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Doxorubicin (DOX) is one of the most effective chemotherapeutic drugs; however, the incidence of cardiotoxicity compromises its therapeutic index. Oxidative stress and apoptosis are believed to be involved in DOX-induced cardiotoxicity. Chitosan oligosaccharides (COS), the enzymatic hydrolysates of chitosan, have been reported to possess diverse biological activities including antioxidant and anti-apoptotic properties. The objective of the present study was to investigate the potential role of COS against DOX-induced cardiotoxicity, and the effects of COS on apoptosis and oxidative stress in rats and H9C2 cells. Furthermore, we also shed light on the involved pathways during the whole process. For this purpose, first, we demonstrated that COS exhibited a significant protective effect on cardiac tissue by not only inducing a decrease in body and heart growth but also ameliorated oxidative damage and ECG alterations in DOX-treated rats. Second, we found that COS reversed the decrease of cell viability induced by DOX, reduced the intracellular reactive oxygen species (ROS), increased the mitochondrial membrane potential (MMP) and Bcl-2/Bax ratio. COS treatment also results in reduced caspase-3 and caspase-9 expressions, and an increase in the phosphorylation of MAPKs (mitogen-activated protein kinases) in DOX-exposed H9C2 cells. Additionally, cellular homeostasis was re-established via stabilization of MAPK mediated nuclear factor erythroid 2-related factor 2/antioxidant-response element (Nrf2/ARE) signaling and transcription of downstream cytoprotective genes. In summary, these findings suggest that COS could be a potential candidate for the prevention and treatment of DOX-induced cardiotoxicity.

Several lines of evidence indicate that reactive oxygen species (ROS) overproduction under the metabolic syndrome condition is the leading cause of cardiovascular events. Calcium is an important stimulus for vasoconstriction and plays a pivotal role in the development of hypertension. Here, we investigate whether a relationship exists between metabolic syndrome-induced mitochondrial ROS overproduction and Ang II-mediated Ca2+ release in vascular smooth muscle cells (VSMC). The effect of mitochondrial ROS on AT1 expression, and Ca2+ and IP3 generation was studied in 2 VSMC models of metabolic syndrome using fura-2/AM probes and ELISA-based assay. Ang II-mediated aortic ring contraction in SD rats fed with high-fat diet (HFD) was measured using a force transducer connected to chart recorder. In the metabolic syndrome, almost 2-fold increased mitochondrial O2 - significantly upregulated AT1 expressions by ~60%, companied by elevated Ca2+ and IP3 levels in VSMC and enhanced aortic rings contraction. All these increments were blocked by rotenone (inhibitor of mitochondrial respiratory chain complex I), ruthenium red (inhibitor of calcium uniporter), cyclosporin A (inhibitor of mitochondrial permeability pore), and N-acetylcysteine. Therefore, in the states of metabolic syndrome, ROS overproduction in mitochondrial complex I enhances Ang II-mediated vascular contraction via an AT1-dependent pathway. In addition, the import of Ca2+ from endoplasmic reticulum to mitochondria via calcium uniporter and mitochondrial permeability pore seems to serve as a mechanism to further aggravate mitochondrial damage and vascular dysfunction that may contribute to the occurrence of hypertension.

Extracellular cysteine (Cys)/cystine (CySS) redox potential (Eh) plays a crucial role in maintaining redox homeostasis and an alteration of redox state occurs in various physiological conditions, including diabetes, cancer, and aging. This study was designed to determine whether a variation in extracellular redox state would alter the function of insulin-resistant PC12 cells. Various redox states were established by providing different extracellular Cys/CySS Eh to insulin-resistant PC12 cells. We intensively investigated the relationship between redox state and catecholamine biosynthesis in PC12 cells, and evaluated the changes in cellular reactive oxygen species (ROS), catecholamine (CA) synthesis, tyrosine hydroxylase (TH) expressions, and the activity of rate-limiting enzyme in CA synthesis by using DCF-fluorescence, HPLC, and the real-time PCR, respectively. We also determined the protein levels of NF-E2-related factor 2 (Nrf2), a redox sensitive transcription factor, using an ELISA assay. We found that the oxidized Cys/CySS Eh (0 mV) pretreatment decreased CA, TH, and Nrf2 levels, but induced ROS overproduction. Insulin induced a significant increase in CA synthesis and ROS production, blocked by more reducing redox conditions. The paradox of CA and TH alterations between insulin and 0 mV groups may be attributed to degree of redox imbalance as evidenced by different ROS levels in 2 groups, which is further confirmed by CA alterations in different concentrations of hydrogen peroxide. Additionally, dithiole-3-thione (D3T, an inducer of Nrf2) corrected 0 mV-induced TH inhibition. In conclusion, CA biosynthesis in insulin-resistant PC12 cells could be influenced by extracellular Cys/CySS redox effects on cellular redox sensitive transcription factors.