Dietary polyphenols and their relationship to the modulation of non-communicable chronic diseases and epigenetic mechanisms: A mini-review.

Chronic Non-Communicable Diseases (NCDs) have been considered a global health problem, characterized as diseases of multiple factors, which are developed throughout life, and regardless of genetics as a risk factor of important relevance, the increase in mortality attributed to the disease to environmental factors and the lifestyle one leads. Although the reactive species (ROS/RNS) are necessary for several physiological processes, their overproduction is directly related to the pathogenesis and aggravation of NCDs. In contrast, dietary polyphenols have been widely associated with minimizing oxidative stress and inflammation. In addition to their antioxidant power, polyphenols have also drawn attention for being able to modulate both gene expression and modify epigenetic alterations, suggesting an essential involvement in the prevention and/or development of some pathologies. Therefore, this review briefly explained the mechanisms in the development of some NCDs, followed by a summary of some evidence related to the interaction of polyphenols in oxidative stress, as well as the modulation of epigenetic mechanisms involved in the management of NCDs.

Neomenthol prevents the proliferation of skin cancer cells by restraining tubulin polymerization and hyaluronidase activity.

Introduction: Neomenthol, a cyclic monoterpenoid, is a stereoisomer of menthol present in the essential oil of Mentha spp. It is used in food as a flavoring agent, in cosmetics and medicines because of its cooling effects. However, neomenthol has not been much explored for its anticancer potential. Additionally, targeting hyaluronidase, Cathepsin-D, and ODC by phytochemicals is amongst the efficient approach for cancer prevention and/or treatment. Objectives: To investigate the molecular and cell target-based antiproliferative potential of neomenthol on human cancer (A431, PC-3, K562, A549, FaDu, MDA-MB-231, COLO-205, MCF-7, and WRL-68) and normal (HEK-293) cell lines. Methods: The potency of neomenthol was evaluated on human cancer and normal cell line using SRB, NRU and MTT assays. The molecular target based study of neomenthol was carried out in cell-free and cell-based test systems. Further, the potency of neomenthol was confirmed by quantitative real-time PCR analysis and molecular docking studies. The in vivo anticancer potential of neomenthol was performed on mice EAC model and the toxicity examination was accomplished through in silico, ex vivo and in vivo approaches. Results: Neomenthol exhibits a promising activity (IC50 17.3 ± 6.49 µM) against human epidermoid carcinoma (A431) cells by arresting the G2/M phase and increasing the number of sub-diploid cells. It significantly inhibits hyaluronidase activity (IC50 12.81 ± 0.01 µM) and affects the tubulin polymerization. The expression analysis and molecular docking studies support the in vitro molecular and cell target based results. Neomenthol prevents EAC tumor formation by 58.84% and inhibits hyaluronidase activity up to 10% at 75 mg/kg bw, i.p. dose. The oral dose of 1000 mg/kg bw was found safe in acute oral toxicity studies. Conclusion: Neomenthol delayed the growth of skin carcinoma cells by inhibiting the tubulin polymerization and hyaluronidase activity, which are responsible for tumor growth, metastasis, and angiogenesis.

The in vitro assessment of a novel vaping technology.

We have developed a novel vaping product (NVP) IS1.0(TT), which utilises a stainless-steel mesh to transfer and vaporise the e-liquid, mitigating some of the potential sources of toxicants that can be generated using the more traditional 'wick and coil' approach. The emissions from IS1.0(TT) have previously been found to have lower levels of toxicants overall when directly compared with a commercial wick and coil e-cig. This current study assessed the toxicological responses to aerosols from this NVP. Responses induced by IS1.0(TT)were compared to those from a 3R4F reference cigarette, using in vitro test methods which included regulatory genetic toxicological assays as well as some more contemporary screening approaches. The experimental conditions were designed to facilitate the testing of aerosol from this vaping product at doses that in most cases greatly exceeded those of the 3R4F comparator showed little to no toxicological responses and demonstrated significantly reduced effects in these in vitro assays when compared to 3R4F. Furthermore, the extreme doses tested in the present study indicate that the toxicant profile of this NVP translates to lower biological activity in vitro, and suggests that the absolute risk hazard level associated with electronic cigarettes can be reduced through continuous improvement as the technology evolves.

Convenient methodology for extraction and subsequent selective propagation of mouse melanocytes in culture from adult mouse skin tissue.

Mouse melanoma B16-BL6 cells are useful cells for cancer metastatic studies. To understand the metastatic principle at molecular levels, it is necessary to carry out experiments in which cancer cells and their normal counterparts are compared. However, unlike normal human melanocytes, preparation of normal mouse melanocytes is quite difficult due to the lack of marketing and insufficient information on an established protocol for primary culture of mouse melanocytes. In this study, we aimed to establish a convenient method for primary culture of mouse melanocytes on the basis of the protocol for human melanocytes. The main obstacles to preparing pure mouse melanocytes are how to digest mouse skin tissue and how to reduce the contamination of keratinocytes and fibroblasts. The obstacles were overcome by collagenase digestion for skin specimens, short time trypsinization for separating melanocytes and keratinocytes, and use of 12-O-Tetradecanoylphorbol 13-acetate (TPA) and cholera toxin in the culture medium. These supplements act to prevent the proliferation of keratinocytes and fibroblasts, respectively. The convenient procedure enabled us to prepare a pure culture of normal mouse melanocytes. Using enriched normal mouse melanocytes and cancerous B16-BL6 cells, we compared the expression levels of melanoma cell adhesion molecule (MCAM), an important membrane protein for melanoma metastasis, in the cells. The results showed markedly higher expression of MCAM in B16-BL6 cells than in normal mouse melanocytes.

Comparative study of anti-inflammatory activity and qualitative-quantitative composition of triterpenoids from ten genotypes of Ugni molinae / Estudio comparativo de la actividad antiinflamatoria y composición química cualitativa y cuantitativa de triterpenoides de diez genotipos de Ugni molinae

The aim of this study was to assess the differences in qualitative-quantitative composition of triterpenoids and total phenolic contents, together with anti-inflammatory activity of Ugni molinae leaves obtained from ten genotypes. The ethyl acetate (EAE) and ethanol extracts (ETE) were obtained and analyzed. The plant genotypes were grown under same soil and climate conditions and under same agronomic management; the leaves were also harvested under the same conditions. Anti-inflammatory activity was evaluated by mice ear edema induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) at a single dose of 200 mg/kg BW of each extract. Composition of triterpenoids and total phenolic contents was determined by HPLC-DAD and Folin-Ciocalteu method, respectively. Ugni molinae leaves of different plant genotypes exhibited significant differences in regard to their anti-inflammatory activity, as well as in qualitative-quantitative composition of triterpenoids and total phenolic content.

El objetivo de este estudio fue establecer las diferencias en la composición cualitativa y cuantitativa de triterpenoides y en los contenidos totales de fenoles, junto con la actividad antiinflamatoria de las hojas de Ugni molinae provenientes de diez genotipos. Los extractos de acetato de etilo (EAE) y etanólicos (ETE) fueron obtenidos y analizados. Los genotipos fueron cultivados bajo las mismas condiciones edafo-climáticas y con el mismo manejo agronómico; las hojas fueron cosechadas bajo las mismas condiciones. La actividad antiinflamatoria fue evaluada en ratones a los que se les indujo un edema en la oreja mediante la aplicación del 12-O-tetradecanoilforbol-13 acetato (TPA) y los extractos fueron evaluados a una dosis única de 200 mg/kg de peso corporal. La composición en triterpenoides y los contenidos de fenoles totales fueron determinados por CLAE-DAD y por el método de Folin-Ciocalteu, respectivamente. Las hojas provenientes de los diferentes genotipos de U. molinae, exhibieron significativas diferencias en sus actividades antiinflamatorias, así como, en el contenido cualitativo y cuantitativo de triterpenoides y en el contenido de fenoles totales.

Activation of conventional and novel protein kinase C isozymes by different diacylglycerol molecular species.

A variety of diacylglycerol (DG) molecular species are produced in stimulated cells. Conventional (α, ßII and γ) and novel (δ, ε, η and θ) protein kinase C (PKC) isoforms are known to be activated by DG. However, a comprehensive analysis has not been performed. In this study, we analyzed activation of the PKC isozymes in the presence of 2-2000 mmol% 16:0/16:0-, 16:0/18:1-, 18:1/18:1-, 18:0/20:4- or 18:0/22:6-DG species. PKCα activity was strongly increased by DG and exhibited less of a preference for 18:0/22:6-DG at 2 mmol%. PKCßII activity was moderately increased by DG and did not have significant preference for DG species. PKCγ activity was moderately increased by DG and exhibited a moderate preference for 18:0/22:6-DG at 2 mmol%. PKCδ activity was moderately increased by DG and exhibited a preference for 18:0/22:6-DG at 20 and 200 mmol%. PKCε activity moderately increased by DG and showed a moderate preference for 18:0/22:6-DG at 2000 mmol%. PKCη was not markedly activated by DG. PKCθ activity was the most strongly increased by DG and exhibited a preference for 18:0/22:6-DG at 2 and 20 mmol% DG. These results indicate that conventional and novel PKCs have different sensitivities and dependences on DG and a distinct preference for shorter and saturated fatty acid-containing and longer and polyunsaturated fatty acid-containing DG species, respectively. This differential regulation would be important for their physiological functions.

The integral membrane protein ITM2A, a transcriptional target of PKA-CREB, regulates autophagic flux via interaction with the vacuolar ATPase.

The PKA-CREB signaling pathway is involved in many cellular processes including autophagy. Recent studies demonstrated that PKA-CREB inhibits autophagy in yeast; however, the role of PKA-CREB signaling in mammalian cell autophagy has not been fully characterized. Here, we report that the integral membrane protein ITM2A expression is positively regulated by PKA-CREB signaling and ITM2A expression interferes with autophagic flux by interacting with vacuolar ATPase (v-ATPase). The ITM2A promoter contains a CRE element, and mutation at the CRE consensus site decreases the promoter activity. Forskolin treatment and PKA expression activate the ITM2A promoter confirming that ITM2A expression is dependent on the PKA-CREB pathway. ITM2A expression results in the accumulation of autophagosomes and interferes with autolysosome formation by blocking autophagic flux. We demonstrated that ITM2A physically interacts with v-ATPase and inhibits lysosomal function. These results support the notion that PKA-CREB signaling pathway regulates ITM2A expression, which negatively regulates autophagic flux by interfering with the function of v-ATPase.

Downregulation of PEA-15 reverses G1 arrest, and nuclear and chromatin changes of senescence phenotype via pErk1/2 translocation to nuclei.

We previously showed that senescent cells respond to TPA with translocation of senescence associated-pErk1/2 (SA-pErk1/2) into nuclei along with reversal of senescence morphology. Here, we describe that the reversal of senescence phenotype was manifested by knockdown of cytoplasmic PEA-15 expression, a sequestrator of cytoplasmic pErk1/2. Transfection of short-interfering RNA to PEA-15 (siPEA-15) significantly induced nuclear translocation of SA-pErk1/2, and siPEA-15 with TPA co-treatment further increased the translocation. Moreover, the reversal of senescence phenotype, such as expressions of SA-ß-galactosidase, p53, p21(WAF1), PML body, 53BP1 and H3K9me2, was modified by either knockdown of PEA-15 or TPA treatment, indicating that nuclear translocation of SA-pErk1/2 might inhibit senescence progression. Indeed, knockdown of PEA-15 or TPA treatment significantly induced progression of G1 arrested cells to S-phase in human diploid fibroblast (HDF) senescent cells, examined by immunocytochemistry, FACS and immunoblot analyses. In conclusion, downregulation of PEA-15 expression reverses senescence phenotypes via nuclear translocation of SA-pErk1/2, which suggests in vivo maintenance of senescence phenotype by sequestration of pErk1/2 in cytoplasm.

2-Arachidonoylglycerol enhances platelet formation from human megakaryoblasts.

Platelets modulate vascular system integrity, and their loss is critical in haematological pathologies and after chemotherapy. Therefore, identification of molecules enhancing platelet production would be useful to counteract thrombocytopenia. We have previously shown that 2-arachidonoylglycerol (2-AG) acts as a true agonist of platelets, as well as it commits erythroid precursors toward the megakaryocytic lineage. Against this background, we sought to further interrogate the role of 2-AG in megakaryocyte/platelet physiology by investigating terminal differentiation, and subsequent thrombopoiesis. To this end, we used MEG-01 cells, a human megakaryoblastic cell line able to produce in vitro platelet-like particles. 2-AG increased the number of cells showing ruffled surface and enhanced surface expression of specific megakaryocyte/platelet surface antigens, typical hallmarks of terminal megakaryocytic differentiation and platelet production. Changes in cytoskeleton modeling also occurred in differentiated megakaryocytes and blebbing platelets. 2-AG acted by binding to CB1 and CB2 receptors, because specific antagonists reverted its effect. Platelets were split off from megakaryocytes and were functional: they contained the platelet-specific surface markers CD61 and CD49, whose levels increased following stimulation with a natural agonist like collagen. Given the importance of 2-AG for driving megakaryopoiesis and thrombopoiesis, not surprisingly we found that its hydrolytic enzymes were tightly controlled by classical inducers of megakaryocyte differentiation. In conclusion 2-AG, by triggering megakaryocyte maturation and platelet release, may have clinical efficacy to counteract thrombocytopenia-related diseases.