Interactions Between Neural Crest-Derived Cells and Extracellular Microenvironment During Cardiovascular Development

G protein coupled receptors (GPCRs) have long been known to play crucial roles in transducing environmental signals to the adult cardiovascular system. In recent years, the roles of G protein-mediated signaling pathways in orchestrating the interactions of different tissues during cardiovascular development have become increasingly evident. To analyze the role of G protein signaling pathways in vivo we have generated mice where the function of the heterotrimeric G alpha subunit Gai can be ablated in a cell type specific manner utilizing the Cre-loxP system. We have mated these mice to two different neural crest-specific Cre lines in order to probe the effects of loss of Gai mediated signaling on the ability of neural crest cells (NCC) to contribute to the developing outflow tract and aortic arch arteries. METHODS: We have generated mice that express the Gai-inhibiting pertussis toxin S1 subunit (PTX) from the ROSA26 locus in a Cre recombination dependent manner (ROSA-PTX mice). These were mated to mice expressing either the Wnt1 Cre or P0 Cre transgene. Wnt1Cre is active in both premigratory and migratory NCC, whereas P0Cre is active only in migratory NCC and their derivatives. RESULTS: P0Cre-ROSA-PTX mice were normal at birth and demonstrated no structural heart defects. In contrast, Wnt1Cre-ROSA-PTX mice were present in normal numbers at late gestation but died perinatally due in part to cardiac outflow tract defects. Excision reporter and in situ hybridization studies suggest this is secondary to a delay/blockage of cardiac NCC migration into the developing outflow tract. NCC migration into the pharyngeal arches was unaffected in these mice and no craniofacial, thymic, or aortic arch abnormalities were observed. CONCLUSIONS: These results indicate that Gai-mediated signaling is required in premigratory or early migratory cardiac NCC for normal development of the outflow tract. In contrast, endothelin A receptor knockout mice (currently the only GPCR knock out with a neural crest phenotype) are thought to exhibit defects of postmigratory NCC function. RNA profiling of NCC for GPCRs involved in this Gai-dependent pathway has revealed several potential candidate receptors, including orphan receptors. Further analysis of these receptors is underway.

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Abstract 3937: Generation and Analysis of Neural Crest- and Endothelial-specific Mutant Mice for Foxc1

Circulation ◽

10.1161/circ.118.suppl_18.s_505-c ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Seungwoon Seo ◽

Hisaki Hayashi ◽

Tsutomu Kume

Keyword(s):

Neural Crest ◽

Corneal Stroma ◽

Cell Types ◽

Mutant Mice ◽

Specific Cell ◽

Cardiovascular Development ◽

Forkhead Transcription Factor ◽

Vascular Abnormalities ◽

Autonomous Function ◽

Conditional Mutant

The forkhead transcription factor Foxc1 has been implicated in craniofacial, ocular and cardiovascular development. Foxc1 is expressed in mesoderm- and neural crest (NC)-derived cells, including endothelial cells (ECs) of the heart and blood vessels and vascular smooth muscle cells. However, the precise role of Foxc1 in specific cell types still remains unclear. Therefore, to define the distinct function of Foxc1 in NC and EC, we have generated conditional mutant mice for Foxc1 crossed with either Tie2-Cre or Wnt1-Cre mice. EC-specific Foxc1 mutants survive until adulthood with no apparent embryonic defects, although they show vascular abnormalities in the adult. By contrast, NC-specific Foxc1 mutants die perinately with haemorrhagic hydrocephalus, rudimentary frontal bones, and abnormal patterning of the aortic arch. NC-derived cells also give rise to the stroma and endothelium of the cornea, an avascular organ whose transparency is critical for vision. Importantly, we found that NC-specific Foxc1 mutants show failure of the formation of the anterior chamber and corneal endothelium in the eye. Mutant corneal stroma is much thicker than normal with increased cell proliferation. Most Intriguingly, NC-specific Foxc1 mutants exhibit ectopic neovascularization in the cornea with significant upregulation of Mmp9, sFlt1, Flt1 and Tek at E15.5, while Vegfa and Fgf expression is not changed. By contrast, the cornea of EC-specific Foxc1 mutants is normally formed and avascular. These data suggest that the cell-autonomous function of Foxc1 in the neural crest is essential for craniofacial and cardiovascular development and that Foxc1 plays an important role in inhibition of vascular formation in the cornea.

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