Zhenya Yordanova

Involvement of ethylene and nitric oxide in cell death in mastoparan-treated unicellular alga Chlamydomonas reinhardtii.

This work demonstrates a contribution of ethylene and NO (nitric oxide) in MP (mastoparan)‐induced cell death in the green algae Chlamydomonas reinhardtii. Following MP treatment, C. reinhardtii showed massive cell death, expressing morphological features of PCD (programmed cell death). A pharmacological approach involving combined treatments with MP and ethylene‐ and NO‐interacting compounds indicated the requirement of trace amounts of both ethylene and NO in MP‐induced cell death. By employing a carbon dioxide laser‐based photoacoustic detector to measure ethylene and a QCL (quantum cascade laser)‐based spectrometer for NO detection, simultaneous increases in the production of both ethylene and NO were observed following MP application. Our results show a tight regulation of the levels of both signalling molecules in which ethylene stimulates NO production and NO stimulates ethylene production. This suggests that, in conjunction with the elicitor, NO and ethylene cooperate and act synchronously in the mediation of MP‐induced PCD in C. reinhardtii. To the best of our knowledge, this is the first report on the functional significance of ethylene and NO in MP‐induced cell death.

Mastoparan-induced programmed cell death in the unicellular alga Chlamydomonas reinhardtii

BACKGROUND AND AIMS Under stress-promoting conditions unicellular algae can undergo programmed cell death (PCD) but the mechanisms of algal cellular suicide are still poorly understood. In this work, the involvement of caspase-like proteases, DNA cleavage and the morphological occurrence of cell death in wasp venom mastoparan (MP)-treated Chlamydomonas reinhardtii were studied. METHODS Algal cells were exposed to MP and cell death was analysed over time. Specific caspase inhibitors were employed to elucidate the possible role of caspase-like proteases. YVADase activity (presumably a vacuolar processing enzyme) was assayed by using a fluorogenic caspase-1 substrate. DNA breakdown was evaluated by DNA laddering and Comet analysis. Cellular morphology was examined by confocal laser scanning microscopy. KEY RESULTS MP-treated C. reinhardtii cells expressed several features of necrosis (protoplast shrinkage) and vacuolar cell death (lytic vesicles, vacuolization, empty cell-walled corpse-containing remains of digested protoplast) sometimes within one single cell and in different individual cells. Nucleus compaction and DNA fragmentation were detected. YVADase activity was rapidly stimulated in response to MP but the early cell death was not inhibited by caspase inhibitors. At later time points, however, the caspase inhibitors were effective in cell-death suppression. Conditioned medium from MP-treated cells offered protection against MP-induced cell death. CONCLUSIONS In C. reinhardtii MP triggered PCD of atypical phenotype comprising features of vacuolar and necrotic cell deaths, reminiscent of the modality of hypersensitive response. It was assumed that depending on the physiological state and sensitivity of the cells to MP, the early cell-death phase might be not mediated by caspase-like enzymes, whereas later cell death may involve caspase-like-dependent proteolysis. The findings substantiate the hypothesis that, depending on the mode of induction and sensitivity of the cells, algal PCD may take different forms and proceed through different pathways.

Revealing the reviving secret of the white dead nettle (Lamium album L.)

Lamium album, commonly known as white dead nettle or non-stinging nettle is a flowering herbaceous plant, native throughout Europe, Western Asia and North Africa. From ancient times this plant has been endowed with revival, curative and culinary virtues. In the past, in the traditional and folk medicine white dead nettle has been used mainly for its anti-inflammatory, astringent and anti-septic activity. Nowadays significant amount of knowledge on the efficacy of extracts and raw material of L. album is accumulated and a number of health-related beneficial activities have been scientifically proven. In vitro analyses conducted in various model systems have demonstrated antiviral, antimicrobial, antioxidant, anticancer, cytoprotective, wound healing and other important pharmacological effects. The present review summarizes the recent information on the phytochemical features of this pharmacologically important species. The findings on the chemical composition, biological activities and the pharmacological properties underlying the revival secret of white dead nettle are described and discussed in the view of potential applications for treatment of human diseases. Trends for further research are outlined.

Mastoparan-Induced Cell Death Signalling in Chlamydomonas Reinhardtii

ABSTRACT The present study was focused on the elucidation of stress-induced cell death signaling events in the unicellular alga Chlamydomonas reinhardtii exposed to treatment with wasp venom mastoparan. By applying pharmacological approach with specific inhibitors, we have investigated the involvement of ethylene, nitric oxide and lipid signaling in MP-treated C. reinhardtii. Superior sensitive laser-based detectors were used for real-time measurement of trace amounts of ethylene and nitric oxide. The morphological features in the cells undergoing programmed cell death were detected by using laser-scanning confocal microscopy. Data showed that mastoparan induces programmed cell death in C. reinhardtii 137 C(+) that is associated with phospholipid signaling, including phospholipases C and D, ethylene and nitric oxide and, the dead cells express apoptotic-like cellular disintegration, involving cytoplasm shrinkage and condensation of the nucleus.

XYLANSE-INDUCED CELL DEATH EVENTS IN DETACHED TOBACCO LEAVES

ABSTRACT Plant-pathogen interactions are associated with plant defense mechanism known as hypersensitive response (HR), which is a form of programmed cell death (PCD). In the present work we have tested the potency of chemicals, proven as PCD inhibitors in other systems, to prevent the spread of cell death in detached tobacco leaves challenged with β- xylanase from Aspergillus awamori K1. Xylanase induced cell death that was accompanied by electrolyte leakage and increased levels of the stress metabolites hydrogen peroxide and malondialdehyde. Lesion development, ion leakage and the production of stress metabolites were suppressed if the infiltration site was pre-infiltrated with broad range caspase inhibitor benzyoxycarbonyl-Asp-2,6-dichlorobenzoyloxymethylketone (Z-Asp-CH2-DCB), cysteine protease inhibitor iodoacetamide (IA), serine protease inhibitor 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF), the antioxidant l-galactone-γ-lactone (L-gal) and the ethylene blocker (S)-trans-2-Amino-4-(2-aminoethoxy)-3-butenoic acid hydrochloride (AVG). Massive ROS accumulation, as determined by staining with 3–3′-diaminobenzidine and 2′,7′-dichlorofluorescein diacetate, occurred in xylanase-infiltrated lesions and was substantially reduced by the inhibitors. To the best of our knowledge, this is the first report showing that β-xylanase produced by Aspergillus awamori K1 induces cell death response in tobacco and proteolysis, ROS and ethylene are involved in the mediation of the signaling.

Genetic transformation of rare Verbascum eriophorum Godr. plants and metabolic alterations revealed by NMR-based metabolomics

ObjectivesTo develop a protocol to transform Verbascum eriophorum and to study the metabolic differences between mother plants and hairy root culture by applying NMR and processing the datasets with chemometric tools.ResultsVerbascum eriophorum is a rare species with restricted distribution, which is poorly studied. Agrobacterium rhizogenes-mediated genetic transformation of V. eriophorum and hairy root culture induction are reported for the first time. To determine metabolic alterations, V. eriophorum mother plants and relevant hairy root culture were subjected to comprehensive metabolomic analyses, using NMR (1D and 2D). Metabolomics data, processed using chemometric tools (and principal component analysis in particular) allowed exploration of V. eriophorum metabolome and have enabled identification of verbascoside (by means of 2D-TOCSY NMR) as the most abundant compound in hairy root culture.ConclusionMetabolomics data contribute to the elucidation of metabolic alterations after T-DNA transfer to the host V. eriophorum genome and the development of hairy root culture for sustainable bioproduction of high value verbascoside.

2nd international symposium on phytochemicals in medicine and food (2-ISPMF)

The 2nd international symposium on phytochemicals in medicine and food (2-ISPMF) will be held from April 7th to 10th, 2017 in Fuzhou, China. 2-ISPMF is organized jointly by the Phytochemical Society of Europe (PSE), Phytochemical Society of Asia (PSA) and International Society for Chinese Medicine (ISCM). It dedicates to creating a stage for exchanging the latest research results in phytochemicals for food and human health. It is jointly organized by Fujian Agriculture and Forestry University, University of Macau, and University of Hong Kong. The local sponsoring institutions contain Kunming University of Science and Technology, Yancheng Institute of Technology, Guiyang Medical University, Nanchang University, Hebei Normal University of Science and Technology, Zhejiang University, Beijing Normal University, Jinan University and so on. Over 250 scientists from 32 counties have registered to attend this conference. Prof. Baodong Zheng from Fujian Agriculture and Forestry University is the Chairman of local organizing committee. The international organizing committee board of 2-ISPMF consists of Yitao Wang (Macau, Honorary Chairman), Jianbo Xiao (China, Executive Chairman), Yanbo Zhang (Hong Kong, Co-Chairman), Miroslav Strnad (Czech), Yoshinori Asakawa (Japan), Bernhard Hennig (USA), Mei Han (China), Milen I. Georgiev (Bulgaria), Amir Reza Jassbi (Iran), Virginia Lanzotti (Italy), Pinarosa Avato (Italy), Veronique Seidel (UK), Rosa Tundis (Italy). The international scientific committee board is comprised of the representatives from China, Chile, Sweden, Romania, Botswana, Pakistan, USA, Italy, Spain, Austria, Turkey, Algeria, Germany, Thailand, UK, Netherlands, and South Korea. The international organizing committee and scientific committee board of 2-ISPMF assembled an exciting and diverse program, featuring 14 keynote lectures, 28 invited lectures, 46 short lectures, a J. Xiao (&) College of Food Science, Fujian Agriculture and Forestry University, Fuzhou 350002, Fujian, People’s Republic of China e-mail: jianboxiao@yahoo.com

Changes in the functional characteristics of tumor and normal cells after treatment with extracts of white dead-nettle.

Lamium album L. is a perennial herb widely used in folk medicine. It possesses a wide spectrum of therapeutic activities (anti-inflammatory, astringent, antiseptic, antibiotic, antispasmodic, antioxidant and anti-proliferative). Preservation of medicinal plant could be done by in vitro propagation to avoid depletion from their natural habitat. It is important to know whether extracts from L. album plants grown in vitro possess similar properties as extracts from plants grown in vivo. For these reasons, it is important to examine changes in the composition of secondary metabolites during in vitro cultivation of the plant and how they affect the biological activity. We used A549 human cancer cell line and normal kidney epithelial cells MDCKII (Madin–Darby canine kidney cells II) as controls in assessing the anti-cancer effect of plant extracts. To elucidate changes in some key functional characteristics, adhesion test, MTT (3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyl tetrazolium bromide), transepithelial resistance (TER), immunofluorescence staining and trypan blue exclusion test were performed. Methanol and chloroform extracts of in vivo and in vitro propagated plants affected differently cancerous and non-cancerous cells. The most pronounced differences were observed in the morphological analysis and in the cell adhesive properties. We also detected suppressed epithelial transmembrane electrical resistance of MDCK II cells, by treatment with plant extracts, compared to non-treated MDCK II cells. A549 cells did not polarize under the same conditions. Altered organization of actin filaments in both cell types were noticed suggesting that extracts from L. album L. change TER and actin filaments, and somehow may block cell mechanisms, leading to the polarization of MDCK II cells.