Congenital heart disease (CHD) is the most common birth defect. Among various CHDs, single ventricle phenotypes resulting from altered ventricular morphogenesis have the poorest clinical prognoses and include Tricuspid Atresia (OMIM# 605067), Pulmonary Atresia (OMIM# 265150), and Hypoplastic Left Heart Syndrome (HLHS; OMIM# 241550, 614435). The single ventricle heart presents with a series circuit such that systemic venous return to the right ventricle and pulmonary arteries combined with the flow from the pulmonary venous return into the left ventricle and out to the body is incompatible with survival. Currently, there is a poor understanding of the molecular mechanisms and cellular etiology causative of the many forms of single ventricle CHD. Human mutations in the cardiac transcription factor genes NKX2.5 and HAND1 have been observed in HLHS patients. Modeling and thus the study of possible HLHS phenotypes have been limited as current systemic and conditional knockouts of Nkx2.5 and Hand1 results in embryonic lethality given the broad expression domains of available Cre lines. The lack of a restricted left ventricle Cre driver prohibits such investigations. Hand1 is expressed within the primary heart field myocardium of the left ventricle. We have isolated the enhancer that regulates Hand1 left ventricular expression and used it to generate a novel left ventricular-specific Cre driver with which interrogation of th cellular and molecular mechanism driving left ventricular morphogenesis can be realized. Our experimental plan is to ablate the Hand1 left ventricular lineage cells from the developing embryonic heart, conditionally delete Nkx2.5 specifically within the left ventricular myocardium, and validate an identified human mutation in HAND1 isolated from 24 unrelated patients as being causative of HLHS. This study of the role that Hand1-lineage myocardium plays during cardiogenesis will shed light on the cell etiology of single ventricle phenotypes and on the molecular programs controlling ventricular maturation, thus expanding the understanding of ventricular morphogenesis as it relates to human disease. Relevance: CHDs resulting in single ventricle phenotypes have the poorest clinical outcomes. Thus, gaining an understanding of the etiology and molecular mechanisms that cause CHDs resulting in a single ventricle heart has the potential to benefit thousands of pediatric patients annually. The Hand1-lineage plays a key role in the genesis of single ventricle phenotypes and gaining insight into the cellular and molecular mechanism of this understudied myocardial population will have a great benefit to developing non-surgical treatments for CHD patients.

Public Health Relevance

This proposal seeks to define the role of the bHLH factor Hand1 and its cell lineage within the left ventricle. We have isolated the Hand1 cardiac enhancer and generated a left ventricle-specific Cre. We will test the hypothesis that the Hand1-lineage, loss of left ventricle Nkx2.5 function, and left ventricle specific expression of an identified human mutation in HAND1 all result in phenotypes that model hypoplastic left heart syndrome.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL122123-04
Application #
9204685
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Schramm, Charlene A
Project Start
2013-12-15
Project End
2017-11-30
Budget Start
2016-12-01
Budget End
2017-11-30
Support Year
4
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Indiana University-Purdue University at Indianapolis
Department
Pediatrics
Type
Schools of Medicine
DUNS #
603007902
City
Indianapolis
State
IN
Country
United States
Zip Code
46202
Firulli, Beth A; Toolan, Kevin P; Harkin, Jade et al. (2017) The HAND1 frameshift A126FS mutation does not cause hypoplastic left heart syndrome in mice. Cardiovasc Res 113:1732-1742
Firulli, Beth A; Milliar, Hannah; Toolan, Kevin P et al. (2017) Defective Hand1 phosphoregulation uncovers essential roles for Hand1 in limb morphogenesis. Development 144:2480-2489
Vincentz, Joshua W; Toolan, Kevin P; Zhang, Wenjun et al. (2017) Hand factor ablation causes defective left ventricular chamber development and compromised adult cardiac function. PLoS Genet 13:e1006922
Iklé, Jennifer M; Tavares, Andre L P; King, Marisol et al. (2017) Nkx2.5 regulates endothelin converting enzyme-1 during pharyngeal arch patterning. Genesis 55:
Li, Jingjing; Miao, Lianjie; Shieh, David et al. (2016) Single-Cell Lineage Tracing Reveals that Oriented Cell Division Contributes to Trabecular Morphogenesis and Regional Specification. Cell Rep 15:158-70
Vincentz, Joshua W; Casasnovas, Jose J; Barnes, Ralston M et al. (2016) Exclusion of Dlx5/6 expression from the distal-most mandibular arches enables BMP-mediated specification of the distal cap. Proc Natl Acad Sci U S A 113:7563-8
Gajula, Rajendra P; Chettiar, Sivarajan T; Williams, Russell D et al. (2015) Structure-function studies of the bHLH phosphorylation domain of TWIST1 in prostate cancer cells. Neoplasia 17:16-31
Reuter, Sean; Soonpaa, Mark H; Firulli, Anthony B et al. (2014) Recombinant neuregulin 1 does not activate cardiomyocyte DNA synthesis in normal or infarcted adult mice. PLoS One 9:e115871
Firulli, Beth A; Fuchs, Robyn K; Vincentz, Joshua W et al. (2014) Hand1 phosphoregulation within the distal arch neural crest is essential for craniofacial morphogenesis. Development 141:3050-61
VanDusen, Nathan J; Vincentz, Joshua W; Firulli, Beth A et al. (2014) Loss of Hand2 in a population of Periostin lineage cells results in pronounced bradycardia and neonatal death. Dev Biol 388:149-58

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