Marine Actinomycetes-Derived Secondary Metabolites Overcome TRAIL-Resistance via the Intrinsic Pathway through Downregulation of Survivin and XIAP.

Resistance of cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis represents the major hurdle to the clinical use of TRAIL or its derivatives. The discovery and development of lead compounds able to sensitize tumor cells to TRAIL-induced cell death is thus likely to overcome this limitation. We recently reported that marine actinomycetes' crude extracts could restore TRAIL sensitivity of the MDA-MB-231 resistant triple negative breast cancer cell line. We demonstrate in this study, that purified secondary metabolites originating from distinct marine actinomycetes (sharkquinone (1), resistomycin (2), undecylprodigiosin (3), butylcyclopentylprodigiosin (4), elloxizanone A (5) and B (6), carboxyexfoliazone (7), and exfoliazone (8)), alone, and in a concentration-dependent manner, induce killing in both MDA-MB-231 and HCT116 cell lines. Combined with TRAIL, these compounds displayed additive to synergistic apoptotic activity in the Jurkat, HCT116 and MDA-MB-231 cell lines. Mechanistically, these secondary metabolites induced and enhanced procaspase-10, -8, -9 and -3 activation leading to an increase in PARP and lamin A/C cleavage. Apoptosis induced by these compounds was blocked by the pan-caspase inhibitor QvD, but not by a deficiency in caspase-8, FADD or TRAIL agonist receptors. Activation of the intrinsic pathway, on the other hand, is likely to explain both their ability to trigger cell death and to restore sensitivity to TRAIL, as it was evidenced that these compounds could induce the downregulation of XIAP and survivin. Our data further highlight that compounds derived from marine sources may lead to novel anti-cancer drug discovery.

An overview of neuroblastoma cell lineage phenotypes and in vitro models.

This review was conducted to present the main neuroblastoma (NB) clinical characteristics and the most common genetic alterations present in these pediatric tumors, highlighting their impact in tumor cell aggressiveness behavior, including metastatic development and treatment resistance, and patients' prognosis. The distinct three NB cell lineage phenotypes, S-type, N-type, and I-type, which are characterized by unique cell surface markers and gene expression patterns, are also reviewed. Finally, an overview of the most used NB cell lines currently available for in vitro studies and their unique cellular and molecular characteristics, which should be taken into account for the selection of the most appropriate model for NB pre-clinical studies, is presented. These valuable models can be complemented by the generation of NB reprogrammed tumor cells or organoids, derived directly from patients' tumor specimens, in the direction toward personalized medicine.

Effect of crystalline phases of titania nanotube arrays on adipose derived stem cell adhesion and proliferation.

The aim of this work was to evaluate the cellular response to titanium nanotube arrays with variable crystalline structure. Cytotoxicity, viability and the ability of the titania nanotube arrays to stimulate adhesion and proliferation of adipose derived stem cells (ADSCs) was evaluated. Titania nanotube arrays were fabricated by electrochemical anodization of titanium in diethyleneglycol/hydrofluoric acid electrolyte at 60 V for 6 h, then annealed at 300, 530 and 630 °C for 5 h. The nanotube arrays were characterized using scanning electron microscopy (SEM), contact angle goniometry, x-ray diffraction (XRD) and protein adsorption. ADSCs were cultured on titania nanotube arrays at a density of 1 × 104 cells/ml. The cells were allowed to adhere and to proliferate for 1, 4 and 7 days. Cell viability was characterized by the CellTiter-Blue® Cell Viability Assay; and cell morphology was characterized by SEM. Cell adhesion, proliferation and morphology were characterized using fluorescence microscopy by staining the cells with DAPI and rhodamine/phalloidin. The results from this study showed that the annealing at 300 and 530 °C formed anatase phase, and annealing at 630 °C formed anatase/rutile phase. The results indicated that the modification of the crystalline structure (i.e. anatase/rutile phase) of titania nanotube arrays influenced the ADSC adhesion and proliferation. Future studies are now directed towards evaluating differentiation of this cellular model in osteoblasts.

Antitumoral effects of Amblyomma sculptum Berlese saliva in neuroblastoma cell lines involve cytoskeletal deconstruction and cell cycle arrest.

The antitumor properties of ticks salivary gland extracts or recombinant proteins have been reported recently, but little is known about the antitumor properties of the secreted components of saliva. The goal of this study was to investigate the in vitro effect of the saliva of the hard tick Amblyomma sculptum on neuroblastoma cell lines. SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32, and CHLA-20 cells were susceptible to saliva, with 80% reduction in their viability compared to untreated controls, as demonstrated by the methylene blue assay. Further investigation using CHLA-20 revealed apoptosis, with approximately 30% of annexin-V positive cells, and G0/G1-phase accumulation (>60%) after treatment with saliva. Mitochondrial membrane potential (Δψm) was slightly, but significantly (p < 0.05), reduced and the actin cytoskeleton was disarranged, as indicated by fluorescent microscopy. The viability of human fibroblast (HFF-1 cells) used as a non-tumoral control decreased by approximately 40%. However, no alterations in cell cycle progression, morphology, and Δψm were observed in these cells. The present work provides new perspectives for the characterization of the molecules present in saliva and their antitumor properties.

In-vitro cell adhesion and proliferation of adipose derived stem cell on hydroxyapatite composite surfaces.

The goal of this work was to enhance the mechanical strength and fracture toughness of brittle hydroxyapatite (HAP) by reinforcing it with nanocomposites such as graphene oxide (GO), carbon nanotubes (CNT) and Titania. The goal was also to evaluate the cytotoxicity and the cellular adhesion/proliferation of these composites. The composites were characterized for their crystallinity, functionality, morphology and mechanical properties. Altering the composition by adding 1wt% GO and CNT significantly altered the wettability, hardness and roughness. Further, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FITR) and X-ray photoelectron spectroscopy (XPS) results confirm the crystal structure, bulk chemical composition and surface elemental composition respectively of the composites. The bulk hardness of HAP with CNT was significantly higher than that of HAP. The wettability of HAP with GO was significantly lower than that of HAP with GO and Titania. Adipose Derived Stem Cells (ADSCs) were used for this study to evaluate cytotoxicity and viability. HAP with CNT and HAP with CNT and Titania were found to be least cytotoxic compared to other composites as evaluated by Lactate Dehydrogenase (LDH) assay and alamarBlue assay. ADSC adhesion and proliferation was investigated after 1, 4 and 7days of culture using fluorescence microscopy. All the composites nurtured ADSC adhesion and proliferation, however, distinct morphological changes were observed by using Scanning Electron Microscopy (SEM). Overall, these composites have the potential to be used as bone graft substitutes.

Antitumoral effects of Amblyomma sculptum Berlese saliva in neuroblastoma cell lines involve cytoskeletal deconstruction and cell cycle arrest / Efeito antitumoral da saliva do carrapato Amblyomma sculptum Berlese em células de neuroblastoma envolve desconstrução do citoesqueleto e parada do ciclo celular

Abstract The antitumor properties of ticks salivary gland extracts or recombinant proteins have been reported recently, but little is known about the antitumor properties of the secreted components of saliva. The goal of this study was to investigate the in vitro effect of the saliva of the hard tick Amblyomma sculptum on neuroblastoma cell lines. SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32, and CHLA-20 cells were susceptible to saliva, with 80% reduction in their viability compared to untreated controls, as demonstrated by the methylene blue assay. Further investigation using CHLA-20 revealed apoptosis, with approximately 30% of annexin-V positive cells, and G0/G1-phase accumulation (>60%) after treatment with saliva. Mitochondrial membrane potential (Δψm) was slightly, but significantly (p < 0.05), reduced and the actin cytoskeleton was disarranged, as indicated by fluorescent microscopy. The viability of human fibroblast (HFF-1 cells) used as a non-tumoral control decreased by approximately 40%. However, no alterations in cell cycle progression, morphology, and Δψm were observed in these cells. The present work provides new perspectives for the characterization of the molecules present in saliva and their antitumor properties.

Resumo As propriedades antitumorais de extratos de glândulas salivares de carrapatos ou proteínas recombinantes foram relatadas recentemente, mas pouco se sabe sobre as propriedades antitumorais dos componentes secretados da saliva. O objetivo deste estudo foi investigar o efeito in vitro da saliva bruta do carrapato duro Amblyomma sculptum sobre as linhagens celulares de neuroblastoma. Células SK-N-SK, SH-SY5Y, Be(2)-M17, IMR-32 e CHLA-20 foram suscetíveis à saliva, com redução de 80% na sua viabilidade em comparação com controles não tratados, como demonstrado pelo ensaio de Azul de Metileno. Investigações posteriores utilizando CHLA-20 revelaram apoptose, com aproximadamente 30% de células positivas para anexina-V, e G0/G1 (> 60%) após tratamento com saliva. O potencial de membrana mitocondrial (Δψm) foi reduzido significativamente (p <0,05), e o citoesqueleto de actina foi desestruturado, como indicado pela microscopia de fluorescência. A viabilidade do fibroblasto humano (células HFF-1), usado como controle não tumoral, diminuiu em aproximadamente 40%. No entanto, não foram observadas alterações na progressão do ciclo celular, morfologia e Δψm nestas células. O presente trabalho fornece novas perspectivas para a caracterização das moléculas presentes na saliva e suas propriedades antitumorais.