4. Discussion
Dilated cardiomyopathy is one of the most serious side effects of doxorubicin treatment, significantly reducing its anticancer value and causing a large societal burden. However, the mechanisms of dilated cardiomyopathy are still unclear, and an effective therapy to prevent the progression of existing cardiac inflammation and fibrosis induced by DOX is urgently needed. Our study aimed to determine the potential effect of LCZ696 and the role of TLR2 in DOX-induced cardiac dysfunction, fibrosis and inflammation, which may help find an effective drug to protect the heart and provide a potential signaling pathway for the treatment of DOX-induced dilated cardiomyopathy.
In our studies, we showed that LCZ696 treatment and TLR2 deficiency attenuated DOX-related dilated heart failure by improving the decreased EF% and increased LVIDd. In addition, in vivo and in vitro , our results showed that DOX stimulated the expression of matrix proteins and inflammatory cytokines in the heart and that LCZ696 treatment and TLR2 knockdown ameliorated these alterations and ultimately improved cardiac function. Furthermore, the underlying mechanisms involved in DOX-induced cardiomyopathy revealed that DOX stimulated the formation of the TLR2-MyD88 complex, which activated the NF-κB pathway, leading to cardiac cell inflammation and fibrosis. This TLR2-MyD88 interaction was TLR4 or MD2 independent and could be disturbed by LZC696, which explains the strong effect of this drug in preventing heart injury caused by DOX.
It is well established that LCZ696, as a novel angiotensin receptor-neprilysin inhibitor, significantly reduced mortality and hospitalization due to heart failure in HF patients with a reduced ejection fraction (HFrEF)(Mann et al., 2020). The results of the PARADIGM-HF trial(McMurray et al., 2014) suggest a second function of sacubitril/valsartan: degrading peptides that regulate the cardiovascular, nervous, inflammatory, and immune systems(D’Elia, Iacovoni, Vaduganathan, Lorini, Perlini & Senni, 2017; Turner, Isaac & Coates, 2001). Recent studies found that LCZ696 increased local BNP/CNP levels, interfered with angiotensin II-mediated signaling, and then reduced the magnitude of cardiac remodeling in animal models of hypertension and myocardial infarction(Oatmen, Zile, Burnett & Spinale, 2018; von Lueder et al., 2015). Regarding the cardiotoxicity induced by DOX, accumulating evidence has discovered the pathophysiological mechanisms, but treatments to mitigate cardiac damage are still limited(Hu et al., 2020; Yang et al., 2020). Xia Y et al found that Drp1 and its Ser-616 phosphorylation were significantly increased in dilated cardiomyopathy (DCM) patients and demonstrated that the use of LCZ696 against DOX-induced cardiac dysfunction is associated with alleviated Drp1-mediated mitochondrial dysfunction(Xia et al., 2017). Similar to our results, we focused on the anti-inflammatory effect of LCZ696. Our data indicated that LCZ696 prevented IκBα degradation, inhibited the nuclear translation of NF-κB and reduced the expression of inflammatory cytokines in vivo and in vitro . More impressively, we found that pretreatment with LCZ696 inhibited the increased formation of the TLR2/MyD88 complex induced by DOX. This finding partly explains the mechanism of the anti-inflammatory effect of LZC696.
Similar to other TLRs, TLR2 is the most characteristic member of pattern recognition receptors (PRRs), which play an important role in innate immune mechanisms(Jacquet & Robinson, 2020). TLRs play different roles in different stages of infection of atherosclerosis-related pathogens such as Chlamydia pneumoniae(Li, Xia & Hu, 2020). Since Nozaki N et al found that TLR2 may play a role in the regulation of inflammatory and apoptotic mediators in the heart after DOX administration(Nozaki, Shishido, Takeishi & Kubota, 2004) in 2004, little research has explored the mechanism by which TLR2 mediates DOX-induced cardiotoxicity. Our current studies indicated that DOX induced the increased formation of the TLR2/MyD88 complex, which leads to the activation of the NF-κB pathway and stimulates the expression of cardiac inflammation and fibrosis. This result partially compensates for the inflammatory mechanism of DOX-induced myocardial injury. Moreover, we found that this interaction of TLR2 and MyD88 induced by DOX is independent of TLR4 or MD2, but the clear role of TLR4 and MD2 in DOX-induced cardiac side effects still needs further research.
Taken together, our results demonstrate that the mechanism by which LCZ696 relieves DOX-induced cardiac inflammation fibrosis and heart failure lies in reducing the formation of TLR2-MyD88 complexes. LCZ696 may be a potential drug to treat DOX-related heart failure, and TLR2-MyD88 could be a parallel target in the prevention and treatment of DOX-related heart injury.