Lidocaine The mitogen activated protein kinase MAPK plays an

The mitogen-activated protein kinase (MAPK) plays an important role in a variety of cell processes by controlling transcriptional or translational regulation [14]. There are three major MAPK family members that have been well characterized: extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 MAPK [15]. MAPK pathways are protein kinase cascades in which signals are relayed through Lidocaine of downstream kinase by activated upstream kinases, leading to the appropriate cellular responses. Apoptosis signal-regulating kinase 1(ASK1) is a member of MAPK kinase kinase (MAPKKK) family and activates both p38 and JNK pathways, resulting in apoptosis [16]. In the last two decades, ASK1 was shown to be involved in various responses, including cell apoptosis, differentiation, and production of inflammatory cytokines. ASK1 is activated in response to various stresses, such as reactive oxygen species (ROS), endoplasmic reticulum stress, lipopolysaccharide, and calcium overload [17], [18]. In a published paper, exposure of podocytes to adriamycin (ADR; doxorubicin) resulted in cell injury, which was preceded by increased ROS generation and ASK1 activation [19]. However, whether ASK1 activation is involved in DOX-induced cardiomyocyte apoptosis is still not known, and the downstream events need a further investigation.
Materials and methods
Result
Discussion DOX is a widely used anticancer drug. However, its clinical use is limited by its toxicity to multiple organs, including the heart. In fact, the toxic mechanism of DOX and its prevention have been the focus of many previous investigations. Accumulated evidence indicated that DOX induces ROS and causes oxidative cell injury [5], [24], [27]. Consistent with these reports, we confirmed that the toxic effect of DOX on cardiomyocytes was closely associated with ROS, as evidenced by the elevated fluorescence intensity level in our experiment (Fig. 4C). Prevention of oxidative stress with antioxidant (Mitotempo) or blockade of oxidative stress induced ASK1/JNK signaling pathways significantly attenuated the cytotoxicity of DOX. IL-33 is reported to have antioxidative actions. For example, in a well-designed study, Lee RT et al. had reported that both angiotensin II and phenylephrine significantly induced ROS generation in cardiomyocytes, which was inhibited by IL-33 [10]. In another study, Rui T et al. also found that pretreatment of myocytes with IL-33 decreased the myocyte oxidant production after anoxia/reoxygenation injury [7]. Here, we tested whether IL-33 can directly protect cardiomyocytes from DOX-induced oxidative stress. The Fig. 4D demonstrated that treatment of cardiomyocytes with IL-33 decreased oxidant production after DOX. The results suggested that IL-33 prevents DOX-induced ASK1/JNK activation through attenuating DOX-induced myocyte oxidative stress. However, how the IL-33 plays its role to attenuate the myocyte oxidative stress is not clear. Further studies are warranted. ROS causes cell injury through directly oxidizing and damaging DNA, proteins, and lipids as well as by activating several stress-sensitive signaling pathways [28], [29]. Among the many kinases activated by ROS, ERK, p38, and JNK have been shown to play a key role in oxidative cell apoptosis [30], [31]. In our study, we also found that DOX can elevate phosphorylation of ERK, p38 and JNK. Interesting, IL-33 only inhibited JNK activation and had a minor effect on p38 and ERK activation. Lee RT et al. also found that co-treatment with IL-33 did not affected ERK phosphorylation, but decreased p38 and JNK phosphorylation induced by angiotensin II [10]. Their research partially confirms our results. Previous studies in cardiomyocytes have demonstrated that suppression of JNK attenuates DOX-induced cardiomyocytes injury [24], [32]. Consistent with these previous reports, we confirmed the mediating role of JNK. Genetic inhibition of JNK significantly attenuated the DOX-induced apoptosis. Interestingly, the protective action of IL-33 was also associated with marked suppression of JNK phosphorylation, suggesting that inhibition of JNK signaling pathway could be involved in the protective effect of IL-33.