eISSN: 1896-9151
ISSN: 1734-1922
Archives of Medical Science
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SCImago Journal & Country Rank
4/2019
vol. 15
 
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abstract:
Basic research

Methylwogonin exerts anticancer effects in A375 human malignant melanoma cells through apoptosis induction, DNA damage, cell invasion inhibition and downregulation of the mTOR/PI3K/Akt signalling pathway

Jiaorong Chen
,
Chunmei Huang
,
Fangfang Liu
,
Zihui Xu
,
Li Li
,
Zheng Huang
,
Hongfeng Zhang

Arch Med Sci 2019; 15 (4): 1056–1064
Online publish date: 2018/04/16
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Introduction
The main purpose of the present research was to study the anticancer effects of methylwogonin in A375 human malignant melanoma cells by evaluating its effects on apoptosis, DNA fragmentation, cancer cell invasion and the mTOR/PI3K/AKT signalling pathway.

Material and methods
Effects on cell cytotoxicity were evaluated by MTT assay while a clonogenic assay determined the effects of methylwogonin on colony formation. Fluorescence microscopy evaluated apoptotic effects of methylwogonin in these cells using acridine orange/propidium iodide and Hoechst 33342 staining dyes. Gel electrophoresis evaluated the effects of methylwogonin on DNA fragmentation while the Matrigel invasion assay evaluated the effects of the drug on cancer cell invasion. Effects of methylwogonin on the mTOR/PI3K/AKT signalling pathway were evaluated by western blot assay.

Results
The results showed that methylwogonin induces concentration-dependent as well as time-dependent growth inhibitory effects inducing significant cytotoxicity in these cancer cells. Methylwogonin led to dose-dependent inhibition of colony formation in A375 human malignant melanoma cells. The number of cell colonies decreased significantly as the methylwogonin dose increased from 0, 50, 150, to 300 µM. Methylwogonin treatment of cells at lower doses led to yellow fluorescence (early apoptosis), which changed to red/orange fluorescence, indicating late apoptosis at higher doses. Similar results were obtained using Hoechst 33342 staining, revealing that 50, 150 and 300 µM doses of methylwogonin led to significant morphological changes including chromatin condensation, fragmented nuclei and cellular shrinkage. DNA ladder formation was also observed, and this effect increased with increasing doses of methylwogonin. Methylwogonin also inhibited cancer cell invasion in a dose-dependent manner.

Conclusions
Different doses of methylwogonin led to concentration-dependent downregulation of phosphorylated PI3K, AKT and mTOR.

keywords:

human melanoma, apoptosis, methylwogonin, fluorescence microscopy, cell invasion

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