# Multi-Lineage Modulation of cardiac neural crest cells during cardiac development

> **NIH NIH R56** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2024 · $506,984

## Abstract

Project Abstract
Congenital heart disease (CHD) is the most common birth defect with complex CHD causing significant morbidity/
mortality. Therapies targeting adult heart failure have been ineffective in treating heart failure in patients with CHD,
which implies different pathomechanisms of congestive heart failure in patients with CHD compared with those with
structurally normal hearts. Therefore, there is an urgent need to understand the genetic basis of CHDs. Genetic analysis
in CHD patients is confounded by the genetic heterogeneity of the human population. Hence, we used large-scale
chemical/N-ethyl-N-nitrosourea (ENU) mutagenesis in inbred mice to recover mutations that cause CHD. We recovered
mutations in ten vesicular/endocytic trafficking proteins, including a missense mutation (C4232R) in the highly
conserved epidermal growth factor (EGF) repeat domain encoding low-density lipoprotein receptor-related protein 1
(LRP1). All Lrp1m/m mutant mice exhibited CHD comprising outflow tract (OFT) malalignment defects and/or
atrioventricular (AV) septal defects. Conditional knockout of Lrp1 in different cell lineages that are crucial in cardiac
development demonstrated ablation of cardiac neural crest cells (CNCC) using Wnt1-Cre driver recapitulate the
phenotypes of ENU-induced mutants. Interestingly, these Cre deletion experiments also showed Lrp1 deficiency in
other cell lineages along the CNCC migratory path generated with Nkx2-5+/cre, Tie2+/cre also yielded either double outlet
right ventricle (DORV), atrioventricular septal defect (AVSD), or ventricular septal defect (VSD) phenotypes. As these are
observed with variable penetrance, they suggest CNCC migration to the heart is supported by redundant signaling to
ensure high fidelity of CNCC deployment to the heart. In addition, the combined double knockout of Nkx2-5+/cre Tie2+/cre:
Lrp1f/f mutants demonstrated synergistic effects which reflect the complex genetics in human CHD. Based on findings
from a large body of preliminary data, we hypothesize that LRP1 is required in the cardiac neural crest lineage and its
migratory path for proper OFT alignment and development of the atrioventricular cushions (AVC). We will investigate
this with a combination of in vivo and in vitro experiments conducted using Wnt1+/cre: Lrp1f/f mutant mice with Lrp1
deletion targeted to the CNCCs and double knockout of Nkx2-5+/cre Tie2+/cre: Lrp1f/f mutants. First, we will investigate the
deployment of CNCCs with in vitro migration studies in combination with in vivo cell lineage fate mapping analysis (Aim
1). We will further investigate the cellular and molecular basis for how Lrp1 deficiency in CNCCs can lead to OFT and AV
valve defects by conducting single-cell RNA sequencing analysis, as well as in vivo and in vitro analysis of endocytic
trafficking (Aim 2). This will include the examination of endocytic trafficking related to cell signaling pathways identified
in the CNCC and its migratory path. We will also inves...

## Key facts

- **NIH application ID:** 11192955
- **Project number:** 1R56HL173608-01
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Jiuann-Huey Ivy Lin
- **Activity code:** R56 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $506,984
- **Award type:** 1
- **Project period:** 2024-09-24 → 2026-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/11192955

## Citation

> US National Institutes of Health, RePORTER application 11192955, Multi-Lineage Modulation of cardiac neural crest cells during cardiac development (1R56HL173608-01). Retrieved via AI Analytics 2026-05-27 from https://api.ai-analytics.org/grant/nih/11192955. Licensed CC0.

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