Psoralidin induces autophagy through ROS generation which inhibits the proliferation of human lung cancer A549 cells.

Wenhui Hao,Wenwen Zhao,Xuenong Zhang,Xiuping Chen

doi:10.7717/peerj.555

Abstract

Psoralidin (PSO), a natural furanocoumarin, is isolated from Psoralea corylifolia L. possessing anti-cancer properties. However, the mechanisms of its effects remain unclear. Herein, we investigated its anti-proliferative effect and potential approaches of action on human lung cancer A549 cells. Cell proliferation and death were measured by MTT and LDH assay respectively. Apoptosis was detected with Hoechst 33342 staining by fluorescence microscopy, Annexin V-FITC by flow cytometry and Western blot analysis for apoptosis-related proteins. The autophagy was evaluated using MDC staining, immunofluorescence assay and Western blot analyses for LC3-I and LC3-II. In addition, the reactive oxygen species (ROS) generation was measured by DCFH2-DA with flow cytometry. PSO dramatically decreased the cell viabilities in dose- and time-dependent manner. However, no significant change was observed between the control group and the PSO-treated groups in Hoechst 33342 and Annexin V-FITC staining. The expression of apoptosis-related proteins was not altered significantly either. While the MDC-fluorescence intensity and the expression ratio of LC3-II/LC3-I was remarkably increased after PSO treatment. Autophagy inhibitor 3-MA blocked the production of LC3-II and reduced the cytotoxicity in response to PSO. Furthermore, PSO increased intracellular ROS level which was correlated to the elevation of LC3-II. ROS scavenger N-acetyl cysteine pretreatment not only decreased the ROS level, reduced the expression of LC3-II but also reversed PSO induced cytotoxicity. PSO inhibited the proliferation of A549 cells through autophagy but not apoptosis, which was mediated by inducing ROS production.

Highlights

Psoralea corylifolia Leguminosae (L.), an herb widely distributed in China and Southeastern Asian countries, has been used as a multi-purpose medicinal plant (Zhao et al, 2005)
Previous screening studies have reported that some extracts and active fractions of P. corylifolia L. exhibited cytotoxicity and inhibition of chemical carcinogenesis (Latha et al, 2000; Latha & Panikkar, 1999; How to cite this article Hao et al (2014), Psoralidin induces autophagy through reactive oxygen species (ROS) generation which inhibits the proliferation of human lung cancer A549 cells
PSO induced cell cycle arrest at G1 phase To measure the underlying mechanism responsible for the anti-proliferation effect of PSO in A549 cells, cell cycle distribution was detected by flow cytometry analysis of DNA content using propidium iodide (PI) staining

Summary

Introduction

Psoralea corylifolia Leguminosae (L.), an herb widely distributed in China and Southeastern Asian countries, has been used as a multi-purpose medicinal plant (Zhao et al, 2005). Previous screening studies have reported that some extracts and active fractions of P. corylifolia L. exhibited cytotoxicity and inhibition of chemical carcinogenesis (Latha et al, 2000; Latha & Panikkar, 1999; How to cite this article Hao et al (2014), Psoralidin induces autophagy through ROS generation which inhibits the proliferation of human lung cancer A549 cells. PSO induced apoptosis in breast cancer cells by inhibiting NOTCH1 signaling (Suman, Das & Damodaran, 2013) It inhibits TNF-mediated survival signaling in androgen independent prostate cancer cells (Srinivasan et al, 2010). We demonstrated that PSO is an anti-proliferative natural compound on human lung cancer A549 cells It induces autophagy rather than apoptosis, which is triggered by increasing intracellular ROS generation

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Conclusion