Sacubitril/valsartan protects against arsenic trioxide induced cardiotoxicity in vivo and in vitro

Abstract

The cardiotoxicity induced by arsenic trioxide (ATO) limits its clinical application in acute promyelocytic leukemia treatment. Sacubitril/valsartan (LCZ696) is an effective drug for the treatment of heart failure. In this study, we aimed to investigate the protective effect and mechanisms of LCZ696 against the ATO-induced cardiotoxicity in mice and H9c2 cells. We found that LCZ696 could alleviate the decrease of ejection fraction and fractional shortening induced by ATO, thereby improving mouse cardiac contractile function. LCZ696 could also reduce the myocardial enzyme, resist oxidative stress, mitigate myocardial fibrosis, and ameliorate myocardial structure, thereby alleviating myocardial damage caused by ATO. In addition, LCZ696 could significantly increase the cell viability and reduce the accumulation of reactive oxygen species in ATO-treated H9c2 cells. Besides, in vivo and in vitro studies have been found that LCZ696 could restore the expression of Bcl-2 and reduce Bax and Caspase-3 levels, inhibiting ATO-induced apoptosis. Meanwhile, LCZ696 decreased the levels of IL-1, IL-6, and TNF-α, alleviating the inflammatory injury caused by ATO. Furthermore, LCZ696 prevented NF-κB upregulation induced by ATO. Our findings revealed that LCZ696 has a considerable effect on preventing cardiotoxicity induced by ATO, which attributes to its capability to suppress oxidative stress, inflammation, and apoptosis.

Keywords: LCZ696; apoptosis; arsenic trioxide; cardiotoxicity; inflammation; oxidative stress.

Fig. 1

LCZ696 ameliorated left ventricular functions after ATO treatment. (A) Representative echocardiogram images of each group; (B) echocardiography indices obtained from the image, expressed as the mean ± SEM, n = 10 (^*P < 0.05 vs. NC group; ^#P < 0.05 vs. ATO group).

Fig. 2

Effect of LCZ696 on the heart function indicators and oxidative homeostasis. (A) CK, LDH, and AST activities in plasma were determined. (B) SOD activity, MDA, and GSH content in cardiac tissue were determined. Values were presented as the mean ± SEM, n = 5 (^*P < 0.05, ^**P < 0.01 vs. the NC group; ^#P < 0.05 vs. the ATO group).

Fig. 3

LCZ696 alleviated ATO-induced myocardial injury and fibrosis in mice hearts. (A) Representative H&E staining micrographs of heart tissues. (B) Masson staining of 4 groups. Collagen area (%) of heart tissues were presented as fibrosis, n = 5. The values were presented as the mean ± SEM (The scale bar represents 50 μm. ^**P < 0.01, vs. the NC group; #P < 0.05 vs. the ATO group).

Fig. 4

Effects of LCZ696 on apoptosis and inflammation in mice heart tissue. (A) Caspase-3, Bcl-2, and Bax levels in mice heart tissue were detected by Western blot, n = 3. (B) IL-1, IL-6, TNF-α, and NF-κB mRNA levels in mice heart tissue were determined by quantitative real-time PCR, n = 3.The relative expression was analyzed by normalization to β-actin. Results are reported as fold changes normalized to the NC group values. Data were presented as the mean ± SEM (^*P < 0.05, ^**P < 0.01, ^***P < 0.001 vs. the NC group; ^#P < 0.05, ^##P < 0.01 vs. the ATO group).

Fig. 5

LCZ696 restored the decline in cell viability induced by ATO. (A) ATO (0–10 μM) decreased H9c2 cell viability. (B) LCZ696 (0–40 μM) alleviated the reduced viability of H9c2 cells induced by 5 μM ATO. (C) The effect of 20 μM LCZ696 and 5 μM ATO on the viability of H9c2 cells. Relative cell viability was determined by the CCK-8 assay. Cells were treated with LCZ696 and ATO for 24 h. Data were expressed as mean ± SEM, n = 5 (^*P < 0.05, ^***P < 0.001 vs. the NC group; ^#P < 0.05 vs. the ATO group).

Fig. 6

Effects of LCZ696 on oxidative stress, apoptosis and inflammation in H9c2 cells. (A) Fluorescence images and the ratio graph of ROS in different treatment groups. The scale bar represents 50 μm, n = 5. (B) Caspase-3, Bcl-2, and Bax levels in H9c2 cells were detected by Western blot, n = 3. (C) IL-1, IL-6, TNF-α, and NF-κB mRNA levels in H9c2 cells were determined by quantitative real-time PCR, n = 3. The relative expression was analyzed by normalization to β-actin. Results are reported as fold changes normalized to the NC group values. Data were presented as the mean ± SEM (^**P < 0.01, ^***P < 0.001 vs. the NC group; ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. the ATO group).