Comparison of Aristolochic acid I derived DNA adduct levels in human renal toxicity models

Abstract

Aristolochic acid (AA) dependent human nephropathy results either from environmental exposure to Aristolochiaceae plant subspecies or their use in traditional phytotherapy. The toxic components are structurally related nitrophenanthrene carboxylic acids, i.e. Aristolochic acid I (AAI) and II (AAII). AAI is considered to be the major cause of Aristolochic acid nephropathy, characterized by severe renal fibrosis and upper urothelial cancer. Following enzymatic activation in kidney and/or liver, AAI metabolites react with genomic DNA to form persistent DNA adducts with purines. To determine whether AAI can be activated in human renal cells to form DNA adducts, we exposed telomerase immortalized renal proximal tubular epithelial cells (RPTEC/TERT1), the human embryonic kidney (HEK293) cell line, as well as primary human kidney cells (pHKC) to AAI in vitro. We modified an isotope dilution ultra-performance liquid chromatography/tandem mass spectrometry (ID-UPLC-MS/MS) based method for the quantification of dA-AAI adducts in genomic DNA. In addition, time dependent accumulation of adducts in renal cortex and bladder tissue from AAI/II treated Eker rats were used to validate the detection method. AAI-induced toxicity in human renal cells was determined by dA-AAI adduct quantification, the impact on cell viability, and NQO1 expression and activity. Our findings demonstrated adduct formation in all cell lines, although only pHKC and RPTEC/TERT1 expressed NQO1. The highest adduct formation was detected in pHKC despite low NQO1 expression, while we observed much lower adduct levels in NQO1-negative HEK293 cells. Adduct formation and decreased cell viability correlated only weakly. Therefore, our data suggested that i.) enzymes other than NQO1 could be at least equally important for AA bioactivation in human renal proximal tubule cells, and ii.) the suggested correlation between adduct levels and viability appears to be questionable.

Mitochondrial Iron Overload-Mediated Inhibition of Nrf2-HO-1/GPX4 Assisted ALI-Induced Nephrotoxicity

“…Aristolactam I (ALI) is one of the potential nephrotoxic AAAs in Asarum species, also the major reductive metabolite of aristolochic acid I (AAI), which is considered to be the most important constituent causing aristolochic acid nephropathy (AAN) (Bastek et al, 2019). Interestingly, it had been reported that the nephrocytotoxicity of ALI might be stronger than that of AAI in vivo (Li et al, 2016).…”

Defining in vivo dose‐response curves for kidney DNA adduct formation of aristolochic acid I in rat, mouse and human by an in vitro and physiologically based kinetic modeling approach

“…The LLC‐PK1 cell line is routinely used to study nephrotoxic effects of chemicals in humans because the cells exhibit many of the enzymatic and transport properties of human proximal tubule cells (Gstraunthaler, Pfaller, & Kotanko, 1985; Hull, Cherry, & Weaver, 1976), which are the cells that represent the direct target of AAI (Lebeau et al, 2005). Furthermore, comparison of the cytotoxicity of AAI in the LLC‐PK1 cells with the AAI cytotoxicity in primary renal human cells and even to the cytotoxicity in other cell types from other species described in the literature (Abdullah et al, 2016; Bastek et al, 2019; Huljic, Bruske, Pfitzenmaier, O'Brien, & Dietrich, 2008) reveals that species differences in dynamics of AAI toxicity may be limited. In addition, in our previous study, in vitro toxicity data on AAI in LLC‐PK1 cells provided adequate input for PBK model‐based prediction of in vivo kidney toxicity of AAI (Abdullah et al, 2016).…”

Editorial: Applications of Herbal Medicine to Control Chronic Kidney Disease

“…First, Xu et al have summarized the mechanism underlying nephrotoxicity of compounds from herbal medicine, which may help in discovering the biomarkers of renal injury and developing preventive strategies. Aristolactam I, an active component from herbs, is considered to be the most important constituent causing aritolochic acid nephropathy ( Bastek et al, 2019 ). Deng et al have shown that mitochondrial iron overload-mediated antioxidant system could inhibit the aristolactam-induced ferroptosis in HK-2 cells.…”