| [Objective]Nanotechnology is an emerging technology. It has been widely used in all aspects of life science research such as discovery of biomarkers, molecular diagnostics, and drug discovery. With the rapid development of nanotechnology, the safety of nanotechnology has caused great concern. It was reported that nanomaterials can distribute into the circulation by different ways, thus blood coagulation may be changed by the reaction with the blood cells. The potential toxicity of nanomaterials has received much research attention, and its effects on blood system have been also studied in depth.In recent years, polymeric micelles have emerged as a potential carrier in the field of drug delivery, with advantages such as better solubilization, long-circulation time and tumor targetting ability. Chitosan oligosaccharide-stearic acid graft copolymer (CSO-SA) can be used to improve drug delivery of anticancer drugs. It can directly react with blood cells and endothelial cells by intravenous administration, therefore, it is important to assess its blood system toxicity.It has been found that a variety of nanoparticles can induce the generation of reactive oxygen species (ROS) and proinflammatory cytokines in both in vitro and in vivo studies, which indicated that oxidative stress and inflammation are the possible mechanisms for the cardiovascular effects of nanomaterials.In this study, the cytotoxicity and oxidative damage induced by CSO-SA on human monocytic cell line THP-1was examined.[Methods]1. CytotoxicityThe morphological change was observed by optical microscopy and transmission electron microscopy(TEM).Improved MTT experiment(CCK-8) was used to study the cytotoxicity of different concentrations(0,10,20,50,100,200μg/ml)of CSO-SA on THP-1cells for24h, and the cytotoxicity of20μg/ml CSO-SA on THP-1cells for0,6,12,24,36,48h.2. Celluar redox statusDCFH-DA was used to detect ROS generation in THP-1cells after treatment with CSO-SA(0,2,10,20μg/ml) for24h.Griess Reagent was applied to detect NO release in THP-1cells after treatment with CSO-SA(0,2,10,20μg/ml) for24h.The activities of superoxide dismutase(SOD), catalase (CAT) and glutathione peroxidase(GSH-Px) after treatment with CSO-SA(0,2,10,20μg/ml) were measured using commercially available kits following the manufacture’s instructions.3. Proinflammatory cytokinesELISA kit was used for detecting TNF-a and IL-1β release from THP-1cells after treatment with CSO-SA(0,2,10,20μg/ml) for24h.4. Pretreatment with NAC Cells were pretreated with NAC(500μM,1h),and then1)assess ROS generation by THP-1cells after treatment with CSO-SA(0,2μg/ml) for24h,2)detect TNF-a and IL-1β release from THP-1cells after treatment with CSO-SA(0,2μg/ml) for24h. All the results were compared with those without NAC pretreatment.[Results]1. CytotoxicityOptical microscope and TEM study showed that CSO-SA was able to induce morphological changes such as cytosolic vacuole formation.CCK-8test revealed that viability of THP-1cells was reduced by treatment of CSO-SA(0-200μg/ml) in a dose-dependent manner, and20μg/ml CSO-SA (0,6,12,24,36,48h) treatment can also reduce viability of THP-1cells in a time-dependent manner.2. Celluar redox status2μg/ml CSO-SA significantly increased intracellular ROS’level. CSO-SA decreased NO level in the supernatant in a dose-dependent manner. CSO-SA altered the activities of SOD, CAT, GSH-Px, but the tendencies were different.3. Proinflammatory cytokinesThe results showed dose-related increases of TNF-a and IL-1β in the supernatants of THP-1cells treated with various doses of CSO-SA(0,2,10,20μg/ml).4. Pretreatment with NACNAC pretreatment can reduce the generation of ROS in THP-1cells with or without treatment of CSO-SA. It can also reduce the levels of TNF-a and IL-1β in the supernatants of THP-1cells with or without treatment of CSO-SA. [Conclusions]CSO-SA can cause cytoxicity on THP-1cells, including the reduction of cell viability, overgeneration of ROS,TNF-α and IL-1β. Oxidative stress may be the main underlying mechanism for its toxicity. |
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