The mA demethylase fat mass and obesity-associated protein mitigates pyroptosis and inflammation in doxorubicin-induced heart failure via the toll-like receptor 4/NF-κB pathway.

BACKGROUND

Doxorubicin (Dox) can induce cardiotoxicity, thereby restricting the utility of this potent drug. Herein, the study ascertained the mechanism of the N6-methyladenosine (mA) demethylase fat mass and obesity-associated protein (FTO) in pyroptosis and inflammation during Dox-induced heart failure (HF).

METHODS

Serum samples were collected from HF patients for detection of the expression of FTO and toll-like receptor 4 (TLR4). Dox-treated H9C2 cardiomyocytes were chosen for HF modeling, followed by measurement of FTO and TLR4 expression. Cardiomyocytes were detected for viability, apoptosis, spatial distribution of NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3), and the levels of lactic dehydrogenase, inflammatory factors, oxidative stress markers, and pyroptosis-related proteins. The mA levels of mRNA were examined. RNA immunoprecipitation (RIP) and mRNA stability measurement were used to determine mRNA and protein expression, and RNA mA dot blot and methylated-RIP assay were performed to detect mA methylation levels. The expression of p-NF-κB p65 and p-IκB-α was measured by western blotting.

RESULTS

In the serum of HF patients, FTO was elevated while TLR4 was decreased. Dox treatment reduced FTO expression and increased mA methylation levels and TLR4 expression in H9C2 cells. Overexpression of FTO and knockdown of TLR4 reduced apoptosis, cytotoxicity, inflammation, pyroptosis, oxidative stress, NLRP3 co-localization, and fluorescence intensity in Dox-induced H9C2 cells. Mechanistically, FTO resulted in reduced binding activity of YTHDF1 to TLR4 mRNA via mA demethylation of TLR4, thus declining TLR4, p-NF-κB p65, and p-IκB-α expression. TLR4 knockdown counteracted the effects of FTO knockdown on Dox-induced H9C2 cells.

CONCLUSIONS

FTO alleviated Dox-induced HF by blocking the TLR4/NF-κB pathway.