Neuroprotective Effects of Ethanol Extract of Polyscias fruticosa (EEPF) against Glutamate-Mediated Neuronal Toxicity in HT22 Cells.

In traditional herbal medicine, the Polyscias fruticosa has been frequently used for the treatment of ischemia and inflammation. Oxidative stress mediated by elevated glutamate levels cause neuronal cell death in ischemia and various neurodegenerative diseases. However, so far, the neuroprotective effects of this plant extract against glutamate-mediated cell death have not been investigated in cell models. The current study investigates the neuroprotective effects of ethanol extracts of Polyscias fruticosa (EEPF) and elucidates the underlying molecular mechanisms of EEPFs relevant to neuroprotection against glutamate-mediated cell death. The oxidative stress-mediated cell death was induced by 5 mM glutamate treatment in HT22 cells. The cell viability was measured by a tetrazolium-based EZ-Cytox reagent and Calcein-AM fluorescent dye. Intracellular Ca2+ and ROS levels were measured by fluorescent dyes, fluo-3 AM and 2',7'-dichlorodihydrofluorescein diacetate (DCF-DA), respectively. Protein expressions of p-AKT, BDNF, p-CREB, Bax, Bcl-2, and apoptosis-inducing factor (AIF) were determined by western blot analysis. The apoptotic cell death was measured by flow cytometry. The in vivo efficacy of EEPF was evaluated using the Mongolian gerbil mouse by surgery-induced brain ischemia. EEPF treatment showed a neuroprotective effect against glutamate-induced cell death. The EEPF co-treatment reduced the intracellular Ca2+ and ROS and apoptotic cell death. Furthermore, it recovered the p-AKT, p-CREB, BDNF, and Bcl-2 levels decreased by glutamate. The EEPF co-treatment suppressed the activation of apoptotic Bax, the nuclear translocation of AIF, and mitogen-activated protein kinase (MAPK) pathway proteins (ERK1/2, p38, JNK). Further, EEPF treatment significantly rescued the degenerative neurons in the ischemia-induced Mongolian gerbil in vivo model. EEPF exhibited neuroprotective properties that suppress glutamate-mediated neurotoxicity. The underlying mechanism of EEPF is increasing the level of p-AKT, p-CREB, BDNF, and Bcl-2 associated with cell survival. It has therapeutic potential for the treatment of glutamate-mediated neuropathology.

Antioxidative and α-glucosidase inhibitory constituents of Polyscias guilfoylei: experimental and computational assessments.

Searching for bioactive agents from medicinal plants, eleven constituents were isolated from Polyscias guilfoylei stem for the first time, including a nucleoside uracil (1), two sterols ß-sitosterol (2) and daucosterol (3), a saponin androseptoside A (4), two lignans (+)-pinoresinol (5) and (+)-syringaresinol (6), four phenolic acids protocatechuic acid (7), methyl protocatechuate (8), caffeic acid (9), and 5-O-caffeoylquinic acid (10), and a flavonoid quercitrin (11). Metabolites 1, 4, and 6-11 have never been observed in genus Polyscias before. Phenolic compounds 7 and 9 possessed the respective IC50 values of 21.33 and 13.88 µg/mL in DPPH (2,2-diphenyl-1-picrylhydrazyl) antioxidative assay, as compared with that of the positive control resveratrol (IC50 = 13.21 µg/mL). From density functional theory (DFT) calculated approach, the DPPH free radical scavenging capacity of two compounds 7 and 9 can be explained by the role of OH groups at carbons C-3 and C-4. Antioxidative actions of these two potential agents are followed HAT (H atom transfer) mechanism by OH bond disruption in gas, but SPLET (sequential proton loss electron transfer) mechanism in solvents water and methanol. Compared to 4-OH group, 3-OH group showed better bond disruption enthalpies and better kinetic energies since it reacted with HOO• and DPPH radicals. Sterols 2-3 and flavonoid 11 induced the IC50 values of < 2.0 µg/mL better than the positive control acarbose (IC50 = 184.0 µg/mL) in α-glucosidase inhibitory assay. Their interactions with human intestinal C- and N-terminal domains of α-glucosidase were explored using molecular docking study. The obtained results proved that compounds 2, 3, and 11 bind relatively stronger with the C-terminal domain than to the N-terminal domain through pivotal residues in the binding site and could be hypothesized as mixed inhibitors.

Kinetic Study on Chlorophyll and Antioxidant Activity from Polyscias fruticosa (L.) Harms Leaves via Microwave-Assisted Extraction.

Polyscias fruticosa (L.) leaves contain significant bioactive compounds with high antioxidant activity such as chlorophylls, total polyphenols, etc. but these have still been underutilized. In this study, the kinetics of chlorophyll and antioxidant activity extraction from P. fruticosa leaves by microwave-assisted extraction (MAE) were investigated. Microwave power was 300, 450, or 600 (W); the ratio of material/solvent varied from 1:40 to 1:80 (g/mL). In this study, the second-order kinetic model successfully predicted the change of chlorophyll and antioxidant activity during MAE. The increase of microwave power or/and the solvent amount increased saturated extraction efficiency and the extraction rate constant. However, the saturated concentration of chlorophyll and antioxidant activity increased with the increment of microwave power and the decrease in solvent amount.

Size-related shifts in carbon gain and growth responses to light differ among rainforest evergreens of contrasting shade tolerance.

Recent work suggests that plant size affects light requirements and carbon balance of juvenile trees, and such shifts may be greater in light-demanding species than in their more shade-tolerant associates. To explore the physiological basis of such shifts, we measured juvenile light interception, carbon gain and growth of four subtropical Australian rainforest trees differing in shade tolerance, comparing individuals ranging from 13 to 238 cm in height, across a wide range of understory environments. We hypothesized that even in a standardized light environment, increasing sapling size would lead to declines in net daily carbon gain of foliage and relative growth rates (RGR) of all species, with declines more pronounced in light-demanding species. Crown architecture of individuals was recorded using a 3-dimensional digitizer, and the YPLANT program was used to estimate the self-shaded fraction of each crown and model net carbon gain. Increased sapling size caused a significant increase in self-shading, and significant declines in net daily carbon gain and RGR of light-demanding species, while such ontogenetic variations were minimal or absent in shade-tolerant species. Additionally, differences in the slope of the relationship between light and RGR led to crossovers in RGR among shade-tolerant and light-demanding species at low light. Our results show that the magnitude of ontogenetic variation in net daily carbon gain and RGR can be substantial and may depend on successional status, making it unsafe to assume that young seedling performance can be used to predict or model responses of larger juvenile trees.

Antibacterial and antioxidant properties of crude extract, fractions and compounds from the stem bark of Polyscias fulva Hiern (Araliaceae).

BACKGROUND: In our previous work, the dichloromethane-methanol (1:1 v/v) extract, fractions and isolated compounds from Polyscias fulva stem bark showed interesting antifungal activity. As a continuity of that work, this study aimed to bring out complementary informations about the antimicrobial properties of P. fulva stem bark that may be useful in the standardization of phytomedicine from this plant. METHODS: The antibacterial activities of the crude extract, fractions (n-hexane, ethyl acetate, n-butanol and residual) and isolated compounds from Polyscias fulva stem bark were assayed by broth microdilution techniques. Their antioxidant activity were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH), pyrogallol (superoxide anion) and ß-carotene - linoleic acid assays. RESULTS: The crude extract presented antibacterial activities against S. typhi (ATCC 6539), E. aerogenes (ATCC 13045), P. aeruginosa (PA01) and E. coli (ATCC 10536) with MIC values of 2000 to 8000 µg/ml. The fractionation led the ethyle acetate and n-butanol fractions relatively more active (MIC = 500 to 1000 µg/ml) as compared to the crude extract. ß-sitosterol and 3-O-α-L- arabinopyranosyl-hederagenin were the most active compounds on the tested bacteria with MIC values ranging from 6.25 to 100 µg/ml. The most sensitive was P. aeruginosa (PA01) on which all the tested compounds were active with MICs ranging from 6.25 to 400 µg/ml. Among all the tested substances, the crude extract (RSa50 = 84.86 µg/ml) and the methyl atrarate (RSa50 = 14.77 µg/ml), showed the highest scavenging activities against DPPH free radicals and those arising from the oxidation of the linoleic acid respectively. CONCLUSION: From this study, the results obtained reveal that the stem bark of P. fulva possesses antibacterial and antioxidant activities. It may then be useful in the development of an antimicrobial phytomedicine with a large spectrum of actvity endowed with antioxidant properties which can be standardised based on the isolated compounds.

Antioxidant and Anti-Inflammatory Mechanisms of Lipophilic Fractions from Polyscias fruticosa Leaves Based on Network Pharmacology, In Silico, and In Vitro Approaches.

Polyscias fruticosa leaf (PFL) has been used in food and traditional medicine for the treatment of rheumatism, ischemia, and neuralgia. However, the lipophilic components of PFL and their biological properties remain unknown. This study, integrating network pharmacology analysis with in silico and in vitro approaches, aimed to elucidate the antioxidant and anti-inflammatory capacities of lipophilic extracts from PFL. A total of 71 lipophilic compounds were identified in PFL using gas chromatography-mass spectrometry. Network pharmacology and molecular docking analyses showed that key active compounds, mainly phytosterols and sesquiterpenes, were responsible for regulating core target genes, such as PTGS2, TLR4, NFE2L2, PRKCD, KEAP1, NFKB1, NR1l2, PTGS1, AR, and CYP3A4, which were mostly enriched in oxidative stress and inflammation-related pathways. Furthermore, lipophilic extracts from PFL offered powerful antioxidant capacities, as evident in our cell-free antioxidant assays. These extracts also provided a protection against oxidative stress by inducing the expression of catalase and heme oxygenase-1 in lipopolysaccharide (LPS)-treated RAW 264.7 cells. Additionally, lipophilic fractions from PFL showed anti-inflammatory potential in downregulating the level of pro-inflammatory factors in LPS-treated macrophages. Overall, these findings provide valuable insights into the antioxidant and anti-inflammatory properties of lipophilic extracts from PFL, which can be used as a fundamental basis for developing nutraceuticals and functional foods.

Polysciasoside B and C: new dammarane-type triterpene glycosides from the leaves of Australian rainforest plant Polyscias australiana (F.Muell.) Philipson (Araliaceae).

Phytochemical investigation of the leaves of Polyscias australiana (F.Muell.) Philipson (family Araliaceae) led to the isolation and identification of two new analogues belonging to the rare dammarane-type triterpene glycosides, polysciasosides B (1) and C (2). Also isolated in high yields from this plant was the known saponin, ß-hedrin (3). The two new polysciasoside analogues exhibited no anti-inflammatory activity (inhibitory effects on NO inhibition and cell viability in RAW 264.7 macrophages) or cytotoxic activity against AGS gastric adenocarcinoma or the MCF7 breast adenocarcinoma cell lines. In contrast, the known compound ß-hedrin exhibited potent anti-inflammatory and cytotoxicity in these biological assays.

Diverse Novel Viruses Coinfecting the Tropical Ornamental Plant Polyscias balfouriana in China.

The viromic profile of Polyscias balfouriana cv. Marginata, a perennial woody and ornamental plant, was determined using ribosomal RNA-depleted total RNA (rRNA-depleted totRNA) sequencing. Five viruses (i.e., polyscias mosaic virus, PoMV; one potential novel rhabdovirus; and three novel viruses of Betaflexiviridae and Closteroviridae) were detected and prevalence-surveyed in Hainan province, China. The genomes of polyscias capillovirus 1 (PCaV-1) and polyscias citrivirus 1 (PCiV-1) of family Betaflexiviridae were completed, and the genomes of polyscias crinivirus 1 (PCrV-1) of Closteroviridae were nearly completed lacking the 5' and 3' termini. PCaV-1 shares 68% genome nucleotide (nt) identity and 66% replicase (Rep) amino acid (aa) identity with homologues in apple stem grooving virus (ASGV). PCiV-1 shares 65% genome nt identity and 64% Rep aa identity with homologs in citrus leaf blotch virus (CLBV). Meeting the species demarcation criteria, PCaV-1 and PCiV-1 were considered to be new species in genera Capillovirus and Citrivirus, respectively. PCrV-1 shares high genome nt identity (62%), heat shock protein 70-like protein (HSP70h) and RNA-dependent RNA polymerase (RdRp) aa identity (78−80%) with homologues in tomato chlorosis virus (ToCV). We tentatively consider PCrV-1 to be an unclassified member of the Crinivirus genus. PoMV, PCaV-1, PCiV-1, and PCrV-1 are the prevalent viruses with >73% occurrence in the Xinglong Tropical Botanical Garden, Hainan, China.

Growth and Oleanolic Acid Accumulation of Polyscias fruticosa Cell Suspension Cultures.

BACKGROUND: Oleanolic acid is an oleanane triterpene found in many plant species all over the world. This compound is also a major saponin in leaves of Polyscias fruticosa and possesses several promising pharmacological activities, such as hepatoprotective effects, and antiinflammatory, antioxidant, or anticancer activities. OBJECTIVE: The objective of the present work is to establish cell suspension culture of P. fruticosa, investigate the influence of several factors such as plant growth regulators and carbon source on cell growth, and determine their oleanolic acid content. METHODS: Cell culture was established by using 2 g fresh weight of 30 day old friable callus derived from in vitro stem segment in 50 mL of liquid medium with a shaking speed of 220 rpm. The culture was then incubated at 25±2ºC with a shaking speed of 120 rpm in the period of 12 h daylight at a light intensity of about 6.75 µmol/m2/s. Cell growth was measured by fresh and dry biomass at 16 h day. Oleanolic acid content was determined using HPLC analysis. RESULTS & DISCUSSION: The study results showed that MS medium containing 2% sucrose as a carbon source, supplemented with 1 mg/L 6-benzylaminopurine and 0.5 mg/L 2,4-dichlorophenoxyacetic acid was the most appropriate growth medium. Cell biomass and oleanolic acid content reached the highest values of 0.43 g dry weight/flask and 25.4 mg/g dry weight, respectively. CONCLUSION: These results indicated the potential production of oleanolic acid, a compound with high pharmacological value, from P. fruticosa cell culture.

In Vitro Response of Polyscias filicifolia (Araliaceae) Shoots to Elicitation with Alarmone-Diadenosine Triphosphate, Methyl Jasmonate, and Salicylic Acid.

The effectiveness of different elicitation variants in combination with alarmone application was studied in shoot cultures of Polyscias filicifolia. The shoots were elicited with 200 µM methyl jasmonate (MeJA) or 50 µM salicylic acid (SA) alone or in combination, and their activity was compared with those treated with the alarmone diadenosine 5',5‴-P1P3-triphosphate (Ap3A), either alone or in combination with SA and/or MeJA. All treatments resulted in significant stimulation of phenolic acid production (chlorogenic and ferulic acids), as well as oleanolic acid (OA) compared to control, with their highest concentration noted under simultaneous elicitation with SA and MeJA. While the maximum content of caffeic acid was detected after treatment with alarmone alone. In each of the culture variants enhanced antioxidant activity was observed, however the level varied according to the treatment. In addition, the SA, Ap3A and Ap3A+SA variants demonstrated additional peroxidase isoforms, as indicated by Native-PAGE, as well as the highest α-tocopherol content. The highest antioxidant capacity of shoot extracts was correlated with the highest abundance of phenolic compounds and OA. The results indicate that ROS induction appears to participate in the signal transduction following Ap3A treatment.

Polysciosides J and K, two new oleanane-type triterpenoid saponins from the leaves of Polyscias fruticosa (L.) harms. cultivating in An Giang Province, Viet Nam.

For the first time, the phytochemical constituents of the leaves of Polyscias fruticosa (L.) Harms. cultivating in An Giang Province, Viet Nam were investigated and led to purify two new oleanane-type triterpenoid saponins, named polyscioside J (1) and polyscioside K (2) together with two known saponins, ladyginoside A (3) and chikusetsusaponin IVa (4) using variously chromatographic methods. Saponin (4) was reported for the first time from this species. Their structures were verified by IR, UV, HR-ESI-MS, NMR 1D and 2D experiments and compared with previous literatures.

Phytoconstituents from Polyscias guilfoylei leaves with histamine-release inhibition activity.

Phytochemical investigation of Polyscias guilfoylei leaves extract (PGE) led to the isolation of nine compounds, that is, ent-labda-8(17),13-diene-15,18-diol (1), stigmasterol (2), spinasterol (3), N-(1,3-dihydroxyoctadecan-2-yl) palmitamide (4), panaxydiol (5), 3-O-ß-d-glucopyranosylstigmasta-5,22-diene-3-ß-ol (6), (8Z)-2-(2 hydroxypentacosanoylamino) octadeca-8-ene-1,3,4-triol (7), 4-hydroxybenzoic acid (8), and tamarixetin 3,7-di-O-α-L-rhamnopyranoside (9). Compound 4 is reported in this study for the first time in nature whereas compound 9 is reported for the second time. Structural elucidation of the compounds was carried out using Nuclear Magnetic Resonance and Electrospray Ionization coupled with Mass Spectrometry spectroscopic analyses. PGE and compounds 4 and 9 exhibited weak cytotoxicity against both MCF-7 and HCT-116 cell lines using 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide assay. The antimicrobial activity of PGE and compounds 4 and 9 was evaluated using the agar diffusion method. Escherichia coli was the most susceptible Gram-negative bacteria toward PGE with a minimum inhibitory concentration value of 9.76 µg/mL, whereas compounds 4 and 9 did not show any antimicrobial activity. Compound 4 exhibited promising inhibition of histamine release using U937 human monocytes with an IC50 value of 38.65 µg/mL.

Ethanol extract of Polyscias fruticosa leaves suppresses RANKL-mediated osteoclastogenesis in vitro and LPS-induced bone loss in vivo.

BACKGROUND: Many bone-related diseases such as osteoporosis and rheumatoid arthritis are commonly associated with the excessive activity of osteoclasts. Polyscias fruticosa has been used as traditional medicine for the treatment of ischemia and inflammation and also eaten as a salad. However, its effect on the bone related diseases has not been investigated yet. PURPOSE: This study aimed to investigate the effect of ethanol extract of P. fruticosa on RANKL-induced osteoclastogenesis in vitro and LPS-induced bone loss in mouse, and evaluate anti-osteoclastogenic activities of its major constituents. METHODS: BMMs or RAW264.7 cells were treated with ethanol extract from P. fruticose leaves (EEPL), followed by an evaluation of cell viability, RANKL-induced osteoclast differentiation, actin-ring formation, and resorption pits activity. Effects of EEPL on RANKL-induced phosphorylation of MAPKs were evaluated by Western blotting. The expression levels of NFATc1 and c-Fos were evaluated by Western blotting or immunofluorescence assay. The expression levels of osteoclast-specific marker genes were evaluated by Western blotting and reverse transcription-qPCR analysis. A LPS-induced murine bone loss model was used to evaluate the protective effect of EEPL on inflammation-induced bone loss. HPLC analysis was performed to identify the major constituents of EEPL. RESULTS: EEPL significantly inhibited RANKL-induced osteoclast differentiation by decreasing the number of osteoclasts, osteoclast actin-ring formation, and bone resorption. EEPL suppressed RANKL-induced phosphorylation of p38 and JNK MAPKs, as well as the expression of c-Fos and NFATc1. EEPL decreased the expression levels of osteoclast marker genes, including MMP-9, TRAP and CtsK. Mice treated with EEPL significantly protected the mice from LPS-induced osteoclast formation and bone destruction as indicated by micro-CT and histological analysis of femurs. We also identified 3-O-[ß-d-glucopyranosyl-(1→4)-ß-d-glucuronopyranosyl] oleanolic acid 28-O-ß-d-glucopyranosyl ester (1) and quercitrin (3) as the active constituents in EEPL for inhibiting RANKL-induced osteoclast differentiation. CONCLUSION: The results showed that EEPL exerted anti-osteoclastogenic activity in vitro and in vivo by inhibiting RANKL-induced osteoclast differentiation and function, and suggested that EEPL could have beneficial applications for preventing or inhibiting osteoclast-mediated bone diseases.

Chemical structures of constituents from the leaves of Polyscias balfouriana.

A new pyrrolidine derivative, (5S)-hydroxyethyl 2-oxopyrrolidine-5-carboxylate (1), a new flavonol glycoside, tamaraxetin 3,7-di-O-α-L-rhamnopyranoside (2), and a new triterpene saponin, polyscioside A methyl ester (3), along with six known compounds (4-9) were isolated from the leaves of Polyscias balfouriana. Their chemical structures were elucidated on the basis of extensive spectroscopic analysis.

Cytotoxicity and modes of action of five Cameroonian medicinal plants against multi-factorial drug resistance of tumor cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Beilschmiedia acuta Kosterm, Clausena anisata (Willd) Hook, Fagara tessmannii Engl., Newbouldia laevis Seem., and Polyscias fulva (Hiern) Harms. are medicinal plants used in Cameroonian traditional medicine in the treatment of various types of cancers. The present study aims at investigating 11 methanolic extracts from the above Cameroonian medicinal plants on a panel of human cancer cell lines, including various drug-resistant phenotypes. Possible modes of action were analyzed for two extracts from Beilschmiedia acuta and Polyscia fulva and alpha-hederin, the representative constituent of Polyscia fulva. MATERIALS AND METHODS: Cytotoxicity was determined using a resazurin assay. Cell cycle, apoptosis, mitochondrial membrane potential (MMP), and reactive oxygen species (ROS) were measured by flow cytometry. Cellular response to alpha-hederin was investigated by a mRNA microarray approach. RESULTS: Prescreening of extracts (40µg/mL) showed that three of eleven plant extracts inhibited proliferation of CCRF-CEM cells by more than 50%, i.e. BAL (73.65%), the bark extract of Beilschmiedia acuta (78.67%) and PFR (68.72%). Subsequent investigations revealed IC50 values below or around 30µg/mL of BAL and PFR in 10 cell lines, including drug-resistant models, i.e. P-glycoprotein-overexpressing CEM/ADR5000, breast cancer resistance protein-transfected MDA-MB-231-BCRP, TP53 knockout cells (HCT116 p53(-/-)), and mutation-activated epidermal growth factor receptor-transfected U87MG.ΔEGFR cells. IC50 values below 5µg/mL of BAL were obtained for HCT116 (p53(-/-)) cells. IC50 values below 10µM of alpha-hederin were found for sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 cells. The BAL and PFR extracts induced cell cycle arrest between G0/G1 and S phases. PFR-induced apoptosis was associated with increased ROS generation and MMP breakdown. Microarray-based cluster analysis revealed a gene expression profile that predicted cellular response to alpha-hederin. CONCLUSION: BAL, PFL and alpha-hederin, an exemplarily taken constituent of Beilschmiedia acuta and Polyscia fulva extracts revealed cytotoxicity towards cancer cell lines. Hence, Beilschmiedia acuta and Polyscia fulva may be valuable to develop drugs against otherwise drug-resistant cancer cells.

The rare cyanogen proteacin, and dhurrin, from foliage of Polyscias australiana, a tropical Araliaceae.

The tyrosine-derived cyanogenic di-glucoside proteacin and related mono-glucoside dhurrin were identified as the cyanogens in foliage of the tropical tree species Polyscias australiana, present in the approximate ratio 9:1. To date cyanogenic glycosides have not been characterised from the Araliaceae or the Apiales. Concentrations of cyanogenic glycosides varied significantly between plant parts and with leaf age, with the highest concentrations in young emerging leaves (mean 2217.1 µg CN g(-1) dry wt), petioles (rachis; 1487.1 µg CN g(-1) dry wt) and floral buds (265.8 µg CN g(-1) dry wt). Between 2% and 10% of nitrogen in emerging leaves and petioles was present as cyanogenic glycosides. With the exception of floral buds, all tissues apparently lack a specific cyanogenic ß-glucosidase to catalyse the first step in the breakdown of these cyanogenic glycosides. Only with the addition of emulsin, an exogenous non-specific ß-glucosidase from almonds, were high concentrations of cyanogenic glycosides detected, as much as 20-fold greater than the low to negligible cyanogenic glycoside concentrations determined in the absence of exogenous enzyme. High concentrations of cyanogens in young tissues may confer protection, but may also be a nitrogen source during leaf expansion.