AGER1 downregulation associates with fibrosis in nonalcoholic steatohepatitis and type 2 diabetes
Ali Dehnad, … , Mohammed Ali, Natalie J. Török
Ali Dehnad, … , Mohammed Ali, Natalie J. Török
Published July 13, 2020
Citation Information: J Clin Invest. 2020;130(8):4320-4330. https://doi.org/10.1172/JCI133051.
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AGER1 downregulation associates with fibrosis in nonalcoholic steatohepatitis and type 2 diabetes

Abstract

Type 2 diabetes is clinically associated with progressive necroinflammation and fibrosis in nonalcoholic steatohepatitis (NASH). Advanced glycation end-products (AGEs) accumulate during prolonged hyperglycemia, but the mechanistic pathways that lead to accelerated liver fibrosis have not been well defined. In this study, we show that the AGEs clearance receptor AGER1 was downregulated in patients with NASH and diabetes and in our NASH models, whereas the proinflammatory receptor RAGE was induced. These findings were associated with necroinflammatory, fibrogenic, and pro-oxidant activity via the NADPH oxidase 4. Inhibition of AGEs or RAGE deletion in hepatocytes in vivo reversed these effects. We demonstrate that dysregulation of NRF2 by neddylation of cullin 3 was linked to AGER1 downregulation and that induction of NRF2 using an adeno-associated virus–mediated approach in hepatocytes in vivo reversed AGER1 downregulation, lowered the level of AGEs, and improved proinflammatory and fibrogenic responses in mice on a high AGEs diet. In patients with NASH and diabetes or insulin resistance, low AGER1 levels were associated with hepatocyte ballooning degeneration and ductular reaction. Collectively, prolonged exposure to AGEs in the liver promotes an AGER1/RAGE imbalance and consequent redox, inflammatory, and fibrogenic activity in NASH.

Authors

Ali Dehnad, Weiguo Fan, Joy X. Jiang, Sarah R. Fish, Yuan Li, Suvarthi Das, Gergely Mozes, Kimberly A. Wong, Kristin A. Olson, Gregory W. Charville, Mohammed Ali, Natalie J. Török

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Figure 2

RAGE and NOX4 mediate oxidative injury in hepatocytes by AGEs.

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RAGE and NOX4 mediate oxidative injury in hepatocytes by AGEs. (A) ROS p...
(A) ROS production was measured by lucigenin chemiluminescence. HiAD-induced ROS production was attenuated by PM treatment in WT mice and in RageHepKO mice on a HiAD. (B) Both Nox4 and Nox2 were induced in WT mice on a HiAD, but only Nox4 induction was attenuated by PM treatment and in RageHepKO mice on a HiAD. (C) ChIP was performed to study Nox4 promoter induction. Significant SMAD3-dependent induction was seen in WT mice on a HiAD, and this was reduced in these mice with PM treatment as well as in RageHepKO mice on a HiAD. (D) Primary WT or Rage-KO hepatocytes were treated with 100 μg/mL AGEs-BSA, in the presence or absence of TGF-β–neutralizing antibody (α–TGF-β, 5 μg/mL), or were transfected with dn-TGF-βR2 or an empty plasmid. SMAD3 phosphorylation (p-SMAD3) was attenuated by the TGF-β antibody or by transfection of dn-TGF-βR2, and no induction of p-SMAD3 was seen in Rage-KO cells. (E) WT and Rage-KO primary mouse hepatocytes were treated with 100 μg/mL AGEs-BSA for 24 hours. The Nox4 luciferase promoter assay indicated AGE-mediated Nox4 promoter activity that was reduced in Rage-KO cells and in those exposed to anti–TGF-β or transfected with the dn-TGF-βR2. (F) ROS production increased in AGE-exposed WT hepatocytes, and this was attenuated in Rage-KO cells, as shown by chemiluminescence assay. Data in A–C represent the mean ± SEM. Data in E and F were combined from 3 replicates. Bars indicate the 25th–75th percentiles, lines indicate the median, and whiskers indicate minimum and maximum values. *P < 0.05, **P < 0.01, and ***P < 0.001, by 1-way ANOVA with Tukey’s post hoc test.