Total coumarins of Hedyotis diffusa induces apoptosis of myelodysplastic syndrome SKM-1 cells by activation of caspases and inhibition of PI3K/Akt pathway proteins
2017
Jiang, Jianping | Wang, Bo | Li, Jing | Ye, Baodong | Lin, Shengyun | Qian, Wenbin | Shan, Letian | Efferth, Thomas
Hedyotis diffusa is an ethno-medicine used for anti-cancer treatment in the clinic of traditional Chinese medicine (TCM). The total coumarins of Hedyotis diffusa (TCHD) was a selected extract with observed antiproliferative activity, which has not been tested in treatment of myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML).This study aimed to evaluate the apoptosis-inducing effect of TCHD on human MDS cell line (SKM-1) and explore its action mechanism in association with caspase family and PI3K/Akt signaling pathway.The chemical constituents and total coumarins content of TCHD were determined by High Performance Liquid Chromatography-tandem mass spectrometry (HPLC-MS/MS) and UV–vis spectrophotometry, respectively. MTT assay, Hoechst 33258 staining, and Annexin V-FITC/PI double labeling were applied to evaluate TCHD's efficacy on SKM-1 cells. Western blot analysis was also used to clarify the action mechanism of TCHD on protein expression level.Two compounds, p-coumaric acid and E-6-O-p-coumaroyl scandoside methyl ester, were identified in TCHD, and its total coumarins content reached 87.4%. By MTT assay, apoptosis-inducing effect of TCHD on SKM-1 cells was found in a dose-dependent manner after 24–48h treatment, with IC50 values of 104.48μg/ml and 100.66μg/ml, respectively. Morphological and flow cytometry observation also confirmed such effect of TCHD. Western blot analysis clarified its action mechanism associating with the activation of caspases and inhibition of PI3K/Akt pathway proteins.This is the first report regarding the apoptosis-inducing efficacy and mechanism of TCHD on SKM-1 cells, providing a promising candidate of TCM for MDS and AML therapy with fewer side effects.
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