Wild imitating vs greenhouse cultivated Dendrobium huoshanense: Chemical quality differences.

Dendrobium huoshanense (D. huoshanense) has been used as functional food supplements and herbal medicines for preventing and managing diseases with a long history in China. Due to its endangered natural resources and huge demand, people tend to cultivate D. huoshanense to protect this species. However, the quality of wild and cultivated herbs of the same species may change. This work quantified and compared the main quality traits and chemical components of wild imitating and greenhouse cultivated D. huoshanense with different growth years. As a result, wild and cultivated D. huoshanense had similar chemical composition, but there are significant differences in the content of many ingredients (polysaccharides, flavonoids, nucleosides, bibenzyls, lignans and volatile compounds). And the contents of many of these components increased with growing years. In addition, multivariate statistical analyses have been applied to classify and evaluate samples from different cultivation modes according to these components. In conclusion, our results demonstrated that the overall quality of greenhouse cultivated D. huoshanense was not as good as wild-grown, but this mode can be a promising and sustainable way of producing D. huoshanense.

Identification of lignans and quality assessment in Dendrobium officinale under different cultivation modes.

RATIONALE: Lignans have attracted much attention from researchers because of their wide distribution and industrial applications in plants, as well as the remarkable diversity of their biological activities. As the literature has mainly focused on the extraction and identification of monomeric compounds of lignans, most lignans in Dendrobium officinale, a traditional Chinese medicine with a long cultivation history and rich sources, have not been detected using quality control methods. The aim of this study was to identify the lignans in Dactilon officinale. METHODS: High-performance liquid chromatography (HPLC) coupled with diode array detection and HPLC multiple-stage tandem mass spectrometry was used to identify the chemical constituents of D. officinale. Simultaneously, the characteristic chromatograms of D. officinale were established. Additionally, a method was established to determine the content of syringaresinol-4,4'-di-O-ß-D-glucoside, syringaresinol-4-O-ß-D-glucoside and syringaresinol. RESULTS: Thirty-three lignans, including 17 tetrahydrofuran lignans, two dibenzylbutane lignans, three aryl tetrahydronaphthalene lignans and 11 8-O-4'-neolignans, were tentatively identified from the methanol extract of the stems of D. officinale. This is the first report of 8-O-4'-neolignans from D. officinale. In addition, a total of eight characteristic peaks were marked in characteristic chromatograms, which were identified as lyoniresinol-9'-O-ß-D-glucoside, syringaresinol-4,4'-di-O-ß-D-glucoside, 8-hydroxy-syringaresinol-4-O-ß-D-glucoside, 5,5'-dimethoxy-lariciresinol-4-O-ß-D-glucoside, syringaresinol-4-O-ß-D-glucoside, 4-hydroxy-3,3',5,5'-tetramethoxy-8,4'-oxyneoligna-7'-ene-9,9'-diol-9-O-ß-D-glucoside, 4-hydroxy-3,3',5,5'-tetramethoxy-8,4'-oxyneoligna-7'-ene-9,9'-diol-4-O-ß-D-glucoside and syringaresinol. Our results showed that no significant difference occurred in lignan composition among the 99 batches of D. officinale from different sources. However, the peak areas of the lignans of D. officinale planted under simulated wild culture were generally higher than those in greenhouses, and showed an upward trend with the increase in growth years. The average contents of syringaresinol-4,4'-di-O-ß-D-glucoside, syringaresinol-4-O-ß-D-glucoside and syringaresinol were 10.112-179.873, 51.227-222.294 and 6.368-120.341 µg/g, respectively. CONCLUSIONS: This study provided a basis for improving the quality control of D. officinale and could provide references for the identification of lignans in other Dendrobium species.

Identification of bibenzyls and evaluation of imitative wild planting techniques in Dendrobium officinale by HPLC-ESI-MSn.

Dendrobium officinale is a traditional Chinese herb with beneficial properties. Modern pharmacological studies show that bibenzyl is one of the antitumor active ingredients, but there is no effective quality control method for identifying ingredients. In this study, the composition of bibenzyls in Dendrobium officinale was studied by high-performance liquid chromatography coupled with electrospray ionization multistage mass spectrometry (HPLC-ESI-MSn ). A total of nine isolated bibenzyls and their glycosides, 22 bis (bibenzyls), and two phenylpropanol bibenzyl derivatives were identified. The results of HPLC characteristic chromatogram analysis and statistical analysis showed that the relative content of bibenzyls in wild imitation cultivation of samples had been significantly higher than that in greenhouse cultivation. In addition, the relative content of bibenzyls increased with the growth of the original plant. This study provided a scientific reference for controlling the quality of bibenzyls in Dendrobium officinale, developing the cultivation technology and improving the quality of Dendrobium officinale. HIGHLIGHTS: HPLC-ESI-MS/MS method for the analysis of bibenzyls and bis (bibenzyls) in Dendrobium officinale. Easy-to-use method facilitating rapid measurement of large sample quantities. The method requires only small volumes of samples for the analysis. Applicable for the establishment of Chinese medicine studies and the quality control standard of Chinese herbs.

Identification of three types of O-glycosylated flavonoids in Dendrobium loddigesii, Dendrobium primulinum, Dendrobium crepidatum, Dendrobium porphyrochilum, and Dendrobium hancockii using mass spectrometry.

RATIONALE: Flavonoids, representing the pharmacologically active ingredients, are found widely in Dendrobium species. The biodiversity of Dendrobium makes the identification of its varieties all the more complicated. Previous studies showed that C-glycosylated flavones and a few O-glycosylated flavonols could be used in the identification of various Dendrobium species. Accordingly, this study further explores the significance of the identification of various types of O-glycosylated flavonoids in Dendrobium species. METHODS: High-performance liquid chromatography coupled with electrospray ionization multistage tandem mass spectrometry (HPLC-ESI-MSn ) was used to identify the chemical constituents in five types of Dendrobium: Dendrobium loddigesii, Dendrobium primulinum, Dendrobium crepidatum, Dendrobium porphyrochilum, and Dendrobium hancockii. RESULTS: A total of 41 O-glycosylated flavonoids and 3 C-glycosylated flavones were identified, among which O-glycosylated dihydroflavones were the main flavonoids in D. loddigesii and D. primulinum, O-Glycosylated flavonols were rich in both D. crepidatum and D. porphyrochilum characterized by the main aglycone, substituted sugars, and their structural characteristics, and O-glycosylated flavones were the main constituents in D. hancockii. CONCLUSIONS: In this study, three types of O-glycosylated flavonoids in the five Dendrobium species were determined to have certain significance. This also provides a reference for the identification of other O-glycosylated flavonoids in Chinese herbs.

Identification of C-glycosyl flavones and O-glycosyl flavones in five Dendrobium species by high-performance liquid chromatography coupled with electrospray ionization multi-stage tandem MS.

RATIONALE: Flavones are widely used in traditional Chinese medicine (TCM) and are the pharmacologically active ingredients of many medicinal plants, such as Dendrobium. With the increasing demand for medicinal Dendrobium, the identification of characteristic flavones that can serve as chemical markers for quality control is critical step for quality assurance and safety in the TCM industry. METHODS: High-performance liquid chromatography coupled with electrospray ionization multi-stage tandem mass spectrometry (HPLC/ESI-MSn ) was used to identify the chemical constituents in five types of Dendrobium: D. crystallinum, D. falconeri, D. strongylanthum, D. moniliforme, and D. gratiosissimum. RESULTS: A total of seventy-six C-glycosyl flavones and three O-glycosyl flavones were identified, of which fifteen C-glycosyl flavones were found in D. crystallinum, twenty four were found in D. falconeri, thirty were found in D. strongylanthum, seven were found in D. moniliforme (also called "Huangtongpi", from Anhui, China), fifteen were found in D. moniliforme (also called "Zitongpi", from Yunnan, China) and seventeen were found in D. gratiosissimum. Additionally, three flavone O-glycosides were all found in D. strongylanthum. CONCLUSIONS: The results of this study may be useful for the quality assessment and for the application of D. crystallinum, D. falconeri, D. strongylanthum, D. moniliforme, and D. gratiosissimum. This study provides comprehensive information for identification of flavones from other Chinese herbs.

Identification of C-glycosyl flavones and quality assessment in Dendrobium nobile.

RATIONALE: Flavones are significant indicators of quality in traditional Chinese medicines (TCMs) and thus play a significant role in the quality control of TCMs in the pharmaceutical industry. Most flavones in Dendrobium nobile Lindl, a TCM with a long cultivation history and rich sources, have not been identified. This study was aimed at identifying the flavones in D. nobile from various habitats. METHODS: High-performance liquid chromatography (HPLC) coupled with diode-array detection and HPLC multiple-stage tandem mass spectrometry was used to identify the chemical constituents of D. nobile from various habitats, and a method was established to determine the content of vicenin II, violanthin and isoviolanthin. Hierarchical cluster analysis, principal component analysis and orthogonal partial least-squares discriminant analysis were used to analyze the variations among 26 batches from different habitats. RESULTS: A total of 33 flavones were tentatively identified. Twenty-five flavones, previously undescribed in D. nobile, were acylated by p-coumaroyl, feruloyl, sinapoyl or 3-hydroxy-3-methylglutaryl. The D. nobile habitats were distinguished by significant differences in their flavone content. The C-glycosyl flavones were demonstrated to be characteristic compounds for evaluating D. nobile from various habitats. In particular, flavones acylated with 3-hydroxy-3-methylglutaryl were specific compounds that were only detected in samples from Yunnan. CONCLUSIONS: The results of this study could be used to improve the quality control of D. nobile and could provide references for the identification of acylated C-glycosyl flavones in other natural products.

HPLC coupled with electrospray ionization multistage MS/MS and TLC analysis of flavones-C-glycosides and bibenzyl of Dendrobium hercoglossum.

Dendrobium hercoglossum Rchb. f. (D. hercoglossum), as one of the origins of medicinal Dendrobium, has been widely used as a health food and nutrient source promoting fluid production. Due to a lack of quality control, it is often counterfeited or mixed with other Dendrobium. In this study, a high-performance liquid chromatography characteristic chromatogram method is established for the quality evaluation of D. hercoglossum. Based on the high-performance liquid chromatography characteristic chromatogram, D. hercoglossum is divided into two classes, each with different flavone peaks. These flavone peaks were identified using high-performance liquid chromatography coupled with electrospray ionization multistage tandem mass spectrometry. Among them, the acylated (3-hydroxy-3-methylglutaryl, p-coumaroyl, feruloyl, or sinapoyl) flavones-C-glycosides are first found in D. hercoglossum in this study. In addition, one unique band was found in D. hercoglossum by thin-layer chromatography, which can be used to distinguish it from other Dendrobium species as a characteristic marker of this plant. Combining the high-performance liquid chromatography characteristic chromatogram and high-performance liquid chromatography coupled with electrospray ionization multistage tandem mass spectrometry, the unique band was identified as 4,5-dihydroxy-3,3'-dimethoxybibenzyl. These analysis methods can be applied for the quality control and identification of D. hercoglossum as well as providing reference for the identification of similar constituents in other Dendrobium species.

Network Analysis of Transcriptome and LC-MS Reveals a Possible Biosynthesis Pathway of Anthocyanins in Dendrobium officinale.

Anthocyanins, a group of flavonoids, are widely present in plants and determine the colors of the peels of stems, fruits, and flowers. In this study, we used UHPLC-ESI-MS to identify anthocyanins in the herbal plant Dendrobium officinale, which has been used for centuries in China. The results indicated that the total anthocyanin content in samples from Guangxi was the highest. Seven anthocyanins were identified, and the fragmentation pathways were proposed from D. officinale. Most of the identified anthocyanins were composed of cyanidin and sinapoyl groups. We also carried out that the sinapoyl group had active sites on breast cancer receptors by using Schrödinger. The relative levels of the 7 anthocyanins in the samples from the three locations were determined. Transcriptomic analysis was used to analyze the sinapoyl anthocyanin synthesis-related genes in plants, such as genes encoding UGTs and serine carboxypeptidase. We speculated that sinapoyl anthocyanin biosynthesis was associated with the activities of certain enzymes, including chalcone flavonone isomerase-like, hydroxycinnamoyltransferase 1, UGT-83A1, UGT-88B1 isoform X1, serine carboxypeptidase-like 18 isoform X3, and serine carboxypeptidase-like 18.

Identification of C-glycosyl flavones by high performance liquid chromatography electrospray ionization mass spectrometry and quantification of five main C-glycosyl flavones in Flickingeria fimbriata.

Flickingeria fimbriata is commonly applied in China as a traditional Chinese medicine (TCM), however the quality control of it is incomplete. In this work, we aim to identify and quantify the structures of C-glycosyl flavones in F. fimbriata. High performance liquid chromatography-diode array detector (HPLC-DAD) and High performance liquid chromatography-electrospray ionization-multiple stage tandem mass spectrometry (HPLC-ESI-MSn) methods were combined to identify C-glycosyl flavones and determine their contents. Twenty acylated C-glycosyl flavones and ten non-acylated C-glycosyl flavones were identified for the first time in F. fimbriata on systematic MSn analysis via HPLC-ESI-MSn. The aglycones of all of these compounds were apigenin or chrysoeriol and were acylated with p-coumaric, ferulic, 3,4-dimethoxycinnamic or 3,4,5-trimethoxycinnamic acids. Furthermore, the quantification result suggest that two C-glycosyl flavones (vicenin-I and vicenin-III) with relative high contents were revealed to be more strongly acylated in F. fimbriata. The method is sufficiently precise, accurate, and sensitive for the qualitative and quantitative analysis of C-glycosyl flavones, which is expected to establish a standard for quality control and identification in this plant.

Structure Identification of ViceninII Extracted from Dendrobium officinale and the Reversal of TGF-ß1-Induced Epithelial⁻Mesenchymal Transition in Lung Adenocarcinoma Cells through TGF-ß/Smad and PI3K/Akt/mTOR Signaling Pathways.

ViceninII is a naturally flavonoid glycoside extracted from Dendrobium officinale, a precious Chinese traditional herb, has been proven to be valuable for cancer treatment. Transforming growth factor-ß1 (TGF-ß1), promotes the induction of epithelial⁻mesenchymal transition (EMT), a process involved in the metastasis of cells that leads to enhanced migration and invasion. However, there is no previously evidence that ViceninII has an inhibitory effect on cancer metastasis, specifically on the TGF-ß1-induced EMT process in lung adenocarcinoma cells. In this experiment, we used UV, ESIMS, and NMR to identify the structure of ViceninII.A549 and H1299 cells were treated with TGF-ß1 in the absence and presence of ViceninII, and subsequent migration and invasion were measured by wound-healing and transwell assays. The protein localization and expressions were detected by immunofluorescence and Western blotting. The results indicated that TGF-ß1 induced spindle-shaped changes, increased migration and invasion, and upregulated or downregulated the relative expression of EMT biomarkers. Meanwhile, these alterations were significantly inhibited when co-treated with ViceninII and inhibitors LY294002 and SB431542. In conclusion, ViceninII inhibited TGF-ß1-induced EMT via the deactivation of TGF-ß/Smad and PI3K/Akt/mTOR signaling pathways.This is the first time that the anti-metastatic effects of ViceninII have been demonstrated, and their molecular mechanisms provided.

Identification of flavonoids in Dendrobium huoshanense and comparison with those in allied species of Dendrobium by TLC, HPLC and HPLC coupled with electrospray ionization multi-stage tandem MS analyses.

Dendrobium huoshanense, a unique species in the genus Orchidaceae, is only found in China and is known as "mihu". Due to the lack of quality control, the use of D. huoshanense in the herbal market has been limited. In this study, methods based on thin-layer chromatography, high-performance liquid chromatography and high-performance liquid chromatography coupled with electrospray ionization multi-stage tandem mass spectrometry were used to identify the flavonoids in D. huoshanense and distinguish this species from other Dendrobium species. Using thin-layer chromatography, a characteristic band was observed for D. huoshanense, and this band was absent from the thin-layer chromatography plates of other Dendrobium species. Then, using high-performance liquid chromatography, nine peaks of flavonoids were observed in the chromatograms of ten batches of D. huoshanense. Ultimately, 22 flavonoids in D. huoshanense were identified by multi-stage tandem mass spectrometry, and 11 of these compounds are being reported from D. huoshanense for the first time. In addition, two compounds both with molecular weights of 710, were identified as being unique to D. huoshanense; one of these compounds, apigenin-6-C-α-L-rhamnosyl-(1→2)-ß-D-glucoside-8-C-α-L-arabinoside, was proven to be responsible for the characteristic thin-layer chromatography band of D. huoshanense. These analysis methods can be applied for the identification and quality control of D. Huoshanense.

Physicochemical and Antioxidant Properties of the Degradations of Polysaccharides from Dendrobium officinale and Their Suitable Molecular Weight Range on Inducing HeLa Cell Apoptosis.

Different molecular weight polysaccharides of Dendrobium officinale (DOPs) have gradually attracted attention because of their broad biological activities. They, however, remain poorly defined whether their antitumor activity is associated with molecular weight. In this study, the physicochemical, antioxidant, and antitumor properties of DOPs, including the crude polysaccharide (DOP) and its six degradation fractions (DOP1-DOP6) extracted from Dendrobium officinale, were determined. Consequently, DOPs were mainly composed of different ratios of mannose and glucose as follows: 5.15 : 1, 4.62 : 1, 4.19 : 1, 4.46 : 1, 4.32 : 1, 4.29 : 1, and 4.23 : 1, and their molecular weights were significantly different ranging from 652.29 kDa to 11.10 kDa. With the concentration increase of DOPs, the scavenging capacity against OH and DPPH free radicals increased. The antitumor ability of DOPs was different that DOP1-DOP5 (Mw: 176.29 kDa-28.48 kDa) exhibited the best antiproliferation activity than DOP (Mw: 652.29 kDa) and DOP6 (Mw: 11.10 kDa) in HeLa cells rather than PC9, A549, and HepG2 cells. Moreover, it is worth mentioning that DOP1 and DOP5 showed stronger capability on inducing apoptosis of HeLa cells than DOP and DOP6 via the mitochondrial pathway by upregulating the ratio of the Bax/Bal-2 mRNA expression. The results demonstrated that DOPs can be used as the potential natural antioxidant and antitumor products in pharmaceutical industries, and the molecular weight is a crucial influential factor of their antitumor activity that 28.48 kDa-176.29 kDa is a suitable range we may refer to.

Correction to: Physicochemical properties of polysaccharides from Dendrobium officinale by fractional precipitation and their preliminary antioxidant and anti-HepG2 cells activities in vitro.

The original version of the article [1] contained a mistake. The Figure legends are right, but the pictures in Figs. 3 and 4 are contrary. The corrected figures are given below.

Physicochemical properties of polysaccharides from Dendrobium officinale by fractional precipitation and their preliminary antioxidant and anti-HepG2 cells activities in vitro.

BACKGROUND: Dendrobium officinale as a precious traditional Chinese herb is widely used in medicines and health supplements. Thus the extraction, purification and biological activities of polysaccharides from the stem of Dendrobium officinale have significant meaning on theory and application value. METHODS: The crude Dendrobium officinale polysaccharide (DOP) was obtained by hot water extraction- ethanol precipitation method, and four new polysaccharide fractions (DOP-40, DOP-50, DOP-60, and DOP-70) were further obtained from the crude DOP by fractional precipitation with ethanol method, then four fractions were further purified by Toyopearl-H65F gel resin. The molecular weight and monosaccharide composition of four purified fractions were determined by high performance anion exchange chromatography and high performance liquid chromatography. The antioxidant activities of them were evaluated by the reducing power assay, and the superoxide anion, 2,2-diphenyl-1-picrylhydrazyl (DPPH), and hydroxyl free radicals scavenging assays, respectively. Finally, the anticancer activities of them were investigated via the MTT assay and the western blot analysis using HepG2 cells. RESULTS: Among these four purified fractions were mainly composed of D-mannose and D-glucose with different molar ratios, and their average molecular weights were 999, 657, 243 and 50.3 kDa, respectively. What's more, DOP-70 always exhibited the strongest antioxidant and anticancer activities, while DOP-40 and DOP-60 showed very close antioxidant and anticancer activities which were better than that of DOP-50. The western blotting analysis also showed that DOP-40, DOP-60, and DOP-70 induced apoptosis in HepG2 human liver cancer cells through the Bcl-2 and Bax-dependent pathway. CONCLUSIONS: Fractional precipitation with ethanol could successfully apply to extract four new polysaccharide fractions from Dendrobium officinale stems, and the polysaccharide fractions possessed efficient antioxidant and anticancer activities, especially DOP-70.

Isoviolanthin Extracted from Dendrobium officinale Reverses TGF-ß1-Mediated Epithelial⁻Mesenchymal Transition in Hepatocellular Carcinoma Cells via Deactivating the TGF-ß/Smad and PI3K/Akt/mTOR Signaling Pathways.

Dendrobium officinale is a precious medicinal herb and health food, and its pharmacological actions have been studied and proved. However, the mechanisms by which its active flavonoid glycosides affect epithelial⁻mesenchymal transition (EMT) in hepatocellular carcinoma (HCC) cells, such as HepG2 and Bel-7402 cells, have not been previously investigated. Therefore, we investigated whether isoviolanthin extracted from the leaves of Dendrobium officinale inhibits transforming growth factor (TGF)-ß1-induced EMT in HCC cells. In this study, the physicochemical properties and structure of isoviolanthin were identified by HPLC, UV, ESIMS, and NMR and were compared with literature data. HCC cells were pretreated with 10 ng/mL TGF-ß1 to induce EMT and then treated with isoviolanthin. Herein, we found that isoviolanthin exhibited no cytotoxic effects on normal liver LO2 cells but notably reduced the migratory and invasive capacities of TGF-ß1-treated HCC cells. Additionally, isoviolanthin treatment decreased matrix metalloproteinase (MMP)-2 and -9 levels, and remarkably altered the expression of EMT markers via regulating the TGF-ß/Smad and PI3K/Akt/mTOR signaling pathways; Western blot analysis confirmed that the effects of the inhibitors SB431542 and LY294002 were consistent with those of isoviolanthin. These findings demonstrate the potential of isoviolanthin as a therapeutic agent for the treatment of advanced-stage metastatic HCC.

Transcriptome Analysis Reveals genes involved in flavonoid biosynthesis and accumulation in Dendrobium catenatum From Different Locations.

In this study, we applied transcriptome and UHPLC-MS technologies to investigate the flavonoids and their biosynthesis- and accumulation-related genes in Dendrobium catenatum from three different locations. Eight flavonoid glycosides were identified using standard references or previously isolated substances with MS data analysis. The total flavonoid contents were determined by reagents, and all the data were analyzed. In total, 23139 unigenes were obtained using the Dendrobium catenatum genome data. Of these, 10398 were annotated in the Gene Ontology (GO) database, 4203 were annotated in the KEGG database, and 10917 were annotated in the EuKaryotic Orthologous Groups (KOG) database. Thirty-one of the unigenes annotated by the KEGG database were involved in flavonoid pathways. The genes involved in bio-modification, accumulation, transportation and the regulation of the flavonoid bio-synthesis process were investigated. In conclusion, the flavonoids in Dendrobium catenatum from three different locations were different in quantitative and qualitative which may contribute to the establishment of quality control method for this herbal plant. These differences were determined by flavonoids biosynthesis process and they were concluded by sorting out the expression level of certain biosynthesis related genes.

Simultaneous identification and determination of flavonoids in Dendrobium officinale.

BACKGROUND: The quality of material medicine resources has had a considerable impact on the development of the health industry, which has created a bottleneck for traditional Chinese medicine (TCM). Dendrobium officinale, which has been widely used for health prevention in TCM, has become a high-nutritive health food that is strongly recommended by many white-collar workers and people who pay more attention to their health. The aim of this study was to develop a method to authenticate and evaluate D. officinale from different origins via simultaneous qualitative and quantitative analyses of flavonoid glycosides. Ultra-high-performance liquid chromatography-electrospray ionization/mass spectrometry was used for the structural elucidation of the compounds. RESULTS: 9 characteristic peaks, including those representing 7 flavonoid C-glycosides and 2 flavonoid O-glycosides, were identified. Additionally, the contents of 5 representative flavonoid glucosides in 25 batches of D. officinale from different sources were determined. To further investigate the different sources of the 25 batch samples, principal component analysis (PCA) and hierarchical cluster analysis (HCA) were carried out. A study on the methodology revealed that all results were reliable. CONCLUSIONS: This method is an efficient tool for the rapid identification of the different geographical origins of D. officinale and provides references for the quality evaluation of other natural products.

Rosmarinic acid protects on rat bone marrow mesenchymal stem cells from hydrogen peroxide-induced apoptosis.

To investigate the anti-oxidant activities and mechanism of rosmarinic acid (RA) on rat bone marrow mesenchymal stem cells (rBMSCs) from ischemia-induced apoptosis in vitro, which was established using H2O2-damage and analyzed for cell viability, cell apoptosis, ROS, morphological changes, and levels of apoptosis proteins. Pretreatment with RA significantly suppressed the generation of ROS, protected the morphological changes of cells, decrease the ratio of cell apoptosis, down-regulated the level of caspase-3, caspase-9, Bax/Bcl-2, and up-regulated the level of p-PI3K. These findings suggest that RA may protect rBMSCs from H2O2-induced apoptosis by partly regulating PI3K/Akt signaling pathway and can be developed as a potential anti-apoptotic agent for therapy in cardiovascular diseases.

Protective effect of gigantol against hydrogen peroxide­induced apoptosis in rat bone marrow mesenchymal stem cells through the PI3K/Akt pathway.

Bone marrow mesenchymal stem cell (BMSC) transplants are promising for the treatment of certain central nervous system diseases. However, oxidative stress is one of the major factors that may limit the survival of the transplanted BMSCs. The present study investigated the effect of pretreatment with gigantol on hydrogen peroxide (H2O2)­induced apoptosis in rat BMSCs (rBMSCs) and the potential underlying mechanisms. The results demonstrated that gigantol pretreatment significantly inhibited H2O2­induced apoptosis of rBMSCs. rBMSCs were incubated with 600 µM H2O2 in the absence or presence of different doses of gigantol (1­100 µM). Cell viability and apoptosis ratios were assessed by MTT assays and flow cytometry, respectively. Morphological alterations and reactive oxygen species were measured by the fluorescent­based methods of Hoechst staining and dichlorodihydrofluorescein diacetate, respectively. Furthermore, the protein levels of phosphorylated­protein kinase B (Akt), B­cell lymphoma­2 (Bcl­2), Bcl­2­associated X (Bax), caspase­3 and caspase­9 were investigated by western blotting. Following incubation with H2O2 for 2 h, gigantol significantly inhibited the H2O2­induced reductions in the cell viability of rBMSCs in a dose­dependent manner. Furthermore, gigantol upregulated Akt phosphorylation and Bcl­2 expression, downregulated Bax expression, and reduced the expression of caspase­3 and caspase­9 in H2O2­treated rBMSCs, whereas an opposite effect was detected when LY294002, an inhibitor of phosphatidylinositol 3­kinase, was administered in combination with gigantol. These results indicate that gigantol may be developed as a promising neuroprotective agent for successful MSC transplantation in ischemic diseases.

Gigantol attenuates the proliferation of human liver cancer HepG2 cells through the PI3K/Akt/NF-κB signaling pathway.

Gigantol is a phenolic substance extracted from plants in the genus Dendrobium and used in traditional Chinese medicine. In the present study, we aimed to investigate the growth inhibition and apoptotic effects of gigantol on human liver cancer cells through the PI3K/Akt/NF-κB signaling pathway. HepG2 cells were treated with different concentrations of gigantol (0-150 µM) for 12, 24 and 48 h. It was found that gigantol significantly inhibited the proliferation and induced apoptosis of the HepG2 cells. The results of fluorescence micrographs showed that a 48-h treatment with gigantol induced typical apoptotic morphological features, which were consistent with the flow cytometric analysis where 20% of apoptotic cells were detected in response to gigantol treatment. In addition, western blot analysis indicated that gigantol enhanced the activities of caspase-3, PARP and p53 and downregulated the expression of p-Akt/Akt. Collectively, the present data suggest that gigantol induces growth inhibition and apoptosis of HepG2 cells via the PI3K/Akt/NF-κB signaling pathway.