| Introduction Cisplatin (cis-diamminedichloroplatinum, CDDP) is one of the primary anticancer drugs widely used in the clinic. It has apparent clinical effects on testicular cancer, ovarian cancer and so on. However, CDDP, as a kind of cytotoxicity drug, also can produce some side effects on kidney, peripheral nerve and other normal tissues, which hinders its clinical uses. Untill now, the mechanisms of CDDP-induced cytotoxicity have been unknown. It has been reported that oxidative stress and lipid peroxidation is one of the mechanisms of CDDP-induced cytotoxicity. CDDP can cause kidney damages by producing reactive oxygen species (ROS), exhausting cellular glutathione (GSH) and inhibiting the activity of antioxidant enzyme; Also, CDDP can bind to DNA base and form a stable Pt-DNA complex which will disturb the structure of DNA, inhibit DNA replication and thus lead to apoptotic evidence. In this study, we will explore the mechanisms of CDDP-induced cytotoxicity by using biochemistry and molecular biology technique. Emodin (EMD), usually extracted from rhubarb, is an active component. It has biologically extensive effects. It has been reported that EMD can elevate ERCC1 gene expression and thus increase the repair of cisplatin-induced DNA damages in human cells. In addition, Ca2+-mediated DNA repair process may be involved in the repair mechanisms of EMD. In this study, we will study mechanisms of EMD's preventive effects on cisplatin-induced cytotoxicity and provide scientific evidence of making wide use of EMD in the future. Methods In this study, human lung diploid fibroblast cells (WI-38) were used as experimental objects. WI-38 cells were seeded in culture plate or flask and incubated at 37 癈 for at the least 24 h prior to use. Then serial amounts of CDDP were added to the wells and flasks. WI-38 cells were incubated continuously at 37℃ for some time. With the help of optical and electron microscopy, morphology and ultrastructurechanges of WI-38 cells were observed. Cytotoxicity was determined by MTT color assay. ROS, lipid peroxidants and GSH were measured by fluorescence method. DNA damages were determined by electrophoresis and flow cytometry. With the help of the methods described above, HMD as an intervenor was added into cultures together with CDDP to explore its preventive effects on oxidative damages and DNA damages induced by cisplatin.Results After WI-38 cells were incubated with cisplatin for 15 h, morphologicalchanges were apparently observed and the changes were alleviated by HMD. AfterWI-38 cells were treated by different concentrations of CDDP for 22 h, CDDPproduced the strongest cytotoxicity and IC5o was 16±3 mg L-1; The cytotoxicityinduced by CDDP was significantly decreased by HMD and IC50 was 34 ± 6 mg L-1. Theproduction of reactive oxygen radicals was significantly increased after WI-38 cellswere treated by CDDP for a period of time and the increase was the most significantafter CDDP treatment for 3 h. At the same time, GSH level was significantly decreased.Lipid peroxidants were also significantly increased after WI-38 cells were treated byCDDP for 22 h. The production of lipid peroxidants and ROS was further increased byHMD and GSH level was decreased. With the help of electron microscopy, we foundthat cellular ultrastructure has been changed. After WI-38 cells were treated by CDDPfor 22 h, larger cytoplasm protuberances turned up on some part of the cell surface,many vacuoles appeared in the cytoplasm and chromatin was peripherally aggregated.The change could be remarkably inhibited by HMD. DNA electrophoresis showed thatthe ladder bands, a typical feature of apoptotic cell, appeared in the groups treated byCDDP for 22 h, and with the increase of dosage the phenomena was more apparent;the DNA damage could be alleviated by HMD. Flow cytometry showed that theapoptotic rate was significantly increased, at the same time, the cell numbers in Sphase were increased and the cell numbers in G0±G1 phase were...
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