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1.
Anticancer Res ; 39(10): 5437-5448, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570438

RESUMO

BACKGROUND/AIM: Epithelial-mesenchymal transition (EMT) is a key multi-step process which enables cancer cells to detach from the epithelial primary tumor mass and allows them to metastasize to distant organs. We immunohistochemically analyzed the expression of the transcription factors (TWIST-1, SLUG, ZEB1, ZEB2) and components of the extracellular matrix (laminin-5, fibronectin) which influence the EMT. MATERIALS AND METHODS: Primary human breast (MDA-MB-231), colon (HT29, HCT116), ovarian (SKOV3, OVCAR3) and head and neck squamous cell carcinoma cell lines (UTSCC2, UTSCC24A) grown as xenografts were immunohistochemically analyzed in vitro and in vivo. RESULTS: A high SLUG expression was observed in every cancer entity both in vitro and in vivo. ZEB1 and ZEB2 showed a high in vivo expression especially in SKOV3 and in in vitro grown MDA-MB-231 cells. CONCLUSION: SLUG expression showed the highest expression in all cancer entities investigated. Hence, it presumably represents the master regulator of EMT in these metastatic tumor entities.


Assuntos
Biomarcadores Tumorais/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Fatores de Transcrição/metabolismo , Linhagem Celular Tumoral , Matriz Extracelular/metabolismo , Células HCT116 , Células HT29 , Humanos , Imuno-Histoquímica/métodos , Homeobox 2 de Ligação a E-box com Dedos de Zinco/metabolismo , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo
2.
Anticancer Res ; 39(10): 5473-5481, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31570441

RESUMO

BACKGROUND/AIM: Aerial parts and seeds of the neem tree (Azadirachta indica) have long been used in traditional medicine such as Ayurveda for health-related purposes. Our interest in neem bioactives lies in their potential use as standalone anticancer agents, or as adjuvants to standard therapy. The aim of the present study was to explore a supercritical CO2 extract (SCNE) of neem leaf and a prominent liminoid in neem leaf, nimbolide, for epigenetic activity. MATERIALS AND METHODS: Human colorectal cancer cell lines (HCT116 and HT29) were cultured for 48 h in the presence of neem extract or nimbolide and evaluated for growth inhibition and evidence of suppression of histone deacetylation and DNA methylation. RESULTS: Both SCNE and nimbolide suppressed the proliferation of colon cancer cells by inducing epigenetic modifications. CONCLUSION: Neem leaf contains bioactive constituents which modify epigenetic activity.


Assuntos
Azadirachta/química , Neoplasias Colorretais/tratamento farmacológico , Epigênese Genética/efeitos dos fármacos , Extratos Vegetais/farmacologia , Folhas de Planta/química , Dióxido de Carbono/química , Dióxido de Carbono/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Metilação de DNA/efeitos dos fármacos , Células HCT116 , Células HT29 , Humanos , Limoninas/farmacologia
3.
Chem Biol Interact ; 310: 108734, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31276661

RESUMO

This work aimed to evaluate the mechanisms involved in the apoptosis induction of isorhamnetin-3-O-glucosyl-pentoside (IGP) in metastatic human colon cancer cells (HT-29). To achieve this, we assessed phosphatidylserine (PS) exposure, cell membrane disruption, chromatin condensation, cell cycle alterations, mitochondrial damage, ROS production, and caspase-dependence on cell death. Our results showed that IGP induced cell death on HT-29 cells through PS exposure (48%) and membrane permeabilization (30%) as well as nuclear condensation (54%) compared with control cells. Moreover, IGP treatment induced cell cycle arrest in G2/M phase. Bax/Bcl-2 ratio increased and the loss of mitochondrial membrane potential (63%) was observed in IGP-treated cells. Finally, as apoptosis is a caspase-dependent cell death mechanism, we used a pancaspase-inhibitor (Q-VD-OPh) to demonstrate that the cell death induced by IGP was caspase-dependent. Overall these results indicated that IGP induced apoptosis through caspase-dependent mitochondrial damage in HT-29 colon cancer cells.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Glicosídeos/farmacologia , Mitocôndrias/efeitos dos fármacos , Opuntia/química , Quercetina/análogos & derivados , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Neoplasias do Colo/patologia , Flavonóis , Glicosídeos/isolamento & purificação , Glicosídeos/uso terapêutico , Células HT29 , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/patologia , Extratos Vegetais/farmacologia , Quercetina/isolamento & purificação , Quercetina/farmacologia , Quercetina/uso terapêutico
4.
Chem Commun (Camb) ; 55(65): 9649-9652, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31339160

RESUMO

Intracellular delivery of bioactive polyphenols is currently evaluated as a protective strategy for cells under pharmaceutical stress. To this end, the 20mer R5 peptide from the marine diatom C. fusiformis was N-terminally modified with a quercetin derivative. This polyphenol-peptide conjugate was used to generate homogeneous silica particles under biomimetic conditions that are efficiently taken up by eukaryotic cells without being cytotoxic. However, not only was accumulation in the cytoplasm of living cells observed via electron and fluorescence microscopy but also translocation into the nucleus. The latter was only seen when the quercetin-peptide conjugate was present within the silica particles and provides a novel targeting option for silica particles to nuclei.


Assuntos
Núcleo Celular/metabolismo , Corantes Fluorescentes/farmacocinética , Fragmentos de Peptídeos/farmacocinética , Quercetina/análogos & derivados , Quercetina/farmacocinética , Dióxido de Silício/farmacocinética , Transporte Ativo do Núcleo Celular , Biomimética , Diatomáceas/química , Corantes Fluorescentes/síntese química , Corantes Fluorescentes/química , Corantes Fluorescentes/toxicidade , Células HT29 , Humanos , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/toxicidade , Quercetina/síntese química , Quercetina/toxicidade , Dióxido de Silício/química , Dióxido de Silício/toxicidade
5.
Anticancer Res ; 39(7): 3815-3822, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262909

RESUMO

BACKGROUND/AIM: Fatty acid synthase (FASN) provides palmitate for cell membrane formation in colorectal cancer (CRC) cells, however, palmitate is also available in the blood of CRC patients. The aim of this study was to examine whether orlistat, a FASN inhibitor, is able to attenuate CRC cell growth despite the availability of extracellular palmitate. MATERIALS AND METHODS: Palmitate concentrations were measured in serum from CRC patients and healthy controls. HT-29 CRC cells were treated with orlistat and palmitate. RESULTS: Treatment of CRC cells with orlistat caused a dose-dependent inhibition of cell proliferation. In turn, delivery of extracellular palmitate at doses lower than those found in the serum of CRC patients reversed inhibition by orlistat concentrations of up to 10 µM. CONCLUSION: Inhibition of CRC cell proliferation by orlistat is reversed by palmitate which is present at high levels in the serum. Therefore, orlistat may be effective in vivo only at high concentrations.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Orlistate/farmacologia , Palmitatos/sangue , Adulto , Idoso , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/sangue , Ácido Graxo Sintase Tipo I/antagonistas & inibidores , Ácido Graxo Sintase Tipo I/genética , Feminino , Células HT29 , Humanos , Masculino , Pessoa de Meia-Idade
6.
Zhongguo Zhong Yao Za Zhi ; 44(11): 2278-2282, 2019 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-31359654

RESUMO

Fourteen chemical constituents, including 5-hydroxy-4-methoxy-1-tetralone(1), 4,8-dihydroxy-1-tetralone(2), 4,5-dihydroxy-α-tetralone(3), blumenol B(4), dehydrovomifoliol(5), megastigm-5-ene-3,9-diol(6), juglanin B(7), blumenol C(8), loliolide(9), oleracone B(10), syringarsinol(11), pinoresinol(12), methyl 4-hydroxy-3-methoxybenzoate(13), and isovanillic acid(14), were isolated from the dichloromethane fraction of 95% methanol extract of green walnut husks by silica gel and MCI column chromatography, and Pre-HPLC. Their structures were determined by spectroscopic methods, such as NMR, MS and so on. Among them, compounds 1, 4-6, 8-13 were isolated from the green walnut husks for the first time, and compounds 4-6, 8, 10, 12, 13 were isolated from the Juglans genus for the first time. All of isolates were detected their inhibitory activities against HeLa, HGC-27 and Ht-29 cell lines by the MTT assay. The result showed that compounds 2, 3, 7, 9 and 11 exhibited inhibitory activity against the tested cell line. The IC_(50) of 7 were 26.5, 9.0, 25.4 µmol·L~(-1), respectively.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Juglans/química , Compostos Fitoquímicos/farmacologia , Extratos Vegetais/química , Antineoplásicos Fitogênicos/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Células HT29 , Células HeLa , Humanos , Estrutura Molecular , Compostos Fitoquímicos/isolamento & purificação
7.
Int J Oncol ; 55(1): 223-242, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31180528

RESUMO

The aim of this study was to examine the effects of 5­fluorouracil (5­FU), anti­epidermal growth factor receptor (EGFR) antibody and aspirin (ASA) on the characteristics of two CRC cell lines, HCT116 and HT29, maintained in a spherical culture system. We observed that the morphology of both the HCT116 and HT29 cell­derived spheres was significantly impaired and the size of the colonospheres was markedly reduced following treatment with the aforementioned three drugs. In contrast to adherent cultures, the spherical cultures were more resistant to the tested drugs, as was reflected by their capacity to re­create the colonospheres when sustained in serum­free medium. Flow cytometric analysis of the drug­treated HCT116 cell­derived spheres revealed changes in the fraction of cells expressing markers of cancer stem cells (CSCs), whereas the CSC phenotype of HT29 cell­derived colonospheres was affected to a lesser extent. All reagents enhanced the percentage of non­viable cells in the colonospheres despite the diminished fraction of active caspase­3­positive cells following treatment of the HT29 cell­derived spheres with anti­EGFR antibody. Increased autophagy, assessed by acridine orange staining, was noted following the incubation of the HT29­colonospheres with ASA and 5­FU in comparison to the control. Notably, the percentage of cyclooxygenase (COX)­2­positive cells was not affected by ASA, although its activity was markedly elevated in the colonospheres incubated with anti­EGFR antibody. On the whole, the findings of this study indicate that all the tested drugs were involved in different cellular processes, which suggests that they should be considered for the combined therapeutic treatment of CRC, particularly for targeting the population of CSC­like cells. Thus, cancer cell­derived spheres may be used as a preferable model for in vitro anticancer drug testing.


Assuntos
Anticorpos Monoclonais/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Aspirina/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Fluoruracila/farmacologia , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Aspirina/administração & dosagem , Autofagia/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/metabolismo , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/imunologia , Proteína Ligante Fas/biossíntese , Fluoruracila/administração & dosagem , Células HCT116 , Células HT29 , Humanos , Esferoides Celulares , Receptor fas/biossíntese
8.
Chem Biol Interact ; 310: 108711, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31207224

RESUMO

Gastrointestinal infections are considered a serious public health problem in view of their high incidence and the increasing antibiotic resistance of the microorganisms involved in their pathogenesis, namely Escherichia coli. Consequently, finding new ways to prevent and/or threat these infections is urgent. In this study we investigated whether a well-characterised polyphenolic red wine extract is able to inhibit the cytotoxic effects induced by a clinical pathogenic Escherichia coli strain (E. coli 270) against HT-29 colon epithelial cells. Firstly, we provide evidences showing that the E. coli strain triggered the death of the intestinal epithelial cells through the production and release of a toxin. Then we support that, in a concentration dependent way, RWE through both, a direct interaction with bacterial exotoxin and the epithelial cells, prevented the action of the toxin on the cells, significantly reducing cell death. This intends to constitute a position paper as detailed mechanisms for the inhibition of E. coli-induced toxicity by polyphenols are yet to be completely unraveled. However, considering that the amount of red wine polyphenols used can be easily achieved in a normal diet, this study suggests that RWE may provide a readily available dietary product with potential benefit for the prevention and/or treatment of intestinal infections induced by intestinal pathogenic bacteria and may open new therapeutic avenues for the development of potential natural antimicrobial agents.


Assuntos
Infecções por Escherichia coli/prevenção & controle , Gastroenteropatias/prevenção & controle , Polifenóis/uso terapêutico , Vinho , Morte Celular/efeitos dos fármacos , Células Epiteliais/patologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/patogenicidade , Infecções por Escherichia coli/tratamento farmacológico , Gastroenteropatias/tratamento farmacológico , Gastroenteropatias/microbiologia , Células HT29 , Humanos , Polifenóis/farmacologia
9.
Int J Nanomedicine ; 14: 3893-3909, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31239663

RESUMO

Background: Photothermal and chemotherapy treatment has been frequently studied for cancer therapy; however, chemotherapy is equally toxic to both normal and cancer cells. The clinical application value of most kinds of photothermal transforming agents remains limited, due to their poor degradation and minimal accumulation in tumors. Materials and methods: We reported the synthesis of photothermal transforming agents (MoS2) and chemotherapeutic (doxorubicin, DOX) co-loaded electrospun nanofibers using blend electrospinning for the treatment of postoperative tumor recurrence. Results: Under the irradiation of an 808 nm laser, the as-prepared chitosan/polyvinyl alcohol/MoS2/DOX nanofibers showed an admirable photothermal conversion capability with a photothermal conversion efficiency of 23.2%. These composite nanofibers are in vitro and in vivo biocompatible. In addition, they could control the sustained release of DOX and the generated heat can sensitize the chemotherapeutic efficacy of DOX via enhancing its release rate. Their chemo-/photothermal combined therapy efficiency was systematically studied in vitro and in vivo. Instead of circulating with the body fluid, MoS2 was trapped by the nanofibrous matrix in the tumor and so its tumor-killing ability was not compromised, thus rendering this composite nanofiber a promising alternative for future clinical translation within biomedical application fields. Conclusion: Chitosan/polyvinyl alcohol/MoS2/DOX nanofibers showed an excellent photothermal conversion capability with a photothermal conversion efficiency of 23.2% and can completely inhibit the postoperative tumor reoccurrence.


Assuntos
Dissulfetos/química , Doxorrubicina/uso terapêutico , Molibdênio/química , Nanofibras/química , Nanotecnologia/métodos , Neoplasias/terapia , Fototerapia , Animais , Materiais Biocompatíveis/farmacologia , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Reagentes para Ligações Cruzadas/química , Doxorrubicina/farmacologia , Liberação Controlada de Fármacos , Células HT29 , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanofibras/ultraestrutura , Recidiva Local de Neoplasia/patologia , Neoplasias/sangue , Neoplasias/patologia , Neoplasias/cirurgia , Padrões de Referência , Resultado do Tratamento
10.
BMC Complement Altern Med ; 19(1): 114, 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31159791

RESUMO

BACKGROUND: Lactobacillus plantarum, a major species of Lactic Acid Bacteria (LAB), are capable of producing postbiotic metabolites (PM) with prominent probiotic effects that have been documented extensively for rats, poultry and pigs. Despite the emerging evidence of anticancer properties of LAB, very limited information is available on cytotoxic and antiproliferative activity of PM produced by L. plantarum. Therefore, the cytotoxicity of PM produced by six strains of L. plantarum on various cancer and normal cells are yet to be evaluated. METHODS: Postbiotic metabolites (PM) produced by six strains of L. plantarum were determined for their antiproliferative and cytotoxic effects on normal human primary cells, breast, colorectal, cervical, liver and leukemia cancer cell lines via MTT assay, trypan blue exclusion method and BrdU assay. The toxicity of PM was determined for human and various animal red blood cells via haemolytic assay. The cytotoxicity mode was subsequently determined for selected UL4 PM on MCF-7 cells due to its pronounced cytotoxic effect by fluorescent microscopic observation using AO/PI dye reagents and flow cytometric analyses. RESULTS: UL4 PM exhibited the lowest IC50 value on MCF-7, RG14 PM on HT29 and RG11 and RI11 PM on HL60 cell lines, respectively from MTT assay. Moreover, all tested PM did not cause haemolysis of human, dog, rabbit and chicken red blood cells and demonstrated no cytotoxicity on normal breast MCF-10A cells and primary cultured cells including human peripheral blood mononuclear cells, mice splenocytes and thymocytes. Antiproliferation of MCF-7 and HT-29 cells was potently induced by UL4 and RG 14 PM respectively after 72 h of incubation at the concentration of 30% (v/v). Fluorescent microscopic observation and flow cytometric analyses showed that the pronounced cytotoxic effect of UL4 PM on MCF-7 cells was mediated through apoptosis. CONCLUSION: In conclusion, PM produced by the six strains of L. plantarum exhibited selective cytotoxic via antiproliferative effect and induction of apoptosis against malignant cancer cells in a strain-specific and cancer cell type-specific manner whilst sparing the normal cells. This reveals the vast potentials of PM from L. plantarum as functional supplement and as an adjunctive treatment for cancer.


Assuntos
Antineoplásicos/metabolismo , Citotoxinas/metabolismo , Lactobacillus plantarum/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Citotoxinas/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Eritrócitos/efeitos dos fármacos , Células HT29 , Humanos , Células MCF-7 , Probióticos
11.
Eur J Med Chem ; 177: 386-400, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158752

RESUMO

We explored the approach of using an analog of E-64, a well-known and hydrophilic cysteine cathepsin (CC) inhibitor, as a potent cysteine cathepsin-trapping agent (CCTA) to improve the tumor retention of low-molecular-weight, receptor-targeted radiopharmaceuticals. The synthesized hydrophilic CCTA-incorporated, NTSR1-targeted agents demonstrated a substantial increase in cellular retention upon uptake into the NTRS1-positive HT-29 human colon cancer cell line. Similarly, biodistribution studies using HT-29 xenograft mice revealed a significant and substantial increase in tumor retention for the CCTA-incorporated, NTSR1-targeted agent. The intracellular trapping mechanism of the CCTA-incorporated agents by macromolecular adduct formation was confirmed using multiple in vitro and in vivo techniques. Furthermore, utilization of the more hydrophilic CCTA greatly increased the hydrophilicity of the resulting NTSR1-targeted constructs leading to substantial decreases in most non-target tissues in contrast to our previously reported dipeptidyl acyloxymethyl ketone (AOMK) constructs. This work further confirms that the CCTA trapping approach can make significant improvements in the clinical potential of NTSR1-and other receptor-targeted radiopharmaceuticals.


Assuntos
Catepsinas/antagonistas & inibidores , Inibidores Enzimáticos/metabolismo , Peptídeos/metabolismo , Compostos Radiofarmacêuticos/metabolismo , Receptores de Neurotensina/metabolismo , Animais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Feminino , Células HT29 , Humanos , Interações Hidrofóbicas e Hidrofílicas , Lutécio/química , Camundongos SCID , Neoplasias/diagnóstico , Peptídeos/síntese química , Peptídeos/química , Radioisótopos/química , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/química , Distribuição Tecidual
12.
Cancer Sci ; 110(8): 2396-2407, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31148360

RESUMO

The tumor microenvironment offers favorable conditions for tumor progression, and activated fibroblasts, known as cancer-associated fibroblasts, play a pivotal role. TP53-deficient cancer cells are known to induce strong fibroblast activation. We aimed to elucidate the oncogenic role of exosomes derived from TP53-deficient colon cancer cells in fibroblast proliferation and tumor growth. Cancer cell-derived exosomes (CDEs) were isolated from the conditioned media of cancer cells using a sequential ultracentrifugation method. The effects of exosomes on tumor growth were evaluated using human cell lines (TP53-WT colon cancer, HCT116; TP53-mutant colon cancer, HT29; and fibroblasts, CCD-18Co and WI-38) and an immune-deficient nude mouse xenograft model. HCT116 (HCT116sh p53 ) cells deficient in TP53 accelerated cocultured fibroblast proliferation compared to TP53-WT HCT116 (HCT116sh control ) cells in vitro. Exosomes from HCT116sh p53 cells suppressed TP53 expression of fibroblasts and promoted their proliferation. Xenografts of HCT116sh p53 cells grew significantly faster than those of HCT116sh control cells in the presence of co-injected fibroblasts, but this difference was diminished by CDE inhibition. Microarray analysis identified upregulation of several microRNAs (miR-1249-5p, miR-6737-5p, and miR-6819-5p) in TP53-deficient CDEs, which were functionally proven to suppress TP53 expression in fibroblasts. Exosomes derived from TP53-mutant HT29 cells also suppressed TP53 expression in fibroblasts and accelerated their growth. The proliferative effect of HT29 on cocultured fibroblasts was diminished by inhibition of these miRNAs in fibroblasts. Our results suggest that CDEs play a pivotal role in tumor progression by fibroblast modification. Cancer cell-derived exosomes might, therefore, represent a novel therapeutic target in colon cancer.


Assuntos
Neoplasias do Colo/genética , Exossomos/genética , MicroRNAs/genética , Proteína Supressora de Tumor p53/genética , Animais , Fibroblastos Associados a Câncer/patologia , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias do Colo/patologia , Progressão da Doença , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Células HT29 , Xenoenxertos/patologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transdução de Sinais/genética , Microambiente Tumoral/genética , Regulação para Cima/genética
13.
Plant Mol Biol ; 101(1-2): 65-79, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31190320

RESUMO

KEY MESSAGE: Overexpression of BoMYB29 gene up-regulates the aliphatic glucosinolate pathway in Brassica oleracea plants increasing the production of the anti-cancer metabolite glucoraphanin, and the toxic and pungent sinigrin. Isothiocyanates, the bio-active hydrolysis products of glucosinolates, naturally produced by several Brassicaceae species, play an important role in human health and agriculture. This study aims at correlating the content of aliphatic glucosinolates to the expression of genes involved in their synthesis in Brassica oleracea, and perform functional analysis of BoMYB29 gene. To this purpose, three genotypes were used: a sprouting broccoli, a cabbage, and a wild genotype (Winspit), a high glucosinolate containing accession. Winspit showed the highest transcript level of BoMYB28, BoMYB29 and BoAOP2 genes, and BoAOP2 expression was positively correlated with that of the two MYB genes. Further analyses of the aliphatic glucosinolates also showed a positive correlation between the expression of BoAOP2 and the production of sinigrin and gluconapin in Winspit. The Winspit BoMYB29 CDS was cloned and overexpressed in Winspit and in the DH AG1012 line. Overexpressing Winspit plants produced higher quantities of alkenyl glucosinolates, such as sinigrin. Conversely, the DH AG1012 transformants showed a higher production of methylsulphinylalkyl glucosinolates, including glucoraphanin, and, despite an up-regulation of the aliphatic glucosinolate genes, no increase in alkenyl glucosinolates. The latter may be explained by the absence of a functional AOP2 gene in DH AG1012. Nevertheless, an extract of DH AG1012 lines overexpressing BoMYB29 provided a chemoprotective effect on human colon cells. This work exemplifies how the genetic diversity of B. oleracea may be used by breeders to select for higher expression of transcription factors for glucosinolate biosynthesis to improve its natural, health-promoting properties.


Assuntos
Brassica/genética , Glucosinolatos/metabolismo , Extratos Vegetais/farmacologia , Fatores de Transcrição/metabolismo , Brassica/química , Brassica/metabolismo , Expressão Gênica , Variação Genética , Células HT29 , Humanos , Imidoésteres/metabolismo , Isotiocianatos/metabolismo , Extratos Vegetais/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Fatores de Transcrição/genética
14.
Fitoterapia ; 137: 104194, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31175954

RESUMO

Four unusual heterodimeric tetrahydroxanthones, usneaxanthones A-D (1-4) were isolated from lichen Usnea aciculifera Vain (Parmeliaceae). Their structures and absolute configurations, particularly the central and axial chiralities, were unambiguously demonstrated by a combination of spectroscopic data (1D, 2D NMR, HRESIMS), electronic circular dichroism (ECD) experiments, and single-crystal X-ray crystallographic analyses. Cytotoxic effects of isolated compounds (1, 2 and 4) were evaluated on HT-29 human colorectal cancer cells. Compound 4 showed potent cytotoxicity against HT-29 with IC50 values of 2.41 µM.


Assuntos
Antineoplásicos/farmacologia , Usnea/química , Xantonas/farmacologia , Antineoplásicos/isolamento & purificação , Células HT29 , Humanos , Estrutura Molecular , Vietnã , Xantonas/isolamento & purificação
15.
World J Microbiol Biotechnol ; 35(6): 85, 2019 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-31134456

RESUMO

Surface properties like hydrophobicity, aggregation ability, adhesion to mucosal surfaces and epithelial cells and transit time are key features for the characterization of probiotic strains. In this study, we used two Lactobacillus paracasei subsp. paracasei strains (BGNJ1-64 and BGSJ2-8) strains which were previously described with very strong aggregation capacity. The aggregation promoting factor (AggLb) expressed in these strains showed high level of binding to collagen and fibronectin, components of extracellular matrix. The working hypothesis was that strains able to aggregate have an advantage to resist in intestinal tract. So, we assessed whether these strains and their derivatives (without aggLb gene) are able to bind or not to intestinal components and we compared the transit time of each strains in mice. In that purpose parental strains (BGNJ1-64 and BGSJ2-8) and their aggregation negative derivatives (BGNJ1-641 and BGSJ2-83) were marked with double antibiotic resistance in order to be tracked in in vivo experiments in mice. Comparative analysis of binding ability of WT and aggregation negative strains to different human intestinal cell lines and mucin revealed no significant difference among them, excluding involvement of AggLb in interaction with surface of intestinal cells and mucin. In vivo experiments showed that surviving and transit time of marked strains in mice did not drastically depend on the presence of the AggLb aggregation factor.


Assuntos
Moléculas de Adesão Celular/metabolismo , Células Epiteliais/microbiologia , Intestinos/microbiologia , Lactobacillus paracasei/crescimento & desenvolvimento , Lactobacillus paracasei/fisiologia , Ligação Proteica , Animais , Aderência Bacteriana/fisiologia , Células CACO-2 , Moléculas de Adesão Celular/fisiologia , Colágeno/metabolismo , Fibronectinas/metabolismo , Células HT29 , Interações entre Hospedeiro e Microrganismos/fisiologia , Humanos , Interações Hidrofóbicas e Hidrofílicas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mucinas/metabolismo , Probióticos , Análise de Onda de Pulso , Propriedades de Superfície
16.
Cell Physiol Biochem ; 52(6): 1503-1516, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31112017

RESUMO

BACKGROUND/AIMS: Zinc Finger Protein 281 (ZNF281) was recently identified as a novel oncogene in several human carcinomas. However, the clinical significance of ZNF281 in colorectal cancer (CRC) and the molecular mechanisms by which ZNF281 promotes the growth and metastasis of CRC remain unknown. METHODS: ZNF281 expression in CRC tissues was assessed, and the outcomes were analyzed to determine the clinical importance of ZNF281 expression. Cell Transwell assays and a wound healing assay were performed to assess the effects of ZNF281 on CRC cell migration and invasion in vitro. Western blotting was applied to analyze the potential mechanisms. RESULTS: ZNF281 mRNA and protein levels were significantly increased in CRC tissues compared with normal colon tissues, and high ZNF281 expression was associated with advanced T stage, N stage, TNM stage and differentiation. Therefore, ZNF281 expression might be an independent prognostic indicator in CRC patients. Moreover, knockdown of ZNF281 expression suppressed cell proliferation, migration and invasion by inhibiting the Wnt/ß-catenin pathway. CONCLUSION: Our study indicates that ZNF281 plays a critical role in the progression and metastasis of CRC and could represent a potential therapeutic target for CRC.


Assuntos
Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Transativadores/metabolismo , Idoso , Linhagem Celular Tumoral , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Neoplasias Colorretais/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HT29 , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Invasividade Neoplásica/fisiopatologia , Prognóstico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , RNA Interferente Pequeno/genética , Transativadores/antagonistas & inibidores , Transativadores/genética , Regulação para Cima , Via de Sinalização Wnt , beta Catenina/metabolismo
17.
Cell Physiol Biochem ; 52(6): 1517-1534, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31120230

RESUMO

BACKGROUND/AIMS: Cadmium (Cd) is a heavy metal contaminant whose toxicity is associated with colorectal cancer (CRC). However, the underlying molecular mechanisms of Cd-induced CRC malignancy remain obscure. METHODS: A monolayer scratch assay was employed to assess the migration of HT-29 human adenocarcinoma cells. Luciferase reporter assay was used to determine cyclooxygenase-2 (COX-2) transcriptional activity, and Western blotting was used to detect p38 Mitogen Activated Protein Kinase (MAPK) and Akt phosphorylation as well as COX-2 expression. Prostaglandin E2 (PGE2) levels were measured using Enzyme Linked Immunosorbent Assay (ELISA) and reactive oxygen species (ROS) formation was assessed using dihydroethidium (DHE) stain. RESULTS: Here, we show that Cd potentiates the migratory capacity of HT-29 CRC cells. Cd caused a time-dependent increase in COX-2 expression. Celecoxib, a COX-2 selective inhibitor, significantly reduced Cd-induced migration. Cd also increased levels of ROS and phosphorylated p38. Importantly, Cd-induced COX-2 expression and migration were significantly abolished by N-Acetyl-Cysteine (NAC), a ROS scavenger, or SB202190, a specific p38 inhibitor. Furthermore, Cd-induced p38 phosphorylation was inhibited by NAC. Cd (100 nM) also increased PGE2 levels, which was abrogated by NAC, SB202190, or celecoxib. Exogenous PGE2 significantly potentiated cell migration. Cd caused a significant increase in Akt phosphorylation in a ROS-mediated pathway. Moreover, Cd-induced migration was significantly attenuated by LY294 002, a phosphatidylinositol-3-kinase inhibitor. CONCLUSION: Taken together, our results suggest that exposure to low levels of Cd promotes a more migratory cancer phenotype in a ROS-p38-COX-2-PGE2 pathway as well as ROS-Akt pathway. Therefore, COX-2, PGE2 receptors or Akt represent potential targets in the treatment of CRC, particularly in patients exposed to Cd.


Assuntos
Adenocarcinoma/induzido quimicamente , Cádmio/toxicidade , Neoplasias do Colo/induzido quimicamente , Ciclo-Oxigenase 2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Movimento Celular/efeitos dos fármacos , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Ciclo-Oxigenase 2/genética , Ativação Enzimática/efeitos dos fármacos , Células HT29 , Humanos , Ativação Transcricional/efeitos dos fármacos
18.
Molecules ; 24(9)2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31058835

RESUMO

In several European countries, especially in Sweden, the seeds of the species Astragalus boeticus L. were widely used as coffee substitutes during the 19th century. Nonetheless, data regarding the phytochemistry and the pharmacological properties of this species are currently extremely limited. Conversely, other species belonging to the Astragalus genus have already been extensively investigated, as they were used for millennia for treating various diseases, including cancer. The current work was addressed to characterize cycloartane glycosides from A. boeticus, and to evaluate their cytotoxicity towards human colorectal cancer (CRC) cell lines. The isolation of the metabolites was performed by using different chromatographic techniques, while their chemical structures were elucidated by nuclear magnetic resonance (NMR) (1D and 2D techniques) and electrospray-ionization quadrupole time-of-flight (ESI-QTOF) mass spectrometry. The cytotoxic assessment was performed in vitro by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays in Caco-2, HT-29 and HCT-116 CRC cells. As a result, the targeted phytochemical study of A. boeticus enabled the isolation of three new cycloartane glycosides, 6-O-acetyl-3-O-(4-O-malonyl)-ß-d-xylopyranosylcycloastragenol (1), 3-O-(4-O-malonyl)-ß-d-xylopyranosylcycloastragenol (2), 6-O-acetyl-25-O-ß-d-glucopyranosyl-3-O-ß-d-xylopyranosylcycloastragenol (3) along with two known compounds, 6-O-acetyl-3-O-ß-d-xylopyranosylcycloastragenol (4) and 3-O-ß-d-xylopyranosylcycloastragenol (5). Importantly, this work demonstrated that the acetylated cycloartane glycosides 1 and 4 might preferentially inhibit cell growth in the CRC cell model resistant to epidermal growth factor receptor (EGFR) inhibitors.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Astrágalo (Planta)/química , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Glicosídeos/farmacologia , Triterpenos/química , Acilação , Antineoplásicos Fitogênicos/química , Células CACO-2 , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Glicosídeos/química , Células HCT116 , Células HT29 , Humanos , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Extratos Vegetais/química , Espectrometria de Massas por Ionização por Electrospray , Suécia
19.
Cell Prolif ; 52(4): e12632, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31094023

RESUMO

OBJECTIVE: Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) play essential roles in the tumour progression. LncRNAs mostly act as competing endogenous RNAs (ceRNAs) by sponging miRNAs. This study aimed to study the association of a novel lncRNA MFI2-AS1 with miR-574-5p/MYCBP axis in the development of colorectal cancer (CRC). METHODS: Ninety-four CRC tissues and paired adjacent non-tumour tissues were included in our study. The relative expression level of MFI2-AS1 was detected, and its relationship with clinico-pathological factors was analysed. Then, the CRC cells lines (LoVo and RKO) were transfected with MFI2-AS1 siRNA, miR-574-5p mimics and inhibitors. Cell proliferation, migration, invasion, cell cycle distribution and DNA damage in response to different transfection conditions were examined. Dual-luciferase reporter assay was performed to identify the target interactions between MFI2-AS1 and miR-574-5p, miR-574-5p and MYCBP. RESULTS: LncRNA MFI2-AS1 and MYCBP were up-regulated in CRC tissues when compared with adjacent non-tumour tissues. The expression levels of MFI2-AS1 were significantly associated with tumour histological grade, lymph and distant metastasis, TNM stage and vascular invasion. Both MFI2-AS1 siRNA and miR-574-5p mimics inhibited proliferation, migration and invasion in LoVo and RKO cells. The transfection of miR-574-5p inhibitor showed MFI2-AS1 siRNA-induced changes in CRC cells. Dual-luciferase reporter assay revealed target interactions between MFI2-AS1 and miR-574-5p, miR-574-5p and MYCBP. CONCLUSIONS: These findings suggested that lncRNA MFI2-AS1 and MYCBP have promoting effects in CRC tissues. LncRNA MFI2-AS1 promoted CRC cell proliferation, migration and invasion through activating MYCBP and by sponging miR-574-5p.


Assuntos
Movimento Celular/genética , Proliferação de Células/genética , Neoplasias Colorretais/genética , Proteínas de Ligação a DNA/genética , MicroRNAs/genética , Invasividade Neoplásica/genética , RNA Longo não Codificante/genética , Fatores de Transcrição/genética , Linhagem Celular , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Células HEK293 , Células HT29 , Humanos , Invasividade Neoplásica/patologia , RNA Interferente Pequeno/genética , Regulação para Cima/genética
20.
Chem Biodivers ; 16(7): e1900170, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31134745

RESUMO

The tumor-suppressor function of p53 makes it an attractive drug target. Efforts were mostly put on stabilization of the functional p53 or reactivation of mutated p53. Previous studies have shown that small molecules targeting Loop1/Sheet3 (L1/S3) can reactivate the R175H-p53 and stabilize p53 in vitro. Since the L1/S3 pocket is shared by the mutate and the wild type (WT) p53, virtual screening is introduced to identify natural products targeting the L1/S3 of WT p53. Considering the high flexibility of Loop1, ensemble docking method is utilized for different clusters of the L1/S3. Seven conformations were chosen for docking. As one of the 181 selected candidates, torilin not only improved p53 activity, but also increased p21 protein expression level, which lies downstream of p53, therefore suppressing HCT116 cancer cell growth. Torilin may covalently bind to Cys124 of p53 by 2-methyl-2-butenal (2M2B) group, as torilin derivatives, which do not contain the 2M2B group, were not able to increase the p53 transcription activity. In conclusion, this study demonstrated that L1/S3 of WT-p53 is a druggable pocket, and torilin has a potential cytotoxicity through activating the p53 pathway.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Descoberta de Drogas , Simulação de Acoplamento Molecular , Proteína Supressora de Tumor p53/antagonistas & inibidores , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Células HCT116 , Células HT29 , Humanos , Conformação Molecular , Simulação de Dinâmica Molecular , Relação Estrutura-Atividade , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/metabolismo
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