Antioxidant and Hypolipidemic Activity of Açai Fruit Makes It a Valuable Functional Food

Several plant extracts are acquiring increasing value because of their antioxidant activity and hypolipidemic properties. Among them, great interest has been recently paid to acai fruit as a functional food. The aim of this study was to test the ability of acai extract in reducing oxidative stress and modulating lipid metabolism in vitro using different cell models and different types of stress. In fact, lipid peroxidation as evaluated in a HepG2 model was reduced five-fold when using 0.25 mu g/mL of extract, and it was further reduced (20-fold) with the concentration increase up to 2.5 mu g/mL. With the nonalcoholic fatty liver disease (NAFLD)in vitro model, all concentrations tested showed at least a two-fold reduced fat deposit. In addition, primary adipocytes challenged with TNF-alpha under hypoxic conditions to mimic the persistent subcutaneous fat, treated with acai extract showed an approximately 40% reduction of fat deposit. Overall, our results show that acai is able to counteract oxidative states in all the cell models analysed and to prevent the accumulation of lipid droplets. No toxic effects and high stability overtime were highlighted at the concentrations tested. Therefore, acai can be considered a suitable support in the prevention of different alterations of lipid and oxidative metabolism responsible for fat deposition and metabolic pathological conditions

Characterization and gene expression of cancer stem cells grown as sarcospheres from human primary sarcomas

This study aimed to identify, isolate and characterize cancer stem cells from human primary sarcomas. We performed cytometric analyses for stemness and differentiation antigens including CD29, CD34, CD44, CD90, CD117 and CD133 on 21 human primary sarcomas on the day of surgery. From sarcoma biopsies, we obtained two chondrosarcoma stabilized cell lines and two osteosarcoma stabilized cell lines on which spheres formation, side population profile, stemness gene expression and in vivo and in vitro assays were performed. On a chondrosarcoma cell line, the whole genoma microarray analyses were performed (sarcospheres versus adherent cells). All samples expressed the CD133, CD44 and CD29 markers. We selected a CD133+ subpopulation from stabilized cell lines that displayed the capacity to grow as sarcospheres able to initiate and sustain tumour growth in NOD/SCID mice, to express stemness genes including OCT3/4, Nanog, Sox2 and Nestin and to differentiate into mesenchymal lineages. Microarray analyses pointed out a huge gene expression difference between sarcospheres and adherent cells. About 2000 genes, including ones related to cell cycle, stemness and epigenetic regulation, resulted to be very differentially expressed in the two population considered. With signed ratio of 2,806 genes were over-expressed and 1,029 under-expressed on sarcospheres when compared with adherent cells. The most highly overexpressed gene were thioredoxin interacting protein (fold +25) that modulates the cellular redox state, growth differentiation factor 15 (fold +10), member of the TGF–β superfamily , histone cluster 1, H2ad (fold +9) and solute carrier family 2 member 14 that facilitate glucose transport (fold +8,9). Our findings evidence the existence of cancer stem cells in human primary bone sarcomas and highlight CD133 as pivotal marker for identification of these cells. This may be of primary importance in the development of new therapeutic strategies and new prognostic procedures against these highly aggressive and metastatic tumours

The role of CD133 in the identification and characterisation of tumour-initiating cells in non-small-cell lung cancer☆☆☆

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Cytotoxic Potential of the Marine Diatom Thalassiosira rotula: Insights into Bioactivity of 24-Methylene Cholesterol

Marine microalgae are receiving great interest as sustainable sources of bioactive metabolites for health, nutrition and personal care. In the present study, a bioassay-guided screening allowed identifying an enriched fraction from SPE separation of the methanolic extract of the marine diatom Thalassiosira rotula with a chemically heterogeneous composition of cytotoxic molecules, including PUFAs, the terpene phytol, the carotenoid fucoxanthin and the phytosterol 24-methylene cholesterol (24-MChol). In particular, this latter was the object of deep investigation aimed to gain insight into the mechanisms of action activated in two tumour cell models recognised as resistant to chemical treatments, the breast MCF7 and the lung A549 cell lines. The results of our studies revealed that 24-MChol, in line with the most studied β-sitosterol (β-SIT), showed cytotoxic activity in a 3-30 µM range of concentration involving the induction of apoptosis and cell cycle arrest, although differences emerged between the two sterols and the two cancer systems when specific targets were investigated (caspase-3, caspase-9, FAS and TRAIL)

Cancer stem cells in head and neck tumors: evidence for metastatic spread and treatment resistance

The major challenge in the management of patients with oral squamous cell carcinomas (OSCC) is the development of resistance to therapy leading to disseminated disease. Since cancer stem cells (CSC) have emerged as important players in OSCC metastasis, our objectives were to explore the implications of CSC in OSCC tumor progression, invasion and response to conventional therapies. Methods: A panel of well-characterized cell lines originated from the most common sites in the head and neck area was used. Cells were cultured as floating spheres or under normal adherent conditions and analyzed for CD44, ALDH, CD24, CD29, CD56 by flow cytometry, PCR arrays for genes related to stemness, metastasis and EMT . We also investigated sLeX expression, known to play a key-role in many cancers metastasis by promoting tumor cells binding to endothelial E-selectin. We analyzed the tumorigenic potential of OSCC cells by invasion assays and in vivo OSCC experimental models comparatively to CSC cells. Moreover resistance to cisplatin and radiation was assessed by annexin V/PI assay and by colony forming assay. Results: The highest levels of sLeX expression were found in cell lines originated from oral cavity (9%-47%) compared to other head and neck locations (0.1%-7%). Cells grown as spheres were 95-100% positive for sLeX compared to 10-40% of adherent counterpart. Although sLeX+ and sLeX- cells were both able to form spheres, sLeX+ spheres were predominant and larger. Flow cytometry and PCR arrays indicated that the spheres were highly enriched in CSC and metastatic markers. Consistently, the spheres showed increased invasive and tumorigenic potential, and resistance to conventional chemotherapy and radiations. Conclusion: these studies are the first to unveil a novel link between sLeX expression, stem cell formation and metastatic spread in OSCC, and provide supportive evidence for CSC resistance to treatment. Understanding the mechanisms of tumor invasion and metastasis will improve patient outcome and survival

Methylation and epigenetic modification by 5’ azacytidine and valproic acid treatment increase stemness attributes in bone sarcoma cell lines

Bone sarcoma is an aggressive malignancy with high mortality rate. Despite recent advances, the prognosis is still extremely poor. Bone sarcomas contain a small cell population with stem cell like properties, referred to as cancer stem cells (CSCs) expressing CD133 (Tirino et al, 2009; 2011). The biological relevance and regulatory mechanism of CD133 expression are not yet understood. The aim of this study is to elucidate mechanisms regulating aberrant expression of CD133 and stemness phenotype. Saos-2, MG63 and BS15 cell lines were treated with 0,5 mM valproic acid (VPA) and 3μM 5’azacytidine (5-AZA) for 48 hours alone and in combination. CD133 and stemness markers expression including OCT4, Sox2 and Nanog were analyzed by flow cytometry and real-time PCR. Vimentin and osteocalcin levels were also tested. Sarcospheres formation rate was assessed as spheres number/seed single cell number. After treatment with 5-AZA or VPA, the expression level of CD133 mRNA as well as of protein was significantly increased in all three cell lines. Also OCT4, Sox2 and Nanog, stemness markers, and vimentin, mesenchymal marker resulted to be upregulated after treatment by real time-PCR. On the contrary, the expression level of osteocalcin remained similar before and after treatment. Interestingly, combined treatment with 5-AZA and VPA induced an increase of CD133 expression in a synergistic manner in all three cell lines. In addition, sarcospheres formation rate was increased after drug treatment compared to untreated cells. Also in this case, the drug combination lead to synergistic increase of formation rate of spheres. In conclusion, our results indicate that DNA methylation is an important determinant of CD133 and stemness profile in human bone sarcomas and this mechanism may be associated with histone deacetilase inhibition

Hyaluronan-Based Gel Promotes Human Dental Pulp Stem Cells Bone Differentiation by Activating YAP/TAZ Pathway.

Background: Hyaluronans exist in different forms, accordingly with molecular weight and degree of crosslinking. Here, we tested the capability to induce osteogenic differentiation in hDPSCs (human dental pulp stem cells) of three hyaluronans forms: linear pharmaceutical-grade hyaluronans at high and (HHA) low molecular weight (LHA) and hybrid cooperative complexes (HCC), containing both sizes. Methods: hDPSCs were treated with HHA, LHA, HCC for 7, 14 and 21 days. The effects of hyaluronans on osteogenic differentiation were evaluated by qRT-PCR and WB of osteogenic markers and by Alizarin Red S staining. To identify the involved pathway, CD44 was analyzed by immunofluorescence, and YAP/TAZ expression was measured by qRT-PCR. Moreover, YAP/TAZ inhibitor-1 was used, and the loss of function of YAP/TAZ was evaluated by qRT-PCR, WB and immunofluorescence. Results: We showed that all hyaluronans improves osteogenesis. Among these, HCC is the main inducer of osteogenesis, along with overexpression of bone related markers and upregulating CD44. We also found that this biological process is subordinate to the activation of YAP/TAZ pathway. Conclusions: We found that HA's molecular weight can have a relevant impact on HA performance for bone regeneration, and we unveil a new molecular mechanism by which HA acts on stem cells. Keywords: YAP/TAZ pathway; dental pulp stem cells; hyaluronic acid; osteogenic differentiation