# Peripheral Receptor Mechanisms in Orofacial Muscle Pain

> **NIH NIH R01** · UNIVERSITY OF MARYLAND BALTIMORE · 2020 · $366,938

## Abstract

PROJECT SUMMARY
DNA methylation, an epigenetic factor, plays an important role in regulating gene expression and alterations in
DNA methylation is a feature associated with a number of human diseases. Inflammation and environmental
factors such as psychophysical stress induces demethylation of pro-nociceptive genes leading to their aberrant
expression. The objective of this renewal application is to investigate how muscle inflammation remotely
regulates DNA methylation of multiple pro-nociceptive genes in trigeminal ganglia (TG) that have been
implicated in pain and hyperalgesia. Our central hypothesis is that masseter muscle inflammation results in
reduced methylation of pro-nociceptive genes in TG leading to their aberrant expression, which contributes to
the development of pain and mechanical hyperalgesia. We further hypothesize that psychophysical stress
potentiates these effects via the excess production of reactive oxygen species (ROS) within TG, which regulate
DNA methylation. In Aim 1, we will determine the role of DNA methylation in inflammatory pain responses.
Specifically, we will examine whether increased DNA methylation via DNMTs at the promoter region of
individual pro-nociceptive genes, prevents inflammatory pain and hyperalgesia. We have confirmed that
pro-nociceptive genes such as TRPV1, TRPA1, P2X3 and PIEZO2 contain CG islands that bind DNMTs and
that the inhibition of DNMT activities increased their expression in TG. In order to determine the role of DNA
methylation in individual genes, we designed and validated a novel DNMT fusion protein complex that targets
the promoter region of a specific gene, using the CRISPR-dCAS9 technology. We expect that the expression of
the fusion protein within TG will prevent the upregulation of the target gene and reveal the relative contribution
of DNA methylation for a specific gene in pain and hyperalgesia under a myositis condition. In Aim 2, we will
investigate the role of intraganglionic ROS in DNA methylation of pro-nociceptive genes. We will examine
whether ROS regulates DNA methylation of TRPV1 and TRPA1 genes in TG and whether stress elevates
intraganglionic ROS, which maintains the reduced level of methylation of the pro-nociceptive genes. Our
preliminary data suggest ROS as a key upstream factor involved in DNA methylation of the two pro-nociceptive
genes. We predict that the blockade of ROS accumulation in TG or targeted methylation of DNA promoters will
prevent stress-mediated potentiation of hyperalgesia and the upregulation of TRPV1 and TRPA1. Successful
achievement of this project should unravel novel mechanisms involving DNA methylation and intraganglionic
oxidative metabolites on functional regulation of multiple pro-nociceptive genes, providing a mechanistic basis
for how inflammation and stress engage sensory ganglia to induce prolonged persistent muscle pain. The
anticipated outcomes should have broad translational implications for the development of therapeutic
approaches...

## Key facts

- **NIH application ID:** 9895424
- **Project number:** 5R01DE016062-13
- **Recipient organization:** UNIVERSITY OF MARYLAND BALTIMORE
- **Principal Investigator:** JIN Y Ro
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $366,938
- **Award type:** 5
- **Project period:** 2005-04-01 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9895424

## Citation

> US National Institutes of Health, RePORTER application 9895424, Peripheral Receptor Mechanisms in Orofacial Muscle Pain (5R01DE016062-13). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9895424. Licensed CC0.

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