icaritin and desmethylicaritin

Epimedium is popularly used in traditional Chinese medicine to treat sexual dysfunction, menstrual irregularity, and osteoporosis. The estrogenic effects of the prenylated flavonoids of Epimedium make it an attractive alternative for hormone replacement therapy. Here, we examined the therapeutic potential of the estrogenic herb extract of Epimedium brevicornum as an alternative to hormone replacement therapy in a breast cancer mouse model. To that end, athymic and ovariectomized female nude mice were subcutaneously injected into the mammary fat pads with MCF-7 breast cancer cells, randomly grouped and fed with soy-free feeds, alone or in combination with ethinyl estradiol or different doses of the estrogenic herb extract of E. brevicornum. Our findings demonstrate that unlike ethinyl estradiol, it did not promote the growth of breast cancer xenograft volume and weight, with the highest dose showing a significant reduction in growth and ERα protein content. Moreover, the extract increased uterine weight at the lowest dose, while higher doses had no effects. Put together, our data shows for the first time that despite the estrogenic activity of E. brevicornum, its action is largely tissue specific and dose-dependent. Our data on E. brevicornum presents in vivo evidence for its selective estrogen receptor modulator effect and warrants exploration of its use as an alternative to hormone replacement therapy in menopausal women.

Topics: Animals; Breast Neoplasms; Dose-Response Relationship, Drug; Epimedium; Estrogen Receptor alpha; Ethinyl Estradiol; Female; Flavonoids; Humans; Medicine, Chinese Traditional; Mice; Mice, Nude; Organ Size; Ovariectomy; Plant Extracts; Selective Estrogen Receptor Modulators; Uterus; Xenograft Model Antitumor Assays

The significant promoting effects of some prenylflavonoids on cardiac differentiation of mouse embryonic stem (ES) cells via reactive oxygen species (ROS) signaling pathway were investigated. The most effective differentiation was facilitated by icariin (ICA), followed by icaritin (ICT), while desmethylicaritin (DICT) displayed the weakest but still significant inducible effect. Contrarily, DICT demonstrated the strongest anti-oxidative activity while ICA displayed only little in vitro, which was well matched with the hydroxyl (OH) numbers and the positions in the molecular structures. Therefore, ROS signaling cascades were assumed to be involved in prenylflavonoids induced cardiomyogenesis. Treatment with ICA, intracellular ROS in embryoid bodies was rapidly elevated, which was abolished by the NADPH-oxidase inhibitor apocynin; elimination of intracellular ROS by vitamin E or pyrrolidine dithiocarbamate (PDTC) inhibited ICA induced cardiomyogenesis; ROS-sensitive extracellular-regulated kinase 1, 2 (ERK1, 2) and p38 activation were further observed, the cardiomyogenesis was significantly inhibited in the presence of ERK1, 2 or p38 inhibitor U0126 or SB203580, indicating the roles of NADPH-ROS-MAPKs signaling cascades in prenylflavonoids induced cardiac differentiation. There was no difference in Nox4 NADPH oxidase expression between ICA and ICT treatments, however, ROS concentration in EBs after ICT administration was lower than that after ICA treatment, followed by less activation of ERK1, 2, and p38. These results revealed that the significant promoting effects of prenylflavonoids on cardiac differentiation was at least partly via ROS signaling cascades, and the facilitating abilities preferentially based on the nature of prenylflavonoids themselves, but anti-oxidative activity determined by the OH numbers and the positions in the structures do influence the cardiomyogenesis in vitro.

Topics: Animals; Cell Differentiation; Cell Line; Embryonic Stem Cells; Enzyme Inhibitors; Flavonoids; MAP Kinase Signaling System; Mice; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Myocytes, Cardiac; NADPH Oxidase 4; NADPH Oxidases; p38 Mitogen-Activated Protein Kinases; Reactive Oxygen Species

The prenyl-flavones, icaritin and desmethylicaritin, are bioactive compounds from the traditional Chinese medicinal herb, Epimedium, extracts of which can enhance bone health in animal models. In order to examine their bioavailability in humans, we have developed and validated a sensitive method to quantify icaritin and desmethylicaritin in human sera, using gas chromatography-mass spectrometry. The serum samples were extracted with ethyl acetate and then derivatized with BSTFA in pyridine (4:1). With genistein as internal standard, calibration curves with good linearity (R(2)>0.99) within the concentration range of 0.15-10nM in the selective ion monitoring mode were obtained. The limits of detection and quantization were 11 and 33 pM for icaritin, and 23 and 70 pM for desmethylicaritin, respectively; inter- and intra-assay variabilities were <15%, and accuracies were between 89 and 110%. Icaritin, but not desmethylicaritin, was detected from 1h, increasing to a peak at 8h (1.51+/-1.6 nM) in sera of human volunteers after ingestion of an aqueous decoction of Epimedium. This sensitive method can be used to quantify serum levels of icaritin and desmethylicaritin for pharmacokinetic studies.

Topics: Administration, Oral; Biological Availability; Calibration; Drugs, Chinese Herbal; Epimedium; Feasibility Studies; Flavonoids; Gas Chromatography-Mass Spectrometry; Genistein; Humans; Menopause; Molecular Structure; Phytotherapy; Plants, Medicinal; Reference Standards; Reproducibility of Results; Sensitivity and Specificity; Spectrometry, Mass, Electrospray Ionization; Trimethylsilyl Compounds

To investigate the possible inducible effects of icariin, icaritin, and desmethylicaritin on the directional differentiation of embryonic stem (ES) cells into cardiomyocytes in vitro.. ES cells were cultivated as embryoid bodies (EBs) in hanging drops with icariin, icaritin, or desmethylicaritin. ES cells treated with retinoic acid and with solvent were used as positive and negative controls, respectively. The cardiomyocytes derived from the ES cells were verified using immunocytochemistry. The expression of cardiac developmental-dependent genes was detected using the reverse transcription-polymerase chain reaction (RT-PCR) method. Cell cycle distribution and apoptosis were analyzed using flow cytometry to determine the partly inducible effect mechanisms involved.. The total percentage of beating EBs treated with 10(-7) mol/L icariin, icaritin, or desmethylicaritin was 87% (P<0.01), 59% (P<0.01), and 49%, respectively. All the beating cardiomyocytes derived from the ES cells expressed cardiac-specific proteins for a-actinin and troponin T. Among them, 10(-7) mol/L icariin treatment resulted in a significantly advanced and increased mRNA level of a-cardiac major histocompatibility complex (MHC) and myosin light chain 2v (MLC-2v) in EBs in the early cardiac developmental stage. Before shifting to the cardiomyocyte phenotype, icariin could evoke the accumulation of ES cells in G0/G1 and accelerate apoptosis of the cell population (P<0.05).. Icariin facilitated the directional differentiation of ES cells into cardiomyocytes at a concentration of 10(-7) mol/L. The promoting effect of icariin on cardiac differentiation was related to increasing and accelerating gene expression of a-cardiac MHC and MLC-2v, as well as regulating the cell cycles and inducing apoptosis.

Topics: Animals; Apoptosis; Cell Cycle; Cell Differentiation; Cells, Cultured; Embryo, Mammalian; Epimedium; Female; Flavonoids; Gene Expression Regulation; Male; Mice; Myocytes, Cardiac; Myosin Heavy Chains; Myosin Light Chains; Plants, Medicinal; RNA, Messenger; Stem Cells

To investigate the estrogen-like activities of icariin (ICA), icaritin (ICT) and desmethylicaritin (DICT) and their structure/activity relationships.. ICT was hydrolyzed from ICA by cellulase and then DICT was demethylated from ICT in boron tribromide and dichloromethane system. Estrogen-sensitive MCF-7 cells and T47D cells were co-incubated with different concentrations of test compounds for 6 and 9 d respectively, and the cell proliferation was measured by MTT.. ICT and DICT both markedly enhanced cell proliferation. Compared with estradiol (10.(-9) mol/L), the proliferative effects of 10.-6 mol/L ICT and DICT on MCF-7 cells were 90.0% and 94.0% (P<0.01), respectively, and those of T47D cells were 65.6% and 50.0%. (P<0.01). But this phenomenon was not observed with ICA. Cell proliferation induced by ICT and DICT was completely antagonized by 10.(-7 )mol/L pure estrogen receptor antagonist, ICI182,780.. ICT and DICT possess estrogen-like activity of enhancing proliferation in MCF-7 and T47D cells. However, ICA appears to have no estrogenicity on MCF-7 and T47D cell lines in vitro.

Topics: Breast Neoplasms; Cell Division; Drugs, Chinese Herbal; Flavonoids; Humans; Phytoestrogens; Tumor Cells, Cultured

The aims of the present study were to determine the estrogenic activities of icariin (ICA) and its derivatives and their structure-estrogenic activity relationship. Therefore, icaritin (ICT) and desmethylicaritin (DICT) were derived from ICA. The estrogenic activities of ICA, ICT and DICT were examined by cell proliferation and progestogen receptor mRNA expression of estrogen-receptor-positive MCF-7 cells. Current studies exhibited that ICT and DICT both markedly enhanced the proliferation of MCF-7 cells; as compared to estradiol (100%), their relative proliferative effects (RPE) were 90% and 94%, respectively. Cell proliferation induced by ICT and DICT was completely antagonized by ICI182,780. ICT and DICT increased progestogen receptor (PR) at mRNA levels at 48 h after treatment, although the effects were not as prominent as 17beta-estradiol (E2). Those phenomena were not observed with ICA. Results demonstrate that ICT and DICT (nonconjugated forms) possess estrogen-like activity; however, ICA appears to have no estrogenicity in the MCF-7 cell line model in vitro.

Topics: Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Drugs, Chinese Herbal; Estrogen Receptor Modulators; Female; Flavonoids; Humans; Receptors, Progesterone; Structure-Activity Relationship; Transcription, Genetic

Icariin, icaritin and desmethylicaritin are constituents of Epimedium with a similar structure to genistein and daidzein. Using the modified MCF-7 cell proliferation assay (E-SCREEN assessment system), these compounds were tested for their estrogen-like activities. Icaritin and desmethylicaritin, but not icariin, strongly stimulated the proliferation of MCF-7/BUS cells. Cell cycle analysis revealed that the proliferation stimulatory effect was associated with a marked increase in the number of MCF-7/BUS cells in S phase and a significant increase in the G2/M population, with effects similar to those of estradiol. These actions were dose dependent (range from 1 nM to 10 microM) and could be significantly inhibited by the specific estrogen receptor antagonist ICI 182,780 [7 alpha-[9(4,4,5,5,5-pentafluoropentyl)sulfinyl]nonyl)-estra-1,3,5(10)-triene-3,17beta-diol)]. The estrogen receptor-regulated progesterone receptor and PS2 mRNA levels were increased by treatment with icaritin or desmethylicaritin within 24 h and the effects were also reversed by ICI 182,780. It was concluded that icaritin and desmethylicaritin are novel phytoestrogens and that the estrogenic effects of icaritin and desmethylicaritin are mediated by the estrogen receptor.

Topics: Cell Cycle; Cell Line, Tumor; Cell Proliferation; Charcoal; Dextrans; Estradiol; Estrogen Antagonists; Estrogens, Non-Steroidal; Female; Flavonoids; Fulvestrant; Humans; Receptors, Estrogen; Receptors, Progesterone; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stimulation, Chemical