Enhanced apoptotic action of trichosanthin in HIV-1 infected cells Yuan-Yuan Wang a,c , Dong-Yun Ouyang a,c , Hai Huang b , Herman Chan b , Siu-Cheung Tam b , Yong-Tang Zheng a, * a Laboratory of Molecular Immunopharmacology, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming 650223, China b Department of Physiology, The Chinese University of Hong Kong, Hong Kong SAR, China c Graduate School of the Chinese Academy of Sciences, Beijing 100039, China Received 17 March 2005 Available online 13 April 2005 Abstract Trichosanthin (TCS) is a type 1 ribosome-inactivating protein (RIP) effective against HIV-1 replication. The mechanism is not clear. Present results suggested that the antiviral action may be partly mediated through enhanced apoptosis on infected cells. TCS induced apoptosis in normal H9 cells and this action was more potent in those infected with HIV-1. In flow cytometry study, TCS induced larger population of apoptotic H9 cells chronically infected with HIV-1 in a dose-dependent manner. At TCS concentration of 25 lg/ml, 8.4% of normal H9 cells were found to be apoptotic whereas the same concentration induced 24.5% in HIV-1 chron- ically infected cells. Such difference was not found in the control experiments without TCS treatment. Two other studies supported this action. Cytotoxic study showed that cell viability was always lower in HIV-1 infected cells after TCS treatment, and DNA frag- mentation study confirmed more laddering in infected cells. The mechanism of TCS induced apoptosis in normal or infected H9 cells is not clear. Results in this study demonstrated that TCS is more effective in inducing apoptosis in HIV-1 infected cells. This may explain in part the antiviral action of TCS. Ó 2005 Elsevier Inc. All rights reserved. Keywords: Trichosanthin; HIV-1; Apoptosis; Infected cells; Anti-HIV activity Trichosanthin (TCS) is a type 1 ribosome-inactivat- ing protein (RIP) isolated from root tuber of Trichosan- thes kirilowii. This single chain polypeptide depurinates A-4324 of the 28S rRNA and inhibits protein synthesis [1]. In its crude form, it was used in ancient China as an abortifacient agent. The active ingredient is now known as trichosanthin. This protein was found to inhi- bit HIV-1 replication in the late 1980s and generated some research interest in the past years [2–8]. Clinical trial of TCS on AIDS patients had been conducted, and one of the major drawbacks was anaphylaxis [6–8]. Recently, several TCS derivatives were designed with less antigenicity while retaining most of the activity [9–11]. These derivatives are therefore potential antiviral agent. TCS inhibits viral replication including herpes sim- plex virus (HSV-1) [12] and human immunodeficiency virus (HIV-1). Less work is done on its antiviral mecha- nism and it is generally presumed that this action is re- lated to ribosome inactivation. Almost all ribosome inactivating proteins are cytotoxic. Type 2 ribosome inactivating proteins like ricin are usually extremely toxic while type 1 like TCS is generally much milder [13]. Since most ribosome inactivating proteins are anti- viral in nature [14], it is logical to explore the role of cytotoxicity as a possible antiviral mechanism. When a virus infects a cell, it will try to ensure survival of the cell long enough for its own replication [15,16]. If cytotoxic effect of TCS is enhanced on infected cells, premature death of infected cells will limit massive viral replication 0006-291X/$ - see front matter Ó 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2005.03.230 * Corresponding author. Fax: +86 871 5191823. E-mail address: zhengyt@mail.kiz.ac.cn (Y.-T. Zheng). www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 331 (2005) 1075–1080 BBRC