Metabolomic approach for the study of stress response of flax mutant for lignans biosynthesis pathway

Les lignanes sont des composés polyphénoliques largement distribués dans le monde végétal. Le lin, cultivé pour son huile ou sa fibre, renferme de grandes quantités d'un lignane de type dibenzylbutane : le sécoisolaricirésinol, qui est stocké dans le tégument de la graine sous forme diglucosylée, dans une macromolécule contenant de nombreux composés phénoliques assemblés par des liaisons esters. De nombreuses études s'intéressent aux effets bénéfiques sur la santé des lignanes et du sécoisolaricirésinol en particulier, du fait de propriétés antioxydantes marquées, néanmoins le rôle de ces molécules in planta n'est pas clairement élucidé ; un rôle potentiel dans les mécanismes de réponse aux stress est toutefois proposé. Une enzyme clé de la biosynthèse du sécoisolaricirésinol est la PLR (Pinorésinol Laricirésinol Réductase). Des lins RNAi PLR ont montré l’absence d'accumulation de SDG dans les graines. Il est donc envisagé dans cette thèse de comparer le contenu en métabolites (semi-)polaires de lins accumulant du SDG ou non, en condition de stress osmotique ou en condition témoin, par des approches métabolomiques en couplant deux méthodes analytiques : la résonance magnétique nucléaire du proton (RMN 1H) et la chromatographie liquide lié à la spectrométrie de masse (LC-MS). Les résultats ont montré que les plantes de lin témoins ainsi que celles transgéniques déficientes en lignanes, ont une réponse métabolique différente après avoir été exposées à des conditions de stress osmotique, cependant, les deux lignées ont montré une capacité d'induire une réponse adaptative au stress osmotique. Ces résultats suggèrent donc l'implication indirecte des lignanes dans la réponse au stress osmotiqueLignans, phenolic plant secondary metabolites, are derived from the phenylpropanoid biosynthetic pathway. Although, being investigated for their health benefits in terms of antioxydant, antitumor, anti-inflammatory and antiviral properties, the role of these molecules in planta remains not completely elucidated; a potential role in stress response mechanisms has been however proposed. In this study, a non-targeted metabolomic analysis of the roots, stems and leaves of wild type and PLR1-RNAi transgenic flax, devoid of (+) Secoisolariciresinol diglucoside ((+) SDG) – the main flaxseed lignan, was performed using 1H-NMR and LC-MS, in order to obtain further insight into the involvement of lignan in the response of plant to osmotic stress. Results showed that wild type and lignan deficient flax plants have a different metabolic response after being exposed to osmotic stress conditions, but they both showed the capacity to induce an adaptative response to osmotic stress. These findings suggest the indirect involvement of lignans in osmotic stress respons

Determination of the Antioxidant and Antiproliferative Properties of Pomegranate Peel Extract Obtained by Ultrasound on HCT-116 Colorectal Cancer Cell Line

International audienceColorectal cancer (CRC) is one of the most common cancers in the world. When treating patients, therapeutic agents have side effects; hence, the use of natural compounds found in medicinal plants including pomegranate. Ultrasound assisted extraction (UAE) is a new technique evolving to the detriment of traditional methods such as maceration. In this study, we investigated the antioxidant and anticancer effect of pomegranate peel extracts obtained by maceration and UAE at three different ultrasonic power levels (P1 = 10 W; P2 = 50 W; P3 = 100 W) on HCT-116 colorectal cancer cells. Phytochemical screening highlighted the presence of primary and secondary metabolites in pomegranate peels. In addition, the ethanolic extract obtained by UAE at 50 W was shown to be the most concentrated in phenolic and flavonoid compounds and have the most powerful antioxidant activity, which reached a maximum activity of 92% as determined by DPPH test. Similarly, the MTT cell viability test showed that the extract obtained by UAE at 50 W had the most potent inhibitory effect compared to the other extracts. In conclusion, the UAE at 50 W was shown to be the most suitable and efficient extraction technique to obtain bioactive compounds from pomegranate peel extracts that can be used in the treatment of CRC

Preventive and Therapeutic Effects of Punica granatum (Pomegranate) in Respiratory and Digestive Diseases: A Review

The pomegranate fruit is made of white to deep purple seeds that are enclosed in a white, spongy, astringent membrane, also known as pericarp, covered by a thick red skin and a crown-shaped calyx. It contains a variety of beneficial ingredients, including flavonoids, ellagitannin, punicalagin, ellagic acid, vitamins, and minerals. Pomegranates possess numerous health benefits, and their use in disease treatment has been widely recognized since antiquity. This fruit was known to exhibit several biological properties, including antibacterial, anti-inflammatory, antioxidant, and anticancer activities. Pomegranate has been used in a variety of medical systems for the treatment and therapy of a wide range of diseases and illnesses. This review summarizes studies highlighting the potential role of pomegranate in the prevention and treatment of diseases related to respiratory and digestive systems

<i>Laurus nobilis</i> Leaves and Fruits: A Review of Metabolite Composition and Interest in Human Health

Laurus nobilis belongs to the family Lauraceae and is characterized by its evergreen nature. It is well known because of its traditional use in cooking and folk medicine. Many studies have been performed regarding its chemical composition and the pharmacological potential it exhibits. It has been reported to have antioxidant, antibacterial, antifungal, antiviral, biocidal, antidiabetic, antiulcerogenic, neuroprotective, analgesic, anti-inflammatory, and anticholinergic properties, making this plant of great clinical importance. The main chemical compounds that show bioactive properties in L. nobilis are terpenoids, phenolics, and fatty acids. The purpose of this review is to describe the main use of L. nobilis, notably of the leaves and fruits, and to relate their metabolite content with their pharmacological and toxicological properties

Study of the antioxidant and anti-pancreatic cancer activities of Anchusa strigosa aqueous extracts obtained by maceration and ultrasonic extraction techniques

International audiencePancreatic cancer is a highly aggressive malignancy and a leading cause of cancer-related deaths worldwide. Moreover, the incidence and mortality rates for pancreatic cancer are projected to keep increasing. A major challenge in the treatment of pancreatic cancer is the lack of effective screening approaches, which contributes to its poor prognosis, indicating the need for new treatment regimens and alternative therapies, such as herbal medicine. The medicinal plant A. strigosa , which is widely distributed in the Eastern Mediterranean region, is a short prickly plant from the Boraginaceae family that has been widely used in traditional medicine for treating various diseases. Nevertheless, its effect on human pancreatic cancer remains poorly investigated. In the present study, we screened the phytochemical content of Anchusa strigosa aqueous extracts obtained by maceration and ultrasound-assisted methods (ASM and ASU, respectively) and evaluated their antioxidant effects. We also investigated their anticancer effects and possible underlying mechanisms. The results show that both extracts were rich in bioactive molecules, with slight differences in their composition. Both extracts exhibited remarkable antioxidant potential and potent radical-scavenging activity in vitro . Additionally, non-cytotoxic concentrations of both extracts attenuated cell proliferation in a time- and concentration-dependent manner, which was associated with a decrease in the proliferation marker Ki67 and an induction of the intrinsic apoptotic pathway. Furthermore, the extracts increased the aggregation of pancreatic cancer cells and reduced their migratory potential, with a concomitant downregulation of integrin β1. Finally, we showed that the ASM extract caused a significant decrease in the levels of COX-2, an enzyme that has been linked to inflammation, carcinogenesis, tumor progression, and metastasis. Taken together, our findings provide evidence that A. strigosa extracts, particularly the extract obtained using the maceration method, have a potential anticancer effect and may represent a new resource for the design of novel drugs against pancreatic cancer

Ethanolic Extract of Origanum Syriacum L. Leaves Exhibits Potent Anti-Breast Cancer Potential and Robust Antioxidant Properties.

International audienceBackground: Breast cancer (BC) is the second most common cancer overall. In women, BC is the most prevalent cancer and the leading cause of cancer-related mortality. Triple-negative BC (TNBC) is the most aggressive BC, being resistant to hormonal and targeted therapies. HYPOTHESIS/PURPOSE: The medicinal plant Origanum syriacum L. is a shrubby plant rich in bioactive compounds and widely used in traditional medicine to treat various diseases. However, its therapeutic potential against BC remains poorly investigated. In the present study, we screened the phytochemical content of an ethanolic extract of O. syriacum (OSEE) and investigated its anticancer effects and possible underlying mechanisms of action against the aggressive and highly metastatic human TNBC cell line MDA-MB-231. METHODS: MTT, trans-well migration, and scratch assays were used to assess cell viability, invasion, or migration, respectively. Antioxidant potential was evaluated in vitro using the DPPH radical-scavenging assay and levels of reactive oxygen species (ROS) were assessed in cells in culture using DHE staining. Aggregation assays were used to determine cell-cell adhesion. Flow cytometry was used to analyze cell cycle progression. Protein levels of markers of apoptosis (BCL-2, pro-Caspase3, p53), proliferation (p21, Ki67), cell migration, invasion, or adhesion (FAK, E-cadherin), angiogenesis (iNOS), and cell signaling (STAT3, p38) were determined by immunoblotting. A chorioallantoic Membrane (CAM) assay evaluated in ovo angiogenesis. RESULTS: We demonstrated that OSEE had potent radical scavenging activity in vitro and induced the generation of ROS in MDA-MB-231 cells, especially at higher OSEE concentrations. Non-cytotoxic concentrations of OSEE attenuated cell proliferation and induced G(0)/G(1) cell cycle arrest, which was associated with phosphorylation of p38 MAPK, an increase in the levels of tumor suppressor protein p21, and a decrease of proliferation marker protein Ki67. Additionally, only higher concentrations of OSEE were able to attenuate inhibition of proliferation induced by the ROS scavenger N-acetyl cysteine (NAC), indicating that the anti-proliferative effects of OSEE could be ROS-dependent. OSEE stimulated apoptosis and its effector Caspase-3 in MDA-MB-231 cells, in correlation with activation of the STAT3/p53 pathway. Furthermore, the extract reduced the migration and invasive properties of MDA-MB-231 cells through the deactivation of focal adhesion kinase (FAK). OSEE also reduced the production of inducible nitric oxide synthase (iNOS) and inhibited in ovo angiogenesis. CONCLUSION: Our findings reveal that OSEE is a rich source of phytochemicals and has robust anti-breast cancer properties that significantly attenuate the malignant phenotype of MD-MB-231 cells, suggesting that O. syriacum may not only act as a rich source of potential TNBC therapeutics but may also provide new avenues for the design of novel TNBC drugs

Optimizing 1D 1H-NMR profiling of plant samples for high throughput analysis: extract preparation, standardization, automation and spectra processing

Introduction Proton nuclear magnetic resonance spectroscopy (H-1-NMR)-based metabolomic profiling has a range of applications in plant sciences. Objectives The aim of the present work is to provide advice for minimizing uncontrolled variability in plant sample preparation before and during NMR metabolomic profiling, taking into account sample composition, including its specificity in terms of pH and paramagnetic ion concentrations, and NMR spectrometer performances. Methods An automation of spectrometer preparation routine standardization before NMR acquisition campaign was implemented and tested on three plant sample sets (extracts of durum wheat spikelet, Arabidopsis leaf and root, and flax leaf, root and stem). We performed H-1-NMR spectroscopy in three different sites on the wheat sample set utilizing instruments from two manufacturers with different probes and magnetic field strengths. The three collections of spectra were processed separately with the NMRProcFlow web tool using intelligent bucketing, and the resulting buckets were subjected to multivariate analysis. Results Comparability of large- (Arabidopsis) and medium-size (flax) datasets measured at 600MHz and from the wheat sample set recorded at the three sites (400, 500 and 600MHz) was exceptionally good in terms of spectral quality. The coefficient of variation of the full width at half maximum (FWHM) and the signal-to-noise ratio (S/N) of two selected peaks was comprised between 5 and 10% depending on the size of sample set and the spectrometer field. EDTA addition improved citrate and malate resonance patterns for wheat sample sets. A collection of 22 samples of wheat spikelet extracts was used as a proof of concept and showed that the data collected at the three sites on instruments of different field strengths and manufacturers yielded the same discrimination pattern of the biological groups. Conclusion Standardization or automation of several steps from extract preparation to data reduction improves data quality for small to large collections of plant samples of different origins

NMR and LC-MS-Based Metabolomics to Study Osmotic Stress in Lignan-Deficient Flax

International audienceLignans, phenolic plant secondary metabolites, are derived from the phenylpropanoid biosynthetic pathway. Although, being investigated for their health benefits in terms of antioxidant, antitumor, anti-inflammatory and antiviral properties, the role of these molecules in plants remains incompletely elucidated; a potential role in stress response mechanisms has been, however, proposed. In this study, a non-targeted metabolomic analysis of the roots, stems, and leaves of wild-type and PLR1-RNAi transgenic flax, devoid of (+) secoisolariciresinol diglucoside ((+) SDG)-the main flaxseed lignan, was performed using H-1-NMR and LC-MS, in order to obtain further insight into the involvement of lignan in the response of plant to osmotic stress. Results showed that wild-type and lignan-deficient flax plants have different metabolic responses after being exposed to osmotic stress conditions, but they both showed the capacity to induce an adaptive response to osmotic stress. These findings suggest the indirect involvement of lignans in osmotic stress response