Polypeptide Fraction fromArca subcrenataInduces Apoptosis and G2/M Phase Arrest in HeLa Cells via ROS-Mediated MAPKs Pathways

Xianjing Hu,Jinghua Cai,Rongmin Yu,Jianhua Zhu,Zhongyi Guo,Liyan Song,Zhang Zhang,Ting Liu

doi:10.1155/2015/930249

Abstract

Arca subcrenata is documented in the literature of marine Traditional Chinese Medicine. Polypeptide fraction from A. subcrenata, coded as P2, was demonstrated to possess significant antitumor activity in our previous study. However, the underlying mechanism remains undefined. The present study was carried out to investigate the underlying antitumor mechanism of P2 in human cervical cancer HeLa cells by MTT, FCM, LSCM, and western blot assays. The results revealed that P2 significantly induced apoptosis of HeLa cells in a concentration- and time-dependent manner. High level of ROS was provoked by P2, which was in turn responsible for induction of apoptosis through activation of intrinsic mitochondrial pathway and JNK1/2, p38 MAPK pathways, as well as inhibition of ERK1/2 pathway, as evidenced by the abrogation of P2's effect on HeLa cells preincubated with the ROS scavenger NAC. P2 also was observed to display significant effect on G2/M phase arrest by downregulating the expression of cyclin B1/cdc2 complex and upregulating the expression of p21. These findings demonstrate that P2 induces apoptosis and G2/M phase arrest in HeLa cells through ROS-mediated MAPKs pathways, suggesting that P2 would be worth investigating as a promising agent within the scope of marine drugs for treatment of cervical cancer.

Highlights

Cervical cancer, the second most common type of tumor in females surpassed only by breast cancer [1], becomes a major health problem worldwide despite advances in screening [2]
Primary antibodies against cleaved caspases 3, 7, and 9, PARP, p-cdc2, p21, cyclin B1, ERK1/2, p-ERK1/2, p38, p-p38, c-Jun N-terminal kinase 1/2 ERK1/2 (JNK1/2), p-JNK1/2, and anti-rabbit or anti-mouse IgG horseradish peroxidase- (HRP-) linked secondary antibodies were purchased from Cell Signaling Technology (Boston, MA, USA)
To investigate the proapoptotic effect of P2, HeLa cells stained with DAPI were selected to analyze the morphological changes of apoptosis such as cell shrinkage, condensed and fragmented chromatin, and annexin V-FITC/PI double staining was used to quantify the percentage of apoptotic cells by Flow cytometry LSCM (FCM)

Summary

Introduction

The second most common type of tumor in females surpassed only by breast cancer [1], becomes a major health problem worldwide despite advances in screening [2]. For early-stage cervical cancer, patients are primarily treated by surgery or a combination of radiotherapy/chemotherapy [1]. Chemotherapeutic drugs are limited in clinical use because of their serious adverse effects, with increased doses and prolonged courses [3]. Newer chemotherapeutic agents with fewer side effects and better efficacy are urgently needed for the treatment of cervical cancer. Marine organisms are known for their capability of producing large amount of bioactive materials, with properties such as antitumor, anti-inflammatory, and antioxidant activities mainly due to the marine environment being characterized with high salinity, high pressure, low temperature, low light intensity, and an oligotrophic condition. Numbers of marine-original compounds have been found to exhibit significant antitumor activity such as antiproliferation of bryostatins on the murine lymphocytic leukemia P388 cells [6], anti-invasion of sungsanpin on A549

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