RESUMO
Hepatocellular carcinoma (HCC) is one of the most common human malignant tumors. It is known that in the cells of many cancers, including HCC, nuclear translocation and accumulation of YB-1 often indicates a poor prognosis. This nuclear translocation is induced by genotoxic stress resulting from administration of anticancer agents. Accumulation of YB-1 in the nucleus induces the expression of many genes related to cancer aggressiveness. Therefore, compounds capable of inhibiting anticancer drug-induced YB-1 nuclear translocation without cytotoxicity will be a powerful tool for cancer chemotherapy. In the present study, we found that indirubin derivative, 7-hydroxyindirubin strongly inhibited the actinomycin D-induced nuclear translocation of YB-1 more efficiently without showing cytotoxicity in HepG2, a human HCC cells. The compound successfully suppressed the nuclear YB-1-mediated expression of genes such as MDR1, MVP, EGFR, and CXCR4, which are known to disturb cancer treatment. 7-Hydroxyindirubin also increased the susceptibility of drug-resistant HepG2 cells to ActD. It was also demonstrated that 7-hydroxyindirubin inhibits the nuclear translocation of YB-1 with or without phosphorylation at the Ser102 residue.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Núcleo Celular/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Proteína 1 de Ligação a Y-Box/metabolismo , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Células Hep G2 , Humanos , Indóis/química , Indóis/farmacologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Transporte Proteico , Receptores CXCR4/metabolismoRESUMO
Obesity elevates the risk of cardiovascular disease and has been strongly associated with increases in the incidence of many metabolic diseases. Therefore, prevention of obesity leads to the prevention of metabolic diseases. In light of this, substances that exert anti-obesity effects are crucial for the prevention of obesity. Indirubin, a 3,2' bisindole isomer of indigo, is the active component of the traditional Chinese medicine used for the treatment of chronic myelocytic leukemia. In particular, indirubin-3'-oxime (1) was shown to inhibit the differentiation of adipocytes. In this study, we investigated the inhibitory effects of nine indirubin-3'-oxime derivatives against lipid accumulation during differentiation in 3T3-L1 cells. Among the compounds tested, 5-methoxyindirubin-3'-oxime (2) and 6-bromoindirubin-3'-oxime (7) at 5 µM exhibited significantly stronger inhibitory activity than indirubin-3'-oxime (1). Furthermore, 5-methoxyindirubin-3'-oxime (2) and 6-bromoindirubin-3'-oxime (7) markedly suppressed the expression of CCAAT/enhancer-binding protein α, peroxisome proliferator activator γ2, and adipocyte protein 2, both of which are key adipogenic regulators at the intermediate stage of adipocyte differentiation. Our results demonstrate that 5-methoxyindirubin-3'-oxime (2) and 6-bromoindirubin-3'-oxime (7) significantly down-regulated lipid accumulation during differentiation of 3T3-L1 cells, suggesting their potential as novel therapeutic drugs against the development of obesity.
Assuntos
Metabolismo dos Lipídeos/efeitos dos fármacos , Células 3T3-L1/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular , Indóis/química , Indóis/farmacologia , Camundongos , Oximas , Extratos Vegetais/farmacologiaRESUMO
In order to develop an efficient organocatalyst for the enantioselective N-H insertion reaction via carbene/carbenoid, the catalytic core of the cinchona alkaloids was investigated. According to our working hypothesis of an eight-membered ring transition state in the N-H insertion reaction, two pairs of enantiomers related to 2-amino-1-phenylethanol were investigated for their chiral inducing potential. Since both (1R,2S)-isomers gave the N-phenyl-1-phenylglycine derivative enriched in the R-form, while their enantiomers gave the S-form, the 2-amino-1-phenylethanol structure is concluded to be the catalytic core of the cinchona alkaloid in the enantioselective N-H insertion reaction via rhodium(II) carbenoid.
Assuntos
Amino Álcoois/química , Alcaloides de Cinchona/química , Álcoois Benzílicos/química , Catálise , Complexos de Coordenação/química , Hidrogênio/química , Nitrogênio/química , Ródio/química , EstereoisomerismoRESUMO
Hepatocellular carcinoma (HCC) is a disease with poor prognosis. Nuclear accumulation of YB-1 is closely related to the malignancy of HCC. Treatment with anticancer agents often induces translocation of YB-1 from cytoplasm to nucleus and activates the expression of multidrug resistance gene 1 (MDR1). Therefore, any effective inhibitor of this phenomenon would be useful for cancer treatment. Here we examined various indirubin derivatives and found that indirubin 3'-oxime inhibits actinomycin D-induced nuclear transport of YB-1 and suppresses the activation of MDR1 gene expression in the human hepatocellular carcinoma cell line HepG2. Furthermore, use of both indirubin 3'-oxime and actinomycin D in combination increased the anticancer effect on HepG2 cells. Indirubin 3'-oxime is a novel and efficient inhibitor of anticancer agent-induced YB-1 nuclear translocation.
Assuntos
Antineoplásicos/administração & dosagem , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Indóis/administração & dosagem , Oximas/administração & dosagem , Proteína 1 de Ligação a Y-Box/metabolismo , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Inibidores da Angiogênese/administração & dosagem , Carcinoma Hepatocelular/patologia , Núcleo Celular/patologia , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células Hep G2 , HumanosRESUMO
Indirubin and its derivatives have been reported to exhibit anti-cancer and anti-inflammatory activities. Recently, some of its derived analogs have been shown to have neuroprotective potential. Endoplasmic reticulum (ER) stress has been demonstrated to contribute to the pathogenesis of various neurodegenerative diseases, whereas the effects of indirubin derivatives on ER stress-induced cell death have not been addressed. In the present study, a series of 44 derivatives of indirubin was prepared to search for a novel class of neuroprotective agents against ER stress-induced neuronal death. The MTT reduction assay indicated that tunicamycin (TM), an inducer of ER stress, significantly decreased the viability of hippocampal neuronal HT22 cells. Among the compounds tested, eight showed significant inhibitory activity against TM-induced cell death. Western blot analysis showed that application of these analogs to the cells simultaneously with TM reduced the TM-induced expression of CHOP, an established mediator of ER stress. Our results suggest that the preventive effect of these indirubin derivatives against ER stress-induced neuronal death may be due, at least in part, to attenuation of the CHOP-dependent signaling system.
Assuntos
Regulação para Baixo/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Substâncias Protetoras/farmacologia , Fator de Transcrição CHOP/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Indóis/química , Indóis/farmacologia , Camundongos , Substâncias Protetoras/química , Relação Estrutura-Atividade , Tunicamicina/toxicidadeRESUMO
Indirubin 3'-oxime (Indox (1b)) suppresses cancer cell growth (IC50: 15µM towards HepG2 cells) and inhibits cell cycle-related kinases such as cyclin-dependent kinases and glycogen synthase kinase-3ß. We have previously reported that the conjugation of 1b with oxirane, a protein-reactive component, enhanced the cytotoxic activity of Indox as determined from the IC50 value (1.7µM) of indirubin 3'-(O-oxiran-2-ylmethyl)oxime (Epox/Ind (1c)). Here we prepared Epox/Ind derivatives with one or two halogen atoms or a methoxy group on the aromatic ring(s) of an Indox moiety and studied the structure-activity relationships of the substituent(s). We found that bromine-substitution at the 5-position on 1c or any Epox/Ind derivative(s) having bromine on the aromatic ring except Epox/6'-Br-Ind was efficient to improving anticancer activity. Of the 22 Epox/Ind derivatives, 5-bromoindirubin 3'-(O-oxiran-2-ylmethyl)oxime (Epox/5-Br-Ind (2c)) was the best anticancer agent in both short- (24h) (IC50: 0.67µM) and extended-duration (72h) cultures. The high anticancer activity of 2c was partly due to it being a poor substrate and a suicide inhibitor for epoxide hydrolase as epoxide hydrolase was identified as the enzyme primarily responsible for the metabolism of 2c.
Assuntos
Antineoplásicos/química , Epóxido Hidrolases/antagonistas & inibidores , Indóis/química , Oximas/química , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Sítios de Ligação , Bromo/química , Sobrevivência Celular/efeitos dos fármacos , Epóxido Hidrolases/metabolismo , Células Hep G2 , Humanos , Indóis/metabolismo , Indóis/farmacologia , Cinética , Simulação de Acoplamento Molecular , Oximas/metabolismo , Oximas/farmacologia , Estrutura Terciária de Proteína , Relação Estrutura-AtividadeRESUMO
S-allyl-l-cysteine (SAC) is known to have neuroprotective properties. We synthesized various SAC derivatives and tested their effects on endoplasmic reticulum stress-induced neurotoxicity in cultured hippocampal neurons (HPNs). Among the compounds tested, S-propyl-l-cysteine (SPC) exhibited the strongest neuroprotective activity in HPNs, followed by S-ethyl-l-cysteine (SEC) and S-methyl-l-cysteine (SMC). Unlike SAC and SMC, SPC and SEC did not have inhibitory activity on µ-calpain, suggesting that the mechanism underlying the protective activity of SPC and SEC differs from that of SAC.
Assuntos
Calpaína/antagonistas & inibidores , Cisteína/análogos & derivados , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fármacos Neuroprotetores , Animais , Células Cultivadas , Cisteína/farmacologia , Estresse do Retículo Endoplasmático/fisiologia , Hipocampo/citologia , Ratos WistarRESUMO
Indirubin inhibits cyclin-dependent kinases by binding to their ATP-binding site, thereby exerting potent cytotoxicity on some tumor cells. We examined the anti-tumor effect of indirubin 3'-epoxide on human neuroblastoma cell lines (IMR-32, SK-N-SH, and NB-39). The results revealed potent cytotoxicity of indirubin 3'-epoxide against the IMR-32 (IC50: 0.16 µM) and SK-N-SH (IC50: 0.07 µM) cells. Furthermore, it also induced an increase of the sub-G1 population in the IMR-32 cells. Examination by Hoechst 33342 staining revealed apoptosis characterized by cell shrinkage, nuclear condensation and nuclear fragmentation in a concentration-dependent manner. Furthermore, annexin V-propidium iodide (PI) double-staining revealed an increase in the percentage of early apoptotic cells following treatment of the cells with indirubin 3'-epoxide without activation of caspases. In addition, significant decreases in the protein level of survivin and poly(ADP-ribose)polymerase (PARP), and increase in that of apoptosis-inducing factor (AIF) were found in the nuclei of the cells. These results suggest that indirubin 3'-epoxide induced caspase-independent apoptosis through mechanisms involving DNA fragmentation and inhibition of DNA repair.
Assuntos
Antineoplásicos/farmacologia , Indóis/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Caspases/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Fragmentação do DNA , Humanos , Neuroblastoma/patologiaRESUMO
A Pseudomonas aeruginosa quorum-sensing system, which produces N-(3-oxododecanoyl)-l-homoserine lactone (3-oxo-C12 -HSL) and N-butanoyl-l-homoserine lactone (C4 -HSL), regulates the virulence factors. In our previous study, 3-oxo-C12 -HSL, encoded by lasI gene, was shown to promote wound healing. However, the effect of C4 -HSL, encoded by rhlI gene, remains to be elucidated. We addressed the effect of C4 -HSL on wounds in P. aeruginosa infection. Wounds were created on the backs of Sprague-Dawley SD rats, and P. aeruginosa PAO1 (PAO1) or its rhlI deletion mutant (ΔrhlI) or lasI deletion mutant (ΔlasI) was inoculated onto the wound. Rats were injected intraperitoneally with anti-C4 -HSL antiserum or treated with C4 -HSL at the wound surface. PAO1 inoculation led to significant acceleration of wound healing, which was associated with neutrophil infiltration and TNF-α synthesis. These responses were reversed, except for TNF-α production, when ΔrhlI was inoculated instead of PAO1 or when rats were co-treated with PAO1 and anti-C4 -HSL antiserum. In contrast, the healing process and neutrophil infiltration, but not TNF-α synthesis, were accelerated when C4 -HSL was administered in the absence of PAO1. This acceleration was not affected by anti-TNF-α antibody. These results suggest that C4 -HSL may be involved in the acceleration of acute wound healing in P. aeruginosa infection by modifying the neutrophilic inflammation.
Assuntos
4-Butirolactona/análogos & derivados , Pseudomonas aeruginosa/enzimologia , Cicatrização/fisiologia , Ferimentos e Lesões/patologia , Ferimentos e Lesões/terapia , 4-Butirolactona/farmacologia , Doença Aguda , Análise de Variância , Animais , Biópsia por Agulha , Modelos Animais de Doenças , Feminino , Homosserina/análogos & derivados , Homosserina/farmacologia , Imuno-Histoquímica , Neutrófilos/metabolismo , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Estatísticas não Paramétricas , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Indirubin is a potent inhibitor of cell cycle-related protein kinases by binding to the ATP-binding site and thus is a promising compound for development as an antitumor drug. We prepared indirubin 3'-(O-oxiran-2-ylmethyl)oxime (Epox/Ind), in which the ATP-binding site orientated part was attached by non-specific alkylating group. The IC50 value of Epox/Ind at 1.7 µM in HepG2 cells is comparable to that of cisplatin (4.0 µM). Furthermore, Epox/Ind was shown to be metabolized by a HepG2 cell lysate into indirubin 3'-(O-2,3-dihydroxypropyl)oxime (E804), the sole extractable metabolite. The lower toxicity of this metabolite may explain the lack of cytotoxicity of 1 µM Epox/Ind observed in HepG2 cells beyond an initial loss of viability in the first 24h of treatment.
Assuntos
Antineoplásicos/farmacologia , Indóis/farmacologia , Oximas/farmacologia , Antineoplásicos/química , Antineoplásicos/metabolismo , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Células Hep G2 , Humanos , Indóis/síntese química , Indóis/química , Estrutura Molecular , Oximas/síntese química , Oximas/química , Relação Estrutura-AtividadeRESUMO
BACKGROUND: Bacteria release a wide variety of small molecules including cell-to-cell signaling compounds. Gram-negative bacteria use a variety of self-produced autoinducers such as acylated homoserine lactones (acyl-HSLs) as signal compounds for quorum sensing (QS) within and between bacterial species. QS plays a significant role in the pathogenesis of infectious diseases and in beneficial symbiosis by responding to acyl-HSLs in Pseudomonas aeruginosa. It is considered that the selection of bacterial languages is necessary to regulate gene expression and thus it leads to the regulation of virulence and provides a growth advantage in several environments. In this study, we hypothesized that RND-type efflux pump system MexAB-OprM of P. aeruginosa might function in the selection of acyl-HSLs, and we provide evidence to support this hypothesis. RESULTS: Loss of MexAB-OprM due to deletion of mexB caused increases in QS responses, as shown by the expression of gfp located downstream of the lasB promoter and LasB elastase activity, which is regulated by a LasR-3-oxo-C12-HSL complex. Either complementation with a plasmid containing wild-type mexB or the addition of a LasR-specific inhibitor, patulin, repressed these high responses to 3-oxo-acyl-HSLs. Furthermore, it was shown that the acyl-HSLs-dependent response of P. aeruginosa was affected by the inhibition of MexB transport activity and the mexB mutant. The P. aeruginosa MexAB-OprM deletion mutant showed a strong QS response to 3-oxo-C10-HSL produced by Vibrio anguillarum in a bacterial cross-talk experiment. CONCLUSION: This work demonstrated that MexAB-OprM does not control the binding of LasR to 3-oxo-Cn-HSLs but rather accessibility of non-cognate acyl-HSLs to LasR in P. aeruginosa. MexAB-OprM not only influences multidrug resistance, but also selects acyl-HSLs and regulates QS in P. aeruginosa. The results demonstrate a new QS regulation mechanism via the efflux system MexAB-OprM in P. aeruginosa.
Assuntos
Acil-Butirolactonas/metabolismo , Proteínas da Membrana Bacteriana Externa/metabolismo , Homosserina/análogos & derivados , Lactonas/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Pseudomonas aeruginosa/fisiologia , Percepção de Quorum , Fusão Gênica Artificial , Proteínas da Membrana Bacteriana Externa/genética , Deleção de Genes , Genes Reporter , Teste de Complementação Genética , Proteínas de Fluorescência Verde/análise , Proteínas de Fluorescência Verde/genética , Homosserina/metabolismo , Proteínas de Membrana Transportadoras/genética , Modelos Biológicos , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/metabolismo , Transdução de Sinais , Vibrio/metabolismo , Vibrio/fisiologiaRESUMO
This paper reports the synthesis of methoxy- and bromo-indirubins, and their antiproliferative activities in human neuroblastoma. Among 20 compounds, 5'-methoxyindirubin induced cell death in human neuroblastoma cells (IMR-32, SK-N-SH and NB-39) without inhibiting normal cells (NHDF and HUVEC). Typical morphologic features of apoptosis were observed in 5'-methoxyindirubin-treated cells by Hoechst 33342 staining. Additional studies by flow cytometry support apoptosis induction. These data suggest that 5'-methoxyindirubin might be an effective drug for treatment of neuroblastoma.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Neuroblastoma/tratamento farmacológico , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Química Sintética , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Indóis/síntese química , Indóis/química , Indóis/farmacologia , Estrutura Molecular , Neuroblastoma/patologia , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
We have examined the potential bactericidal activities of several tetramic acids derived from Pseudomonas autoinducers against Clostridium difficile, a cause of antibiotic-associated pseudomembranous colitis. Clinical isolates of C. difficile (n=4) were incubated in broth with a chemically synthesized Pseudomonas autoinducer and its tetramic acid derivatives. The structure-activity relationship and the mechanisms of action were examined by a time-killing assay and by determination of the morphological/staining characteristics. The use of some tetramic acids derived from N-3-oxododecanoyl L-homoserine lactone resulted in more than 3-log reductions in the viability of C. difficile within 30 min at 30 microM. The outer membrane was suggested to be one of the targets for the bactericidal activity of tetramic acid, because disturbance of the bacterial outer surface was demonstrated by alteration of the Gram-staining characteristic and electron microscopy. The data for the tetramic acid derivatives demonstrate that the keto-enol structure and the length of the acyl side chain of tetramic acid may be essential for the antibacterial activity of this molecule. These results suggest the potential for tetramic acid derivatives to be novel agents with activity against C. difficile.
Assuntos
Antibacterianos/farmacologia , Clostridioides difficile/efeitos dos fármacos , Homosserina/análogos & derivados , Lactonas/química , Pirrolidinonas/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Clostridioides difficile/ultraestrutura , Homosserina/química , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Transmissão , Pirrolidinonas/síntese química , Pirrolidinonas/química , Percepção de Quorum/fisiologiaRESUMO
Cinchona alkaloids are effective additives for enantioselective O-H insertion of alpha-phenyldiazoacetate and water by rhodium(II) complexes. Addition of silica gel promotes O-H insertion in the reaction rate and the reaction proceeds smoothly at less than the freezing point of water, e.g., -10 degrees C, and provided mandelate in up to 50% ee. The results reported here are the highest asymmetric inductions obtained to date for O-H insertions via a Rh-carbenoid.
Assuntos
Alcaloides de Cinchona/química , Complexos de Coordenação/química , Fenilacetatos/química , Ródio/química , Água/química , Catálise , Estrutura Molecular , EstereoisomerismoRESUMO
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with high invasive and metastatic potential. We generated a spontaneous PDAC mouse model and examined the therapeutic potential of indirubin 3'-oxime (Indox) against PDAC bearing mouse in vivo. METHODS: Randomized 3-month-old LSL-KrasG12D/+;Trp53flox/+;Pdx-1-cre (KPCflox) mice were intraperitoneally injected with 40 mg/kg Indox (n = 9) or a vehicle (n = 10) twice a week. At the end point, tumor status including proliferation, direct invasion, and distant metastasis was analyzed histopathologically. The inhibitory potentials of Indox for proliferation, migration/invasion, and the phosphorylation of target molecules were determined in KPCflox-derived PDAC cells in vitro. RESULTS: Prolonged survival by Indox via intraperitoneal administration was observed in the KPCflox mice. Indox inhibited tumor proliferation accompanied with low levels of nuclear phosphorylated cyclin-dependent kinase (p-CDK) and cyclin B1 in vivo. Furthermore, Indox inhibited the migration/invasive activities of PDAC via down-regulation of matrix metalloproteinase (MMP)-9 in vitro and in vivo. Antibody array and immunoblotting analysis revealed that Indox inhibited the phosphorylation of multiple molecules, including key upstream proteins of MMP-9 in RAF/extracellular signal-regulated kinase (ERK), AKT, and stress-activated protein kinase/c-Jun-N-terminal kinase (SAPK/JNK) pathways. CONCLUSION: Indox inhibited the proliferative, invasive, and metastatic potentials of PDAC in vitro and in vivo. Therefore, Indox could a therapeutic candidate for treating spontaneously occurring PDAC via blocking the RAF/ERK, AKT and SAPK/JNK pathways.
RESUMO
Metallothionein-3 (MT-3), also known as growth inhibitory factor (GIF), was originally identified in the brain. An essential step in elucidating the potential roles of MT-3 is to evaluate its expression levels in organs other than the brain. In this present study, we carried out RT-PCR, Western blot and immunohistochemical analyses to quantify MT-3 mRNA and its protein in the cerebrum, eye, heart, kidney, liver, prostate, testis, tongue, and muscle in male Wistar rats. MT-3 mRNA was detected in the cerebrum, the dorsolateral lobe of the prostate, testis, and tongue. Using a monoclonal anti-MT-3 antibody, we detected MT-3 in the cerebrum, the dorsolateral lobe of the prostate, testis, and tongue as a single band on an immunoblot. Immunohistochemical staining showed MT-3 in some astrocytes in the deep cortex, ependymal cells, and choroidal cells in the cerebrum. MT-3 was also detected in some cells of the glomerulus and the collective tubules in the kidney, some cells in the glandular epithelium of the dorsolateral lobe of the prostate, some Sertoli cells and Lydig cells in the testis, and taste bud cells in the tongue. Although MT-3 immunopositivity was obviously demonstrated in the kidney by the immnunohistochemical method, the expression of MT-3 was not fully detectable by RT-PCR and Western blot analyses. Interestingly, only a subset of cells showed positivity for MT-3, not all cells in all tissues. The localization of MT-3 in peripheral organs outside the brain suggests that MT-3 has roles in these tissues besides its role in growth inhibition of neurites.
Assuntos
Encéfalo/enzimologia , Células Epiteliais/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Vísceras/enzimologia , Animais , Astrócitos/citologia , Astrócitos/metabolismo , Western Blotting , Encéfalo/anatomia & histologia , Plexo Corióideo/citologia , Plexo Corióideo/metabolismo , Epêndima/citologia , Epêndima/metabolismo , Células Epiteliais/citologia , Inibidores do Crescimento/metabolismo , Imuno-Histoquímica , Rim/citologia , Rim/metabolismo , Masculino , Metalotioneína 3 , Proteínas do Tecido Nervoso/análise , Proteínas do Tecido Nervoso/genética , Especificidade de Órgãos , Próstata/citologia , Próstata/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Papilas Gustativas/citologia , Papilas Gustativas/metabolismo , Testículo/citologia , Testículo/metabolismo , Língua/citologia , Língua/metabolismo , Vísceras/citologiaRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with a poor prognosis. To identify potential effective therapeutic drugs for PDAC, we established a screening system based on spheroid formation using 170#3 mouse PDAC cells with or without fibroblasts. We found that indirubin 3'-oxime (Indox) and 5-methoxyindirubin 3'-oxime (5MeOIndox) inhibited PDAC cell proliferation. Furthermore, PDAC xenograft growth was also inhibited in BALB/c nu/nu mice after administration of Indox and 5MeOIndox. Both phosphorylated CDK1 and cyclin B1 levels in 170#3 cells were significantly reduced by treatment with Indox and 5MeOIndox in vitro and in vivo. Cell cycle analysis revealed that 5MeOIndox, but not Indox, induced G2/M arrest. Annexin V-propidium iodide double-staining analysis demonstrated that Indox induced abundant non-apoptotic cell death of 170#3 cells, while 5MeOIndox predominantly induced early apoptosis, indicating that the cytotoxicity of 5MeOIndox is lower than that of Indox. These results suggest that one mechanism of 5MeOIndox is to induce G2/M arrest of PDAC cells via inhibition of CDK1/cyclin B1 levels, thereby leading to apoptosis. Our findings suggest 5MeOIndox as a potential useful anticancer agent in PDAC.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Ductal Pancreático/tratamento farmacológico , Indóis/farmacologia , Oximas/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Animais , Proteína Quinase CDC2 , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Ciclina B1/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Relação Dose-Resposta a Droga , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Fosforilação , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Quorum-sensing systems have been reported to play a critical role in the pathogenesis of several bacterial infections. Recent data have demonstrated that Pseudomonas N-3-oxododecanoyl-L-homoserine lactone (3-oxo-C12-homoserine lactone, 3-oxo-C12-HSL), but not N-butanoyl-L-homoserine lactone (C4-HSL), induces apoptosis in macrophages and neutrophils. In the present study, the effects of active immunization with 3-oxo-C12-HSL-carrier protein conjugate on acute P. aeruginosa lung infection in mice were investigated. Immunization with 3-oxo-C12-HSL-BSA conjugate (subcutaneous, four times, at 2-week intervals) elaborated significant amounts of specific antibody in serum. Control and immunized mice were intranasally challenged with approximately 3 x 10(6) c.f.u. P. aeruginosa PAO1, and survival was then compared. All control mice died by day 2 post bacterial challenge, while 36 % of immunized mice survived to day 4 (P<0.05). Interestingly, bacterial numbers in the lungs did not differ between control and immunized groups, whereas the levels of pulmonary tumour necrosis factor (TNF)-alpha in the immunized mice were significantly lower than those of control mice (P<0.05). Furthermore, the extractable 3-oxo-C12-HSL levels in serum and lung homogenate were also significantly diminished in the immunized mice. Immune serum completely rescued reduction of cell viability by 3-oxo-C12-HSL-mediated apoptosis in macrophages in vitro. These results demonstrated that specific antibody to 3-oxo-C12-HSL plays a protective role in acute P. aeruginosa infection, probably through blocking of host inflammatory responses, without altering lung bacterial burden. The present data identify a promising potential vaccine strategy targeting bacterial quorum-sensing molecules, including autoinducers.
Assuntos
4-Butirolactona/análogos & derivados , Homosserina/análogos & derivados , Pneumonia Bacteriana/prevenção & controle , Infecções por Pseudomonas/prevenção & controle , Pseudomonas aeruginosa , Soroalbumina Bovina/administração & dosagem , Vacinação , 4-Butirolactona/administração & dosagem , 4-Butirolactona/análise , 4-Butirolactona/imunologia , Animais , Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular , Contagem de Colônia Microbiana , Homosserina/administração & dosagem , Homosserina/análise , Homosserina/imunologia , Soros Imunes/farmacologia , Injeções Subcutâneas , Pulmão/metabolismo , Pulmão/microbiologia , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos BALB C , Pneumonia Bacteriana/sangue , Pneumonia Bacteriana/metabolismo , Infecções por Pseudomonas/sangue , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/imunologia , Pseudomonas aeruginosa/isolamento & purificação , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/metabolismo , Vacinas Conjugadas/administração & dosagem , Vacinas SintéticasRESUMO
Ban-Lan-Gen is the common name for the dried roots of indigo plants, including Polygonum tinctorium, Isatis indigotica, Isatis tinctoria, and Strobilanthes cusia. Ban-Lan-Gen is frequently used as an anti-inflammatory and an anti-viral for the treatment of hepatitis, influenza, and various types of inflammation. One of the cytochrome P450 (CYP) enzymes, CYP3A4, is responsible for the metabolism of a wide variety of xenobiotics, including an estimated 60% of all clinically used drugs. In this study, we investigated the effect of Ban-Lan-Gen on the transcriptional activation of the CYP3A4 gene. Ban-Lan-Gen extract increased CYP3A4 gene reporter activity in a dose-dependent manner. Indirubin, one of the biologically active ingredients in the Ban-Lan-Gen, also dose-dependently increased CYP3A4 gene reporter activity. Expression of short hairpin RNA for the human pregnane X receptor (hPXR-shRNA) inhibited CYP3A4 gene reporter activity, and overexpression of human PXR increased indirubin- and rifampicin-induced CYP3A4 gene reporter activity. Furthermore, indirubin induced CYP3A4 mRNA expression in HepG2 cells. Taken together, these results indicate that indirubin, a component of Ban-Lan-Gen, activated CYP3A4 gene transcription through the activation of the human PXR.
Assuntos
Citocromo P-450 CYP3A/genética , Receptores de Esteroides/metabolismo , Ativação Transcricional/efeitos dos fármacos , Citocromo P-450 CYP3A/metabolismo , Relação Dose-Resposta a Droga , Células Hep G2 , Humanos , Indóis/farmacologia , Receptor de Pregnano X , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Relação Estrutura-Atividade , Ativação Transcricional/genéticaRESUMO
Organocatalysis and cooperative catalysis are fast growing research areas. In these fields, cinchona alkaloids and their derivatives play a major role. The ready availability of both pseudo-enantiomeric pairs from natural sources makes them outstanding catalysts for reactions including oxidation, alkylation, cycloaddition, and carbene insertion. This short review focuses on recent achievements in the field of asymmetric carbon-nitrogen atom bond formation reactions using cinchona alkaloids and their derivatives.