Euphol from Tapinanthus sp. Induces Apoptosis and Affects Signaling Proteins in Glioblastoma and Prostate Cancer Cells

BACKGROUND: Plants play an important role in cancer therapy. They are source of natural molecules which can induce apoptosis in cancer cells by affecting molecular mechanisms implicated in cancer progression. The MAP Kinase/ERK1/2 and PI3K/AKT signaling pathways are two classical signaling pathways implicated in cancer progression and constitute therapeutic targets against cancer. This study aimed to evaluate the effect of euphol on MAP Kinase/ERK1/2 and PI3K/AKT signaling pathways in glioblastoma and prostate cancer cells. Euphol is a tetracyclique triterpene alcohol isolated from Tapinanthus sp. which is a hemi parasitic plant belonging to Loranthaceae family. METHODS: Plant powder was extracted by maceration and euphol was isolated and described using respectively column chromatography separation on silica gel and spectroscopic data. Cytotoxic effect of euphol was evaluated using XTT assay and its effect on MAP Kinase/ERK1/2 and PI3K/AKT protein expression was investigated by Western immunoblot analysis. Apotosis was analyzed by evaluating caspase-3/7 activity. RESULTS: Our investigations demonstrated that this compound has an important cytotoxic effect on C6 and U87 MG glioblastoma (GBM) cells and PC-3 prostate cancer cells. Furthermore, euphol-induced apoptosis revealed by elevated caspase 3/7 activity, was correlated with a significant inhibition of MAP kinase/Erk 1/2 and PI3K/Akt signaling pathway in glioblastoma U87 MG cells. The reverse effect was observed in C6 glioblastoma cells, where apoptosis was correlated with a long-lasting activation of Erk 1/2.  In PC-3 cells, euphol had no or limited effect on Erk 1/2 and Akt activity. CONCLUSION: These results indicate that euphol induces cell death in glioblastoma and prostate cancer cells and regulates significantly Erk1/2 and Akt activity in glioblastoma cells.

α­rhamnrtin­3­α­rhamnoside exerts anti­inflammatory effects on lipopolysaccharide­stimulated RAW264.7 cells by abrogating NF­κB and activating the Nrf2 signaling pathway.

α­rhamnrtin­3­α­rhamnoside (ARR) is the principal compound extracted from Loranthus tanakae Franch. & Sav. However, its underlying pharmacological properties remain undetermined. Inflammation is a defense mechanism of the body; however, the excessive activation of the inflammatory response can result in physical injury. The present study aimed to investigate the effects of ARR on lipopolysaccharide (LPS)­induced RAW264.7 macrophages and to determine the underlying molecular mechanism. A Cell Counting Kit­8 assay was performed to assess cytotoxicity. Nitric oxide (NO) production was measured via a NO colorimetric kit. Levels of prostaglandin E2 (PGE2) and proinflammatory cytokines, IL­1ß and IL­6, were detected using ELISAs. Reverse transcription­quantitative (RT­q)PCR analysis was performed to detect the mRNA expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase­2 (COX­2), IL­6 and IL­1ß in LPS­induced RAW246.7 cells. Western blotting, immunofluorescence and immunohistochemistry analyses were performed to measure the expression levels of NF­κB and nuclear factor­erythroid 2­related factor 2 (Nrf2) signaling pathway­related proteins to elucidate the molecular mechanisms of the inflammatory response. The results of the cytotoxicity assay revealed that doses of ARR ≤200 µg/ml exhibited no significant effect on the viability of RAW264.7 cells. The results of the Griess assay demonstrated that ARR inhibited the production of NO. In addition, the results of the ELISAs and RT­qPCR analysis discovered that ARR reduced the production of the proinflammatory cytokines, IL­1ß and IL­6, as well as the proinflammatory mediators, PGE2, iNOS and COX­2, in LPS­induced RAW264.7 cells. Immunohistochemical analysis demonstrated that ARR inhibited LPS­induced activation of TNF­associated factor 6 (TRAF6) and NF­κB p65 signaling molecules, while reversing the downregulation of the NOD­like receptor family CARD domain containing 3 (NLRC3) signaling molecule, which was consistent with the results of the western blotting analysis. Immunofluorescence results indicated that ARR reduced the increase of NF­κB p65 nuclear expression induced by LPS. Furthermore, the results of the western blotting experiments also revealed that ARR upregulated heme oxygenase­1, NAD(P)H quinone dehydrogenase 1 and Nrf2 pathway molecules. In conclusion, the results of the present study suggested that ARR may exert anti­inflammatory effects by downregulating NF­κB and activating Nrf2­mediated inflammatory responses, suggesting that ARR may be an attractive anti­inflammatory candidate drug.

Host-mediated volatile polymorphism in a parasitic plant influences its attractiveness to pollinators.

Host-plants can mediate the interactions between herbivores and their mutualists and also between parasitic plants and their mutualists. The present study reveals how a hemiparasitic plant parasitizing three host species gives rise to three distinct hemiparasite-host neighborhoods which differ in terms of volatile composition and pollinator attractiveness. The study was performed in a population of the mistletoe Tristerix verticillatus infecting three different species of hosts occurring in sympatry within a small area, thus exposing all individuals studied to similar abiotic conditions and pollinator diversity; we assessed the effect of hosts on the hemiparasites' visual and olfactory cues for pollinator attraction. During the study period, the hemiparasite individuals were flowering but the hosts were past their flowering stage. We collected volatile organic compounds from the hemiparasite and its hosts, measured floral display characteristics and monitored bird and insect visitors to inflorescences of T. verticillatus. We showed that: (1) floral patches did not differ in terms of floral display potentially involved in the attraction of pollinators, (2) hosts and hemiparasites on each host were discriminated as distinct chemical populations in terms of their volatile chemical profiles, (3) insect visitation rates differed between hemiparasites parasitizing different hosts, and (4) volatile compounds from the host and the hemiparasite influenced the visitation of hemiparasite flowers by insects. The study showed that a species regarded as "ornithophilic" by its floral morphology was actually mostly visited by insects that interacted with its sexual organs during their visits and carried its pollen, and that host-specific plant-volatile profiles within the T. verticillatus population were associated with differential attractiveness to pollinating insects.

Evaluation of phytochemical and antimicrobial properties of leaf extract of Tapinanthus sessilifolius (P. Beauv) van Tiegh.

Leaf extracts of T. sessilifolius growing on five different host plants (Psidium guajava, Citrus lemon, Vernonia amygdalina, Persea americana and Jatropa curcas) were evaluated for antimicrobial activity of the plant. Powdered leaves of T. sessilifolius collected from each host plant was divided into two portions. One portion was used for aqueous infusion and the other portion was successively extracted with hexane, ethylacetate and methanol. Infusion of aqueous extract of powdered leaves did not show antimicrobial effect even at the concentration of 1000 and 2000 microg/ml on test microorganisms (Staph. aureus, E. coli, Bacillus subtilis, Pseudomonas aeruginosa and Candida albicans). However in broth culture, methanolic and hexane extract had MIC range of 62.5-500 microg/ml and ethylacetate extract had 250-500 microg/ml. Phytochemical screening of leaf samples of T. sessilifolius collected from different host plants showed positive test for hydrolysable tannins, saponins, flavonoids, terpenes, cardiac glycoside, reducing sugars and proteins. LD50 concentration was found to be > 1.500 mg/kg for samples from P. guajava; 489.89 mg/kg for J. curcas and C. lemon; and 692 mg/kg for V. amydalina in mice.

A comparative study of aluminium and nutrient concentrations in mistletoes on aluminium-accumulating and non-accumulating hosts.

Mistletoes offer a unique model to study interactions among Al and nutrients in vascular plants, because they grow and reproduce on hosts with distinct Al uptake strategies. We investigated Al distribution and nutrient relations of mistletoes on Al-accumulating and non-accumulating hosts. We hypothesised that mistletoes would exhibit similar leaf nutrient and Al concentrations as their host plants, but a strong compartmentalisation of Al when growing on Al-accumulators. We measured concentrations of N, P, K, Ca, Mg, Cu, Fe, Mn, Zn in leaves and Al in leaves, seeds and branches of Phthirusa ovata and Psittacanthus robustus infecting Miconia albicans, an Al-accumulator, and Ph. ovata infecting Byrsonima verbascifolia, a non-Al-accumulator. High leaf concentrations of Al in Ph. ovata only occurred while parasitizing the Al-accumulating host; there was no accumulation in branches or seeds. In P. robustus, large concentrations of Al were found in leaves, branches and seeds. Mistletoe seed viability and leaf nutrient concentrations were not affected by Al accumulation. Passive uptake of Al, Ca, Mg, Mn and Cu in mistletoes was evidenced by significant correlations between mistletoes and host leaf concentrations, but not of N, P and K. Al was retranslocated to different plant organs in P. robustus, whereas it was mostly restricted to leaves in Ph. ovata. We suggest that Al might have some specific function in P. robustus, which only parasitizes Al-accumulator hosts, while the host generalist Ph. ovata can be considered a facultative Al-accumulator.

Evaluation of phytochemical and antimicrobial properties of leaf extract of Tapinanthus sessilifolius (P. Beauv) van Tiegh.

Leaf extracts of T. sessilifolius growing on five different host plants (Psidium guajava, Citrus lemon, Vernonia amygdalina, Persea americana and Jatropa curcas) were evaluated for antimicrobial activity of the plant. Powdered leaves of T. sessilifolius collected from each host plant was divided into two portions. One portion was used for aqueous infusion and the other portion was successively extracted with hexane, ethylacetate and methanol. Infusion of aqueous extract of powdered leaves did not show antimicrobial effect even at the concentration of 1000 and 2000 microg/ml on test microorganisms (Staph. aureus, E. coli, Bacillus subtilis, Pseudomonas aeruginosa and Candida albicans). However in broth culture, methanolic and hexane extract had MIC range of 62.5-500 microg/ml and ethylacetate extract had 250-500 microg/ml. Phytochemical screening of leaf samples of T. sessilifolius collected from different host plants showed positive test for hydrolysable tannins, saponins, flavonoids, terpenes, cardiac glycoside, reducing sugars and proteins. LD50 concentration was found to be > 1.500 mg/kg for samples from P. guajava; 489.89 mg/kg for J. curcas and C. lemon; and 692 mg/kg for V. amydalina in mice.