#3589 Renal AhR controls blood pressure regulation and cysteine metabolism in an <i>in vivo</i> model of Obstructive Sleep Apnea

Abstract Background and Aims Arterial hypertension (HTN) associated with Obstructive Sleep Apnea (OSA), often drug-resistant, poses a challenge in treatment and needs innovative therapeutic approaches. Chronic intermittent hypoxia (CIH) is the main clinical feature of OSA and is associated with several disease mechanisms, including inflammation and oxidative stress. Our group was pioneer in linking the CIH-induced drug-resistant HTN with the overactivation of the aryl hydrocarbon receptor (AHR) in the kidney, which was reverted by the treatment with a selective AhR antagonist (CH-223191). Moreover, we identified a new AhR activator (cystine, an oxidized form of cysteine) and a new AhR antagonist (Hydroxytyrosol – OHTyr – a NRF2 agonist) in vitro and in vivo, respectively. As a ligand-activated transcription factor from the Per-Arnt-Sim (PAS) superfamily, the AhR shares its dimerization partner with the hypoxia-inducible factors (HIFs). As the role of the AhR in CIH-induced changes is not well characterized, in this work, we aimed at identifying putative genes under the AhR control associated with HTN development, as well as understanding the impact of both CIH, when HTN is already established, and AhR blockade in the AhR-HIF signaling and cysteine metabolism, to better understand the effect of CIH exposure in the kidney, and the role of the AhR in these changes. Method Briefly, Male Wistar rats (n ≥ 4) were subjected to CIH (21%–5% O2, 5.6 cycles/h, 10.5 h/day, during the animals’ inactive period) for 21 days, to establish HTN, and from the 22nd day they were treated orally with the AhR antagonist CH-223191 (5 mg/kg/day in vegetable oil, gavage) or hydroxytyrosol (OHTyr, 15 mg/kg/day in vegetable oil, gavage) for 14 days under CIH. Animals under 35 days of normoxia (Nx) or CIH, treated or not with the drug's vehicle, were used as controls. Blood pressure was monitored during active and inactive periods using radiotelemetry. The renal cortex was collected and used for Western Blot quantification of AHR-related signaling (e.g., CYP1A1, PON1, HIFs) and cysteine/H2S metabolic enzymes (e.g., CBS, PDO). The cysteine-related thiols (e.g., GSH, Cys) were quantified by HPLC-FD. Transcriptomics analyses of kidney cortex by RNA-Seq. was performed in CH-223191 and OHTyr-treated animals. The study has the Ethical approval by DGAV and NMS ethic committee. Results The blockade of the AhR by both CH-223191 and OHTyr was able to revert the CIH-induced increase in blood pressure, coincident with a normalization of renal protein levels of CYP1A1 (hallmark of AhR activation). CIH exposure altered the expression of AHR-related signaling proteins and cysteine metabolic enzymes, which were at least partially reverted by the AhR blockade. OHTyr treatment was also able to revert the CIH-induced increase in oxidized cysteine levels. In turn, we found that CH-223191 selectively altered 10 differentially expressed genes (DEGs) in CIH which are primarily involved in metabolic stress, mitochondrial dysfunction, lipid accumulation, inflammation, and heme metabolism (Alad). Conclusion Altogether, our findings show that, in the kidney, CIH affect the AhR-HIF axis and cysteine metabolism, which are normalized upon AhR blockade. Moreover, we identify AHR pharmacological pathways, putatively relevant to OSA-HTN.