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Abstract and Figures
Polygonum multiflorum is an important medicinal plant. Hairy roots systems obtained by transforming plant tissues with the natural genetic engineer Agrobacterium rhizogenes can produce valuable biological active substances, which have immense potential in the pharmaceutical industry.
To optimize the inductive and cultural conditions of P. multiflorum hairy roots and to identify the major active secondary metabolites in hairy roots.
P. multiflorum hairy root were mediated with A. rhizogenes R1601 to induce hairy roots. Four combinations, including Murashige-Skoog (MS), 1/2 MS, B5, and White, were investigated to optimize the culture medium. MS medium was selected for the growth measurement. The qualitative and quantitative determinations of free anthraquinone in hairy roots were compared with the calli and aseptic plantlets using high-performance liquid chromatography.
The inductive rates of hairy roots by leaves were higher than for any other explants. The presence of agropine in the P. multiflorum hairy roots confirmed that they were indeed transgenic. MS medium was the most suitable of the four media for hairy root growth. Meanwhile, the growth kinetics and nutrient consumption results showed that the hairy roots displayed a sigmoidal growth curve and that their optimal inoculation time was 18-21 days. The determination of the anthraquinone constituents indicated that the rhein content of the hairy roots reached 2.495 μg g(-1) and was 2.55-fold higher than that of natural plants.
Transgenic hairy roots of P. multiflorum could be one of the most potent materials for industrial-scale production of bioactive anthraquinone constituents.
To read the full-text of this research, you can request a copy directly from the authors.
... Eclipta prostrata (L.) L. produces phenolic acids such as chlorogenic acid and its derivative 3,5-di-O-caffeoylquinic acid (3,5-diCQA) (Lee et al. 2010). Chlorogenic acid and its derivatives have demonstrated several biological activities, such as antioxidant, antibiosis, anti-inflammatory, antivirus, and antitumor activities (Huang et al. 2014;Ali et al. 2017;Liu et al. 2018;Devrnja et al. 2020). More recently, studies by molecular docking using a serine-type protease (Mpro), a SARS-CoV-2 virus polyprotein, showed that the obtained binding energies of 3,5-diCQA were closed to remdesivir (antiviral compound), indicating stronger binding to Mpro protein and promising antiviral compounds that could treat Covid-19 (Shah et al. 2021). ...
... Obtained results indicated the 21st day of culture growth as the most effective day for starting elicitation. The growth profile of E. prostrata (L.) L. hairy roots resemble Polygonum multiflorum hairy roots where it was found that the ideal time for the induction of anthraquinones was between the 18th and 21st days of elicitation (Huang et al. 2014). Similarly, studies have reported the 21st day of elicitation as the end of the exponential growth phase of Artemisia annua hairy roots (Sivakumar et al. 2010). ...
Eclipta prostrata (L.) L. is widely used in traditional medicine for treatment of hepatitis, poisoning from snake bites and viral infections. Pharmacological studies confirmed its antioxidant, anti-inflammatory and anticancer activities. The efficacy of E. prostrata (L.) L. extracts has been correlated to phenylpropanoids such as flavonoids, coumestans and caffeoylquinic acid derivatives. In this work, the production of wedelolactone, demethylwedelolactone and 3,5-di-O-caffeoylquinic acid (3,5-diCQA) in hairy root cultures of E. prostrata (L.) L. C19 clone was increased after addition of eliciting agents jasmonic acid (JA) or methyl jasmonate (MeJA) at multiple concentrations. Cultures elicited with 100 μM of JA saw a 5.2 fold increase in wedelolactone (from 0.72 to 3.72 mg/g d.w.), a 1.6 fold increase in demethylwedelolactone (from 5.54 to 9.04 mg/g d.w.) and a 2.47 fold increase in 3,5-diCQA (from 18.08 to 44.71 mg/g d.w.). Obtained data validate the potential of E. prostrata (L.) L. hairy root cultures as a production system of wedelolactone, demethylwedelolactone and especially 3,5-diCQA, which has recently been reported to possess activity against coronavirus disease (Covid-19) by in silico computational studies.
The online version contains supplementary material available at 10.1007/s11240-021-02201-4.
... Meanwhile, the hairy roots growth kinetics and nutrient consumption findings revealed that they had a sigmoidal curve and that the best inoculation period for them was 18-21 days. The analysis of the anthraquinone component shown that the hairy roots had a rhein concentration of 2.495 g/g, which was 2.55 times greater than that of natural plants . ...
Medicinal plants are an inevitable source of pharmaceutical drugs and most of the world population depends on these plants for health benefits. The increasing global demand for bioactive compounds from medicinal plants has posed a great threat to their existence due to overexploitation. Adventitious root and hairy root culture systems are an alternative approach to the conventional method for mass production of valuable compounds from medicinal plants owing to their rapid growth, biosynthetic and genetic stability. The main purpose of this review is to investigate the recent scientific research published worldwide on the application of adventitious and hairy root cultures to produce valuable compounds from medicinal plants. Furthermore, a comparison of adventitious root vs. hairy root cultures to produce valuable compounds has also been discussed. Various aspects such as medium composition, carbon source, pH, amount of macronutrients, optimization strategy, scale-up cultures, and use of biotic abiotic and nano-elicitors at various concentrations are the topic of discussion in this review. Several studies on adventitious and hairy root cultures of Polygonum multiflorum¸ Withania somnifera¸ Echinacea purpurea and Ajuga bracteosa have been discussed in detail which highlights the importance of elicitation strategies and bioreactor system, presenting commercial applications.
... We used rhizome cuttings from 20 R. japonica clones and 13 R. × bohemica clones, with approximately ten rhizome fragments, each containing two nodes and thus one intact internode, from each clone. After removal of all ne roots, the rhizome fragments were surface-sterilized using the method described by (Huang et al., 2014). ...
There is increasing evidence that microbes play a key role in some plant invasions. A diverse and widespread but little understood group of plant-associated microbes are the fungal root endophytes of the order Sebacinales. They are associated with exotic populations of invasive knotweed ( Reynoutria ssp.) in Europe, but their effects on the invaders are unknown. We used the recently isolated Sebacinales root endophyte S erendipita herbamans to experimentally inoculate invasive knotweed and study root colonisation and effects on knotweed growth under different environmental conditions. We verified the inoculation success and fungal colonisation through immunofluorescence microscopy and qPCR. We found that S. herbamans strongly colonized invasive knotweed in low-nutrient and shade environments, but much less under drought or benign conditions. At low nutrients, the endophyte had a positive effect on plant growth, whereas the opposite was true under shaded conditions. Our study demonstrates that the root endophyte S. herbamans has the potential to colonize invasive knotweed fine roots and impact its growth, and it could thus also play a role in natural populations. Our results also show that effects of fungal endophytes on plants can be strongly environment-dependent, and may only be visible under stressful environmental conditions.
... In another study, Ho et al. (2018b) tested the effect of elicitors like methyl jasmonate (MeJA), salicylic acid (SA), yeast extract (YE), and chitosan on the biomass and metabolite accumulation in adventitious root cultures of PM and reported that lower concentrations of elicitors, i.e., 50 μM MeJA and SA and 50 mg/LYE improved metabolite accumulation. Among the elicitor treatments, Lin et al. (2003) Cell suspension cultures 2,3,5,4′-tetrahydroxystilbene-2-O-β-D-glucoside (TSG) Shao et al. (2012) Emodin, physcoin, and phenolic compounds Thiruvengadam et al. (2016) Hairy root cultures Rhein, emodin, physcion Yu et al. (2006) Rhein, emodin, chrysophanol Huang et al. (2014) Emodin, physcion, and phenolic compounds Thiruvengadam et al. (2014) Anthraquinones and phenolic compounds Ho et al. (2018a, b) Adventitious root cultures Anthraquinones and phenolic compounds Ho et al. (2017) Anthraquinones and phenolic compounds Ho et al. (2018a, b) MeJA treatment (50 μM) was responsible for 3.12-fold increment in physcion, 1.89-fold ferulic acid, 1.35-fold increment in p-hydroxybenzoic acid, and 2.42-fold increment in biochanin. The above reports suggest that shoot, cell/hairy root/adventitious root suspension cultures of PM are useful for the production of bioactive compounds. ...
A vast array of plant-based compounds has enriched red biotechnology to serve the human health and food. A peculiar medicinal plant which was an element of traditional Chinese medicine for centuries as a liver and kidney tonic, for life longevity and hair blackening, is Polygonum multiflorum Thunb. (PM) which is popularly known as “He shou wu” or “Fo-ti” and is rich in chemical components like stilbenes, quinones, and flavonoids which have been used as anti-aging, anti-alopecia, anti-cancer, anti-oxidative, anti-bacterial, anti-hyperlipidemia, anti-atherosclerosis, and immunomodulating and hepatoprotective agents in the modern medicine. The health benefits from PM are attained since long through commercial products such as PM root powder, extract, capsules, tincture, shampoo, and body sprays in the market. Currently, the production of these pharmaceuticals and functional foods possessing stilbenes, quinones, and flavonoids is through cell and organ cultures to meet the commercial demand. However, hepatotoxic effects of PM-based products are the stumbling blocks for its long-term usage. The current review encompasses a comprehensive account of bioactive compounds of PM roots, their biological activities as well as efficacy and toxicity issues of PM ingredients and future perspectives.
... Although qualitatively 23 types of polyphenolic compounds were detected in both NT and hairy root cultures, hairy root of P. multiflorum showed higher amount of total phenolic content including higher amount of pyrogallol, hesperidin, naringenin, and formononetin, as well as significantly higher flavonoid content and antioxidant and antimicrobial activity in optimized condition compared to untransformed roots (Thiruvengadam et al. 2014). Similarly, optimum growth (i.e., up to 19.30-fold) and significantly higher accumulation of anthraquinone constituents (Rhein 2.495 μg g À1 ; 2.55-fold than in wildtype plant) were obtained when hairy root of P. multiflorum culture in full-strength MS medium was compared to other half-strength MS, B 5 , and White media (Huang et al. 2014a). ...
In the past three decades, differentiated hairy root culture-related researches gained a great attention due to the equal or greater bio-production capacity of low amount, high-value secondary metabolites as compared to their parent plants with several advantages over undifferentiated cell suspension cultures in plants. This was mainly because hairy roots are capable of auxin-independent rapid growth and are genetically and biochemically stable, with high productivity and suitability for adaptation to large-scale systems. Nowadays, hairy root cultures of various plant species offer a novel promising opportunity and great prospects for in vitro mass production of economically important bioactive metabolites. At present, the productivity of desired compounds by hairy root cultures is generally too low to fulfill the demands of pharmaceutical industry owing to various biological and technological limitations. Screening and selection for high-yielding root lines and optimization of the culture media and the culture conditions like type of nutrient medium, salt strength, source of carbon and concentration, source of nitrogen and the ratio of NH4⁺/NO3⁻, concentration of phosphate, inoculum density, hydrogen ion concentration, temperature, and light intensity and quality have been taken as yield enhancement strategies among others, to produce desired secondary metabolites using hairy root cultures. Feasibility of commercial application of hairy root culture in bioreactors requires several optimization steps. This review highlights some of the recent progress and outlines future prospects for metabolite production and yield enhancement approaches in hairy root cultures for producing bioactive substances.
... Some of the advantages provided by hairy root cultures include high growth rates without the need for plant growth regulators, genetically and biochemically stable cultures, and a similar capacity for production of secondary metabolites than cell suspension cultures (Guillon et al. 2006;Georgiev et al. 2012;Mora-Pale et al. 2014). A wide range of natural products has been produced using this system, including lignans, steroids, anthraquinones and alkaloids (Doma et al. 2012;Huang et al. 2014;Pandey et al. 2014;Wawrosch et al. 2014). However, metabolite 1 3 production requires that the compound of interest be one that is normally synthesized within the roots of the source plant, so this limits the versatility of the hairy root system (Ochoa-Villarreal et al. 2016). ...
Plant tissue culture as an important tool for the continuous production of active compounds including secondary metabolites and engineered molecules. Novel methods (gene editing, abiotic stress) can improve the technique. Humans have a long history of reliance on plants for a supply of food, shelter and, most importantly, medicine. Current-day pharmaceuticals are typically based on plant-derived metabolites, with new products being discovered constantly. Nevertheless, the consistent and uniform supply of plant pharmaceuticals has often been compromised. One alternative for the production of important plant active compounds is in vitro plant tissue culture, as it assures independence from geographical conditions by eliminating the need to rely on wild plants. Plant transformation also allows the further use of plants for the production of engineered compounds, such as vaccines and multiple pharmaceuticals. This review summarizes the important bioactive compounds currently produced by plant tissue culture and the fundamental methods and plants employed for their production.
... Taxol (plaxitaxol): The bark of the taxus tree (Taxus wallichiana) produces a complex diterpene alkaloid which is a potent anticancer agent because of its unique mode of action on Artemisia annua Artemisinin Callus  Artemisia Annua Artemisinin Callus  Aspidosperma ramiflorum Ramiflorin Callus  Azadirachta indica Azadirachtin Suspension  Azadirachta indica Azadirachtin Anther culture  Azadirachta indica Azadirachtin Suspension  Azadirachta indica Azadirachtin Hairy root  Azadirachta indica Azadirachtin Hairy root  Brucea javanica Cathin Suspension  Bupleurum falcatum Saikosaponins Root  Bupleurum chinensis Saikosaponins Root  Camellia chinensis Flavones Callus  Capsicum annum Capsiacin Callus  Capsicum chinense Capsiacin Suspension  Capsicum chinense Capsiacin Cotyledon  Capsicum chinense Capsiacin Suspension  Cassia acutifolia Anthraquinones Suspension  Aloe barbadensis Anthraquinone Callus  Morinda citrifola Anthraquinone Suspension  ( ...
Bioactive natural products are economically important as drugs, fragrances, pigments, food additives and pesticides. The biotechnological tools are important to select, multiply, improve and analyze medicinal plants for production of such products. The utilization of medicinal plant cells for the production of natural or recombinant compounds of commercial interest has gained increasing attention over the past decades. Plant tissue culture systems are possible source of valuable medicinal compounds, fragrances and colorants, which cannot be produced by microbial cells or chemical synthesis. In vitro production of bioactive natural products in plant cell suspension culture has been reported from various medicinal plants and bioreactors are the key step towards commercial production. Genetic transformation is a powerful tool for enhancing the productivity of novel products; especially by Agrobacterium tumefacians . Combinatorial biosynthesis is another approach in the generation of novel natural products and for the production of rare and expensive natural products. Recent advances in the molecular biology, enzymology and bioreactor technology of plant cell culture suggest that these systems may become a viable source of important secondary metabolites. Genetic fingerprinting could be a powerful tool in the field of medicinal plants to be used for correct germplasm identification. In addition, when linked to emerging tools such as metabolomics and proteomics, providing fingerprints of the plant’s metabolites or protein composition, it gives data on phenotypic variation, caused by growth conditions or environmental factors, and also yield data on the genes involved in the biosynthesis. DNA profiling techniques like DNA microarrays serve as suitable high throughput tools for the simultaneous analysis of multiple genes and analysis of gene expression that becomes necessary for providing clues about regulatory mechanisms, biochemical pathways and broader cellular functions. New and powerful tools in functional genomics can be used in combination with metabolomics to elucidate biosynthetic pathways of natural products.
There is increasing evidence that microbes play a key role in some plant invasions. A diverse and widespread but little understood group of plant-associated microbes are the fungal root endophytes of the order Sebacinales. They are associated with exotic populations of invasive knotweed (Reynoutria ssp.) in Europe, but their effects on the invaders are unknown. We used the recently isolated Sebacinales root endophyte Serendipita herbamans to experimentally inoculate invasive knotweed and study root colonisation and effects on knotweed growth under different environmental conditions. We verified the inoculation success and fungal colonisation through immunofluorescence microscopy and qPCR. We found that S. herbamans strongly colonized invasive knotweed in low-nutrient and shade environments, but much less under drought or benign conditions. At low nutrients, the endophyte had a positive effect on plant growth, whereas the opposite was true under shaded conditions. Our study demonstrates that the root endophyte S. herbamans has the potential to colonize invasive knotweed fine roots and impact its growth, and it could thus also play a role in natural populations. Our results also show that effects of fungal endophytes on plants can be strongly environment-dependent, and may only be visible under stressful environmental conditions.
The current trends of consumer-driven demands for natural therapeutics and the availability of evidence-based phytopharmaceuticals from traditional knowledge has once again brought the medicinal plants into forefront of health. In 2019, World Health Organization global report on traditional and complementary medicine has also substantiated the revival of herbal medicine including its convergence with conventional medicine for the management and prevention of diseases. It means these industries need plenty of plant materials to meet the unprecedented demands of herbal formulations. However, it is pertinent to mention here that around 70–80% medicinal plants are sourced from the wild and most of such highly acclaimed plants are listed under Rare, Endangered and Threatened species by IUCN. Additionally, over 30% traditional health formulations are based on underground plant parts, which lead to the uprooting of plants. Overharvesting from limited plant populations, meager conventional cultivation and a rising fondness for natural products exerting enormous pressure on natural habitats. Therefore, the nondestructive means of phytochemical production employing biotechnological tools could be used for sustainable production and consumption patterns. In recent years, a number of reports described the use of adventitious roots induced under in vitro conditions for the extraction of phytochemicals on a sustainable basis. In this article, efforts are made to review recent developments in this area as well as understand the induction mechanisms of adventitious roots, their in vitro cultivation, probable factors that affect the growth and metabolite production, and assess the possibility of industrial scale production to meet the rising demands of natural herbs.
Medicinal plants produce valuable secondary metabolites with anticancer, analgesic, anticholinergic or other activities, but low metabolite levels and limited available tissue restrict metabolite yields. Transformed root cultures, also called hairy roots, provide a feasible approach for producing valuable secondary metabolites. Various strategies have been used to enhance secondary metabolite production in hairy roots, including increasing substrate availability, regulating key biosynthetic genes, multigene engineering, combining genetic engineering and elicitation, using transcription factors (TFs), and introducing new genes. In this review, we focus on recent developments in hairy roots from medicinal plants, techniques to boost production of desired secondary metabolites, and the development of new technologies to study these metabolites. We also discuss recent trends, emerging applications, and future perspectives.
Polygonum chinense L (Fam. – Polygonaceae) is an erect herb, height 1.8 m, of which leaves are oblong‐lanceolate to elliptic in shape; flowers white or pink or purplish red in color, inflorescence – cymose or corymbose, distributed in India and warm temperate regions of Asia. The aerial and underground parts of this species are used in the treatment of diarrhea, dyspepsia, and hemorrhoids. Polygonum amplexicaule is used in treatment of bone fractures, rheumatoid arthritis, muscle injury, and pain as prescribed in Chinese traditional system of medicine. Callus and suspension cultures were established from roots and shoots of Polygonum hydropiper, and the polygodial production was increased. Similarly, the cell cultures of Polygonum multiflorum were established on Murashige and Skoog medium with supplementation of 6‐benzylaminopurine and kinetin.
The review highlights the history of studying the phenomenon of hairy roots and biodiversity strains of Agrobacterium rhizogenes used to create these roots. Briefly discusses the features of the interaction of A. rhizogenes with plants and the process of hairy roots formation. The main emphasis in this review is given to the practical use of hairy roots as producers of the biologically active substances, including producing raw materials for further chemical synthesis. In recent years hairy roots became commercial application, so part of the review devoted to the description of a special bioreactors developed for the industrial cultivation and the development of the methods for long-term preservation of the hairy roots‘ cultures. At the same time hairy roots remain one of the major models in molecular biology and physiology of plants, but their potential for fundamental research is not fully exploited. References amounted to more than 450 publications covering a 115-year period.
An easily self-assembled and gelated octa-peptide FHFDFHFD was chosen as a novel drug delivery system (DDS) for both tanshinone IIA and total tanshinone extract. The DDS showed increased loading capacity, sustained drug release and better anticancer capability. Our research proved that a hydrogel DDS of other traditional Chinese medicines is possible.
Plants have been exploited as a source of medicinal substances for years. Nowadays, achievements of modern science, including molecular biotechnology, allow their huge potential to be utilized. They have become a promising platform for the production of valuable compounds such as biopharmaceuticals. Among the various plant systems used for this purpose, hairy root cultures are also applied for the production of recombinant proteins and secondary metabolites. For this purpose plant cells of selected species are genetically transformed using different strains of Agrobacterium rhizogenes carrying the desired genes. The next steps of this process include stable and efficient expression of these genes. Hairy root cultures exhibit a number of features which make them attractive compared to various pro- and eukaryotic cell systems including other plant models. Their main advantages are: relatively low production costs, ease of scale-up, production of compounds typical for eukaryotic cells with post-translational modifications, biological safety, and in many cases there is no need for complex purification techniques of the final product. Several compounds that are successfully obtained using this production strategy are valuable pharmaceuticals. This group includes selected cytokines, vaccine antigens and antibodies.
Recalcitrance of most large-seeded legumes, such as peanut, to regeneration and genetic transformation has hampered studies on gene function and efforts for genetic improvement. Agrobacterium rhizogenes-mediated transformation provides a system for rapid and efficient transformation of plant tissues. In this study, embryonic axes along with cotyledons of peanut were injected with a suspension culture of A. rhizogenes using microliter syringes. The influence of several factors such as plant genotype, A. rhizogenes culture stage, co-culture period of A. rhizogenes, and acetosyringone concentration in the co-cultivation medium have been evaluated. It is found that A. rhizogenes-mediated transformation of peanut is genotype-independent. Up to 61% transformation was recorded when embryonic axes were co-cultivated with 5 × 107A. rhizogenes cells from logarithmic phase for 2 days on co-culture medium containing 50 μmol l−1 acetosyringone. Composite plants with transgenic roots were harvested after 45 days of treatment. Furthermore, this method was applied to assess the insecticidal activity of a synthetic cry8Ea1 gene against Holotrichia parallela in transgenic roots of peanut.
Cell suspension cultures of Gymnema sylvestre treated with four different elicitors, methyl jasmonate (MJ), yeast extract, chitin and pectin were studied for the production
of gymnemic acid as gymnemagenin equivalent, that was analyzed by high performance liquid chromatography (HPLC). All the four
tested elicitors induced gymnemic acid production in cell suspension cultures. Highest gymnemic acid content was achieved
following treatment with yeast extract (100.47 ± 0.28 mg/l), this was followed by MJ (70.43 ± 0.26 mg/l), pectin (64.19 ± 0.23 mg/l)
and chitin (62.72 ± 0.13 mg/l). The addition of elicitors has shown a significant influence on cell growth that affected cell
growth compared to respective controls. The highest gymnemic acid production was obtained after 20 days of elicitation in
cultures treated with 0.5 g l−l yeast extract, it was 5.25-folds greater than in control. These results suggest that the addition of an elicitor to Gymnema sylvestre cell suspension cultures could stimulate and enhance gymnemic acid production. In our present study we could able to overproduce
gymnemic acid up to 51.97 ± 0.26 mg l−l (dry weight basis) in yeast extract treated cell suspension cultures.
Rutagraveolens L. is a source of pharmacologically active compounds such as coumarins, furanocoumarins and furoquinolone alkaloids. Hypocotyls,
callus and shoots of R.graveolens were inoculated with bacteria from two Agrobacteriumrhizogenes strains. Hairy root cultures were established after inoculation of hypocotyls with wild A.rhizogenes strain LBA 9402. The transgenic nature of the regenerated tissue was confirmed by PCR amplification. Coumarins, furanocoumarins
and alkaloids present in the hairy root tissue were identified by GC and GC-MS and compared with those present in invitro
shoot cultures. The level of pinnarin and rutacultin, bergapten, isopimpinelin and xanthotoxin was approximately twofold higher
in hairy root than in shoot cultures. Two additional coumarins: osthole and osthenol, never been found in R.graveolens, were identified in hairy root tissue. Besides coumarins, alkaloids were identified: dictamnine, skimmianine, kokusaginine,
rybalinine and an isomer of rybalinine. The levels of nearly all coumarins and alkaloids in hairy roots cultured in the darkness
were higher than those accumulated under a photoperiod mode.
Spontaneous shoot regeneration was observed from Agrobacterium rhizogenes-induced hairy roots of Plumbago indica when these were incubated in liquid MS medium for a period of 3weeks under continuous light. Insertion of the rolB gene in putative transformed plants was confirmed by PCR and sequencing. Transformed plants grown for a period of 1week
on solid MS medium containing 0.5mgl−1 6-benzyladenine and then transplanted to growth regulator-free medium showed better overall growth than control plants. Transformed
plants had a higher root bio-biomass and an increased plumbagin content relative to non-transformed plants.
Stilbene glycosides were isolated from the ethanol extract of the roots of Polygonum multiflorum Thunb. Two samples were obtained; a fraction separated by macroporous resin and pure crystals of 2,3,5,4′-tetrahydroxystilbene 2-O-β-glucopyranoside. The antioxidant activities of these two samples were evaluated using antioxidant tests of rats in vivo. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of the serum and the organs (liver, heart and brain of rats) of d-galactose induced senile rats which were fed with stilbene glycoside, were increased; however, the content of 2-thiobarbituric acid-reactive substances (TBARS) was decreased. It is concluded that the stilbene glycoside from Polygonum multiflorum Thunb possesses high in vivo antioxidant activity.
In this study, the effects of ploidy level and
culture medium were studied on the production of tropane
alkaloids. We have successfully produced stable tetraploid
hairy root lines of Hyoscyamus muticus and their ploidy
stability was confirmed 30 months after transformation.
Tetraploidy affected the growth rate and alkaloid accumulation
in plants and transformed root cultures of Egyptian
henbane. Although tetraploid plants could produce 200%
higher scopolamine than their diploid counterparts, this
result was not observed for corresponding induced hairy root
cultures. Culture conditions did not only play an important
role for biomass production, but also significantly affected
tropane alkaloid accumulation in hairy root cultures. In spite
of its lower biomass production, tetraploid clone could
produce more scopolamine than the diploid counterpart
under similar growth conditions. The highest yields of scopolamine
(13.87 mg l-1) and hyoscyamine (107.7 mg 1-1)
were obtained when diploid clones were grown on medium
consisting of either Murashige and Skoog with 60 g/l
sucrose or Gamborg’s B5 with 40 g/l sucrose, respectively.
Although the hyoscyamine is the main alkaloid in the
H. muticus plants, manipulation of ploidy level and culture
conditions successfully changed the scopolamine/hyoscyamine
ratio towards scopolamine. The fact that hyoscyamine
is converted to scopolamine is very important due to the
higher market value of scopolamine.
We report on the transformation and expression in sugar beet (Beta vulgaris) hairy roots of a Nicotiana alata NaPI gene encoding a serine proteinase inhibitor (PI) that has been shown to effectively reduce the population of a number of insect pests. Using in-gel analysis, two PI protein activities were detected at approximately 24- and/or 28-kDa in hairy roots generated via Agrobacterium rhizogenes-mediated gene transfer. Immunoblot analysis revealed the presence of the expected ~40 kDa precursor, and in some transformants, a ~20 kDa processing intermediate and the mature 6-kDa PIs. In general, processing of the precursor in the clonal lines was reduced or not detected. The reduced efficiency of post-translational processing of the N. alata PI precursor may be attributed to modification and/or altered folding of the recombinant protein or distinct post-translational machinery functioning in sugar beet hairy roots and Nicotiana.
The T-DNA regions of the root-inducing (Ri) plasmid pRiA4b of Agrobacterium rhizogenes were characterized. Two regions, designated TL-DNA and TR-DNA, were found to be integrated and stably maintained in the plant genome. The TL-DNA spanned a 15- to 20-kilobase region of pRiA4b and was separated from the TR-DNA region by at least 15 kilobases of nonintegrated plasmid DNA. The TR-DNA region also spanned a 15- to 20-kilobase region of pRiA4b and included a region of homology to the tms morphogenic loci of the tumor-inducing (Ti) plasmid of Agrobacterium tumefaciens. Eighteen deletions and 95 transposon insertions were generated in the T-DNA regions and tested for alterations in virulence. Insertions into four loci in the TL-DNA affected the morphology of root formation of Kalanchoë diagremontiana leaves and stems, but had no visible effects on other host plants. Insertions into two loci (tms-1 and tms-2) in the TR-DNA eliminated virulence symptoms on all plants tested, with the exception of K. diagremontiana stems, where sparse root formation occurred. Complementation experiments with Ri and Ti plasmid T-DNA mutations indicate that the tms genes of the two plasmids serve similar functions and suggest a functional relationship between one or more genes of the TL-DNA and the cytokinin synthesis locus tmr of the Ti plasmid.
High-speed counter-current chromatography methods, combined with solvent partition, were applied to the systematic separation and purification of chemical components from Chinese medicinal herb Polygonum multiflorum extract. The aim of this paper is summing up the rules of solvent system selection for diverse fractions of herbal extract, and establishing the systematic pattern to screen the bioactive constituents rapidly. Nine compounds including emodin, chrysophanol, rhein, 6-OH-emodin, emodin-8-beta-D-glucoside, polygonimitin B, 2,3,5,4'-tetrahydroxystilbene-2-beta-D-glucoside, gallic acid and an unknown glycoside, which differed in quantity and polarity remarkably, were obtained. The purities of them were all above 97% as determined by high-performance liquid chromatography (HPLC), and their structures were identified by 1H NMR and electrospray ionization mass spectrometry (ESI-MS). The results demonstrated that HSCCC is a speedy and efficient technique for systematic isolation of bioactive components from traditional medicinal herbs.
Withania somnifera (L.) Dunal. (Indian ginseng) is an important medicinal plant which yields pharmaceutically active compounds called withanolides. The present work deals with optimization of parameters of hairy root culture of W. somnifera for the production of biomass and withanolide A. We also investigated the effects of carbon source [sucrose, glucose, fructose, maltose, glucose + fructose (1:1), fructose + sucrose (1:1) and sucrose + glucose (1:1)], sucrose concentration (1%, 2%, 3%, 4%, 6% and 8%) and the initial medium pH (4.0, 4.5, 5.0, 5.5, 5.8, 6.0 and 6.5) on growth and production of withanolide A in hairy root cultures of W. somnifera. We found that biomass accumulation and production of withanolide A was highest when sucrose was used as the carbon source (11.92 g l−1 DW and 11.96 mg g−1 DW of withanolide A). Further 3% sucrose concentration was found to be optimal for biomass accumulation (11.92 g l−1 DW) and 4% sucrose favoured the production of withanolide A (13.28 mg g−1 DW) in the tested range of concentrations (1–8%). The biomass of hairy roots was optimal when the initial medium pH was 5.8 (12.1 g l−1 DW) and the withanolide A production was highest in the medium pH set at 6.0 (13.84 mg g−1 DW).
The neuroprotective effects of Polygonum multiflorum extract (PME) and its two fractions, ethanol-soluble PME (PME-I) and -insoluble PME (PME-II), on the degeneration of nigrostriatal dopaminergic neurons induced by a combination of paraquat and maneb (PQMB) were investigated in male C57BL/6 mice. The mice were treated twice a week for 6 weeks with intraperitoneal injections of PQMB. This combination caused a reduction of spontaneous locomotor activity, motor incoordination, and declines of dopamine level in the striatum and tyrosine hydroxylase-positive neurons in the substantia nigra. Administration of PME and PME-I once daily for 47 days during 6 weeks of PQMB treatment and last 8 days after PQMB significantly attenuated the impairment of behavioral performance and the decrease in striatal dopamine level and substantia nigral tyrosine hydroxylase-positive neurons in the PQMB-treated animals, whereas the administration of PME-II had no effect on these behavioral, neurochemical and histological indices. The present findings suggest that PME has a beneficial influence on parkinsonism induced by PQMB and that the effects of PME are attributable to some substance(s) included in the ethanol-soluble fraction of PME (PME-I).
The leaf, stem and root of fresh Polygonum multiflorum plant were investigated. The contents of bioactive compounds and their antioxidant potential were studied by several radicals scavenging assay, and the antioxidative substances measured. The total phenolic contents were 179.0, 70.3 and 118.0 mg/g, whereas flavonoids contents were 7.3, 8.0 and 37.5 mg/g for water extracts from root (R), stem (S) and leaf (L), respectively. Further HPLC analysis revealed that the leaf and stem of this functional plant contained quercetin at 13,469 and 1,095 mg/kg, respectively. Furthermore, the root and stem contained rich emodin-related compounds that were 6,620 and 1,245 mg/kg, respectively. And different concentrations of emodin-related compounds and quercetin in root, stem and leaf showed different antioxidative ability. We observed that the DPPH scavenging activity, total antioxidant activity (TEAC), reducing power and NO scavenging ability of the three parts of the plant samples and found that the activities were R > L > S except that was S > L > R on superoxide anion scavenging effect. Our results indicated that the radical scavenging abilities of the three sample parts were more potent than that of emodin and quercetin. P. multiflorum contains rich emodin-related compounds and quer-cetin, indicating that these compounds played important roles in antioxidative effect. We suggested that the antioxidative effect of the fresh P. multiflorum plant correlate-well with emodin-related compounds and quercetin.
Prolonged application of ammonium as a source of nitrogen leads to serious physiological and morphological disorders in many plants, including mustard (Sinapis alba L,) seedlings (ammonium toxicity syndrome). Ammonium tolerance was previously observed in mustard seedlings in the presence of considerable amounts of nitrate in the medium. In the present study, the question was addressed as to what extent accumulation of nitrate and ammonium occurs in the mustard seedling and how this relates to ammonium toxicity and tolerance. Emphasis was on light control of accumulation in the attached cotyledons. Both NQ3 and NH4 became strongly accumulated in the mustard cotyledons once the concentration in the medium exceeded 1 mM, In the cotyledons, we measured concentrations > 30 mM in the case of nitrate and > 50 mM in the case of ammonium 4 days after sowing. Accumulation of inorganic nitrogen in the mustard cotyledons did not depend on photosynthesis nor on intact chloroplasts. However, the rate of nitrate accumulation was strongly stimulated by light, operating through phytochrome, while ammonium accumulation was not affected by light in short-term experiments, i,e, within 24 h and only weakly (and probably indirectly) in long-term light.
Primary hairy root tissues as well as aseptic hairy root culture lines contain specific compounds that have been biologically characterized as opines. These substances are agropine, mannopine, mannopinic acid, and agropinic acid; they have been synthesized and their electrophoretic behavior has been studied. Hairy root tissues also contain agrocinopines. According to the opine content of hairy root tissues, two types of Agrobacterium rhizogenes strains have been identified. Agropine-type strains (A4, 15834, HRI) elicit roots containing agropine, mannopine, mannopinic acid, and agropinic acid, whereas mannopine-type strains (8196, TR7, TR101) elicit roots containing only mannopine, mannopinic acid and agropinic acid. A. rhizogenes strains catabolize the opines whose synthesis they induce in the hairy root tissues. However, strain HRI only catabolizes agropine. Except for strain HRI, all A. rhizogenes strains studied contain three plasmids, of which the largest appears to be a cointegrate of the two others. Transconjugants of A. rhizogenes plasmids in A. tumefaciens have been obtained by selection on opines. Their properties have been studied and related to their plasmid content. In the mannopine strain C58C1(pRi8196), the virulence functions and the opine-related functions are located on the same plasmid (pRi8196). In agropine strains the catabolic functions are dissociated: agropine degradation is specified by the virulence plasmid, which also specifies opine synthesis in hairy root tissue, however, mannopine, mannopinic acid and agropinic acid degradation are specified by the smaller plasmid. Strain HRI contains only the virulence plasmid, which explains its inability to degrade mannopine, mannopinic acid, and agropinic acid.
Centella asiatica is a herbaceous plant used in medicine for its wound-healing and anti-inflammatory properties. Its bioactive compounds are
ursane-type triterpene saponins known as centellosides. With the aim of increasing the biotechnological production of these
compounds, C. asiatica cell suspensions were established and treated with two concentrations (100 and 200μM) of methyl jasmonate (MeJA). The maximum
centelloside production was observed in the stationary growth phase, reaching 0.16mgg−1 dry weight (DW) at day 25 of the culture in the control and 1.11mg−1 g DW at day 15 in the MeJA-elicited cultures. The elicitor did not change the centelloside pattern, with madecassoside being
the main compound, followed by asiaticoside. Reverse transcription polymerase chain reaction (RT-PCR) analysis of the β-amyrin
synthase gene (CabAS, the specific oxidosqualene cyclase that leads to centelloside formation) showed higher levels of expression in the elicited
cultures than in the control. The maximum content of centellosides was obtained at day 15, with a time lag between gene activation
and centelloside biosynthesis. In the cultures elicited with 200μM MeJA, the centelloside production did not increase compared
to the control. Both elicitor concentrations decreased the content of phytosterols. Thus, MeJa elicitation in this type of
culture was dose-dependent and its inducing role was apparent at low concentrations.
KeywordsAsiaticoside-Cell suspension cultures-
Rheum ribes L. (Polygonaceae) is the source of one of the most important crude drugs in Asiatic regions .The medicinal character of rhubarb is due to its anthraquinone content . Different parts of sterile seedlings were cultured on MS medium to study the generation of callus. The explants were cultured with different ranges of plant growth regulators and the best range of plant growth regulators for generation of callus was IBA (1 mg l–1) and BA (1 mg l–1). The content of anthraquinones were determined by HPLC . The concentration of sucrose, vitamins and Myo-inositol and ratio of NO3 to NH4 in the medium was changed and growth rate and content of 2 anthraquinones was determined . The growth rate of callus declined with increased rate of secondary metabolites production. Myo-inositol 100 mg l–1 in the medium increased anthraquinone content and in medium that had NO3/NH4:1/1 the maximum content of anthraquinone was obtained.
In this study, methyl jasmonate (MJ)-induced changes of triterpene saponins in ginseng (Panax ginseng C.A. Meyer) hairy roots and expression profiling of relevant responsive genes were analyzed. The transcription of PgSS (squalene synthase), PgSE (squalene epoxidase), and PNA (dammarenediol synthase-II) genes in hairy root cultures elicited by MJ treatment increased as compared with the controls,
whereas that of PNX (cycloartenol synthase) decreased slightly. In order to select candidate genes encoding for cytochrome P450-dependent hydroxylase
or glucosyltransferase associated with triterpene biosynthesis, RT-PCR analysis was conducted following MJ elicitation. No
differences were observed in any expression among the five genes associated with the cytochrome P450 family, when compared
to that of control. For candidates of the glucosyltransferase gene,expression of EST IDs PG07020C06, PG07025D04, and PG07029G02
was upregulated. In an effort to assess the effects of MJ elicitation on the biosynthesis of triterpene saponin, protopanaxadiol
saponin (Rb group) and protopanaxatriol saponin (Rg group) contents in hairy roots were evaluated by HPLC analysis. With 7
days of MJ elicitation, levels of all ginseonsides of the two-groups increased much higher than that observed in the control.
In particular, protopanaxadiol-type saponin contents increased by 5.5–9.7 times that of the control, whereas protopanaxatriol-type
saponin contents were increased by 1.85–3.82-fold. In the case of Rg1 ginsenoside after MJ elicitation, the content was affected
negatively in ginseng hairy root cultures.
To study the production of secondary metabolites of Maesalanceolata and Medicagotruncatula, hairy root cultures of both plant species were established. Because maintenance of large numbers of cultures is laborious
and costly, we developed a cryopreservation protocol and stored different isolated lines over time. Using encapsulation-dehydration,
high survival rates were observed for both Maesa and Medicago hairy roots. Root tips were isolated and encapsulated in calcium-alginate beads, containing 0.1M sucrose. The encapsulated
hairy roots were precultured for 3days using basal medium containing high sucrose concentrations. Medicago root tip growth during the preculturing time lead to unwanted outgrowth which could be tempered by addition of plant growth
inhibitors. After preculturing, the beads were dehydrated in the air flow of a laminar flow until 35–40% of the initial bead
weight was reached. Dehydrated beads were plunged into liquid nitrogen and after different storage times thawed in a water
bath at 40°C. The survival rates were 90% for Maesa and 53% for Medicago, which are sufficient to allow implementation in large storage experimental set-ups.
Cotyledon explants of two Oriental melons produced hairy roots when cultured on Murashige and Skoog basal medium after infection
by the Agrobacterium
rhizogenes. Hairy roots were produced from the wounded surface of the cotyledon explants of Cucumis melo L. cv. Geumssaragi-euncheon on Murashige and Skoog selective medium and 86% of the GUS stained hairy roots were positive
for the expression of beta-glucuronidase. The insertion of the gfp-gus fusion gene in the genomic DNA and the presence of the gfp-gus-specific transcript in the total RNAs of transgenic hairy roots were confirmed by PCR and RT-PCR, respectively. An immunoblot
analysis of the transgenic hairy root extract revealed 97kDa single bands coincident with the molecular weight of the GFP-GUS
fusion proteins. ELISA demonstrated that the highest level of GFP-GUS fusion protein expression was 0.47% of the total soluble
protein in a transgenic hairy root. The MS medium showed the fastest growth among three media types tested. Infection of the
hairy roots with a root-knot nematode resulted in the development of a mature egg mass about 4–5weeks after inoculation.
The highest number of egg mass was obtained on the hairy roots cultured in SH medium containing 0.3% agar.
An acidic polysaccharide, designated AEPS-1, was fractionated from the exopolysaccharide (EPS) produced by a medicinal fungus Cordyceps sinensis Cs-HK1 in mycelial culture. The molecular structure of AEPS-1 was characterised and elucidated by spectral and chromatographic analyses, and through derivatization by periodate oxidation, Smith degradation and methylation. AEPS-1 was composed of glucopyranose (Glcp) and pyrano-glucuronic acid (GlcUp) in an 8:1 M ratio plus a trace amount of mannose, having an average molecular weight of about 36 kDa. AEPS-1 had a linear backbone of (1→3)-linked α-d-Glcp residues with two branches, α-d-Glcp and α-d-GlcUp, attached to the main chain by (1→6) glycosidic bonds at every seventh α-d-Glcp unit. Atomic force microscopy revealed that AEPS-1 formed large networks in water that are connected primarily with triple helical strands. In Raw264.7 macrophage cell cultures, AEPS-1, at suitable doses between 25 and 250 μg/ml, significantly stimulated the release of several major cytokines, demonstrating an immunomodulatory property.
In the past three decades, hairy roots research for the production of valuable biological active substances has received a lot of attention. The addition of knowledge to enhance the yields of desired substances and the development of novel tools for biomass engineering offer new possibilities for large-scale cultivation of the plant hairy root. Hairy roots can also produce recombinant proteins through the transfer of Agrobacterium T-DNA into the plant genome, and thereby hold immense potential for the pharmaceutical industry. This review highlights some of the significant progress made in the past few years and outlines future prospects for exploiting the potential utility of hairy root cultures as "chemical factories" for producing bioactive substances.
Polygonum multiflorum has traditionally been used for treating patients suffering from baldness and hair loss in East Asia.
The present study sought to investigate the hair growth promoting activities of Polygonum multiflorum and its mechanism of action.
The Polygonum multiflorum extract was topically applied to the shaved dorsal skin of telogenic C57BL6/N mice. To determine the effect of Polygonum multiflorum extract in telogen to anagen transition, the expression of β-catenin and Sonic hedgehog (Shh) was determined by immunohistochemistry analysis.
Polygonum multiflorum extract promoted hair growth by inducing anagen phase in telogenic C57BL6/N mice. In Polygonum multiflorum extract treated group, we observed increase in the number and the size of hair follicles that are considered as evidence for anagen phase induction. Immunohistochemical analysis revealed that earlier induction of β-catenin and Shh were observed in Polygonum multiflorum extract treated group compared to that in control group.
These results suggest that Polygonum multiflorum extract promotes hair growth by inducing anagen phase in resting hair follicles.
Transgenic hairy root system is important in several recalcitrant plants, where Agrobacterium tumefaciens-mediated plant transformation and generation of transgenic plants are problematic. Jute (Corchorus spp.), the major fibre crop in Indian subcontinent, is one of those recalcitrant plants where in vitro tissue culture has provided a little success, and hence, Agrobacterium-mediated genetic transformation remains to be a challenging proposition in this crop. In the present work, a system of transgenic hairy roots in Corchorus capsularis L. has been developed through genetic transformation by Agrobacterium rhizogenes harbouring two plasmids, i.e. the natural Ri plasmid and a recombinant binary vector derived from the disarmed Ti plasmid of A. tumefaciens. Our findings indicate that the system is relatively easy to establish and reproducible. Molecular analysis of the independent lines of transgenic hairy roots revealed the transfer of relevant transgenes from both the T-DNA parts into the plant genome, indicating the co-transformation nature of the event. High level expression and activity of the gusA reporter gene advocate that the transgenic hairy root system, thus developed, could be applicable as gene expression system in general and for root functional genomics in particular. Furthermore, these transgenic hairy roots can be used in future as explants for plantlet regeneration to obtain stable transgenic jute plants.
To investigate the protective effects of preconditioning human umbilical vein endothelial cells (HUVECs) with Polygonum multiflorum stilbeneglycoside (PMS) under anoxia/reoxygenation (A/R), and the mechanism of protection.
Prior to A/R, HUVECs were incubated with PMS (0.6 x 10(-11), 1.2 x 10(-11), or 2.4 x 10(-11) mol/L) for 3 h. Cell injury was subsequently evaluated by measuring cell viability with an MTT assay and lactate dehydrogenase (LDH) release, whereas lipid peroxidation was assayed by measuring malondialdehyde (MDA) content. Antioxidant capacity was quantified by superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity. Nitric oxide (NO) production was determined by nitrite accumulation. Endothelial NO synthase (eNOS) and inducible NOS (iNOS) protein expression was detected by Western blotting. Guanylate cyclase activity and cyclic GMP (cGMP) activity were assessed by an enzyme immunoassay kit.
PMS incubation attenuated A/R-induced injury in a concentration-dependent manner, as evidenced by a decrease in LDH activity and an increase in cell viability. PMS exerted its protective effect by inhibiting the A/R-mediated elevation of MDA content, as well as by promoting the recovery of SOD and GSH-Px activities. Additionally, PMS incubation enhanced NO and cGMP formation by increasing iNOS expression and guanylate cyclase activity. The protective effects of PMS were markedly attenuated by NOS inhibitor L-NAME, soluble guanylate cyclase inhibitor ODQ or PKG inhibitor KT5823.
PMS preincubation resulted in the enhancement of antioxidant activity and anti-lipid peroxidation. The NO/cGMP/cGMP-dependent protein kinase (PKG) signaling pathway was involved in the effect of PMS on HUVECs.
Due to their fast growth rates and biochemical stability, 'hairy root' cultures remain unsurpassed as the choice for model root systems and have promise as a bioprocessing system. Applications are wide-ranging, from the production of natural products and foreign proteins to a model for phytoremediation of organic and metal contaminants. Hairy roots will have a continuing role as an experimental model in plant metabolic engineering.
Dihydroartemisinic acid hydroperoxide (2) was isolated for the first time as a natural product from the plant Artemisia annua in a 29% yield. Its structure was identified by (1)H and (13)C NMR spectroscopy. Compound 2 is known as an intermediate of the photochemical oxidation of dihydroartemisinic acid (1) leading to artemisinin (3). The presence of 1 and 2 in the plant and the conditions under which 1 can be converted into 2, which can very easily oxidize to 3, provide evidence for a nonenzymatic, photochemical conversion of 1 into 3, in vivo, in the plant.
The in vivo antioxidant action of a lignan-enriched extract of the fruit of Schisandra chinensis (FS) and an anthraquinone-containing extract of the root of Polygonum multiflorum (PME) was compared with their respective active constituents schisandrin B (Sch B) and emodin by examining their effect on hepatic mitochondrial glutathione antioxidant status in control and carbon tetrachloride (CCl4)-intoxicated mice. FS and PME pretreatments produced a dose-dependent protection against CCl4 hepatotoxicity, with the effect of FS being more potent. Pretreatment with Sch B, emodin or α-tocopherol (α-Toc) also protected against CCl4 hepatotoxicity, with the effect of Sch B being more potent. The extent of hepatoprotection afforded by FS/Sch B and PME/emodin pretreatment against CCl4 toxicity was found to correlate well with the degree of enhancement in hepatic mitochondrial glutathione antioxidant status, as evidenced by increases in reduced glutathione level and activities of glutathione reductase, glutathione peroxidase as well as glutathione S-transferases, in both control and CCl4-intoxicated mice. α-Toc, which did not enhance mitochondrial glutathione antioxidant status, seemed to be less potent in protecting against CCl4 hepatotoxicity. The ensemble of results indicates that FS/PME produced a more potent in vivo antioxidant action than α-Toc by virtue of their ability to enhance hepatic mitochondrial glutathione antioxidant status and that the differential potency of FS and PME can be attributed to the difference in in vivo antioxidant potential between Sch B and emodin.
FS:lignan-enriched extract of Schisandra fruit
PME:anthraquinone-containing fraction of Polygonum root
Sch B:schisandrin B
Polygonum multiflorum stilbeneglycoside (PMS) is a water-soluble fraction of Polygonum multiflorum Thunb., one of the most famous tonic traditional Chinese medicines, that has protective effects on the cardiovascular system. The purpose of the present study is to elucidate the effects of PMS on macrophage-derived foam cell functions and the reduction of severity of atherosclerosis in hypercholesterolemic New Zealand White (NZW) rabbits. NZW rabbits were fed for 12 weeks with a normal diet, a high cholesterol diet, or a high cholesterol diet associated with irrigation with different doses of PMS (25, 50, or 100 mg/kg). Treatment of NZW rabbits fed with high cholesterol diet with 100 mg/kg PMS attenuated the increase in plasma cholesterol, low-density lipoprotein cholesterol, very low-density lipoprotein cholesterol, and plasma triglyceride. Treatment with 50 and 100 mg/kg PMS caused 43% and 60% decrease in atherosclerotic lesioned area ratio to total surface area, respectively. In U937 foam cells, PMS could decrease the high expression of intercellular adhesion molecule (ICAM)-1 protein and the vascular endothelial growth factor (VEGF) protein levels in the medium induced by oxidized lipoprotein when analyzed by flow cytometry. The results proved that PMS is a powerful agent against atherosclerosis and that PMS action could possibly be through the inhibition of the expression of ICAM-1 and VEGF in foam cells.
Callus culture of Polygonum multiforum and the production of anthraquinones
A revised medium for rapid growth and bioassays with tobacco tissue cultures