Abstract
Purpose :
ACE2 plays an important role in inflammation mainly by degrading the proinflammatory angiotensin II (AngII) and preventing excessive levels of this peptide in various tissues. The ACE2/ AngII axis is understudied in the cornea and may play an important role in corneal inflammatory diseases. To explore this axis further, we took advantage of mice deficient in ACE2 and studied their responses to corneal perturbation.
Methods :
The inflammatory status, cytokine and gene expression profiles were analyzed in ACE2-/- mice by immunohistochemistry (IHC), q-PCR and ELISA. For mechanistic studies, ACE2 was depleted in hTCEpi cells, a limbal epithelial cell line, and primary human limbal epithelial cells (HLECs). To provoke a mild perturbation, small 1mm epithelial debridement wounds were made in the central corneas of ACE2-/- and age-matched WT mice.
Results :
IHC detected ACE2 and AngII in limbal and corneal epithelial basal cells in humans and mice. More than half of adult ACE2-/- mice (n = 14, 60-82 wks) had marked corneal inflammation observed clinically as corneal haze. Haze occupied the central cornea, accompanied by corneal edema and neovascularization. The corneal epithelium had a squamous metaplasia phenotype and the stroma contained a large number of CD11c, CD68, and CD3 positive cells. Interestingly, many adult ACE2-/- mice had no evidence of haze or inflammation. Therefore, we posited that the haze was due to a prior injury. To test this idea, we wounded young (6-10 wk old) ACE2-/- mice that had clear corneas with no signs of haze or inflammation and compared them with WT mice. After wounding, the ACE2-/- mice had: (i) a brisk polymorphonuclear stromal infiltrate; (ii) large numbers of CD11c, CD68 and CD3 positive cells; and (iii) dramatically increased expression of cytokines (e.g. iNOS, CCL2). In contrast, WT mice had little morphological or immunological evidence of corneal inflammation. To explore the mechanism(s) underlying the induction of inflammation, hTCEpi cells and HLECs deficient in ACE2 demonstrated increased expression of IL1a, IL6, TNF-a, CCL2 and CXCL8, along with a significant activation of NF-kB. These phenotypes were rescued by treatment with the angiotensin II type 1 receptor (AT1R) antagonist, losartan.
Conclusions :
Our study establishes a pivotal role of ACE2 in cornea and identifies AngII as a potential new target for corneal inflammation.
This is a 2020 ARVO Annual Meeting abstract.