Abstract
Purpose: To determine the potential embryotoxicity of Radix scutellariae (RS) extract using an embryonic stem cell test (EST) and to evaluate its effect on the differentiation of mouse embryonic stem (ES) cells.Methods: All the test samples were obtained by water extraction method. The embryotoxicity of RS was assessed with cytotoxicity assays, namely, embryonic stem (ES) cells (IC50ES) and 3T3 fibroblasts (IC503T3), as well as cardiac differentiation inhibition assay (ID50ES). The expression of specific genes and proteins was analyzed by quantitative reverse transcription – polymerase chain reaction (RT-PCR) and Western blot.Results: RS was weakly embryotoxic with IC50ES, IC503T3 and ID50ES of 0.1524, 0.1061, and 0.4169 mg/ml, respectively. Also RS had discordant effects on the expression of tissue-specific genes and proteins in three germ layers, promoting differentiation of the ectoderm (⋆p < 0.05; ⋆⋆p < 0.01) and endoderm (⋆p < 0.05; ⋆⋆p < 0.01; ⋆⋆⋆p < 0.001), while inhibiting mesoderm differentiation (⋆p < 0.05; ⋆⋆p < 0.01; ⋆⋆⋆p < 0.001). The effect of RS on the embryonic development of the three germ layers was concentration-dependent.Conclusion: These results indicate that RS possesses weak embryotoxicity. The variability in the effects of RS may be responsible for its weak embryotoxicity.Keywords: Embryonic stem test, Radix scutellariae, Embryotoxicity, Cardiac differentiation inhibition assay, Ectoderm, Endoderm, Mesoderm
Highlights
Women are frequent users of natural herbs worldwide [1]
A cell viability assay was used to assess the cytotoxic effect of Radix scutellariae (RS) on the BALB/c 3T3 fibroblasts, representing adult tissues, and embryonic stem (ES) cells, representing embryonic tissues
RS extract stimulated the proliferation of the ES cells at low concentration levels
Summary
Women are frequent users of natural herbs worldwide [1]. It is well known that some herbs have powerful pharmacological effects, and significant side effects have been reported in recent years. 750 ES cells in 20 μL of differentiation medium (without mouse LIF) were placed on the lid of a petri dish filled with PBS on day 0. They were incubated for 2 days at 37 °C under 5 % CO2 and 95 % humidity in the presence of a concentration range of the test compound. On day 5, the EBs were transferred singly into wells of a 6-well tissue culture plate (containing the appropriate concentration of test compound) to allow adherence and outgrowth of the EBs and development of spontaneously beating myocardial cells. Specific oligonucleotide primers were designed to produce 100–200 bp products
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