Abstract
The ureohydrolase, type-II arginase (Arg-II), is a mitochondrial enzyme metabolizing L-arginine into urea and L-ornithine and is highly expressed in renal proximal tubular cells (PTC) and upregulated by renal ischemia. Recent studies reported contradictory results on the role of Arg-II in renal injury. The aim of our study is to investigate the function of Arg-II in renal epithelial cell damage under hypoxic conditions. Human renal epithelial cell line HK2 was cultured under hypoxic conditions for 12–48 h. Moreover, ex vivo experiments with isolated kidneys from wild-type (WT) and genetic Arg-II deficient mice (Arg-II–/–) were conducted under normoxic and hypoxic conditions. The results show that hypoxia upregulates Arg-II expression in HK2 cells, which is inhibited by silencing both hypoxia-inducible factors (HIFs) HIF1α and HIF2α. Treatment of the cells with dimethyloxaloylglycine (DMOG) to stabilize HIFα also enhances Arg-II. Interestingly, hypoxia or DMOG upregulates transforming growth factor β1 (TGFβ1) levels and collagens Iα1, which is prevented by Arg-II silencing, while TGFβ1-induced collagen Iα1 expression is not affected by Arg-II silencing. Inhibition of mitochondrial complex-I by rotenone abolishes hypoxia-induced reactive oxygen species (mtROS) and TGFβ1 elevation in the cells. Ex vivo experiments show elevated Arg-II and TGFβ1 expression and the injury marker NGAL in the WT mouse kidneys under hypoxic conditions, which is prevented in the Arg-II–/– mice. Taking together, the results demonstrate that hypoxia activates renal epithelial HIFs-Arg-II-mtROS-TGFβ1-cascade, participating in hypoxia-associated renal injury and fibrosis.
Highlights
Sufficient supply of oxygen is prerequisite for normal organ function and hypoxia related to many conditions such as high altitude, ischemia, or hypoxemia plays important roles in organ damage and development of diseases including heart disease, renal diseases, neurodegenerative disease, etc. (Lee et al, 2019; Schodel and Ratcliffe, 2019; Burtscher et al, 2021)
The results suggest that hypoxia may enhance arginase type-II (Arg-II) through hypoxia-inducible factors (HIFs)
Using cultured human renal epithelial cell line HK-2 or ex vivo experiments with isolated kidney tissues in culture exposed to hypoxic conditions, we demonstrate that hypoxia causes renal epithelial injury as demonstrated by elevated NGAL levels, the epithelial injury marker, accompanied by upregulation of collagen Ia1, transforming growth factor β1 (TGFβ1), and Arg-II levels
Summary
Sufficient supply of oxygen is prerequisite for normal organ function and hypoxia related to many conditions such as high altitude, ischemia, or hypoxemia plays important roles in organ damage and development of diseases including heart disease, renal diseases, neurodegenerative disease, etc. (Lee et al, 2019; Schodel and Ratcliffe, 2019; Burtscher et al, 2021). While the β-subunit is constitutively expressed, the α-subunits including HIF-1α and HIF-2α are rapidly degraded in the presence of sufficient oxygen (Ivan and Kaelin, 2017). HIF-1α and HIF-2α escape hydroxylation and degradation and are stabilized and translocated into the nucleus to form heterodimeric complex with the constitutive β-subunit and regulate gene expression, which is critical for cellular adaptation to hypoxic conditions (Ivan and Kaelin, 2017). TGFβ1 has been known to be the important player in renal tubulointerstitial fibrosis and can be produced by PTCs in response to insults including hypoxia (Meng et al, 2016; Cho et al, 2019). Whether mitochondrial dysfunction i.e., mtROS under hypoxic conditions plays a causal role in TGFβ1 production in renal PTCs is not known
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