Fluoride exposure impairs lysosomal biogenesis and autophagic flux in myocardial injury: Involvement of the FNIP1/mTOR/TFEB pathway

As the mechanism of myocardial injury caused by fluoride is still unclear, the study is to investigate whether the effects of fluorosis on heart involve changes in lysosomal biogenesis and autophagic flux. Electrocardiogram (ECG) was detected in population in endemic fluorosis area and formation of myocardial fibrosis by Masson staining in rats. Aspartate aminotransferase (AST) activity and apoptosis rate were detected by biochemical methods; protein or mRNA levels of folliculin-interacting protein (FNIP1), mammalian rapamycin target protein (mTOR), transcription factor EB (TFEB), lysosome-associated membrane protein 2 (LAMP2), cathepsin D (CTSD), microtubule-associated protein 1 light chain 3 (LC3) and P62 by Western blotting or real-time PCR or immunofluorescence analysis; pH value of acidic lysosomes by fluorescent probe; autophagy flux by transfecting adenovirus vector; autophagosomes in cells by transmission electron microscopy; FNIP1 overexpression transfected with the related lentivirus into H9c2 cells. The abnormal ECG, raised serum fluoride level and AST activity in patients with dental fluorosis were observed. In rat myocardial tissues or H9c2 cells exposed to fluoride, the serum AST activity and collagen fibers between myocardial cells were increased; the expressions of FNIP1, TFEB nuclear translocation, LAMP2 and CTSD obviously decreased, and p-mTOR, P62 and ratio of LC3II/I raised; the degradation of autophagic lysosomes declined, pH values and apoptosis increased. Significantly, overexpression of FNIP1 via inhibition of mTOR/TFEB pathway in vitro reversed the disorder of lysosomal biogenesis and the block of autophagic flux induced by fluoride. Fluoride exposure can induce myocardial damage and alterations in lysosomes biosynthesis and autophagic flux in myocardial cells, in which FNIP1 might be considered as one of the key regulatory factors involved in the mechanism. • Abnormal change of ECG in patients with dental fluorosis was observed. • Collagen fibers between myocardial cells of rats exposed to fluoride were increased. • Decreased FNIP1, TFEB nuclear translocation, LAMP2 and CTSD were induced by fluorosis. • Changed p-mTOR, ratio of LC3II/I and P62, and autophagic lysosomes pH were found. • Overexpression of FNIP1 in vitro reversed the myocardial changes induced by fluorosis.