Nrf2/NRF1 signaling activation and crosstalk amplify mitochondrial biogenesis in the treatment of triptolide-induced cardiotoxicity using calycosin.

Abstract

Nuclear factor erythroid 2-related factor 2 (Nrf2) regulates both oxidative stress and mitochondrial biogenesis. Our previous study reported the cardioprotection of calycosin against triptolide toxicity through promoting mitochondrial biogenesis by activating nuclear respiratory factor 1 (NRF1), a coregulatory effect contributed by Nrf2 was not fully elucidated. This work aimed at investigating the involvement of Nrf2 in mitochondrial protection and elucidating Nrf2/NRF1 signaling crosstalk on amplifying the detoxification of calycosin. Results indicated that calycosin inhibited cardiomyocytes apoptosis and F-actin depolymerization following triptolide exposure. Cardiac contraction was improved by calycosin through increasing both fractional shortening (FS%) and ejection fraction (EF%). This enhanced contractile capacity of heart was benefited from mitochondrial protection reflected by ultrastructure improvement, augment in mitochondrial mass and ATP production. NRF1 overexpression in cardiomyocytes increased mitochondrial mass and DNA copy number, whereas NRF1 knockdown mitigated calycosin-mediated enhancement in mitochondrial mass. For nuclear Nrf2, it was upregulated by calycosin in a way of disrupting Nrf2-Keap1 (Kelch-like ECH associated protein 1) interaction, followed by inhibiting ubiquitination and degradation. The involvement of Nrf2 in mitochondrial protection was validated by the results that both Nrf2 knockdown and Nrf2 inhibitor blocked the calycosin effects on mitochondrial biogenesis and respiration. In the case of calycosin treatment, its effect on NRF1 and Nrf2 upregulations were respectively blocked by PGCα/Nrf2 and NRF1 knockdown, indicative of the mutual regulation between Nrf2 and NRF1. Accordingly, calycosin activated Nrf2/NRF1 and the signaling crosstalk, leading to mitochondrial biogenesis amplification, which would become a novel mechanism of calycosin against triptolide-induced cardiotoxicity.