Transcriptome analysis of the pathophysiological significance of adrenomedullin 2 in the mouse choroidal neovascularization model

Abstract

Purpose : We have reported that peptides belonging to the calcitonin superfamily, such as adrenomedullin (AM) and CGRP, have various biological activities, including vasodilation, angiogenesis, and anti-inflammatory effects that play critical roles in interocular vascular diseases. Intermedin, a newly added peptide to the calcitonin superfamily, is also called adrenomedullin 2 (AM2) because of its structural similarity to AM. In our previous study using a laser-induced choroidal neovascularization (LI-CNV) model, we found that pathological angiogenesis, inflammation, and fibrosis were exacerbated in AM2 knockout mice (AM2-/-). In contrast, exogenous administration of AM2 ameliorated these pathological conditions (ARVO 2020, 2021). In the present study, we performed transcriptome analysis to elucidate the mechanism of the ameliorating effects of AM2 on LI-CNV.

Methods : Wild-type mice (WT) were subjected to LI-CNV and systemic administration of PBS, AM, or AM2 using an osmotic pump. One week later, total RNA was extracted from retinal pigment epithelium-choroid-sclera complex specimens, and transcriptome analysis was performed using the Clariom™S Array. We then compared gene expression levels between WT and AM2-/- by quantitative real-time PCR analysis of angiogenesis-, inflammation-, and fibrosis-related genes found in the transcriptome analysis.

Results : There were 113 genes in the AM-treated group and 82 genes in the AM2-treated group whose expression was significantly changed by more than twofold or less than one-half. Of these, 15 genes, including Meox2, were commonly altered in the AM and AM2-treated groups, and 63 genes were differently altered in the two groups. In quantitative real-time PCR analysis, in samples from the early phase of LI-CNV, the expression of CD68, IL-6, VCAM-1, PAI-1, and fibronectin was significantly up-regulated in AM2-/- compared to WT, and the expression of Meox2 and Angpt-1 was significantly down-regulated.

Conclusions : Meox2, a homeobox transcription factor known to repress epithelial-mesenchymal transition (EMT), was up-regulated by AM2 treatment and conversely down-regulated in AM2-/-. Therefore, Meox2 could be a potential factor in the ameliorating effects of AM2 on LI-CNV. Further transcriptome analysis is expected to reveal functional similarities and differentiation between AM and AM2.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.