Plant miR171 modulates mTOR pathway in HEK293 cells by targeting GNA12.

Plant microRNAs have shown the capacity to regulate mammalian systems. The potential bioactivity of miR171vr, an isoform of the plant miR171, on human embryonic kidney 293 (HEK293) cells was investigated. Bioinformatics simulations revealed that human G protein subunit alpha 12 (GNA12) transcript could represent an excellent target for miR171vr. To confirm this prediction, in vitro experiments were performed using a synthetic microRNA designed on miR171vr sequence. MiR-treated cells showed a significant decrease of GNA12 mRNA and protein levels, confirming the putative cross-kingdom interaction. In addition, miR171vr determined the modulation of GNA12 downstream signaling factors, including mTOR, as expected. Finally, the effect of the plant miRNA on HEK293 cell growth and its stability in presence of several stressors, such as those miming digestive processes and procedures for preparing food, were evaluated. All this preliminary evidence would suggest that miR171vr, introduced by diet or as supplement in gene therapies, could potentially influence human gene expression, especially for treating disorders where GNA12 is over-expressed (i.e. oral cancer, breast and prostate adenocarcinoma) or mTOR kinase is down-regulated (e.g. obesity, type 2 diabetes, neurodegeneration).

Antimicrobial and anti-inflammatory activities of three halophyte plants from Algeria and detection of some biomolecules by HPLC-DAD.

Antimicrobial activity of hydroalcoholic extracts (30/70) from leaves and stems of three halophytes (Tamarix africana, Arthrocnemum macrostachyum and Suaeda fruticose) was investigated. In vivo toxicological study and anti-inflammatory activity of leaf extract of T. africana were tested on carrageenan-induced inflammatory paw edema. T. africana possessed significant anti-inflammatory activity at 150 and 300 mg/kg confirmed by histological study of inflamed tissues. Six phenolic acids and 10 flavonoids where identified by HPLC-DAD. Gallic acid, Rutin and Kaempferol-3-O-glucoside were the major compounds. For the antibiotic assays, S. fruticosa leaf extract exhibited strong bactericidal power against S. aureus with MBC of 1.25 mg/mL whereas T. africana leaf and stem samples exhibited a significant bactericidal activity against S. aureus and B. subtilis compared to the negative control (Ampicillin and Chloramphenicol). Crude leaf and stem extracts from T. africana and stem extract from S. fruticosa exhibited a strong antifungal effect against C. albicans.

Oregano Phytocomplex Induces Programmed Cell Death in Melanoma Lines via Mitochondria and DNA Damage.

Plant secondary metabolites possess chemopreventive and antineoplastic properties, but the lack of information about their exact mechanism of action in mammalian cells hinders the translation of these compounds in suitable therapies. In light of this, firstly, Origanum vulgare L. hydroalcoholic extract was chemically characterized by spectrophotometric and chromatographic analyses; then, the molecular bases underlying its antitumor activity on B16-F10 and A375 melanoma cells were investigated. Oregano extract induced oxidative stress and inhibited melanogenesis and tumor cell proliferation, triggering programmed cell death pathways (both apoptosis and necroptosis) through mitochondria and DNA damage. By contrast, oregano extract was safe on healthy tissues, revealing no cytotoxicity and mutagenicity on C2C12 myoblasts, considered as non-tumor proliferating cell model system, and on Salmonella strains, by the Ames test. All these data provide scientific evidence about the potential application of this food plant as an anticancer agent in in vivo studies and clinical trials.

Plant defense factors involved in Olea europaea resistance against Xylella fastidiosa infection.

Olive quick decline syndrome (OQDS) is a dangerous plant disease, caused by the bacterium Xylella fastidiosa, which targets olive (Olea europaea). Since field observations suggested that some olive cultivars (i.e. Leccino) were more resistant to OQDS than others (i.e. Cellina di Nardò), the plant defense strategies adopted by olive to contrast X. fastidiosa infection were investigated. In the present study, ELISA and genetic approaches were used to confirm plant infection, while microbial colonization mechanism and distribution in host plant tissues and reactive oxygen species (ROS) levels were examined by light, scanning electron and confocal microscopy analyses. Spectrophotometric and chromatographic techniques were performed to measure secondary metabolites content and qPCR assay was carried out for monitoring plant gene expression variation. Our analysis showed that X. fastidiosa caused accumulation of ROS in Leccino samples compared to Cellina di Nardò. Moreover, the infection induced the up-regulation of defense-related genes, such as NADPH oxidase, some protein kinases, pathogen plant response factors and metabolic enzymes. We also found that Leccino plants enhanced the production of specific antioxidant and antimicrobial molecules, to fight the pathogen and avoid its spreading into xylem vessels. We provided new information on OQDS resistance mechanism applied by Leccino cultivar. In particular, we evidenced that high concentrations of ROS, switching on plant defence signalling pathways, may represent a key factor in fighting X. fastidiosa infection.

Hydroalcoholic extract from Origanum vulgare induces a combined anti-mycobacterial and anti-inflammatory response in innate immune cells.

In nature, many plants or their extracted compounds have been found to possess anti-inflammatory features and therapeutic properties against infectious as well as non-infectious diseases, including cancer. In this study, we analysed the immunomodulatory effects on innate immune cells of hydroalcoholic extract from Origanum vulgare L. ssp. hirtum (HyE-Ov), a plant traditionally known for its anti-oxidative properties. The effects of HyE-Ov were tested on human monocyte derived dendritic cells (DC), type-1 (M1) and type-2 macrophages (M2) infected with M. bovis Bacille Calmette-Guérin (BCG), used as a model of persistent intracellular bacterium. DC, M1 and M2 treated with HyE-Ov significantly enhanced their mycobactericidal activity, which was associated with phagosomal acidification in M1 and M2 and increase of phagosomal, but not mitochondrial ROS production in M1, M2, and DC. Treatment of BCG-infected DC with HyE-Ov significantly reduced TNF-α and IL-12 production and increased TGF-ß synthesis. Finally, experiments were repeated using eight different HPLC fractions of HyE-Ov. Results showed that the capability to activate anti-microbial and anti-inflammatory response is shared by different fractions, suggesting that diverse bioactive molecules are present within the hydroalcoholic extract. Altogether, these results show that HyE-Ov promotes anti-mycobacterial innate immunity and limits inflammatory response in vitro and suggest that this plant extract may be exploitable as phytocomplex or nutraceutical for novel host-directed therapeutic approaches.

Hydroalcoholic extract of Spartium junceum L. flowers inhibits growth and melanogenesis in B16-F10 cells by inducing senescence.

BACKGROUND: Ultraviolet light exposure generates, in human tissues, radical species, which represent the main cause of photo-aging, DNA damage and skin cancer onset. On the other hand, Mediterranean plants, being continuously subjected to high solar radiation levels, are naturally adapted to take on this type of abiotic stress, thanks to the production of antioxidant secondary metabolites. For these reasons, several plant extracts were documented to be excellent antineoplastic drugs. PURPOSE: We investigated the potential antitumor activity of the flower extract obtained by Spartium junceum L., a Mediterranean shrub, correlating it with the plant metabolic profile. STUDY DESIGN: After selecting the best extraction method to obtain as more secondary metabolites as possible from S. junceum flowers, we characterized the extract metabolic content. Then, by in vitro analyses, the antioxidant profile and the antineoplastic activity on B16-F10 murine melanoma cell of our extract were investigated. METHODS: Spectrophotometric assays, HPLC-DAD and GC-MS analyses provided us information about flower extract composition and antioxidant activity. MTT assay and Trypan Blue exclusion test were performed to assess the extract toxicity and the viability, after treatments, of B16-F10 cancer cells and of C2C12 murine myoblasts. In vitro experiments (i.e. cytofluorimetry, protein analysis and qPCR) allowed us to analyze the effect of the plant extract on B16-F10 cell redox state, melanogenesis and cell cycle. Senescence induction was investigated by using a specific kit. RESULTS: We observed that the hydroalcoholic extract of S. junceum flowers (HFE) strongly inhibited B16-F10 murine melanoma cell proliferation, while just a feeble effect was observed on C2C12 murine myoblasts. Moreover, we found that HFE exerted a pro-oxidant activity on melanoma cells, inhibited melanogenesis and caused cell cycle arrest in G2/M phase, inducing senescence. These anti-cancer properties of HFE could be related to the rich metabolic profile of the extract that we characterized by HPLC-DAD and GC-MS analyses. CONCLUSION: This evidence suggests that S. junceum phytocomplex can be used as a selective, nontoxic, economic and easily available anticancer drug.

Nanodiamonds coupled with 5,7-dimethoxycoumarin, a plant bioactive metabolite, interfere with the mitotic process in B16F10 cells altering the actin organization.

For the first time, we coupled reduced detonation nanodiamonds (NDs) with a plant secondary metabolite, citropten (5,7-dimethoxycoumarin), and demonstrated how this complex was able to reduce B16F10 tumor cell growth more effectively than treatment with the pure molecule. These results encouraged us to find out the specific mechanism underlying this phenomenon. Internalization kinetics and quantification of citropten in cells after treatment with its pure or ND-conjugated form were measured, and it was revealed that the coupling between NDs and citropten was essential for the biological properties of the complex. We showed that the adduct was not able to induce apoptosis, senescence, or differentiation, but it determined cell cycle arrest, morphological changes, and alteration of mRNA levels of the cytoskeletal-related genes. The identification of metaphasic nuclei and irregular disposition of ß-actin in the cell cytoplasm supported the hypothesis that citropten conjugated with NDs showed antimitotic properties in B16F10 cells. This work can be considered a pioneering piece of research that could promote and support the biomedical use of plant drug-functionalized NDs in cancer therapy.

Temperamental and character profiles of preschool children with ODD, ADHD, and anxiety disorders.

BACKGROUND: The aim of the present research was to identify profiles of Cloninger's temperament and character dimensions associated with anxiety disorders, oppositional defiant disorder (ODD) and attention- deficit/hyperactive disorder (ADHD) in preschoolers. METHOD: The parents of 120 children (mean age=4.65 years; S.D.=.88) completed the Preschool Temperament and Character Inventory (PsTCI). The sample consisted of 4 groups (n=30 per group): ADHD, anxious, ODD and control children. To diagnose the different disorders, the Preschool Age Psychiatric Assessment and Child Behavior Checklist 1.5-5 was administered to the parents. RESULTS: The discriminant analysis showed that three temperamental dimensions (Harm Avoidance, Novelty Seeking and Persistence) enabled the correct classification of 75% of cases within their own group, which demonstrated an adequate accuracy rate. The ADHD children showed a temperamental profile that was characterized by high Novelty Seeking, low Reward Dependence and low Persistence, while the anxious children obtained high scores in Harm Avoidance. The profiles of the ODD children shared some common features (high Novelty Seeking) with the ADHD children, but the ODD children were characterized by higher Persistence and Harm Avoidance compared with ADHD children. CONCLUSIONS: The present results indicate that Cloninger's temperamental dimensions allow to differentiate the three most frequent psychiatric disorders in preschoolers.

Identification and characterization of the defensin-like gene family of grapevine.

Defensins are a class of small and diverse cysteine-rich proteins found in plants, insects, and vertebrates, which share a common tertiary structure and usually exert broad-spectrum antimicrobial activities. We used a bioinformatic approach to scan the Vitis vinifera genome and identified 79 defensin-like sequences (DEFL) corresponding to 46 genes and allelic variants, plus 33 pseudogenes and gene fragments. Expansion and diversification of grapevine DEFL has occurred after the split from the last common ancestor with the genera Medicago and Arabidopsis. Grapevine DEFL localization on the 'Pinot Noir' genome revealed the presence of several clusters likely evolved through local duplications. By sequencing reverse-transcription polymerase chain reaction products, we could demonstrate the expression of grapevine DEFL with no previously reported record of expression. Many of these genes are predominantly or exclusively expressed in tissues linked to plant reproduction, consistent with findings in other plant species, and some of them accumulated at fruit ripening. The transcripts of five DEFL were also significantly upregulated in tissues infected with Botrytis cinerea, a necrotrophic mold, suggesting a role of these genes in defense against this pathogen. Finally, three novel defensins were discovered among the identified DEFL. They inhibit B. cinerea conidia germination when expressed as recombinant proteins.

Childhood maltreatment predicts unfavorable course of illness and treatment outcome in depression: a meta-analysis.

OBJECTIVES: Evidence suggests that childhood maltreatment may negatively affect not only the lifetime risk of depression but also clinically relevant measures of depression, such as course of illness and treatment outcome. The authors conducted the first meta-analysis to examine the relationship between childhood maltreatment and these clinically relevant measures of depression. METHOD: The authors conducted searches in MEDLINE, PsycINFO, and Embase for articles examining the association of childhood maltreatment with course of illness (i.e., recurrence or persistence) and with treatment outcome in depression that appeared in the literature before December 31, 2010. Recurrence was defined in terms of number of depressive episodes. Persistence was defined in terms of duration of current depressive episode. Treatment outcome was defined in terms of either a response (a 50% reduction in depression severity rating from baseline) or remission (a decrease in depression severity below a predefined clinical significance level). RESULTS: A meta-analysis of 16 epidemiological studies (23,544 participants) suggested that childhood maltreatment was associated with an elevated risk of developing recurrent and persistent depressive episodes (odds ratio=2.27, 95% confidence interval [CI]=1.80­2.87). A meta-analysis of 10 clinical trials (3,098 participants) revealed that childhood maltreatment was associated with lack of response or remission during treatment for depression (odds ratio=1.43, 95% CI=1.11­1.83). Meta-regression analyses suggested that the results were not significantly affected by publication bias, choice of outcome measure, inclusion of prevalence or incidence samples, study quality, age of the sample, or lifetime prevalence of depression. CONCLUSIONS: Childhood maltreatment predicts unfavorable course of illness and treatment outcome in depression.