RESUMEN

Rutin (RUT) is considered one the most attractive flavonoids from a therapeutic perspective due to its multispectral pharmacological activities including antiradical, anti-inflammatory, antiproliferative, and antimetastatic among others. Still, this compound presents a low bioavailability what narrows its clinical applications. To overcome this inconvenience, the current paper was focused on the synthesis, characterization, and toxicological assessment of two RUT bioconjugates obtained by enzymatic esterification with oleic acid (OA) and linoleic acid (LA)-rutin oleate (RUT-O) and rutin linoleate (RUT-L), as flavonoid precursors with improved physicochemical and biological properties. Following the enzymatic synthesis in the presence of Novozyme® 435, the two bioconjugates were obtained, their formation being confirmed by RAMAN and FT-IR spectroscopy. The in vitro and in ovo toxicological assessment of RUT bioconjugates (1-100 µM) was performed using 2D consecrated cell lines (cardiomyoblasts - H9c2(2-1), hepatocytes-HepaRG, and keratinocytes-HaCaT), 3D reconstructed human epidermis tissue (EpiDerm™), and chick chorioallantoic membranes, respectively. The results obtained were test compound, concentration-and cell-type dependent, as follows: RUT-O reduced the viability of H9c2(2-1), HepaRG, and HaCaT cells at 100 µM (to 77.53%, 83.17%, and 78.32%, respectively), and induced cell rounding and floating, as well as apoptotic-like features in the nuclei of all cell lines, whereas RUT-L exerted no signs of cytotoxicity in all cell lines in terms of cell viability, morphology, and nuclear integrity. Both RUT esters impaired the migration of HepaRG cells (at 25 µM) and lack irritative potential (at 100 µM) in vitro (tissue viability >50%) and in ovo (irritation scores of 0.70 for RUT-O, and 0.49 for RUT-L, respectively). Computational predictions revealed an increased lipophilicity, and reduced solubility, drug-likeness and drug score of RUT-O and RUT-L compared to their parent compounds-RUT, OA, and LA. In conclusion, we report a favorable toxicological profile for RUT-L, while RUT-O is dosage-limited since at high concentrations were noticed cytotoxic effects.

RESUMEN

Photoinduced hyperthermia with nanomaterials has been proven effective in photothermal therapy (PTT) of tumor tissues, but a precise control in PTT requires determination of the molecular-level mechanisms. In this paper, we determined the mechanisms responsible for the action of photoexcited gold shell-isolated nanoparticles (AuSHINs) in reducing the viability of MCF7 (glandular breast cancer) and especially A549 (lung adenocarcinoma) cells in vitro experiments, while the photoinduced damage to healthy cells was much smaller. The photoinduced effects were more significant than using other nanomaterials, and could be explained by the different effects from incorporating AuSHINs on Langmuir monolayers from lipid extracts of tumoral (MCF7 and A549) and healthy cells. The incorporation of AuSHINs caused similar expansion of the Langmuir monolayers, but Fourier-transform infrared spectroscopy (FTIR) data of Langmuir-Schaefer films (LS) indicated distinct levels of penetration into the monolayers. AuSHINs penetrated deeper into the A549 extract monolayers, affecting the vibrational modes of polar groups and carbon chains, while in MCF7 monolayers penetration was limited to the surroundings of the polar groups. Even smaller insertion was observed for monolayers of the healthy cell extract. The photochemical reactions were modulated by AuSHINs penetration, since upon irradiation the surface area of A549 monolayer decreased owing to lipid chain cleavage by oxidative reactions. For MCF7 monolayers, hydroperoxidation under illumination led to a ca. 5% increase in surface area. The monolayers of healthy cell lipid extract were barely affected by irradiation, consistent with the lowest degree of AuSHINs insertion. In summary, efficient photothermal therapy may be devised by producing AuSHINs capable of penetrating the chain region of tumor cell membranes.

Asunto(s)

Oro , Nanopartículas , Membrana Celular , Oro/farmacología , Membranas , Oxidación-Reducción

RESUMEN

Lichens represent an important resource for common traditional medicines due to their numerous metabolites that can exert diverse pharmacological activities including anticancer effects. To find new anticancer compounds with fewer side effects and low tumor resistance, a bioprospective study of Usnea barbata (L.) F.H. Wigg. (U. barbata), a lichen from the Calimani Mountains (Suceava county, Romania) was performed. The aim of this research was to investigate the anticancer potential, morphologic changes, wound healing property, clonogenesis, and oxidative stress biomarker status of four extracts of U. barbata in different solvents (methanol, ethanol, acetone, and ethyl acetate), and also of usnic acid (UA) as a positive control on the CAL-27 (ATCC® CRL-2095™) oral squamous carcinoma (OSCC) cell line and V79 (ATCC® CCL-93™) lung fibroblasts as normal cells. Using the MTT assay and according to IC50 values, it was found that the most potent anticancer property was displayed by acetone and ethyl acetate extracts. All U. barbata extracts determined morphological modifications (losing adhesion capacity, membrane shrinkage, formation of abnormal cellular wrinkles, and vacuolization) with higher intensity in tumor cells than in normal ones. The most intense anti-migration effect was established in the acetone extract treatment. The clonogenic assay showed that some U. barbata extracts decreased the ability of cancer cells to form colonies compared to untreated cells, suggesting a potential anti-tumorigenic property of the tested extracts. Therefore, all the U. barbata extracts manifest anticancer activity of different intensity, based, at least partially, on an imbalance in antioxidant defense mechanisms, causing oxidative stress.

RESUMEN

Mitochondria are essential organelles for healthy eukaryotic cells. They produce energyrich phosphate bond molecules (ATP) through oxidative phosphorylation using ionic gradients. The presence of mitophagy pathways in healthy cells enhances cell protection during mitochondrial damage. The PTEN-induced putative kinase 1 (PINK1)/Parkin-dependent pathway is the most studied for mitophage. In addition, there are other mechanisms leading to mitophagy (FKBP8, NIX, BNIP3, FUNDC1, BCL2L13). Each of these provides tethering of a mitochondrion to an autophagy apparatus via the interaction between receptor proteins (Optineurin, p62, NDP52, NBR1) or the proteins of the outer mitochondrial membrane with ATG9-like proteins (LC3A, LC3B, GABARAP, GABARAPL1, GATE16). Another pathogenesis of mitochondrial damage is mitochondrial depolarization. Reactive oxygen species (ROS) antioxidant responsive elements (AREs) along with antioxidant genes, including pro-autophagic genes, are all involved in mitochondrial depolarization. On the other hand, mammalian Target of Rapamycin Complex 1 (mTORC1) and AMP-dependent kinase (AMPK) are the major regulatory factors modulating mitophagy at the post-translational level. Protein-protein interactions are involved in controlling other mitophagy processes. The objective of the present review is to analyze research findings regarding the main pathways of mitophagy induction, recruitment of the autophagy machinery, and their regulations at the levels of transcription, post-translational modification and protein-protein interaction that appeared to be the main target during the development and maturation of neurodegenerative disorders.

Asunto(s)

Membranas Mitocondriales , Mitofagia , Autofagia , Mitocondrias , Membranas Mitocondriales/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Ubiquitina-Proteína Ligasas

RESUMEN

Amyloid-ß (Aß), a peptide implicated in Alzheimer's disease, was shown to cause specific fragmentation of lamin proteins, which was mediated by an unidentified protease named nuclear scaffold protease (NSP) independently of caspase-6. Because caspase-6 is responsible for the fragmentation process in many other damage-induced apoptosis, here we further investigated possible involvement of caspase-6 in Aß-induced lamin fragmentation under various conditions. We found that lamin A fragment generated by NSP (named fragment b) disappeared in cells incubated with Aß42 for prolonged periods and this product was preserved by a caspase-6 inhibitor. Furthermore, caspase-6 could remove fragment b in nuclei isolated from Aß42-treated cells (ANU). Lamin B in ANU was fragmented by caspase-6 only after treatment with an alkaline phosphatase. The caspase-mediated fragmentation of lamin B was also achieved with nuclei isolated from cells incubated with Aß42 plus a Cdk5 inhibitor. The results indicate that Aß42 induces NSP-mediated fragmentation of lamin A and the following removal process of fragment b by caspase-6 and an Aß-induced phosphorylation prevents the fragmentation of lamin B by caspase-6. The pathway leading to lamin protein fragmentation in this investigation appears to be specific for Aß and thus the data will provide novel insights into the toxicity of the peptide.

RESUMEN

The concept of hormesis includes a biphasic cellular dose-response to a xenobiotic stimulus defined by low dose beneficial and high dose inhibitory or toxic effects. In the present study, an attempt has been made to help elucidate the beneficial and detrimental effects of thymol on different cell types by evaluating and comparing the impact of various thymol doses on cancerous (AGS) and healthy (WS-1) cells. Cytotoxic, genotoxic, and apoptotic effects, as well as levels of reactive oxygen species and glutathione were studied in both cell lines exposed to thymol (0-600 µM) for 24 h. The results showed significant differences in cell viability of AGS compared to WS-1 cells exposed to thymol. The differences observed were statistically significant at all doses applied (P ≤ 0.001) and revealed hormetic thymol effects on WS-1 cells, whereas toxic effects on AGS cells were detectable at all thymol concentrations. Thymol at low concentrations provides antioxidative protection to WS-1 cells in vitro while already inducing toxic effects in AGS cells. In that sense, the findings of the present study suggest that thymol exerts a dose-dependent hormetic impact on different cell types, thereby providing crucial information for future in vivo studies investigating the therapeutic potential of thymol.

Asunto(s)

Adenocarcinoma/tratamiento farmacológico , Proliferación Celular/efectos de los fármacos , Neoplasias Gástricas/tratamiento farmacológico , Timol/farmacología , Adenocarcinoma/patología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Fibroblastos/efectos de los fármacos , Hormesis/efectos de los fármacos , Humanos , Especies Reactivas de Oxígeno/metabolismo , Neoplasias Gástricas/patología

RESUMEN

Electron microscopic study of cardiomyocytes taken from healthy Wistar and OXYS rats and naked mole rats (Heterocephalus glaber) revealed mitochondria in nuclei that lacked part of the nuclear envelope. The direct interaction of mitochondria with nucleoplasm is shown. The statistical analysis of the occurrence of mitochondria in cardiomyocyte nuclei showed that the percentage of nuclei with mitochondria was roughly around 1%, and did not show age and species dependency. Confocal microscopy of normal rat cardiac myocytes revealed a branched mitochondrial network in the vicinity of nuclei with an organization different than that of interfibrillar mitochondria. This mitochondrial network was energetically functional because it carried the membrane potential that responded by oscillatory mode after photodynamic challenge. We suggest that the presence of functional mitochondria in the nucleus is not only a consequence of certain pathologies but rather represents a normal biological phenomenon involved in mitochondrial/nuclear interactions.

Asunto(s)

Núcleo Celular/fisiología , Microscopía Electrónica/métodos , Mitocondrias Cardíacas/fisiología , Membrana Nuclear/fisiología , Animales , Microscopía Confocal , Modelos Animales , Ratas Topo , Ratas , Ratas Wistar

RESUMEN

In this paper, we demonstrate that the antimicrobial activity of L. plantarum PBS067 strain against antagonist microorganisms was mediated by the production of a bacteriocin-like compound secreted at the stationary phase of the growth. The novel bacteriocin-like compound, designed plantaricin P1053, was identified by using sorption-desorption method, butanol extraction and SEC-HPLC. The molecular mass of plantaricin P1053 was shown to be 1053 Da by ESI-MS analysis. Plantaricin P1053 exhibited a broad-spectrum antimicrobial activity against Gram-positive bacteria as S. aureus and Gram-negative bacteria as E. coli. In addition to the antimicrobial activity, the isolated bacteriocin-like compound showed effects on normal and cancerogenic epithelial intestinal cell lines through an enhancing of viability of healthy cells and a proliferation reduction of cancer cells. Moreover, in this paper we demonstrate that the isolated bacteriocin-like compound acts on healthy cells through the epidermal growth factor receptor (EGFR) pathways. In conclusion, plantaricin P1053 isolated from L. plantarum PBS067 strain could represent one of the first multifunctional bacteriocin-like compound acting on human epithelial intestinal cells.

RESUMEN

The cancer stem cell hypothesis has gained currency in recent times but concerns remain about its scientific foundations because of significant gaps that exist between research findings and comprehensive knowledge about cancer stem cells (CSCs). In this light, a mathematical model that considers hematopoietic dynamics in the diseased state of the bone marrow and peripheral blood is proposed and used to address findings about CSCs. The ensuing model, resulting from a modification and refinement of a recent model, develops out of the position that mathematical models of CSC development, that are few at this time, are needed to provide insightful underpinnings for biomedical findings about CSCs as the CSC idea gains traction. Accordingly, the mathematical challenges brought on by the model that mirror general challenges in dealing with nonlinear phenomena are discussed and placed in context. The proposed model describes the logical occurrence of discrete time delays, that by themselves present mathematical challenges, in the evolving cell populations under consideration. Under the challenging circumstances, the steady state properties of the model system of delay differential equations are obtained, analyzed, and the resulting mathematical predictions arising therefrom are interpreted and placed within the framework of findings regarding CSCs. Simulations of the model are carried out by considering various parameter scenarios that reflect different experimental situations involving disease evolution in human hosts. Model analyses and simulations suggest that the emergence of the cancer stem cell population alongside other malignant cells engenders higher dimensions of complexity in the evolution of malignancy in the bone marrow and peripheral blood at the expense of healthy hematopoietic development. The model predicts the evolution of an aberrant environment in which the malignant population particularly in the bone marrow shows tendencies of reaching an uncontrollable equilibrium state. Essentially, the model shows that a structural relationship exists between CSCs and non-stem malignant cells that confers on CSCs the role of temporally enhancing and stimulating the expansion of non-stem malignant cells while also benefitting from increases in their own population and these CSCs may be the main protagonists that drive the ultimate evolution of the uncontrollable equilibrium state of such malignant cells and these may have implications for treatment.

Asunto(s)

Médula Ósea/patología , Modelos Teóricos , Células Madre Neoplásicas/patología , Proliferación Celular , Simulación por Computador , Hematopoyesis , Humanos , Modelos Biológicos

RESUMEN

Patients diagnosed with osteosarcoma are currently treated with intravenous injections of anticancer agents after tumor resection. However, due to remaining neoplastic cells at the site of tumor removal, cancer recurrence often occurs. Successful bone regeneration combined with the control of residual cancer cells presents a challenge for tissue engineering. Cyclodextrins loaded with chemotherapeutic drugs reversibly release the drugs over time. Hydroxyapatite bone biomaterials coated with doxorubicin-loaded cyclodextrin should release the drug with time after implantation directly at the original tumor site and may be a way to eliminate residual neoplastic cells. In the present study, we have carried out in vitro studies to evaluate such a drug-delivery system and have shown that doxorubicin released from cyclodextrin-coated hydroxyapatite retained biological activity and exhibited longer and higher cytotoxic effects on both cancer (osteosarcoma cells) and healthy cells (primary osteoblasts and endothelial cells) compared to biomaterials without cyclodextrin loaded with doxorubicin. Furthermore, doxorubicin released from biomaterials with cyclodextrin moderately induced the expression of tumor suppressor protein p53 whereas p21 expression was similar to control cells. In addition, hypoxic conditions, which occur after implantation until blood-flow to the area is regenerated, protected endothelial cells and primary osteoblasts from doxorubicin-induced cytotoxicity. This chemo-protective effect was far less prominent for the osteosarcoma cells. These findings indicate that a hydroxyapatite-cyclodextrin-doxorubicin chemotherapeutic strategy may enhance the drug-targeting effect on tumor cells while protecting the more sensitive healthy cells for a period of time after implantation. A successful integration of such a drug delivery system might allow healthy cells to initially survive during the doxorubicin exposure period, while eliminating residual neoplastic cells.

Asunto(s)

Antibióticos Antineoplásicos , Neoplasias Óseas/tratamiento farmacológico , Doxorrubicina , Sistemas de Liberación de Medicamentos/métodos , Osteosarcoma/tratamiento farmacológico , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Antibióticos Antineoplásicos/farmacología , Neoplasias Óseas/metabolismo , Neoplasias Óseas/patología , Ciclodextrinas/química , Ciclodextrinas/farmacocinética , Ciclodextrinas/farmacología , Doxorrubicina/química , Doxorrubicina/farmacocinética , Doxorrubicina/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Durapatita/química , Durapatita/farmacocinética , Durapatita/farmacología , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/patología , Humanos , Osteoblastos/metabolismo , Osteoblastos/patología , Osteosarcoma/metabolismo , Osteosarcoma/patología , Cuidados Posoperatorios/métodos

RESUMEN

SCOPE: Searching for correlations between dietary polyphenols and risk of chronic diseases has been a challenge due to the lack of quantitative evaluation methods of long-term exposure. We previously observed substantial DNA methylation changes in human cancer cells upon treatment with polyphenols of the stilbenoid class. When induced in normal cells, such molecular changes may persist and reflect chronic exposure. METHODS AND RESULTS: Illumina 450K microarray is used to delineate a genome wide DNA methylation landscape in MCF10A human immortalized mammary epithelial cells exposed to resveratrol (RSV) at noncytotoxic 15 µM dose for 9 days. Subtle alterations are observed suggesting remodeling of DNA methylation patterns rather than switch on/off changes. Using pyrosequencing, DNA methylation is quantitatively measured at eight CpG sites located within KCNJ4, RNF169, BCHE, DAOA, HOXA9, RUNX3, KRTAP2-1, and TAGAP, upon exposure to RSV or pterostilbene and shows similar differences induced by both stilbenoids. Two of the probes, Runx3 and Kcnj4, are successfully verified in whole blood DNA from healthy rats on diets supplemented with stilbenoids. CONCLUSIONS: The study provides strong support for testing the utility of polyphenol-mediated changes in DNA methylation as quantitative measures of long-term dietary exposures in nutritional epidemiology and clinical trials.