探討芳香烴受體與組織蛋白去乙醯酶在糖尿病視網膜病變之治療機制

Background: Exacerbating diabetic retinopathy (DR) might result in the loss of vision. DR was a complicated disease with lifestyle, genetic, and environmental factors all contributing to its etiology. In DR, HIF1A and VEGFR2 are essential for angiogenesis. Histone deacetylase (HDAC) controls proliferation, differentiation, and cell cycle regulation in various cells. Aryl hydrocarbon receptor (AHR) significance in microvascular complications and retinal disorders remains unclear, disputed, and understudied. Furthermore, the involvement of azatyrosine-phenylbutyric hydroxamide (AZP) as an HDAC inhibitor and AHR activation to protect the outer blood-retina barrier influencing DR development is unknown.
Methods: The R programming language and Gene Set Enrichment Analysis (GSEA) were used to analyse the Gene Expression Omnibus (GEO) NCBI database. An in-silico study was conducted to identify potential molecular targets of AZP and used molecular docking to evaluate interactions between AZP and salient potential targets. Western blotting was used to assess protein levels related to epithelial mesenchymal transition (EMT), HDAC, AHR, and biomarkers of DR in AZP treat high glucose (HG)-induced human retinal pigment epithelial cells (ARPE-19). The DR animal model used db/db mice were treated with AZP. We also used AHR heterozygous (AHR+/-) and homozygous or double knockout (AHR-/-) mice. Optical coherence tomography (OCT), fundus fluorescein angiography (FFA), and fundus photography (FP) were used to evaluate yellowish-white spots, hyperfluorescence, vascular leakage, new vessels, and reduced retinal thickness in the retina of mice. Expression of tissue proteins detected by immunofluorescence and immunohistochemistry.
Results: EMT is a pathological signature, and HDAC6 is upregulated in the fibrovascular membrane of DR patients. AZP interact with HDAC4,6. HG cause increased protein levels of mesenchymal features and HDAC1,3,4,6 in ARPE-19 cells. AZP reverse protein levels related to the EMT process and decreased levels of HDAC1,3,4,6. In addition, AZP treatment improves retinal vasculature and retinal thickness. Increased VEGFR2, integrin subunit alpha 5 (ITGA5), and inflammatory factor (ICAM1) expression in HG-induced ARPE-19 cells. Meanwhile, AZP administration inhibited the increasing of VEGFR2, ITGA5, and ICAM1 in HG-induced ARPE-19 cells. The synergy between two cell surface receptor systems, growth factor receptors, and integrins were illustrated by these results. We found that AHR+/- mice showed the severity of DR, including morphological changes, anatomic lesions, retinal thickness, retinal detachment, and neovascular compared to AHR-/- mice at 24 weeks. Our investigation showed that loss of AHR impairs the outer blood-retinal barrier with reduced expression of tight junction proteins, increased expression of mesenchymal features, HIF1A and VEGFR2 in HG-induced ARPE-19 cells.
Conclusions: AZP, an HDAC inhibitor and AHR activator, meets the required safety and good drug criteria. AZP restore EMT in DR. AZP reversed increasing HIF1A and VEGFR2 expression in HG-induced RPE cells. AHR proved to protect tight junction on the outer blood-retinal barrier. Deletion of AHR in mice could be developed as a novel animal model of DR.