Investigating The Mechanism Of Noncanonical NF-?B Signaling Pathway Mediating DOX-induced Cardiotoxicity

Doxorubicin(DOX) belongs to anthracyclines antibiotics, which functions as anticancer drug. In clinical therapy, DO X was greatly applied to inhibit the development of solid tumor, leukemia, lymphoma, breast cancer with strong and enhanced efficiency. However, cardiac side effect such as cardiomyopathy and heart failure caused by DOX treatment extremely limited the widely application of DOX in clinic.A large number of studies have illustrated the mechanisms underlying DOX- induced cardiotoxicity, which mainly included oxidative stress, cell apoptosis, calcium homeostasis imbalance and extracellular matrix remodeling etc. A kind of flavonoids called quercetin(QCT) has been found to fight against cancer and scavenge reactive oxidative species, contributing to its clinical application extensively. O ur previous research work demonstrated QCT could ameliorate DOX- induced cardiotoxicity and related molecular mechanisms still needed to be elucidated clearly.We found the expression of MMP2/9 in zebrafish and H9C2 cardiac cell increased significantly when treating with DO X alone, and returned to almost basal level when co-administered with QCT. MMP2/9 are the members of matrix metalloproteins and their aberrant expression could lead to a variety of cardiovascular disease such as myocardial fibrosis, myocardial hypertension, atherosclerosis and myocardial infarction. MMP2/9 were regulated by different transcription factors, including NF-?B, AP-1, STAT and so on, in which NF-?B signaling pathway attracted more attention in cardiovascular research. Numerous investigations have indicated the ca nonical NF-?B pathway mediated by nucleus transcription factor-p65 played a role in regulating MMP2/9, furthermore affecting the function of heart. However the newly identified noncanonical NF-?B signaling pathway mediated by RelB and p100/p52 has never been found to regulate cardiac function. O ur study focused on noncanonical NF-?B pathway and found its role in reducing DOX-induced cardiotoxicity by QCT.Through Realtime PCR, Western blotting, CO-IP, Immunofluorescence, siRN A and other biological technologies, results showed the expression of key protein kinase NIK in noncanonical NF-?B pathway increased, meanwhile its downstream nucleus transcription factors RelB was proved to enter nucleus and the process from p100 to p52 also augmented, all of which above suggested the noncanonical NF-?B signaling could lead to DOX-induced cardiotoxicity. To some extent, QCT could inhibit the alterations of RelB and p52, mitigating the activation of this pathway. In addition, silencing RelB and p100 respectively also influenced the upregulation of MMP2/9 induced by DOX. To step forward, we also explored the membrane protein and associated factors which are likely to activate noncanonical NF-?B pathway. Results indicated the possible role of RANK and TRAF2 played in this pathway through affecting NIK expression, subsequently regulating the transcription and expression of MMP2/9.These results made us convinced that noncanonical NF-?B signaling pathway is able to regulate cardiac function, moreover serving as one of main mechanisms underlying QCT reducing DOX-induced cardiotoxicity.