# Investigating the role of FoxO transcriptional regulation of sGC in smooth muscle

> **NIH NIH F31** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2022 · $46,752

## Abstract

PROJECT SUMMARY/ABSTRACT
Hypertension constitutes the most preventable risk factor for cardiovascular disease, affecting nearly 1 out of every 3
Americans. Cardiovascular diseases are the leading causes of death each year across the globe, creating an average
economic burden of nearly $1 billion dollars/year in direct healthcare costs and loss of economic productivity. Healthy
vascular function is maintained by the nitric oxide (NO) signaling pathway, which plays a necessary role in allowing
for proper dilation of blood vessels in response to higher blood pressure. Within vascular smooth muscle cells (SMC),
the NO receptor, soluble guanylyl cyclase (sGC) plays a pivotal role in producing the second messenger molecule,
cyclic guanosine-3’,5’-monophosphate (cGMP), to induce downstream relaxation of SMC. Despite the integral role
that sGC plays in the regulation of vascular tone, the mechanisms that regulate sGC gene expression in SMC remain
unknown. We recently found evidence that the Forkhead box class O (FoxO) transcription factors are capable of
binding on both the sGCa and b promoters. Our preliminary data indicate that pharmacological inhibition of FoxO
transcription factors in rat aortic SMC with AS1842856 results in a 90-95% loss of sGCα and sGCβ mRNA expression,
a 70-80% loss of sGCα and sGCβ protein expression, and a 90% loss of cGMP production after stimulation by the
NO-donor, DEA-NONOate. Likewise, treatment of isolated murine aortic rings with the FoxO inhibitor resulted in a 48%
loss of sGCβ protein expression as assessed by immunofluorescence and significantly blunted NO-dependent
vasorelaxation. Therefore, we sought to identify which of the three FoxO transcription factors expressed in SMC
(FoxO1, FoxO3a, and FoxO4) is responsible for the regulation of sGC expression in SMC by developing adenoviral
FoxO shRNA constructs to transiently knock down expression of each transcription factor. Our preliminary data show
that sGC expression increases following treatment by either FoxO1 or FoxO3a shRNA. Conversely, treatment of rat
aortic SMC with FoxO4 shRNA resulted in a 50% loss of sGCα and sGCβ mRNA and sGCβ protein expression. These
preliminary data have led to the formation of the following aims: 1) Elucidate whether FoxO4 is responsible for sGC
transcriptional regulation in SMC and how this pathway is modulated, and 2) Determine the consequences of
manipulating FoxO4 expression on NO-mediated vasoreactivity ex vivo vasorelaxation. In Aim 1, we will attempt to
1.1) determine the downstream sGC signaling effects following FoxO4 shRNA knockdown, 1.2) generate sGC
promoter-luciferase constructs and utilize chromatin immunoprecipitation (ChIP) experiments to determine whether
FoxO4 is capable of binding and initiating transcription of sGC within SMC, and 1.3) determine if transient
knockdown/rescue of FoxO proteins is sufficient to modulate sGC mRNA and protein expression. To assess Aim 2,
we will 2.1) treat isolated mouse aortic rings with ...

## Key facts

- **NIH application ID:** 10388277
- **Project number:** 5F31HL151173-03
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Joseph Carl Galley
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $46,752
- **Award type:** 5
- **Project period:** 2020-04-01 → 2023-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10388277

## Citation

> US National Institutes of Health, RePORTER application 10388277, Investigating the role of FoxO transcriptional regulation of sGC in smooth muscle (5F31HL151173-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10388277. Licensed CC0.

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