FoxO3 activation in hypoxic tubules prevents chronic kidney disease
Ling Li, … , Qais Al-Awqati, Fangming Lin
Ling Li, … , Qais Al-Awqati, Fangming Lin
Published March 26, 2019
Citation Information: J Clin Invest. 2019;129(6):2374-2389. https://doi.org/10.1172/JCI122256.
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Research Article Cell biology Nephrology

FoxO3 activation in hypoxic tubules prevents chronic kidney disease

Abstract

Acute kidney injury (AKI) can lead to chronic kidney disease (CKD) if injury is severe and/or repair is incomplete. However, the pathogenesis of CKD following renal ischemic injury is not fully understood. Capillary rarefaction and tubular hypoxia are common findings during the AKI-to-CKD transition. We investigated the tubular stress response to hypoxia and demonstrated that a stress-responsive transcription factor, FoxO3, was regulated by prolyl hydroxylase (PHD). Hypoxia inhibited FoxO3 prolyl hydroxylation and FoxO3 degradation, leading to FoxO3 accumulation and activation in tubular cells. Hypoxia-activated HIF-1α contributed to FoxO3 activation and functioned to protect kidneys, as tubular deletion of HIF-1α decreased hypoxia-induced FoxO3 activation and resulted in more severe tubular injury and interstitial fibrosis following ischemic injury. Strikingly, tubular deletion of FoxO3 during the AKI-to-CKD transition aggravated renal structural and functional damage, leading to a much more profound CKD phenotype. We show that tubular deletion of FoxO3 resulted in decreased autophagic response and increased oxidative injury, which may explain renal protection by FoxO3. Our study indicates that in the hypoxic kidney, stress-responsive transcription factors can be activated for adaptions to counteract hypoxic insults, thus attenuating CKD development.

Authors

Ling Li, Huimin Kang, Qing Zhang, Vivette D. D’Agati, Qais Al-Awqati, Fangming Lin

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Figure 3

HIF-1α contributes to FoxO3 activation and CKD protection.

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HIF-1α contributes to FoxO3 activation and CKD protection. (A) HIF-1α pr...
(A) HIF-1α protein levels increased during CKD development. n = 4. (B–C) HIF-1α was deleted using the Pax8-rtTA and Tet-O-Cre system by giving mice doxycycline for 2 weeks prior to primary culture (Hif-1atub). Vehicle-treated mice (Hif-1actl) were used as controls. (B) The hypoxia-induced increase in FoxO3 protein was partially dependent on HIF-1α in primary cultures. n = 5 with duplicates. ††P < 0.01 and *P ˂ 0.05 comparing Hif-1atub with Hif-1actl at 0 hours and 1 hour, respectively. (C) HIF-1α deletion abolished the increase in FoxO3 mRNA in response to hypoxia with 1% O2. n = 3–8 with duplicates. **P < 0.01 and ***P < 0.001 compared with cells prior to exposure to 1% O2 (0 min) in the Hif-1actl or Hif-1atub group. (D) Nuclear expression of FoxO3 (red) was higher in proximal tubules (labeled with LTA in green) of Hif-1actl kidneys 4 weeks after IRI. n = 4. **P < 0.01. (E–I) Tubular HIF-1α was deleted from day 8 to day 21 following a 35-minute left IRI and right nephrectomy, and kidneys were analyzed 4 weeks after IRI. Tubular deletion of HIF-1α led to more severe tubular atrophy (arrowheads) and cast formation (arrows) (E, PAS staining, n = 5), higher urinary albumin and creatinine (Alb/Cr) levels (F, n = 3 or 5 for normal controls; n = 5 or 6 for 4 weeks after IRI), higher urinary NGAL excretion (G, n = 3 or 5 for normal controls; n = 5 or 6 for 4 weeks after IRI), and more interstitial inflammation and fibrosis (H). For the images in H, the top panel shows more infiltrating CD45-expressing leukocytes (red, n = 5), the middle panel shows more fibrosis with collagen I (Col I) deposition (green, n = 5), and the bottom panel shows no significant differences in the density of capillaries labeled with endomucin (green, n = 4) compared with Hif-1actl kidneys. (I) Blots show lower levels of FoxO3 protein in Hif-1atub kidneys compared with levels in Hif-1actl kidneys, but no significant difference in the LC3II/LC3I ratio was detected. n = 6. *P ˂ 0.05 and **P ˂ 0.01, comparing Hif-1atub with Hif-1actl (E, F, H, and I). Scale bars: 100 μm (E), 50 μm (top and bottom panels of H), and 20 μm (D and middle panel of H). Statistical significance was determined by 2-tailed Student’s t test (B, D, F, H, and I), Wilcoxon-Mann-Whitney U test (E), and 1-way ANOVA followed by Dunnett’s post hoc test for multiple comparisons (C).