Background: Flavonoids, a group of compounds mainly derived from vegetables and herbal medicines, chemically resemble estrogen and some have been used as estrogen substitutes. Kaempferol, a flavonol derived from the rhizome of Kaempferia galanga L., is a well-known phytoestrogen possessing osteogenic effects that is also found in a large number of plant foods. The herb K. galanga is a popular traditional aromatic medicinal plant that is widely used as food spice and in medicinal industries. In the present study, both the estrogenic and osteogenic properties of kaempferol are evaluated. Methods: Kaempferol was first evaluated for its estrogenic properties, including its effects on estrogen receptors. The osteogenic properties of kaempferol were further determined its induction effects on specific osteogenic enzymes and genes as well as the mineralization process in cultured rat osteoblasts. Results: Kaempferol activated the transcriptional activity of pERE-Luc (3.98 ± 0.31 folds at 50 μM) and induced estrogen receptor α (ERα) phosphorylation in cultured rat osteoblasts, and this ER activation was correlated with induction and associated with osteoblast differentiation biomarkers, including alkaline phosphatase activity and transcription of osteoblastic genes, e.g., type I collagen, osteonectin, osteocalcin, Runx2 and osterix. Kaempferol also promoted the mineralization process of osteoblasts (4.02 ± 0.41 folds at 50 μM). ER mediation of the kaempferol-induced effects was confirmed by pretreatment of the osteoblasts with an ER antagonist, ICI 182,780, which fully blocked the induction effect. Conclusion: Our results showed that kaempferol stimulates osteogenic differentiation of cultured osteoblasts by acting through the estrogen receptor signaling.

Astragali Radix (AR), a Chinese materia medica (CMM) known as Huangqi, is an important medicine prescribed in herbal composite formulae (Fufang) by Traditional Chinese medicine (TCM) practitioners for thousands of years. According to the literature, AR is suggested for patients suffering from “Qi”- and “Blood”-deficiencies, and its clinical effects are reported to be related to anti-cancer cell proliferation, anti-oxidation, relief of complications in cardiovascular diseases, etc. The underlying cell signaling pathways involved in the regulation of these various diseases are presented here to support the mechanisms of action of AR. There are two botanical sources recorded in China Pharmacopoeia (CP, 2015): Astragalus membranaceus (Fisch.) Bge. Var. mongohlicus, (Bge.) Hsiao, and Astragalus membranaceus (Fisch.) Bge. (Fam. Leguminosae), whose extracts of dried roots are processed via homogenization-assisted negative pressure cavitation extraction. Geographic factors and extraction methods have impacts on the pharmaceutical and chemical profiles of AR. Therefore, the levels of the major bioactive constituents of AR, including polysaccharides, saponins, and flavonoids, may not be consistent in different batches of extract, and the pharmaceutical efficacy of these bioactive ingredients may vary depending on the source. Therefore, the present review mainly focuses on the consistency of the available sources of AR and extracts and on the investigation of the biological functions and mechanisms of action of AR and of its major bioactive constituents. Furthermore, it will also include a discussion of the most popular AR composite formulae to further elucidate their chemical and biological profiles and understand the pharmaceutical value of AR.

Ginkgetin, a biflavone from Ginkgo biloba leaf, and resveratrol, a polyphenol found in grape and wine, are two phytochemicals being identified for its binding to vascular endothelial growth factor (VEGF): the binding, therefore, resulted in the alteration of the physiological roles of VEGF-mediated angiogenesis. The bindings of ginkgetin and resveratrol were proposed on different sites of VEGF, but both of them suppressed the angiogenic properties of VEGF. The suppressive activities of ginkgetin and resveratrol in VEGF-mediated angiogenesis were supported by several lines of evidence including (i) inhibiting the formation of sub-intestinal vessel in zebrafish embryos and microvascular sprouting in rat aortic ring; and (ii) suppressing the phosphorylations of VEGFR2, Akt, eNOS, and Erk as well as expressions of matrix metalloproteinases (MMPs), MMP-2, and MMP-9 in human umbilical vein endothelial cells (HUVECs). Here, we showed the synergy of ginkgetin and resveratrol in suppressing the VEGF-induced endothelial cell proliferation, migration, invasion, and tube formation. The synergy of ginkgetin and resveratrol was further illustrated in HT-29 colon cancer xenograft nude mice. Ginkgetin and resveratrol, when applied together, exerted a synergistic anti-tumor effect of 5-fluorouracil with decreasing microvessel density of tumors. In parallel, the combination of ginkgetin and resveratrol synergistically relieved the 5-fluorouracil-induced inflammatory response by suppressing expressions of COX-2 and inflammatory cytokines. Thus, the anti-angiogenic roles of ginkgetin and/or resveratrol could provide effective therapeutic strategy in cancer, similar to that of Avastin, in suppressing the VEGF-mediated angiogenesis during cancer development.

This article exemplifies a systematic approach to revealing the complexity of Chinese herbal medicine formulae through three levels of scientific research: standardization of herbs, verification of ancient formulae and mechanism studies. We use Danggui Buxue Tang (DBT) as an example for this approach. Among thousands of traditional Chinese medicine herbal formulae, almost all of which consist of multiple herbs, DBT is one of the simplest. Containing only two herbs, namely Radix Astragali (RA) and Radix Angelicae Sinensis (RAS), DBT is traditionally used to treat ailments in women. The weight ratio of RA to RAS in DBT was prescribed to be 5:1 as early as in 1247 AD. In addition to advanced chemical analysis of herbal constituents, DNA genotyping techniques have been developed for reliable standardization of RA and RAS. Chemical evaluation shows that main active constituents in DBT, including astragaloside IV, calycosin, formononetin and ferulic acid, were most abundant after extraction at the RA to RAS ratio of 5:1, whereas other tested RA to RAS ratios only gave sub-optimal levels of the active constituents. Biological evaluation indicates that bioactivities of DBT, e.g. immuno-modulatory, oesteotropic and estrogenic effects are also best exerted at the RA to RAS ratio of 5:1. Correlation analysis demonstrates statistically significant relationship between the tested chemical constituents and tested bioactivities. Up- and down-regulation of expression of some genes as potential biomarkers has been detected by using gene chip technology. This systematic approach on the basis of herbal standardization, chemical and biological verification and mechanism studies, as exemplified in this article, will be useful to reveal the complexity of not only DBT but also other Chinese medicine herbal formulae.

Xiao Cheng Qi (XCQ) decoction, an ancient Chinese herbal mixture, has been used in treating slow-transit constipation (STC) for years. The underlying action mechanism in relieving the clinical symptoms is unclear. Several lines of evidence point to a strong link between constipation and gut microbiota. Short-chain fatty acids (SCFAs) and microbial metabolites have been shown to affect 5-HT synthesis by activating the GPR43 receptor localized on intestinal enterochromaffin cells, since 5-HT receptors are known to influence colonic peristalsis. The objective of this study was to evaluate the efficacy of XCQ in alleviating clinical symptoms in a mouse model of STC induced by loperamide. The application of loperamide leads to a decrease in intestinal transport and fecal water, which is used to establish the animal model of STC. In addition, the relationship between constipation and gut microbiota was determined. The herbal materials, composed of Rhei Radix et Rhizoma (Rhizomes of Rheum palmatum L., Polygonaceae) 55.2 g, Magnoliae Officinalis Cortex (Barks of Magnolia officinalis Rehd. et Wils, Magnoliaceae) 27.6 g, and Aurantii Fructus Immaturus (Fruitlet of Citrus aurantium L., Rutaceae) 36.0 g, were extracted with water to prepare the XCQ decoction. The constipated mice were induced with loperamide (10 mg/kg/day), and then treated with an oral dose of XCQ herbal extract (2.0, 4.0, and 8.0 g/kg/day) two times a day. Mosapride was administered as a positive drug. In loperamide-induced STC mice, the therapeutic parameters of XCQ-treated mice were determined, i.e., (i) symptoms of constipation, composition of gut microbiota, and amount of short-chain fatty acids in feces; (ii) plasma level of 5-HT; and (iii) expressions of the GPR43 and 5-HT4 receptor in colon. XCQ ameliorated the constipation symptoms of loperamide-induced STC mice. In gut microbiota, the treatment of XCQ in STC mice increased the relative abundances of Lactobacillus, Prevotellaceae_UCG_001, Prevotellaceae_NK3B31_group, Muribaculaceae, and Roseburia in feces and decreased the relative abundances of Desulfovibrio, Tuzzerella, and Lachnospiraceae_ NK4A136_group. The levels of SCFAs in stools from the STC group were significantly lower than those the control group, and were greatly elevated via treatment with XCQ. Compared with the STC group, XCQ increased the plasma level of 5-HT and the colonic expressions of the GPR43 and 5-HT4 receptor, significantly. The underlying mechanism of XCQ in anti-constipation could be related to the modulation of gut microbiota, the increase in SCFAs, the increase in plasma 5-HT, and the colonic expressions of the GPR43 and 5-HT4 receptor. Our results indicate that XCQ is a potent natural product that could be a therapeutic strategy for constipation.