Abstract
Isoniazid (INH), an effective first-line drug for tuberculosis treatment, has been reported to be associated with hepatotoxicity for decades, but the underlying mechanisms are poorly understood. N-acetyltransferase 2 (NAT2) is a Phase II enzyme that specifically catalyzes the acetylation of INH, and NAT2 expression/activity play pivotal roles in INH metabolism, drug efficacy, and toxicity. In this study, we systematically investigated the regulatory roles of microRNA (miRNA) in NAT2 expression and INH-induced liver injury via a series of in silico, in vitro, and in vivo analyses. Four mature miRNAs, including hsa-miR-15a-3p, hsa-miR-628-5p, hsa-miR-1262, and hsa-miR-3132, were predicted to target the NAT2 transcript, and a negative correlation was observed between hsa-miR-15a-3p and NAT2 transcripts in liver samples. Further experiments serially revealed that hsa-miR-15a-3p was able to interact with the 3′-untranslated region (UTR) of NAT2 directly, suppressed the endogenous NAT2 expression, and then inhibited INH-induced NAT2 overexpression as well as INH-induced liver injury, both in liver cells and mouse model. In summary, our results identified hsa-miR-15a-3p as a novel epigenetic factor modulating NAT2 expression and as a protective module against INH-induced liver injury, and provided new clues to elucidate the epigenetic regulatory mechanisms concerning drug-induced liver injury (DILI).
Highlights
Drug-induced liver injury (DILI) refers to the liver impairment caused by naturally existing or manufactured hepatotoxic substances, which enter the liver for biotransformation mainly through gastrointestinal tract and blood circulation, and their metabolites or themselves make the liver suffering from varying degrees of damage (Fontana, 2014; Tolosa et al, 2018)
We further calculated the correlations between the miRNA expression and RNA levels of N-acetyltransferase 2 (NAT2) in 419 liver samples obtained from the The Cancer Genome Atlas (TCGA) database, and found a negative correlation (r –0.192, p < 0.001) between hsa-miR-15a-3p and NAT2 (NM_000015)
Consistent with the studies mentioned above, neither alanine aminotransferase (ALT) nor lactate dehydrogenase (LDH) activity was increased by INH treatment, this conclusion is consistent with the articles on isoniazid-induced liver injury; we observed that aspartate aminotransferase (AST) activity was significantly increased in the 90 and 135 mg/kg·d INH group, compared to the control group (Figures 5B–D and Supplementary Figures S1B–S1D)
Summary
Drug-induced liver injury (DILI) refers to the liver impairment caused by naturally existing or manufactured hepatotoxic substances, which enter the liver for biotransformation mainly through gastrointestinal tract and blood circulation, and their metabolites or themselves make the liver suffering from varying degrees of damage (Fontana, 2014; Tolosa et al, 2018). Isoniazid (INH), one main first-line anti-tuberculosis drug, was reported to be associated with DILI since its introduction into the market in 1952. As one of the ancient human diseases, tuberculosis is still one of the leading causes of death among all infectious diseases (Bloom et al, 2017; Natarajan et al, 2020), but clinical treatment is usually accompanied by a high incidence of INH-induced liver injury. The intermediates formed by CYP2E1 are detoxified by GSTs. The general pathogenesis of DILI was considered to be initiated by an imbalance between chemical metabolism activation and biological detoxification process (Gu and Manautou, 2012); the underlying mechanism of INH-induced liver injury remains unclear
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