Tryptophan Breakdown Product 3-Hydroxyanthranillic Acid Promotes AKI Recovery under Hypoxic Conditions

Background: We and others have demonstrated that hypoxic preconditioning is an effective strategy to protect against renal ischemia reperfusion injury (IRI). While more clinically relevant, however, the effect of hypoxia exposure after IRI remains undefined. Here, by profiling the effects of systemic hypoxia exposure on kidney repair after IRI, we identified the tryptophan metabolite 3-hydroxyanthranillic acid (3HAA) as a metabolite promoting adaptive repair. Beneficial effects of 3HAA reported include oxidative stress reduction via NRF2 activation, anti-ferroptosis, ROS-scavenging and enhanced longevity. Methods: Male C57/BL6 mice were subjected to unilateral IRI and 6 hours later were placed in hypoxic chamber at 8% O2 or normoxia for 72 hours post-IRI. Kidneys and serum collected for RNA-seq and HPLC metabolite profiling. HK2 cells were maintained under defined growth medium, treated with 3HAA (100μM or 200μM) and/or IL1β (10ng/mL) for 24 hours. Results: 72 hours post-IRI, Hx mice had significantly lower injury scores histologically (2.2±0.4 Hx vs 3.3±0.5 Nx, P=0.02) and significantly lower KIM1 mRNA (0.8±0.2 Hx vs 2±0.5 Nx, P=0.0002). RNA-seq analysis followed by KEGG pathway analysis showed significant upregulation of tryptophan metabolism and glutathione metabolism, while inflammatory response was the top downregulated pathway. TRRUST implicated NRF2 as a key TF (P=3e-8), regulating many highly upregulated genes such as glutathione S-transferase alpha (GSTA) genes. Metabolite profiles indicated 3HAA as a key metabolite increased by hypoxia in serum (8.6-fold, P=0.02) and kidney (2.2-fold, P=0.005) compared to normoxia. In HK2 cells, 24-hour treatment with 3HAA 100μM increased GSTA1 expression by 3.1-fold (P=6e-4 vs untreated). Under IL1β-stimulation, 3HAA 100μM or 200μM reduced CCL2 expression by 4.3-fold (P=0.03) and 17.6-fold (P=0.01), respectively, compared to IL1β-stimulation alone. Conclusion: Systemic hypoxia after IRI promotes adaptive kidney repair and reduces inflammation, a response that is associated with increased 3HAA and NRF2 activation. In vitro work indicated 3HAA alone counteracts inflammation in tubular epithelial cells and promotes anti-oxidative defense. Our findings identify a hypoxia/3HAA axis that can be exploited as therapeutic strategy in AKI, warranting further mechanistic investigation. Funding: NIDDK Support