20-Hydroxyecdysone Improves Neuronal Differentiation of Adult Hippocampal Neural Stem Cells in High Power Microwave Radiation-Exposed Rats / 生物医学与环境科学（英文）

Objective@#The hippocampus is thought to be a vulnerable target of microwave exposure. The aim of the present study was to investigate whether 20-hydroxyecdysone (20E) acted as a fate regulator of adult rat hippocampal neural stem cells (NSCs). Furthermore, we investigated if 20E attenuated high power microwave (HMP) radiation-induced learning and memory deficits.@*Methods@#Sixty male Sprague-Dawley rats were randomly divided into three groups: normal controls, radiation treated, and radiation+20E treated. Rats in the radiation and radiation+20E treatment groups were exposed to HPM radiation from a microwave emission system. The learning and memory abilities of the rats were assessed using the Morris water maze test. Primary adult rat hippocampal NSCs were isolated in vitro and cultured to evaluate their proliferation and differentiation. In addition, hematoxylin & eosin staining, western blotting, and immunofluorescence were used to detect changes in the rat brain and the proliferation and differentiation of the adult rat hippocampal NSCs after HPM radiation exposure.@*Results@#The results showed that 20E induced neuronal differentiation of adult hippocampal NSCs from HPM radiation-exposed rats via the Wnt3a/β-catenin signaling pathway in vitro. Furthermore, 20E facilitated neurogenesis in the subgranular zone of the rat brain following HPM radiation exposure. Administration of 20E attenuated learning and memory deficits in HPM radiation-exposed rats and frizzled-related protein (FRZB) reduced the 20E-induced nuclear translocation of β-catenin, while FRZB treatment also reversed 20E-induced neuronal differentiation of NSCs in vitro.@*Conclusion@#These results suggested that 20E was a fate regulator of adult rat hippocampal NSCs, where it played a role in attenuating HPM radiation-induced learning and memory deficits.

Compared activity of agonist molecules towards ecdysone receptor in insect cell-based screening system / Comparação da atividade de moléculas agonistas em relação ao receptor de ecdisona em um sistema de triagem baseado em linhagens celulares de insetos

The ecdysone receptor, naturally activated by steroidal hormones, is a key protein for molting and reproduction processes of insects. Artificial activation of such receptor by specific pesticides induces an anomalous process of ecdysis, causing death of insects by desiccation and starvation. In this paper, we established a protocol for screening agonistic molecules towards ecdysone receptor of insect cells line S2 (Diptera) and Sf9 (Lepidoptera), transfected with the reporter plasmid ere.b.act.luc. Therefore, we set dose-response curves with the ecdysteroid 20-hydroxyecdysone, the phytoecdysteroid ponasterone-A, and tebufenozide, a pesticide belonging to the class of diacylhydrazines. In both cell lines, the median effective concentration values on reporter gene induction (EC50) of ponasterone-A was the smallest, meaning the most active agonist molecule. In Sf9 cells, tebufenozide had as smaller EC50 than 20-hydroxyecdysone, indicating the high agonistic capability and lepidopteran specificity. The protocol established in this study can be useful for a quick screening and rational research of site-specific pesticides.(AU)

O receptor de ecdisona, naturalmente ativado por hormônios esteroidais, é uma proteína-chave nos processos de muda e reprodução de insetos. A ativação artificial desse receptor por meio de pesticidas específicos induz um processo de ecdise anômala, levando o inseto à morte por dessecação e inanição. Neste trabalho, foi estabelecido um protocolo para a triagem de moléculas agonistas em relação ao receptor de ecdisona nas linhagens celulares responsivas S2 (Diptera) e Sf9 (Lepidoptera), transfectadas com o plasmídeo repórter ere.b.act.luc. Para tanto, curvas de dose-resposta foram estabelecidas com o ecdisteroide 20-hidroxiecdisona, o fitoecdisteroide ponasterona-A e tebufenozida, um pesticida pertencente à classe das diacilhidrazinas. Em ambas linhagens celulares, os valores médios de concentração efetiva para indução gênica (EC50) ponasterona-A foram menores, significando que este é o agonista mais potente. Em células Sf9, a tebufenozida apresentou EC50 menor que a 20-hidroxiecdisona, indicando uma alta atividade agonista e especificidade deste inseticida a lepidópteros. O protocolo estabelecido neste trabalho pode ser utilizado para uma rápida triagem e busca racional de pesticidas de alvo bioquímico específico.(AU)

20-Hydroxyecdysone from Dacrycarpus imbricatus bark inhibits the proliferation of acute myeloid leukemia cells

OBJECTIVE@#To investigate the anti-proliferative effects of 20-hydroxyecdysone isolated from the bark of Dacrycarpus imbricatus (Blume) de Laub.@*METHODS@#Column chromatography was used for isolation of compounds from plant material. The structure of the isolated compound was identified by mass spectrometry and nuclear magnetic resonance techniques, including HSQC, HMBC, NOE-difference experiments. The isolated compound was tested for its anti-proliferative activity in acute myeloid leukemia (AML) and OCI-AML cells.@*RESULTS@#Compound 1 was isolated from the ethyl acetate fraction of Dacrycarpus imbricatus barks by column chromatography. Its chemical structure was identified as 20-hydroxyecdysone (20HE), a cholestane-type ecdysteroid, by a combination of mass spectrometry and nuclear magnetic resonance spectrometric analyses. Our goal was to test the anti-proliferative activity of 20HE using the OCI-AML cell line. 20HE significantly decreased OCI cell number at a concentration of 1 mg/mL, whereas lower concentrations were ineffective. Moreover, this decrease was due to partial blockage of the G/S phase of the cell cycle, with a reduction of cells in the GM phase, not due to increased apoptosis.@*CONCLUSIONS@#This indicates that 20HE significantly decreases the number of cells in the G/S phase of the cell cycle in human AML cells. This is the first time that the anti-proliferative activity of 20HE against a human tumor cell line has been reported.

20-Hydroxyecdysone from Dacrycarpus imbricatus bark inhibits the proliferation of acute myeloid leukemia cells

Objective To investigate the anti-proliferative effects of 20-hydroxyecdysone isolated from the bark of Dacrycarpus imbricatus (Blume) de Laub. Methods Column chromatography was used for isolation of compounds from plant material. The structure of the isolated compound was identified by mass spectrometry and nuclear magnetic resonance techniques, including HSQC, HMBC, NOE-difference experiments. The isolated compound was tested for its anti-proliferative activity in acute myeloid leukemia (AML) and OCI-AML cells. Results Compound 1 was isolated from the ethyl acetate fraction of Dacrycarpus imbricatus barks by column chromatography. Its chemical structure was identified as 20-hydroxyecdysone (20HE), a cholestane-type ecdysteroid, by a combination of mass spectrometry and nuclear magnetic resonance spectrometric analyses. Our goal was to test the anti-proliferative activity of 20HE using the OCI-AML cell line. 20HE significantly decreased OCI cell number at a concentration of 1 mg/mL, whereas lower concentrations were ineffective. Moreover, this decrease was due to partial blockage of the G

Desiccation stress induces developmental heterochrony in Drosophila melanogaster.

Stressful environments are known to perturb developmental patterns in insects. In the purview of desiccation as a stressor, relatively little is known about the developmental consequences linked with desiccation tolerance. In this study, we have particularly focused on the exploration of the temporal profile of postembryonic development in response to desiccation exposure in Drosophila melanogaster and the associated trade-offs. We document a correlation between variations in 20-hydroxyecdysone levels and the altered timing of metamorphic events during the life cycle. Following desiccation, we observed an extension in the larval longevity whereas the duration of the pupal and adult stages was significantly shortened. Alternately, feeding of 20-hydroxyecdysone apparently led to the restoration of the normal temporal pattern of development in the desiccated group. In spite of the desiccation-responsive heterochronic shifts in development, the overall lifespan post recovery remained almost unaltered among the desiccated and undesiccated groups suggesting plasticity in developmental control. This observation reminisces ‘canalization-like’ phenomenon that buffers alterations in the overall lifespan. We thus identified a desiccationresponsive period in the lifespan of D. melanogaster during which variations in ecdysone levels are capable to alter the temporal course of development.

Sida tuberculata (Malvaceae): a study based on development of extractive system and in silico and in vitro properties

Sida tuberculata (Malvaceae) is a medicinal plant traditionally used in Brazil as an antimicrobial and anti-inflammatory agent. Here, we aimed to investigate the different extractive techniques on phytochemical parameters, as well as to evaluate the toxicity and antioxidant capacity of S. tuberculata extracts using in silico and in vitro models. Therefore, in order to determine the dry residue content and the main compound 20-hydroxyecdysone (20E) concentration, extracts from leaves and roots were prepared testing ethanol and water in different proportions. Extracts were then assessed by Artemia salina lethality test, and toxicity prediction of 20E was estimated. Antioxidant activity was performed by DPPH and ABTS radical scavenger assays, ferric reducing power assay, nitrogen derivative scavenger, deoxyribose degradation, and TBARS assays. HPLC evaluation detected 20E as main compound in leaves and roots. Percolation method showed the highest concentrations of 20E (0.134 and 0.096 mg/mL of extract for leaves and roots, respectively). All crude extracts presented low toxic potential on A. salina (LD50 >1000 µg/mL). The computational evaluation of 20E showed a low toxicity prediction. For in vitro antioxidant tests, hydroethanolic extracts of leaves were most effective compared to roots. In addition, hydroethanolic extracts presented a higher IC50 antioxidant than aqueous extracts. TBARS formation was prevented by leaves hydroethanolic extract from 0.015 and 0.03 mg/mL and for roots from 0.03 and 0.3 mg/mL on egg yolk and rat tissue, respectively (P<0.05). These findings suggest that S. tuberculata extracts are a considerable source of ecdysteroids and possesses a significant antioxidant property with low toxic potential.

Protection of 20-hydroxyecdysone on SH-SY5Y cells from 1-methy-4-phenylpyridinium-induced injury / 中国药学杂志

OBJECTIVE: To investigate the protective effect and probable mechanism of 20-Hydroxyecdysone on SH-SY5Y cells injured by MPP+. METHODS: MPP+ induced SH-SY5Y cells injury model was established. 20-Hydroxyecdysone was added into the culture to test its protective effect. The morphological changes of cells were observed. Cell viability was detected by method of MTT. The contents of LDH, MDA and activity of SOD were inspected by kits. Apoptosis rate of cells was detected by flow cytometry. RESULTS: Compared with MPP+ group, 20-Hydroxyecdysone group (50, 100, 150 μmol·L-1) alleviated the damage of SH-SY5Y cells induced by MPP+, significantly improved cell survival rate, reduced the release of LDH, increased the activity of SOD, decreased the contents of MDA and reduced the apoptosis of cells. CONCLUSION: 20-Hydroxyecdysone has a significant protective effect on SH-SY5Y cells injured by MPP+. The probable mechanism may be anti-oxidation and anti-apoptosis.

Chemical constituents from roots tubers of Serratula chinensis / 中草药

Objective: To study the chemical constituents from the roots tubers of Serratula chinensis. Methods: The chemical constituents were separated and purified by silica gel, Sephadex LH-20, ODS column chromatographies, and preparative HPLC. Their structures were determined by physicochemical properties and spectral data. Results: Sixteen compounds (ecdysteroids) were isolated from the n-butanol fraction of 95% ethanol extract in the root tubers of S. chinensis, and their structures were identified as 20-hydroxyecdysone (1), polypodine B (2), carthamosterone (3), 20-hydroxyecdysone-20, 22-monoacetonide (4), 24-epi-abutasterone (5), polypodine C (6), coronatasterone (7), 20-hydroxyecdysone 2-O-β-D-glucopyranoside (8), 20-hydroxyecdysone 25-O-β-D-glucopyranoside (9), polypodine B 20, 22-acetonide (10), shidasterone (11), 2-O-acetyl-20-hydroxyecdysone (12), 3-O-acetyl-20-hydroxyecdysone (13), 20-hyderoxyecdysone-20, 22-butylidene acetal (14), 24-methylene-shidasterone (15), and ajugasterone D (16). Conclusion: Compounds 2, 4, 5, 7-10, 12, 15, and 16 are isolated from this plant for the first time, and compounds 5, 7-10, and 16 are found in the plants of genus Serratula L. for the first time.

Larvicidal and Brine Shrimp Activities of Vitex Schiliebenii Extracts and Isolated Phytoecdysteroids on Anopheles gambiae Giles S S Larvae.

Acetone, methanol and aqueous extracts of the leaves, stem bark and root bark of Vitex schiliebenii belonging to the family Verbenaceae were evaluated for their larvicidal activity against late 3rd/early 4th Anopheles gambiae Giles s.s. larvae (Diptera: Culicidae). The extracts of the acetone leaves and stem bark were active with LC50 values of 14.6 and 17.4 ppm respectively at 24 hrs. These extracts exhibited low toxicity to brine shrimps with LC50 values of 180.9 and 154.4 ppm respectively. The constituents in these extracts were isolated and evaluated and the phytoecdysteroids 20-hydroxyecdysone (1) and stigmasterol (2) were identified as the active principles in the acetone stem bark while γ-sitosterol (3) was the active principle of the acetone leaf extract. The methanol leaf extract, the stem bark aqueous extract and the acetone root bark also showed potency against the mosquito species.

The study of ecdysteroid-containing plants in mongolian flora / Монголын Анагаах Ухаан

BackgroundPhytoecdysteroids are secondary metabolites produced by many plants. Ecdysteroid containing plants have widely been used in medicine and show pharmacological effects such as: anabolic, adaptogenic, genoprotection, hepatoprotection, antiradiation, anti-inflammatory, antidiabetic and anticancerous.GoalThe aim of this study was to determination, isolation and purification of phytoecdysteroids in the Mongolian flora.Materials and МetodsPlants collected during flowering in western and central regions of Mongolia in 2006-2008 and its flowering stage. Plants speciesclassification was identified by Professor D. Oyunchimeg (Department of botany, Hovd university, Mongolia), Professor A. I Pyak and A. L. Ebeli (Department of botany, Tomsk State University, Russia). The main ecdysteroid components were identified in Silene repens by HPLC using the UV spectrometry. The absorption maximum is due to the presence of a conjugated ketone and occurs near 240-250 nm.Results The ecdysteroid containing plants of Mongolian flora has been studied for the first time. Total 297 species from 171 genera in 50 families for ecdysteroid were screened. Most of them represent a random sampling of plants growing in the selected regions. The highest concentration of ecdysteroids was detected in the family of Caryophyllaceae (genera Silene, Elisanthe, Melandrium), Chenopodiaceae (genera Chenopodium and Axyris) and Asteraceae (genera Rhaponticum and Serratula). Ecdysteroids were investigated in 16 from 297 species of the families Сaryophyllaсеае, Asteraceae and Chenopodiaceae of Mongolian flora, but in 6 of them: Silene ichebogdo, Silene mongolica, Elisanthe aprica, Serratula marginata, Chenopodium frutescens, Axyris prostrate at the first time. Ecdysteroid profile has isolated from aerial parts of Silene repens 5 phytoecdysteroids such as: 2-deoxyecdysone, 2-deoxy-20-hydroxyecdysone, polypodine B, 20-hydroxyecdysone, integristerone A and identified by column chromatography and HPLC methods.ConclusionEcdysteroid profile of 297 plant species was investigated. Five ecdysteroids were isolated, their structure were analyzed by HPLC analysis and UV spectrometry in Silene repens. The basic esdysteroids concentrations in plants of Silene jenissensis, Silene repens, Serratula marginata, Stemmacantha uniflorum and Axyris prostrate L. comprise 2.2%, 1.1%, 1.0%, 1.3% and 0.9% dry weight, respectively. Regarding those three species, there appeared a good source for bioactive substance and a new class of pharmaceutical preparation from its aboveground parts is being developed.

Screening of Central and South American plant extracts for antimycobacterial activity by the Alamar Blue test / Triagem de extratos de plantas das Américas Central e do Sul para atividade antimicobacteriana pelo teste do Alamar Blue

Forty eight ethanolic crude extracts and fractions (hexane, dichloromethane, ethyl acetate and n-butanol) from ten Brazilian plants (Leguminosae, Monimiaceae and Verbenaceae), 1 from Costa Rica (Verbenaceae) and 1 from Argentina (Verbenaceae) were screened for anti-mycobacterium activity against Mycobacterium tuberculosis (ATCC-27294H37Rv), by the Alamar Blue test, at a fixed concentration of 100 µg/mL. Out of the forty eight, seven were active at this concentration, corresponding to Lantana trifolia (hexane and dichloromethane extracts from leaves), Vitex cooperi (methanol:water, 1:1 extract from barks), Lippia lacunosa (hexane and dichloromethane extracts from leaves) and Lippia rotundifolia (hexane and dichloromethane extracts from leaves), all from the Verbenaceae family.

Quarenta e oito extratos brutos etanólicos e frações (em hexano, diclorometano, acetato de etila e n-butanol) de dez plantas brasileiras pertencentes às famílias Leguminosae, Monimiaceae e Verbenaceae; uma da Costa Rica (Verbenaceae) e uma da Argentina (Verbenaceae) foram ensaiados para verificação da atividade anti-micobacteriana contra Mycobacterium tuberculosis (ATCC-27294H37Rv), pelo teste do Alamar Blue, a uma concentração fixa de 100 µg/mL. Dentre os quarenta e oito extratos e frações estudados, sete mostraram-se ativos na concentração ensaiada - frações em hexano e diclorometano de folhas de Lantana trifolia, extrato em metanol:água, 1:1 de cascas de Vitex cooperi, frações em hexano e diclorometano de folhas de Lippia lacunosa e de Lippia rotundifolia, sendoque todas essas plantas pertencem à família Verbenaceae.