Showing papers by "Maria Teresa Cruz published in 2017"

Dendritic cell-based immunotherapy: a basic review and recent advances

Abstract: Dendritic cells (DCs) are considered a very promising arm to activate the immune system in immunotherapeutic strategies against cancer. DCs are the most powerful antigen-presenting cells (APCs), being highly efficient at generating robust immune responses. They are also considered the center of the immune system, since they provide a crucial link between both innate and adaptive immune responses. Thus, DC-based cancer immunotherapy aims to take advantage of these unique characteristics of DCs to better fight cancer. During the last decade, they have been the subject of numerous studies intending to develop immunotherapeutic strategies against cancer through vaccination. For this purpose, it is essential to gain a better insight into DC immunobiology, regulation of innate and adaptive immune systems, and tumor microenvironment, as well as applying the latest advances in science in order to boost their enormous anti-tumor immunotherapeutic potential. In this review, we will hold focus on DC immunobiology (from their origin, location, and special properties and distinct subsets to the innate and adaptive immunity), on the new concept of cancer immunoediting, and on the knowledge given by clinical trials using DC vaccines. Finally, future perspectives for this emerging field are highlighted.

Valorization of Lipids from Gracilaria sp. through Lipidomics and Decoding of Antiproliferative and Anti-Inflammatory Activity.

Abstract: The lipidome of the red seaweed Gracilaria sp., cultivated on land-based integrated multitrophic aquaculture (IMTA) system, was assessed for the first time using hydrophilic interaction liquid chromatography-mass spectrometry and tandem mass spectrometry (HILIC-MS and MS/MS). One hundred and forty-seven molecular species were identified in the lipidome of the Gracilaria genus and distributed between the glycolipids classes monogalactosyl diacylglyceride (MGDG), digalactosyl diacylglyceride (DGDG), sulfoquinovosyl monoacylglyceride (SQMG), sulfoquinovosyl diacylglyceride (SQDG), the phospholipids phosphatidylcholine (PC), lyso-PC, phosphatidylglycerol (PG), lyso-PG, phosphatidylinositol (PI), phosphatidylethanolamine (PE), phosphatic acid (PA), inositolphosphoceramide (IPC), and betaine lipids monoacylglyceryl- and diacylglyceryl-N,N,N-trimethyl homoserine (MGTS and DGTS). Antiproliferative and anti-inflammatory effects promoted by lipid extract of Gracilaria sp. were evaluated by monitoring cell viability in human cancer lines and by using murine macrophages, respectively. The lipid extract decreased cell viability of human T-47D breast cancer cells and of 5637 human bladder cancer cells (estimated half-maximal inhibitory concentration (IC50) of 12.2 μg/mL and 12.9 μg/mL, respectively) and inhibited the production of nitric oxide (NO) evoked by the Toll-like receptor 4 agonist lipopolysaccharide (LPS) on the macrophage cell line RAW 264.7 (35% inhibition at a concentration of 100 μg/mL). These findings contribute to increase the ranking in the value-chain of Gracilaria sp. biomass cultivated under controlled conditions on IMTA systems.

Antioxidant and anti-inflammatory activities of Geranium robertianum L. decoctions

Abstract: Geranium robertianum L., commonly known as Herb Robert, is an herbaceous plant popularly known for its functional properties including antioxidant and anti-inflammatory activities. In this study, the phenolic profile of leaf and stem decoctions of Geranium robertianum L. was elucidated by UHPLC-DAD-ESI-MSn analysis, and their antioxidant and anti-inflammatory potentials were assessed in vitro. Importantly, and envisaging the use of these extracts in human diets, the potential toxicity of bioactive concentrations was also addressed in macrophages and hepatocytes. Both extracts revealed high amounts of ellagitannins, although a slight prevalence of these compounds was observed in that originating from leaves. High radical scavenging activities against DPPH˙, ABTS˙+ and OH˙ were observed either for the leaf or the stem extract, as well as good activities towards ferric reducing antioxidant power, lipid peroxidation and oxygen radical absorbance capacity. In addition, both extracts were very effective at scavenging NO˙, as measured in a chemical model, while only the stem extract was able to decrease the production of this radical by stimulated macrophages. On the other hand, none of the extracts was able to modulate the activity of lipoxygenase or the expression of the inducible nitric oxide synthase. Overall, these data allowed us to conclude that G. robertianum L. stem and leaf decoctions are particularly rich in tannins. The strong scavenging effects displayed by the stem extract suggest that its anti-inflammatory activity may partially result from its anti-radical capacities towards NO˙.

Antioxidant, Anti-Inflammatory, and Analgesic Activities of Agrimonia eupatoria L. Infusion

Abstract: Agrimony (Agrimonia eupatoria L.) (Ae) is used in traditional medicine to treat inflammatory and oxidative related diseases. Therefore, this study focuses on the anti-inflammatory and analgesic potential of Ae infusion (AeI). Phenolic compounds characterization was achieved by HPLC-PDA-ESI/MS n . To evaluate antioxidant potential, 2,2-diphenyl-1-picrylhydrazyl (DPPH), superoxide anion, hydroxyl radical, and SNAP assays were used. In vitro anti-inflammatory activity of AeI was investigated in LPS-stimulated macrophages by measuring the NO production. In vivo anti-inflammatory activity was validated using the mouse carrageenan-induced paw edema model. Peripheral and central analgesic potential was evaluated using the acetic acid-induced writhing and hot-plate tests, respectively, as well as the formalin assay to assess both activities. The safety profile was disclosed in vitro and in vivo, using MTT and hematoxylin assays, respectively. Vitexin, quercetin O-galloyl-hexoside, and kaempferol O-acetyl-hexosyl-rhamnoside were referred to in this species for the first time. AeI and mainly AePF (Ae polyphenolic fraction) showed a significant antiradical activity against all tested radicals. Both AeI and AePF decreased NO levels in vitro, AePF being more active than AeI. In vivo anti-inflammatory and analgesic activities were verified for both samples at concentrations devoid of toxicity. Agrimony infusion and, mainly, AePF are potential sources of antiradical and anti-inflammatory polyphenols.

Urolithins impair cell proliferation, arrest the cell cycle and induce apoptosis in UMUC3 bladder cancer cells

Abstract: Ellagitannins have been gaining attention as potential anticancer molecules. However, the low bioavailability of ellagitannins and their extensive metabolization in the gastrointestinal tract into ellagic acid and urolithins suggest that the health benefits of consuming ellagitannins rely on the direct effects of their metabolites. Recently, chemopreventive and chemotherapeutic activities were ascribed to urolithins. Nonetheless, there is still a need to screen and evaluate the selectivity of these molecules and to elucidate their cellular mechanisms of action. Therefore, this work focused on the antiproliferative effects of urolithins A, B and C and ellagic acid on different human tumor cell lines. The evaluation of cell viability and the determination of the half-maximal inhibitory concentrations indicated that the sensitivity to the studied urolithins varied markedly between the different cell lines, with the bladder cancer cells (UMUC3) being the most susceptible. In UMUC3 cells, urolithin A was the most active molecule, promoting cell cycle arrest at the G2/M checkpoint, increasing apoptotic cell death and inhibiting PI3K/Akt and MAPK signaling. Overall, the present study emphasizes the chemopreventive/chemotherapeutic potential of urolithins, highlighting the stronger effects of urolithin A and its potential to target transitional bladder cancer cells.

Lipophilic Fraction of Cultivated Bifurcaria bifurcata R. Ross: Detailed Composition and In Vitro Prospection of Current Challenging Bioactive Properties.

Abstract: Macroalgae have been seen as an alternative source of molecules with promising bioactivities to use in the prevention and treatment of current lifestyle diseases. In this vein, the lipophilic fraction of short-term (three weeks) cultivated Bifurcaria bifurcata was characterized in detail by gas chromatography–mass spectrometry (GC-MS). B. bifurcata dichloromethane extract was composed mainly by diterpenes (1892.78 ± 133.97 mg kg−1 dry weight (DW)), followed by fatty acids, both saturated (550.35 ± 15.67 mg kg−1 DW) and unsaturated (397.06 ± 18.44 mg kg−1 DW). Considerable amounts of sterols, namely fucosterol (317.68 ± 26.11 mg kg−1 DW) were also found. In vitro tests demonstrated that the B. bifurcata lipophilic extract show antioxidant, anti-inflammatory and antibacterial activities (against both Gram-positive and Gram-negative bacteria), using low extract concentrations (in the order of µg mL−1). Enhancement of antibiotic activity of drug families of major clinical importance was observed by the use of B. bifurcata extract. This enhancement of antibiotic activity depends on the microbial strain and on the antibiotic. This work represents the first detailed phytochemical study of the lipophilic extract of B. bifurcata and is, therefore, an important contribution for the valorization of B. bifurcata macroalgae, with promising applications in functional foods, nutraceutical, cosmetic and biomedical fields.

Inflammasome in Dendritic Cells Immunobiology: Implications to Diseases and Therapeutic Strategies.

Abstract: Background: An intricate interplay between innate and adaptive immune cells is crucial for an effective immune response during disease, infection and vaccination. This interplay is mainly performed by dendritic cells (DCs), which are professional antigen presenting cells with unparalleled capacity to translate innate to adaptive immunity. They effectively recognize and uptake antigens, migrate to lymphoid tissues, and activate naive T-cells. Indeed, DCs have numerous germline encoded pattern recognition receptors (PRR) that recognize conserved pathogen associated molecular patterns (PAMPs) or danger associated molecular patterns (DAMPs). While some PRRs like Toll-like receptors (TLRs) recognize PAMPs and DAMPs at the cell surface and in endosomal/lysosomal compartments, others, such as NOD-like receptors (NLRs), act as cytosolic sensors. NLRs activation through recognition of PAMPs and DAMPs leads to the assembly of signaling multimeric protein complexes named inflammasomes. Inflammasomes are important regulators of caspase 1, the enzyme responsible for the proteolytically cleavage of precursors’ pro-IL-1β and pro-IL-18 into their active form. Objective: To unveil how inflammasomes are related to maturation, migration, antigen presenting function and DCs ability to fine tune adaptive immune responses. Conclusion: Several studies show that in danger/infectious scenarios NLR and TLR synergize to expand DCs maturation, migration, antigen presenting function and adaptive immune system activation. However, in the absence of a danger scenario, and without TLR engagement, inflammasome activation stimulates an immunosuppressive profile on DCs. Overall, it is clear from literature that activation of the inflammasome in DCs should not be viewed in isolation but rather considering its interconnections with the various PPRdriven pathways. Due to the increasing evidences of inflammasome involvement in multiple inflammatory and immune diseases, this information is of utmost importance since precise inflammasome pharmacological targeting could lead to considerable clinical utility through fine-tuned targeted therapies.

Contact dermatitis: in pursuit of sensitizer's molecular targets through proteomics.

Abstract: Protein haptenation, i.e., the modification of proteins by small reactive chemicals, is the key step in the sensitization phase of allergic contact dermatitis (ACD). Despite the research effort in past decades, the identification of immunogenic hapten-protein complexes that trigger a relevant pathogenic immune response in ACD, as well as the haptenation reaction molecular site, and the elements of a potentially conditioning environment during each of these stages, remain poorly understood. These questions led us to employ a proteomics-based approach to identify modified proteins in the dendritic-like cell line THP-1 sensitized with fluorescein isothiocyanate (FITC), through a combination of 2D-gel electrophoresis, nano-LC and mass spectrometry. A specific set of 39 targeted proteins was identified and comprised proteins from various cellular locations and biological functions. One of FITC targets was identified as MLK, a member of the mixed-lineage kinase family known to act as a mitogen-activated protein kinase kinase kinase and to control the activity of specific mitogen-activated protein kinase pathways, namely p38 and JNK pathways. Haptenated in the vicinity of its active site, our results point to MLK being a relevant target due to a consistent non-activation at early time points of these pathways upon FITC sensitization in THP-1 cells. Moreover, FITC pre-treatment significantly decrease phospho-p38 and phospho-JNK levels induced upon exposure to a classical activator such as lipopolysaccharide or to the sensitizer 2,4-dinitrofluorobenzene. Overall, our data point to specific amino acid residues haptenation within critical proteins as the key step in the subsequent signaling pathways modulation responsible for DC activation and maturation events.

Chemical composition of Laurencia obtusa extract and isolation of a new C15-acetogenin

Abstract: A new C15-acetogenin, sagonenyne (20), exhibiting an unusual single tetrahydropyran ring was isolated from an ethyl acetate extract of Laurencia obtusa collected on the Corsican coastline. Its structure was established by detailed NMR spectroscopic analysis, mass spectrometry, and comparison with literature data. Twenty-three known compounds were identified in the same extract by means of column chromatography steps, using a 13C-NMR computer aided method developed in our laboratory. In addition to sesquiterpenes, which represent the main chemical class of this extract, diterpenes, sterols, and C15-acetogenins were identified. The crude extract was submitted to a cytotoxicity assay and was particularly active against THP-1 cells, a human leukemia monocytic cell line.

In Vitro Dendritic Cell-Based Test for Skin Sensitizers Identification and Potency Estimation

Abstract: Ethical and economic concerns and the recent European legislative framework triggered intensive research efforts towards the development and validation of alternative methods. We developed an in vitro predictive test to assess the skin sensitizing potential of chemicals. The test was based on the analysis and integration of intracellular signalling pathways evoked by chemicals concomitantly with gene expression modulation in skin-derived dendritic cells. In a first approach, cells were treated with four known sensitizers and two non-sensitizers, and the effects on the expression of 20 candidate genes and on the activation of several signalling pathways were analysed by real-time RT-PCR and Western blot, respectively. The genes Trxr1, Hmox1, Nqo1 and Cxcl10 and the signalling pathways p38 MAPK and JNK were identified as good predictor variables and used to construct a dichotomous classifier. For validation of the model, a large number of chemicals were afterwards analysed in a blinded assay. From the total 18 compounds tested, 17 were correctly classified, representing a concordance of 94%, with a sensitivity of 92% and a specificity of 100%. Additionally, we also analysed the feasibility to predict the sensitizer’s potency using in vitro-generated data and several in silico-calculated descriptors. A strong correlation with LLNA EC3 values was obtained (Pearson correlation coefficient r = 0.85, p < 0.001, n = 12). Overall, our results indicate that the analysis of proposed gene and signalling pathway signatures in a mouse skin-derived dendritic cell line represents a valuable model to be integrated in a future in vitro test platform.