Enhanced Production of Nitrogenated Metabolites with Anticancer Potential in Aristolochia manshuriensis Hairy Root Cultures.

Aristolochia manshuriensis is a relic liana, which is widely used in traditional Chinese herbal medicine and is endemic to the Manchurian floristic region. Since this plant is rare and slow-growing, alternative sources of its valuable compounds could be explored. Herein, we established hairy root cultures of A. manshuriensis transformed with Agrobacterium rhizogenes root oncogenic loci (rol)B and rolC genes. The accumulation of nitrogenous secondary metabolites significantly improved in transgenic cell cultures. Specifically, the production of magnoflorine reached up to 5.72 mg/g of dry weight, which is 5.8 times higher than the control calli and 1.7 times higher than in wild-growing liana. Simultaneously, the amounts of aristolochic acids I and II, responsible for the toxicity of Aristolochia species, decreased by more than 10 fold. Consequently, the hairy root extracts demonstrated pronounced cytotoxicity against human glioblastoma cells (U-87 MG), cervical cancer cells (HeLa CCL-2), and colon carcinoma (RKO) cells. However, they did not exhibit significant activity against triple-negative breast cancer cells (MDA-MB-231). Our findings suggest that hairy root cultures of A. manshuriensis could be considered for the rational production of valuable A. manshuriensis compounds by the modification of secondary metabolism.

A Brief Overview of Potential Treatments for Viral Diseases Using Natural Plant Compounds: The Case of SARS-Cov.

The COVID-19 pandemic, as well as the more general global increase in viral diseases, has led researchers to look to the plant kingdom as a potential source for antiviral compounds. Since ancient times, herbal medicines have been extensively applied in the treatment and prevention of various infectious diseases in different traditional systems. The purpose of this review is to highlight the potential antiviral activity of plant compounds as effective and reliable agents against viral infections, especially by viruses from the coronavirus group. Various antiviral mechanisms shown by crude plant extracts and plant-derived bioactive compounds are discussed. The understanding of the action mechanisms of complex plant extract and isolated plant-derived compounds will help pave the way towards the combat of this life-threatening disease. Further, molecular docking studies, in silico analyses of extracted compounds, and future prospects are included. The in vitro production of antiviral chemical compounds from plants using molecular pharming is also considered. Notably, hairy root cultures represent a promising and sustainable way to obtain a range of biologically active compounds that may be applied in the development of novel antiviral agents.

Cell-wall polysaccharide composition and glycanase activity of Silene vulgaris callus transformed with rolB and rolC genes.

The aim of this research is to investigate the effects of the Agrobacterium rhizogenes rol genes on the composition of cell-wall polysaccharides and glycanase activity in the campion callus. The expression of the rolC gene reduces the yield of campion pectin, while the expression of the rolB or rolC gene inhibits the volumetric production of both pectin and intracellular arabinogalactan. The rol genes are involved in regulating the activity of glycanases and esterases, thereby contributing to the modification of polysaccharide structures, their molecular weight (Mw) and the degree of pectin methyl esterification (DE). The increase in pectin arabinose residue appears to be connected to a decrease in intracellular and extracellular α-l-arabinofuranosidase activity in transgenic campion calluses. In transgenic calluses expressing the rolB and rolC genes, the increase in pectin galactose residue is likely due to a decrease in ß-galactosidase activity. The decrease in the Mw of pectin and its d-galacturonic acid content appears to be connected to an increase in extracellular polygalacturonase activity. Finally, the increase in pectinesterase activity causes a decrease in the DE of pectin. Thus, the expression of rolB and rolC genes in campion callus has a considerable effect on pectin's sugar composition, DE and Mw, while it appears to have an insignificant influence on intracellular and extracellular arabinogalactans.

Polyphenolic compounds from callus cultures of Iris pseudacorus.

A callus culture of Iris pseudacorus L. (Iridaceae) was established from plant leaves using a modified Murashige and Skoog medium. A derivative of cinnamic acid (lavandoside) (1), a neolignan (dehydrodiconiferyl alcohol-4-O-beta-D-glucopyranoside) (2) as well as three isoflavonoids, tectoridin (3), tectorigenin (4), and iristectorigenin A (5) were isolated from the callus culture. Under normal conditions, the calli accumulated 0.4% DW of polyphenols. The addition of phenylalanine to a concentration of 1 mM resulted in a 1.5-fold increase in isoflavonoid production, allowing the accumulation of 0.69% of polyphenols in the callus dry weight. Tectorigenin, a promising chemotherapeutic and chemopreventive agent for the treatment of carcinomas, was produced in I. pseudacorus calli in high quantities (0.3% DW).

The rolB gene suppresses reactive oxygen species in transformed plant cells through the sustained activation of antioxidant defense.

The rolB (for rooting locus of Agrobacterium rhizogenes) oncogene has previously been identified as a key player in the formation of hairy roots during the plant-A. rhizogenes interaction. In this study, using single-cell assays based on confocal microscopy, we demonstrated reduced levels of reactive oxygen species (ROS) in rolB-expressing Rubia cordifolia, Panax ginseng, and Arabidopsis (Arabidopsis thaliana) cells. The expression of rolB was sufficient to inhibit excessive elevations of ROS induced by paraquat, menadione, and light stress and prevent cell death induced by chronic oxidative stress. In rolB-expressing cells, we detected the enhanced expression of antioxidant genes encoding cytosolic ascorbate peroxidase, catalase, and superoxide dismutase. We conclude that, similar to pathogenic determinants in other pathogenic bacteria, rolB suppresses ROS and plays a role not only in cell differentiation but also in ROS metabolism.

High rabdosiin and rosmarinic acid production in Eritrichium sericeum callus cultures and the effect of the calli on masugi-nephritis in rats.

During an investigation of plant cell cultures that might be useful in the treatment of renal disorders, we established a vigorously-growing E-4 callus culture of Eritrichium sericeum that produced large amounts of caffeic acid metabolites, (-)-rabdosiin (1.8% dry wt) and rosmarinic acid (4.6% dry wt). Elicitation of the calli by methyl jasmonate induced a 38% increase in total polyphenol production. The most efficient method of eliciting (-)-rabdosiin biosynthesis was through the treatment of E-4 calli with cuprum glycerate, which induced an increase in (-)-rabdosiin production of as much as 4.1% dry wt. Oral administration of E-4 callus biomass (100 mg/kg/d for 30 d) to rats with induced Masugi-nephritis caused an increase in diuresis and lowered creatinine excretion and proteinuria levels as compared with Masugi-nephritis untreated rats. While all of the Masugi-nephritis untreated rats began to suffer, near a quarter of the E-4 treated rats remained in good health. This result indicates that the E-4 culture has the potential to alleviate the symptoms associated with nephritis.

The rolC gene induces expression of a pathogenesis-related beta-1,3-glucanase in transformed ginseng cells.

The Agrobacterium rhizogenes rolC oncogene is capable of stimulating production of secondary metabolites in transformed plant cells that suggest its possible involvement in plant defense reactions. We tested whether the gene could also affect production of pathogenesis-related proteins. Using a well-known group of PR-proteins, such as beta-1,3-glucanases, we observed a 10-fold increase in total beta-1,3-glucanase activity in rolC-transformed Panax ginseng cells compared with normal cells. The increase was due to the production of a salicylic acid-activated beta-1,3-glucanase isoform. We isolated cDNA of the corresponding beta-1,3-glucanase gene (Pg-glu1), which shared 38-60% sequence identity with previously reported sequences of plant beta-1,3-glucanases at the protein level. Levels of Pg-glu1 mRNA transcripts were tightly correlated with expression of the rolC gene. Our data, together with previously reported information, indicate that A. rhizogenes can activate plant defense reactions via expression of T-DNA oncogenes.

Caffeic acid metabolites from Eritrichium sericeum cell cultures.

Eritrichium sericeum (Boraginaceae) callus and root cultures were established and analyzed for caffeic acid metabolite (CAM) production. Two substances, (-)-rabdosiin and rosmarinic acid, were identified as main CAMs produced by these cultures. The E. sericeum Er-1 root culture accumulated up to 1.5 % and 4.5 % DW of (-)-rabdosiin and rosmarinic acid, respectively. Rabdosiin in the Lithospermum erythrorhizon callus cultures was produced exclusively as the (+)-enantiomer while in both Eritrichium cultures it occurred as the (-)-enantiomer. The E. sericeum Er-1 culture accumulated 3-fold higher levels of CAMs than the L. erythrorhizon culture. A new compound, named eritrichin, was isolated from the cultured E. sericeum cells. The structure of this compound was established as (2R)-3-(3,4-dihydroxyphenyl)-2-[4-(3,4-dihydroxyphenyl)-6,7-dihydroxy-2-naphthoyloxy]propanoic acid on the basis of spectral data.