CSE deficient mice there was a significant amelioration of burn-induced accumulation of myeloperoxidase levels in heart, lung, liver and kidney and significantly lower degree of malon dialdehyde accumulation in the heart, lung and kidney than in wild-type mice. CSE deficient mice, compared to wild-type mice, showed a significant attenuation of the burn-induced elevation in circulating alkaline aminotransferase and blood urea nitrogen and creatinine levels, indicative of protective effects of CSE deficiency against burn-induced hepatic, and renal functional impairment. Multiple burn-induced inflammatory mediators (TNF-α, IL-1ß, IL-4, IL-6, IL-10 and IL-12) were significantly lower in the plasma of CSE-/- animals after burn than in the plasma of wild-type controls subjected to burns. In conclusion, CSE deficiency improves organ function and attenuates the inflammatory response in a murine model of burn.
Assuntos
Queimaduras/complicações , Cistationina gama-Liase/deficiência , Inflamação/metabolismo , Insuficiência de Múltiplos Órgãos/metabolismo , Análise de Variância , Animais , Biomarcadores/metabolismo , Western Blotting , Queimaduras/metabolismo , Cistationina gama-Liase/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Sulfeto de Hidrogênio/metabolismo , Masculino , CamundongosRESUMO
Hydrogen sulfide (H2S) has been proposed to exert pro- as well as anti-inflammatory effects in various models of critical illness. In this study, we compared bacterial lipopolysaccharide (LPS)induced changes in inflammatory mediator production, indices of multiple organ injury and survival in wildtype (WT) mice and in mice with reduced expression of one of the three H2Sproducing enzymes, cystathionine-γ-lyase (CSE), cystathionine-ß-synthase (CBS) or 3-mercaptopyruvate sulfurtransferase (3MST). Mice were injected intraperitoneally (i.p.) with LPS (10 mg/kg). After 6 h, the animals were sacrificed, blood and organs were collected and the following parameters were evaluated: blood urea nitrogen (BUN) levels in blood, myeloperoxidase (MPO) and malondialdehyde (MDA) in the lung, cytokine levels in plasma and the expression of the three H2Sproducing enzymes (CBS, CSE and 3MST) in the spleen, lung, liver and kidney. LPS induced a tissuedependent upregulation of some of the H2Sproducing enzymes in WT mice (upregulation of CBS in the spleen, upregulation of 3MST in the liver and upregulation of CBS, CSE and 3MST in the lung). Moreover, LPS impaired glomerular function, as evidenced by increased BUN levels. Renal impairment was comparable in the CSE/ and Δ3MST mice after LPS challenge; however, it was attenuated in the CBS+/ mice. MPO levels (an index of neutrophil infiltration) and MDA levels (an index of oxidative stress) in lung homogenates were significantly increased in response to LPS; these effects were similar in the WT, CBS+/, CSE/ and Δ3MST mice; however, the MDA levels tended to be lower in the CBS+/ and CSE/ mice. LPS induced significant increases in the plasma levels of multiple cytokines [tumor necrosis factor (TNF)α, interleukin (IL)1ß, IL6, IL10, IL12 and interferon (IFN)γ] in plasma; TNFα, IL10 and IL12 levels tended to be lower in all three groups of animals expressing lower levels of H2Sproducing enzymes. The survival rates after the LPS challenge did not show any significant differences between the four animal groups tested. Thus, the findings of this study indicate that a deficiency in 3MST does not significantly affect endotoxemia, while a deficiency in CBS or CSE slightly ameliorates the outcome of LPS-induced endotoxemia in vivo.
Assuntos
Cistationina beta-Sintase/deficiência , Cistationina gama-Liase/deficiência , Endotoxemia/genética , Sulfurtransferases/deficiência , Animais , Biomarcadores , Nitrogênio da Ureia Sanguínea , Cistationina beta-Sintase/genética , Cistationina gama-Liase/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Endotoxemia/imunologia , Endotoxemia/metabolismo , Endotoxemia/mortalidade , Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Sulfeto de Hidrogênio/metabolismo , Lipopolissacarídeos/efeitos adversos , Masculino , Malondialdeído/metabolismo , Camundongos , Camundongos Knockout , Óxido Nítrico/metabolismo , Estresse Oxidativo , Peroxidase/metabolismo , Sulfurtransferases/genéticaRESUMO
Hydrogen sulfide (H2S) exerts beneficial as well as deleterious effects in various models of critical illness. Here we tested the effect of two different pharmacological interventions: (a) inhibition of H2S biosynthesis using the cystathionine-beta-synthase (CBS)/cystathionine-gamma-lyase (CSE) inhibitor aminooxyacetic acid (AOAA) and the mitochondrially targeted H2S donor [10-oxo-10-[4-(3-thioxo-3H-1,2-dithiol-5-yl)phenoxy]decyl]triphenyl-phosphonium (AP39). A 30% body surface area burn injury was induced in anesthetized mice; animals were treated with vehicle, AOAA (10mg/kg i.p. once or once a day for 6days), or AP39 (0.3mg/kg/day once or once a day for 6days). In two separate groups, animals were sacrificed, at 24h post-burn or on Day 7 post-burn, blood and lungs were collected and the following parameters were evaluated: myeloperoxidase (MPO) and malondialdehyde (MDA) in lung homogenates, plasma cytokines (Luminex analysis) and circulating indicators of organ dysfunction (Vetscan analysis). Lung MPO levels (an index of neutrophil infiltration) and MDA levels (an index of oxidative stress) were significantly increased in response to burn injury both at 24h and at 7days; both AOAA and AP39 attenuated these increases. From a panel of inflammatory cytokines (TNFα, IL-1ß, IL-6, IL-10, MCP-1, MIP-2, VEGF and IFNγ) in the plasma, IL-6 and IL-10 levels were markedly elevated at 24h and VEGF was slightly elevated. IL-6 remained highly elevated at 7days post-burn while IL-10 levels decreased, but remained slightly elevated over baseline 7days post-burn. The changes in cytokine levels were attenuated both by AP39 and AOAA at both time points studied. The burn-induced increases in the organ injury markers ALP and ALT, amylase and creatinine were reduced by both AOAA and AP39. We conclude that both H2S biosynthesis inhibition (using AOAA) and H2S donation (using AP39) suppresses inflammatory mediator production and reduces multi-organ injury in a murine model of burn injury, both at an early time point (when systemic H2S levels are elevated) and at a later time point (at which time systemic H2S levels have returned to baseline). These findings point to the complex pathogenetic role of H2S in burns.
Assuntos
Ácido Amino-Oxiacético/farmacologia , Queimaduras/tratamento farmacológico , Queimaduras/metabolismo , Sulfeto de Hidrogênio/metabolismo , Mitocôndrias/efeitos dos fármacos , Compostos Organofosforados/farmacologia , Substâncias Protetoras/farmacologia , Tionas/farmacologia , Animais , Cistationina beta-Sintase/metabolismo , Cistationina gama-Liase , Citocinas/metabolismo , Modelos Animais de Doenças , Inflamação/metabolismo , Malondialdeído/metabolismo , Camundongos , Mitocôndrias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peroxidase/metabolismoRESUMO
Mammalian cells can utilize hydrogen sulfide (H2S) to support mitochondrial respiration. The aim of our study was to explore the potential role of S-sulfhydration (a H2S-induced posttranslational modification, also known as S-persulfidation) of the mitochondrial inner membrane protein ATP synthase (F1F0 ATP synthase/Complex V) in the regulation of mitochondrial bioenergetics. Using a biotin switch assay, we have detected S-sulfhydration of the α subunit (ATP5A1) of ATP synthase in response to exposure to H2S in vitro. The H2S generator compound NaHS induced S-sulfhydration of ATP5A1 in HepG2 and HEK293 cell lysates in a concentration-dependent manner (50-300µM). The activity of immunocaptured mitochondrial ATP synthase enzyme isolated from HepG2 and HEK293 cells was stimulated by NaHS at low concentrations (10-100nM). Site-directed mutagenesis of ATP5A1 in HEK293 cells demonstrated that cysteine residues at positions 244 and 294 are subject to S-sulfhydration. The double mutant ATP synthase protein (C244S/C294S) showed a significantly reduced enzyme activity compared to control and the single-cysteine-mutated recombinant proteins (C244S or C294S). To determine whether endogenous H2S plays a role in the basal S-sulfhydration of ATP synthase in vivo, we compared liver tissues harvested from wild-type mice and mice deficient in cystathionine-gamma-lyase (CSE, one of the three principal mammalian H2S-producing enzymes). Significantly reduced S-sulfhydration of ATP5A1 was observed in liver homogenates of CSE-/- mice, compared to wild-type mice, suggesting a physiological role for CSE-derived endogenous H2S production in the S-sulfhydration of ATP synthase. Various forms of critical illness (including burn injury) upregulate H2S-producing enzymes and stimulate H2S biosynthesis. In liver tissues collected from mice subjected to burn injury, we detected an increased S-sulfhydration of ATP5A1 at the early time points post-burn. At later time points (when systemic H2S levels decrease) S-sulfhydration of ATP5A1 decreased as well. In conclusion, H2S induces S-sulfhydration of ATP5A1 at C244 and C294. This post-translational modification may be a physiological mechanism to maintain ATP synthase in a physiologically activated state, thereby supporting mitochondrial bioenergetics. The sulfhydration of ATP synthase may be a dynamic process, which may be regulated by endogenous H2S levels under various pathophysiological conditions.
Assuntos
Trifosfato de Adenosina/metabolismo , Metabolismo Energético/fisiologia , Sulfeto de Hidrogênio/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Animais , Linhagem Celular , Linhagem Celular Tumoral , Cistationina gama-Liase/metabolismo , Cisteína/metabolismo , Células HEK293 , Células Hep G2 , Humanos , Fígado/metabolismo , Fígado/fisiologia , Masculino , Camundongos , Mutagênese Sítio-Dirigida/métodos , Processamento de Proteína Pós-Traducional/fisiologiaRESUMO
Colon cancer cells contain high levels of cystathionine-beta-synthase (CBS). Its product, hydrogen sulfide (H2S) promotes the growth and proliferation of colorectal tumor cells. In order to improve the antitumor efficacy of the prototypical CBS inhibitor aminooxyacetic acid (AOAA), we have designed and synthesized YD0171, a methyl ester derivative of AOAA. The antiproliferative effect of YD0171 exceeded the antiproliferative potency of AOAA in HCT116 human colon cancer cells. The esterase inhibitor paraoxon prevented the cellular inhibition of CBS activity by YD0171. YD0171 suppressed mitochondrial respiration and glycolytic function and induced G0/G1 arrest, but did not induce tumor cell apoptosis or necrosis. Metabolomic analysis in HCT116 cells showed that YD0171 affects multiple pathways of cell metabolism. The efficacy of YD0171 as an inhibitor of tumor growth was also tested in nude mice bearing subcutaneous HCT116 cancer cell xenografts. Animals were treated via subcutaneous injection of vehicle, AOAA (1, 3 or 9 mg/kg/day) or YD0171 (0.1, 0.5 or 1 mg/kg/day) for 3 weeks. Tumor growth was significantly reduced by 9 mg/kg/day AOAA, but not at the lower doses. YD0171 was more potent: tumor volume was significantly inhibited at 0.5 and 1 mg/kg/day. Thus, the in vivo efficacy of YD0171 is 9-times higher than that of AOAA. YD0171 (1 mg/kg/day) attenuated tumor growth and metastasis formation in the intracecal HCT116 tumor model. YD0171 (3 mg/kg/day) also reduced tumor growth in patient-derived tumor xenograft (PDTX) bearing athymic mice. YD0171 (3 mg/kg/day) induced the regression of established HCT116 tumors in vivo. A 5-day safety study in mice demonstrated that YD0171 at 20 mg/kg/day (given in two divided doses) does not increase plasma markers of organ injury, nor does it induce histological alterations in the liver or kidney. YD0171 caused a slight elevation in plasma homocysteine levels. In conclusion, the prodrug approach improves the pharmacological profile of AOAA; YD0171 represents a prototype for CBS inhibitory anticancer prodrugs. By targeting colorectal cancer bioenergetics, an emerging important hallmark of cancer, the approach exemplified herein may offer direct translational opportunities.
RESUMO
Cecal ligation and puncture (CLP)-induced sepsis is a serious medical condition, caused by a severe systemic infection resulting in a systemic inflammatory response. Recent studies have suggested the therapeutic potential of donors of hydrogen sulfide (H2S), a novel endogenous gasotransmitter and biological mediator in various diseases. The aim of the present study was to assess the effect of H2S supplementation in sepsis, with special reference to its effect on the modulation of regional blood flow. We infused sodium hydrosulfide (NaHS), a compound that produces H2S in aqueous solution (1, 3, or 10âmg/kg/h, for 1 h at each dose level) in control rats or rats 24âh after CLP, and measured blood flow using fluorescent microspheres. In normal control animals, NaHS induced a characteristic redistribution of blood flow, and reduced cardiac, hepatic, and renal blood flow in a dose-dependent fashion. In contrast, in rats subjected to CLP, cardiac, hepatic, and renal blood flow was significantly reduced; infusion of NaHS (1âmg/kg/h and 3âmg/kg/h) significantly increased organ blood flow. In other words, the effect of H2S on regional blood flow is dependent on the status of the animals (i.e., a decrease in blood flow in normal controls, but an increase in blood flow in CLP). We have also evaluated the effect of delayed treatment with NaHS on organ dysfunction and the inflammatory response by treating the animals with NaHS (3âmg/kg) intraperitoneally (i.p.) at 24âh after the start of the CLP procedure; plasma levels of various cytokines and tissue indicators of inflammatory cell infiltration and oxidative stress were measured 6âh later. After 24âh of CLP, glomerular function was significantly impaired, as evidenced by markedly increased (over 4-fold over baseline) blood urea nitrogen and creatinine levels; this increase was also significantly reduced by treatment with NaHS. NaHS also attenuated the CLP-induced increases in malondialdehyde levels (an index of oxidative stress) in heart as well as in liver and myeloperoxidase levels (an index of neutrophil infiltration) in heart and lung. Plasma levels of IL-1ß, IL-5, IL-6, TNF-α, and HMGB1 were attenuated by NaHS. Treatment of NaHS at 3âmg/kg i.p. (but not 1âmg/kg or 6âmg/kg), starting 24âh post-CLP, with dosing repeated every 6âh, improved the survival rate in CLP animals. In summary, treatment with 3âmg/kg H2S-when started in a delayed manner, when CLP-induced organ injury, inflammation and blood flow redistribution have already ensued-improves blood flow to several organs, protects against multiple organ failure, and reduces the plasma levels of multiple pro-inflammatory mediators. These findings support the view that H2S donation may have therapeutic potential in sepsis.
Assuntos
Sulfeto de Hidrogênio/uso terapêutico , Ligadura/efeitos adversos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Choque Séptico/tratamento farmacológico , Animais , Sulfeto de Hidrogênio/farmacologia , Inflamação/sangue , Inflamação/tratamento farmacológico , Inflamação/etiologia , Inflamação/imunologia , Interleucina-5/sangue , Interleucina-6/sangue , Masculino , Infiltração de Neutrófilos/fisiologia , Punções/efeitos adversos , Ratos , Ratos Sprague-Dawley , Choque Séptico/sangue , Choque Séptico/etiologia , Choque Séptico/imunologia , Fatores de TempoRESUMO
This study evaluated the effects of AP39 [(10-oxo-10-(4-(3-thioxo-3H-1,2-dithiol-5yl) phenoxy)decyl) triphenyl phosphonium bromide], a mitochondrially targeted donor of hydrogen sulfide (H2S) in an in vitro model of hypoxia/oxidative stress injury in NRK-49F rat kidney epithelial cells (NRK cells) and in a rat model of renal ischemia-reperfusion injury. Renal oxidative stress was induced by the addition of glucose oxidase, which generates hydrogen peroxide in the culture medium at a constant rate. Glucose oxidase (GOx)-induced oxidative stress led to mitochondrial dysfunction, decreased intracellular ATP content, and, at higher concentrations, increased intracellular oxidant formation (estimated by the fluorescent probe 2, 7-dichlorofluorescein, DCF) and promoted necrosis (estimated by the measurement of lactate dehydrogenase release into the medium) of the NRK cells in vitro. Pretreatment with AP39 (30-300ânM) exerted a concentration-dependent protective effect against all of the above effects of GOx. Most of the effects of AP39 followed a bell-shaped concentration-response curve; at the highest concentration of GOx tested, AP39 was no longer able to afford cytoprotective effects. Rats subjected to renal ischemia/reperfusion responded with a marked increase (over four-fold over sham control baseline) blood urea nitrogen and creatinine levels in blood, indicative of significant renal damage. This was associated with increased neutrophil infiltration into the kidneys (assessed by the myeloperoxidase assay in kidney homogenates), increased oxidative stress (assessed by the malondialdehyde assay in kidney homogenates), and an increase in plasma levels of IL-12. Pretreatment with AP39 (0.1, 0.2, and 0.3âmg/kg) provided a dose-dependent protection against these pathophysiological alterations; the most pronounced protective effect was observed at the 0.3âmg/kg dose of the H2S donor; nevertheless, AP39 failed to achieve a complete normalization of any of the injury markers measured. The partial protective effects of AP39 correlated with a partial improvement of kidney histological scores and reduced TUNEL staining (an indicator of DNA damage and apoptosis). In summary, the mitochondria-targeted H2S donor AP39 exerted dose-dependent protective effects against renal epithelial cell injury in vitro and renal ischemia-reperfusion injury in vivo. We hypothesize that the beneficial actions of AP39 are related to the reduction of cellular oxidative stress, and subsequent attenuation of various positive feed-forward cycles of inflammatory and oxidative processes.
Assuntos
Injúria Renal Aguda/prevenção & controle , Citoproteção/fisiologia , Sulfeto de Hidrogênio/metabolismo , Compostos Organofosforados/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Tionas/farmacologia , Animais , Hipóxia Celular/efeitos dos fármacos , Hipóxia Celular/fisiologia , Células Cultivadas , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Células Epiteliais/efeitos dos fármacos , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Compostos Organofosforados/administração & dosagem , Compostos Organofosforados/uso terapêutico , Ratos Sprague-Dawley , Tionas/administração & dosagem , Tionas/uso terapêuticoRESUMO
Severe thermal injury induces a pathophysiological response that affects most of the organs within the body; liver, heart, lung, skeletal muscle among others, with inflammation and hyper-metabolism as a hallmark of the post-burn damage. Oxidative stress has been implicated as a key component in development of inflammatory and metabolic responses induced by burn. The goal of the current study was to evaluate several critical mitochondrial functions in a mouse model of severe burn injury. Mitochondrial bioenergetics, measured by Extracellular Flux Analyzer, showed a time dependent, post-burn decrease in basal respiration and ATP-turnover but enhanced maximal respiratory capacity in mitochondria isolated from the liver and lung of animals subjected to burn injury. Moreover, we detected a tissue-specific degree of DNA damage, particularly of the mitochondrial DNA, with the most profound effect detected in lungs and hearts of mice subjected to burn injury. Increased mitochondrial biogenesis in lung tissue in response to burn injury was also observed. Burn injury also induced time dependent increases in oxidative stress (measured by amount of malondialdehyde) and neutrophil infiltration (measured by myeloperoxidase activity), particularly in lung and heart. Tissue mononuclear cell infiltration was also confirmed by immunohistochemistry. The amount of poly(ADP-ribose) polymers decreased in the liver, but increased in the heart in later time points after burn. All of these biochemical changes were also associated with histological alterations in all three organs studied. Finally, we detected a significant increase in mitochondrial DNA fragments circulating in the blood immediately post-burn. There was no evidence of systemic bacteremia, or the presence of bacterial DNA fragments at any time after burn injury. The majority of the measured parameters demonstrated a sustained elevation even at 20-40 days post injury suggesting a long-lasting effect of thermal injury on organ function. The current data show that there are marked time-dependent and tissue-specific alterations in mitochondrial function induced by thermal injury, and suggest that mitochondria-specific damage is one of the earliest responses to burn injury. Mitochondria may be potential therapeutic targets in the future experimental therapy of burns.
Assuntos
Queimaduras/metabolismo , Queimaduras/patologia , DNA Mitocondrial/metabolismo , Mitocôndrias/metabolismo , Animais , Dano ao DNA , Modelos Animais de Doenças , Metabolismo Energético , Peroxidação de Lipídeos , Fígado/metabolismo , Fígado/patologia , Pulmão/metabolismo , Pulmão/patologia , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/patologia , Mitocôndrias Cardíacas/metabolismo , Mitocôndrias Hepáticas/metabolismo , Miocárdio/metabolismo , Miocárdio/patologia , Infiltração de Neutrófilos , Estresse Oxidativo , Peroxidase/metabolismoRESUMO
Toll-like receptors (TLRs) are signaling receptors in the innate immune system that is specific immunologic response to systemic bacterial infection and injury. TLRs contribute to the initial induction of neuroinflammation in the CNS. In spinal cord injury (SCI) intricate immune cell interactions are triggered, typically consisting of a staggered multiphasic immune cell response, which can become deregulated. The present study aims to evaluate the role of TLR4 signaling pathway in the development of secondary damage in a mouse model of SCI using TLR4-deficient (TLR4-KO) mice such as C57BL/10ScNJ and C3H/HeJ mice. We evaluated behavioral changes, histological, immunohistochemistry and molecular assessment in TLR4-KO after SCI. SCI was performed on TLR4-KO and wild-type (WT) mice by the application of vascular clips (force of 24g) to the dura via a four-level T5-T8 laminectomy. Mice were sacrificed at 24h after SCI to evaluate the various parameters. SCI TLR4 KO mice developed severer hind limb motor dysfunction and neuronal death by histological evaluation, myeloid differentiation primary response 88 (Myd88) expression as well as an increase in nuclear factor NF-κB activity, tumor necrosis factor (TNF)-α and interleukin (IL)-1ß levels, glial fibrillary acidic protein (GFAP), microglia marker (CD11ß), inducible nitric oxide synthases (iNOS), poly-ADP-ribose polymerase (PARP) and nitrotyrosine expression compared to WT mice. Moreover, the absence of TLR4 also caused a decrease in phosphorylated interferon regulatory transcription factor (p-IRF3) and interferon (IFN-ß) release. In addition, SCI TLR4 KO mice showed in spinal cord tissues a more pronounced up-regulation of Bax and a down-regulation of Bcl-2 compared to SCI WT mice. Finally, we clearly demonstrated that TLR4 is important for coordinating post-injury sequel and in regulating inflammation after SCI.
Assuntos
Inflamação/imunologia , Transdução de Sinais/imunologia , Traumatismos da Medula Espinal/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Apoptose/genética , Apoptose/imunologia , Antígeno CD11b/biossíntese , Regulação da Expressão Gênica , Proteína Glial Fibrilar Ácida , Membro Posterior/patologia , Fator Regulador 3 de Interferon/metabolismo , Interferon beta/metabolismo , Interleucina-1beta/biossíntese , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/biossíntese , Proteínas do Tecido Nervoso/biossíntese , Neuroimunomodulação/imunologia , Infiltração de Neutrófilos/genética , Infiltração de Neutrófilos/imunologia , Óxido Nítrico Sintase Tipo II/biossíntese , Poli(ADP-Ribose) Polimerases/biossíntese , Receptor 4 Toll-Like/genética , Fator de Transcrição RelA/biossíntese , Fator de Necrose Tumoral alfa/biossíntese , Proteína X Associada a bcl-2/biossínteseRESUMO
The aim of this study was to investigate the efficacy of PEA+silymarin as a combination treatment in a mouse model of renal I/R and to verify whether PEA+silymarin could exert more potent effects compared to the single substances even if administered at lower doses. Mice were subjected to bilateral renal artery occlusion (30min) and reperfusion (6h) and received intraperitoneally silymarin (100, 30 and 10mg/kg) or PEA (1mg/kg) or PEA (1mg/kg)+silymarin (10mg/kg) 15min before release of clamps. Specific indicators of renal dysfunction, tubular injury, myeloperoxidase activity and malondialdehyde levels were measured. The nuclear factor κB pathway and apoptotic mechanisms were also investigated. The treatment with silymarin reduced kidney dysfunction, histological damage, neutrophil infiltration and oxidative stress in a dose dependent manner. Furthermore, PEA+silymarin showed a significant potentiated effect. Therefore, NF-κB and apoptosis pathways were also significantly inhibited. Our results clearly demonstrate that PEA+silymarin treatment attenuated the degree of renal inflammation.
Assuntos
Etanolaminas/farmacologia , Rim/efeitos dos fármacos , Rim/lesões , Ácidos Palmíticos/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Silimarina/farmacologia , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Amidas , Animais , Apoptose/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Quimases/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Interações Medicamentosas , Etanolaminas/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Masculino , Camundongos , Nitratos/metabolismo , Nitritos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ácidos Palmíticos/uso terapêutico , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Fatores de Risco , Silimarina/uso terapêutico , Fator de Transcrição RelA/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
The aim of this study was to investigate the effects of palmitoylethanolamide (PEA), an endogenous fatty acid amide belonging to the family of the N-acylethanolamines (NAEs), in rats subjected to endotoxin-induced uveitis (EIU). EIU was induced in male rats by a single footpad injection of 200µg lipopolysaccharide (LPS). PEA was administered intraperitoneally at 1h before and 7h after injection of LPS. Another group of animals was treated with vehicle. Dexamethasone (DEX) was administered as a positive control. Rats were sacrificed 16h after injection and the eyes tissues were collected for histology, immunohistochemical and western blot analyses. The histological evaluation of the iris-ciliary body showed an increase of neutrophilic infiltration and nuclear modification of vessel of endothelial cells. PEA treatment decreased the inflammatory cell infiltration and improved histological damage of eye tissues. In addition, PEA treatment reduced pro-inflammatory tumor necrosis factor (TNF-α) levels, protein extravasion and lipid peroxidation. Immunohistochemical analysis for intracellular adhesion molecule (ICAM)-1 and nitrotyrosine showed a positive staining from LPS-injected rats. The degree of staining for ICAM-1 and nitrotyrosine was significantly reduced in eye sections from LPS-injected rats treated with PEA. In addition, an increase of inducible nitric oxide synthase (iNOS) and nuclear factor (NF-κB) was also evaluated in inflammed ocular tissues by western blot. PEA strongly inhibited iNOS expression and nuclear NF-κB translocation. Thus, in this study we demonstrated that PEA reduces the degree of ocular inflammation in a rat model of EIU.
Assuntos
Anti-Inflamatórios/farmacologia , Etanolaminas/farmacologia , Ácidos Palmíticos/farmacologia , Úvea/efeitos dos fármacos , Uveíte/prevenção & controle , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Amidas , Animais , Dexametasona/farmacologia , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Mediadores da Inflamação/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Lipopolissacarídeos , Masculino , Malondialdeído/metabolismo , Infiltração de Neutrófilos/efeitos dos fármacos , Óxido Nítrico Sintase Tipo II/metabolismo , Ratos Endogâmicos Lew , Fator de Transcrição RelA/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo , Úvea/metabolismo , Úvea/patologia , Uveíte/induzido quimicamente , Uveíte/metabolismo , Uveíte/patologiaRESUMO
Hydrogen sulfide (H2S), as a reducing agent and an antioxidant molecule, exerts protective effects against hyperglycemic stress in the vascular endothelium. The mitochondrial enzyme 3-mercaptopyruvate sulfurtransferase (3-MST) is an important biological source of H2S. We have recently demonstrated that 3-MST activity is inhibited by oxidative stress in vitro and speculated that this may have an adverse effect on cellular homeostasis. In the current study, given the importance of H2S as a vasorelaxant, angiogenesis stimulator and cellular bioenergetic mediator, we first determined whether the 3-MST/H2S system plays a physiological regulatory role in endothelial cells. Next, we tested whether a dysfunction of this pathway develops during the development of hyperglycemia and µmol/L to diabetes-associated vascular complications. Intraperitoneal (IP) 3-MP (1 mg/kg) raised plasma H2S levels in rats. 3-MP (10 1 mmol/L) promoted angiogenesis in vitro in bEnd3 microvascular endothelial cells and in vivo in a Matrigel assay in mice (0.3-1 mg/kg). In vitro studies with bEnd3 cell homogenates demonstrated that the 3-MP-induced increases in H2S production depended on enzymatic activity, although at higher concentrations (1-3 mmol/L) there was also evidence for an additional nonenzymatic H2S production by 3-MP. In vivo, 3-MP facilitated wound healing in rats, induced the relaxation of dermal microvessels and increased mitochondrial bioenergetic function. In vitro hyperglycemia or in vivo streptozotocin diabetes impaired angiogenesis, attenuated mitochondrial function and delayed wound healing; all of these responses were associated with an impairment of the proangiogenic and bioenergetic effects of 3-MP. The antioxidants DL-α-lipoic acid (LA) in vivo, or dihydrolipoic acid (DHLA) in vitro restored the ability of 3-MP to stimulate angiogenesis, cellular bioenergetics and wound healing in hyperglycemia and diabetes. We conclude that diabetes leads to an impairment of the 3-MST/H2S pathway, and speculate that this may contribute to the pathogenesis of hyperglycemic endothelial cell dysfunction. We also suggest that therapy with H2S donors, or treatment with the combination of 3-MP and lipoic acid may be beneficial in improving angiogenesis and bioenergetics in hyperglycemia.
Assuntos
Endotélio Vascular/fisiologia , Metabolismo Energético/fisiologia , Sulfeto de Hidrogênio/metabolismo , Redes e Vias Metabólicas , Neovascularização Fisiológica , Sulfurtransferases/metabolismo , Animais , Linhagem Celular , Proteínas Quinases Dependentes de GMP Cíclico/metabolismo , Cisteína/administração & dosagem , Cisteína/análogos & derivados , Cisteína/farmacologia , Diabetes Mellitus/metabolismo , Modelos Animais de Doenças , Células Endoteliais , Endotélio Vascular/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Sulfeto de Hidrogênio/sangue , Hiperglicemia/tratamento farmacológico , Hiperglicemia/metabolismo , Masculino , Camundongos , Mitocôndrias/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Consumo de Oxigênio , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Sulfurtransferases/genética , Ácido Tióctico/farmacologia , Vasodilatadores/administração & dosagem , Vasodilatadores/farmacologiaRESUMO
BACKGROUND & AIMS: The beneficial properties of the flavonoid fraction of bergamot juice (BJe) have been raising interest and have been the subject of recent studies, considering the potentiality of its health promoting substances. Flavonoids have demonstrated radical-scavenging and anti-inflammatory activities. The aim of the present study was to examine the effects of BJe in mice subjected to experimental colitis. METHODS: Colitis was induced in mice by intracolonic instillation of dinitrobenzene sulfonic acid (DNBS). BJe was administered daily orally (at 5, 10 and 20 mg/kg). RESULTS: Four days after DNBS administration, colon nuclear factor NF-κB translocation and MAP kinase phospho-JNK activation were increased as well as cytokine production such as tumor necrosis factor (TNF)-α and interleukin (IL)-1ß. Neutrophil infiltration, by myeloperoxidase (MPO) activity, in the mucosa was associated with up-regulation of adhesion molecules (ICAM-1 and P-selectin). Immunohistochemistry for nitrotyrosine and poly ADP-ribose (PAR) also showed an intense staining in the inflamed colon. Treatment with BJe decreased the appearance of diarrhea and body weight loss. This was associated with a reduction in colonic MPO activity. BJe reduced nuclear NF-κB translocation, p-JNK activation, the pro-inflammatory cytokines release, the appearance of nitrotyrosine and PAR in the colon and reduced the up-regulation of ICAM-1 and P-selectin. In addition, colon inflammation was also associated with apoptotic damage. Treatment with BJe caused a decrease of pro-apoptotic Bax expression and an increase of anti-apoptotic Bcl-2 expression. CONCLUSIONS: The results of this study suggested that administration of BJe induced, partly specified, anti-inflammatory mechanisms, which potentially may be beneficial for the treatment of IBD in humans.
Assuntos
Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Citrus/química , Colite/tratamento farmacológico , Doenças Inflamatórias Intestinais/tratamento farmacológico , Extratos Vegetais/farmacologia , Administração Oral , Animais , Apoptose/efeitos dos fármacos , Benzenossulfonatos , Bebidas/análise , Colite/induzido quimicamente , Colo/efeitos dos fármacos , Colo/metabolismo , Modelos Animais de Doenças , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Masculino , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Infiltração de Neutrófilos/efeitos dos fármacos , Selectina-P/genética , Selectina-P/metabolismo , Peroxidase/metabolismo , Poli Adenosina Difosfato Ribose/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismo , Regulação para Cima , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
The aim of this study was to investigate the effect of costunolide (CS) and dehydrocostuslactone (DCE) a well-known sesquiterpene lactones contained in many plants, in a model of lung injury induced by carrageenan administration in the mice. Injection of carrageenan into the pleural cavity of mice elicited an acute inflammatory response characterized by fluid accumulation in the pleural cavity which contained a large number of polymorphonuclear cells (PMNs) as well as an infiltration of PMNs in lung tissues and increased production of tumour necrosis factor α (TNF-α). All parameters of inflammation were attenuated by CS and DCE (15mg/kg 10% DMSO i.p.) administered 1h before carrageenan. Carrageenan induced an up regulation of the intracellular adhesion molecules-1 (ICAM-1) and P-selectin, as well as nitrotyrosine and poly (ADP-ribose) (PAR) as determined by immunohistochemical analysis of lung tissues. The degree of staining for the ICAM-1, P-selectin, nitrotyrosine and PAR was reduced by CS and DCE. Additionally we show that this inflammatory events were associated with NF-κB and STAT3 activation and these sesquiterpenes down-regulated it. Taken together, ours results clearly shown that CS and DCE may offer a novel therapeutic approach for the management of inflammatory diseases.