# Combining Targeted Demethylation with Noncoding RNA-mediated mRNA Stabilization as a Strategy for Therapeutic Arteriogenesis in the Aged

> **NIH NIH R01** · OCEAN STATE RESEARCH INSTITUTE, INC. · 2022 · $49,190

## Abstract

ABSTRACT:
Aging and age-related diseases like peripheral artery disease (PAD) lead to considerable morbidity and
mortality. Aging is associated with impaired inflammatory arteriogenesis responses to injury. We defined a
macrophage signaling axis that activates the mRNA stabilizing protein, HuR, to promote VEGF-A expression
required for arteriogenesis. We seek to understand the effects of aging on this pathway. Moderately aged (52-
week-old) mice demonstrated reduced blood flow recovery and decreased arteriogenesis relative to young (12-
week-old) mice in a femoral artery ligation model of ischemia. In aged mice, ischemic muscle tissue and
macrophages revealed reduced VEGF-A expression. Aged macrophages demonstrated increased global DNA
methylation, and though macrophage HuR expression was normal, there was reduced HuR binding to VEGF-A
mRNA with consequent shortened VEGF-A mRNA half-life. Somewhat surprisingly, Dicer1, previously
established as destabilizing for VEGF-A mRNA, was downregulated in aged macrophages. The DNMT
inhibitor, RG108, led to increased Dicer1 and VEGF-A expression and increased HuR binding to VEGF-A
mRNA. miR-29, as a 3p miRNA, appears to be particularly sensitive to changes in Dicer1 expression. Aged
macrophages had decreased expression of miR-29, whose seeding site in the 3′-UTR of VEGF-A is adjacent
to the HuR binding site. Transfection of macrophages with miR-29 mimic increased VEGF-A expression.
Myeloid Dicer1-deleted mice were phenotypically similar to aged mice, having decreased blood flow recovery,
decreased VEGF-A expression, and decreased HuR binding to VEGF-A mRNA with consequent shortened
mRNA half-life. Our hypothesis is that aging acquired methylation of Dicer1 with consequent reductions in
Dicer1 dose-sensitive microRNAs (i.e. miR-29-3p) results in reduced binding of HuR to VEGF-A mRNA and
reductions in both VEGF-A expression and consequent VEGF-A dependent angio/ arteriogenesis. Our aims
seek to 1) define Dicer1 promoter methylation in aged mice to be a major mechanism of impaired VEGF-A-
mediated arteriogenesis with aging; and 2) define the molecular mechanisms whereby the Dicer1 dose-
sensitive microRNA, miR-29-3p, promotes HuR-binding to VEGF-A mRNA with consequent message
stabilization. Our studies will lead to a paradigm shift from Dicer1 as a negative regulator of VEGF-A to that of
a positive regulator. The rescue of macrophage VEGF-A expression by demethylation of Dicer1 or noncoding
RNA (i.e. miR-29) mimics, may have profound implications in the development of treatment strategies that can
promote arteriogenesis and associated tissue preservation in the setting of severe age-related vasculopathies.

## Key facts

- **NIH application ID:** 10631563
- **Project number:** 3R01HL163005-01S1
- **Recipient organization:** OCEAN STATE RESEARCH INSTITUTE, INC.
- **Principal Investigator:** Alan Ross Morrison
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $49,190
- **Award type:** 3
- **Project period:** 2022-09-01 → 2026-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10631563

## Citation

> US National Institutes of Health, RePORTER application 10631563, Combining Targeted Demethylation with Noncoding RNA-mediated mRNA Stabilization as a Strategy for Therapeutic Arteriogenesis in the Aged (3R01HL163005-01S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10631563. Licensed CC0.

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