Congenital diaphragmatic hernia (CDH) is a common, life-threatening birth defect in which the abdominal organs enter the thorax through an abnormal opening in the diaphragm. In about 20% of CDH cases, the herniated viscera are covered with a membranous sac. Our long-term goal is to identify the morphogenetic processes and molecular pathways altered in sac CDH development. Three distinct models of sac hernia formation have been published. These models vary significantly with regards to the process, tissue, and/or timing of the primary defect. Our preliminary data suggests that each of these models accurately describes one or more generalizable features of sac hernia development. Based on this data, we propose a unifying model of sac hernia development in which, (A) a primary defect in mesothelial fold progression causes the diaphragm to remain attached to the liver and (B) halts the migration of neuromuscular tissues and the development of the diaphragmatic vasculature. This leads to (C) regional hypoxia, (D) thinning of the diaphragm, and (E) changes in cell proliferation and apoptosis.
In Specific Aim #1 we will use state-of-the-art three-dimensional micro-CT imaging technology to test individual elements of this model in a recently developed Gata4flox/flox;Prx1-Cre mouse model. These mice have a high incidence (90%) of anterior sac hernias making them ideal for such studies. We will also determine the critical time window for sac hernia formation by ablating Gata4 using a tamoxifen inducible Cre. In humans, haploinsufficiency of GATA4 causes CDH. Since GATA4 encodes a retinoic acid responsive transcription factor, it likely functions to regulate the transcription of key genes during diaphragm development. Although several CDH-related genes have been identified that act upstream of GATA4 in the retinoic acid pathway, GATA4?s CDH-related target genes have not been identified. Our preliminary data suggests that Fras1 may be one of these target genes. FRAS1 is an extracellular matrix protein that forms a self-stabilizing ternary complex with FREM1 and FREM2. We have shown that deficiencies of FRAS1, FREM1 and FREM2 each cause sac hernias that are indistinguishable from those caused by GATA4 deficiency.
In Specific Aim #2, we will confirm that GATA4 modulates the expression of the FRAS1/FREM1/FREM2 protein complex in the developing diaphragm through its primary action on Fras1 transcription. We will then use an unbiased RNAseq-based strategy combined with an innovative bioinformatic approach to identify other CDH-related genes that are regulated by GATA4. Determining the morphogenetic processes that are altered in sac CDH formation and the genes and pathways that are regulated by GATA4 in the developing diaphragm will aid us in the development of therapeutic/preventative interventions for sac CDH. Our studies will also reveal new CDH-related genes whose discoveries will have an immediately impact our ability to provide molecular diagnoses to individuals with CDH.

Public Health Relevance

In this study, we will examine embryonic mice to determine how abnormalities in diaphragm development cause diaphragmatic hernias, a common birth defect in humans. We will also determine which genes are being incorrectly activated or deactivated when these hernias form. Our results will help us understand why some people are born with diaphragm hernias and may ultimately lead to new ways of treating or preventing them.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Small Research Grants (R03)
Project #
5R03HD093962-02
Application #
9747333
Study Section
National Institute of Child Health and Human Development Initial Review Group (CHHD)
Program Officer
Toyama, Reiko
Project Start
2018-07-20
Project End
2020-06-30
Budget Start
2019-07-01
Budget End
2020-06-30
Support Year
2
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Baylor College of Medicine
Department
Genetics
Type
Schools of Medicine
DUNS #
051113330
City
Houston
State
TX
Country
United States
Zip Code
77030