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Conditional Creation and Rescue of Nipbl -Deficiency in Mice Reveals Multiple Determinants of Risk for Congenital Heart Defects

View Article: PubMed Central - PubMed

ABSTRACT

Elucidating the causes of congenital heart defects is made difficult by the complex morphogenesis of the mammalian heart, which takes place early in development, involves contributions from multiple germ layers, and is controlled by many genes. Here, we use a conditional/invertible genetic strategy to identify the cell lineage(s) responsible for the development of heart defects in a Nipbl-deficient mouse model of Cornelia de Lange Syndrome, in which global yet subtle transcriptional dysregulation leads to development of atrial septal defects (ASDs) at high frequency. Using an approach that allows for recombinase-mediated creation or rescue of Nipbl deficiency in different lineages, we uncover complex interactions between the cardiac mesoderm, endoderm, and the rest of the embryo, whereby the risk conferred by genetic abnormality in any one lineage is modified, in a surprisingly non-additive way, by the status of others. We argue that these results are best understood in the context of a model in which the risk of heart defects is associated with the adequacy of early progenitor cell populations relative to the sizes of the structures they must eventually form.

No MeSH data available.


Related in: MedlinePlus

Relationships between Nipbl genotype, embryo size, heart size, and ASDs.A. Table summarizing genotypes, heart size, body size and incidence of ASDs in different crosses. B. Rescued NipblFLEX/+;cTnt-Cre embryos (n = 14) resembled their NipblFLEX/+ littermates (n = 22) in body size and were smaller than control littermates (cTnt-Cre n = 18, wildtype n = 31). C. Similar results were observed in rescued NipblFLEX/+;Sox17-2A-iCre embryos (n = 22; Sox17-2A-iCre n = 16; NipblFLEX/+n = 18; wildtype n = 25). D. NipblFlox/+;cTnt-Cre (n = 15) were similar in overall body size to littermate controls (NipblFlox/+n = 14, cTnt-Cre n = 30, wildtype n = 21). E. Similar results were observed in NipblFlox/+;Sox17-2A-iCre (n = 13) when compared to littermate controls (Sox17-2A-iCre, n = 20; NipblFlox/+n = 10; wildtype n = 19). Note that individual weights for each cross in B–E were normalized to the mean weight of cTnt-Cre controls (B, D), or Sox17-2A-iCre controls (C, E); black bars indicate normalized mean weight for each genotype. F. Ventricular volume analyses (graphed as box plots) show that the overall heart size of rescued NipblFLEX/+;cTnt-Cre embryos (n = 7) were similar in size to NipblFLEX/+ heart size (n = 9) (Mann-Whitney U, p > 0.05). Control hearts (cTnt-Cre, n = 6) were significantly larger than the hearts of their NipblFLEX/+ and NipblFLEX/+;cTnt-Cre littermates (asterisks: Mann-Whitney U, p < 0.05). G. Rescued NipblFLEX/+;Sox17-2A-iCre embryo hearts (n = 5) were also similar in size to NipblFLEX/+ littermate heart size (n = 6) (Mann-Whitney U, p > 0.05) and significantly smaller than control hearts (Sox17-2A-iCre, n = 5) (asterisk: Mann-Whitney U, p < 0.05). H. Ventricular volume analysis show that the ventricle size of NipblFlox/+;cTnt-Cre embryos (n = 9), which display a high frequency of heart defects, were similar in size to control hearts (cTnt-Cre, n = 9) (Mann-Whitney U, p > 0.05). I. NipblFlox/+;Sox17-2A-iCre mutant hearts (n = 5), which also display a high frequency of heart defects, were also similar in size to control hearts (Sox17-2A-iCre, n = 5) (Mann-Whitney U, p > 0.05).
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pbio.2000197.g007: Relationships between Nipbl genotype, embryo size, heart size, and ASDs.A. Table summarizing genotypes, heart size, body size and incidence of ASDs in different crosses. B. Rescued NipblFLEX/+;cTnt-Cre embryos (n = 14) resembled their NipblFLEX/+ littermates (n = 22) in body size and were smaller than control littermates (cTnt-Cre n = 18, wildtype n = 31). C. Similar results were observed in rescued NipblFLEX/+;Sox17-2A-iCre embryos (n = 22; Sox17-2A-iCre n = 16; NipblFLEX/+n = 18; wildtype n = 25). D. NipblFlox/+;cTnt-Cre (n = 15) were similar in overall body size to littermate controls (NipblFlox/+n = 14, cTnt-Cre n = 30, wildtype n = 21). E. Similar results were observed in NipblFlox/+;Sox17-2A-iCre (n = 13) when compared to littermate controls (Sox17-2A-iCre, n = 20; NipblFlox/+n = 10; wildtype n = 19). Note that individual weights for each cross in B–E were normalized to the mean weight of cTnt-Cre controls (B, D), or Sox17-2A-iCre controls (C, E); black bars indicate normalized mean weight for each genotype. F. Ventricular volume analyses (graphed as box plots) show that the overall heart size of rescued NipblFLEX/+;cTnt-Cre embryos (n = 7) were similar in size to NipblFLEX/+ heart size (n = 9) (Mann-Whitney U, p > 0.05). Control hearts (cTnt-Cre, n = 6) were significantly larger than the hearts of their NipblFLEX/+ and NipblFLEX/+;cTnt-Cre littermates (asterisks: Mann-Whitney U, p < 0.05). G. Rescued NipblFLEX/+;Sox17-2A-iCre embryo hearts (n = 5) were also similar in size to NipblFLEX/+ littermate heart size (n = 6) (Mann-Whitney U, p > 0.05) and significantly smaller than control hearts (Sox17-2A-iCre, n = 5) (asterisk: Mann-Whitney U, p < 0.05). H. Ventricular volume analysis show that the ventricle size of NipblFlox/+;cTnt-Cre embryos (n = 9), which display a high frequency of heart defects, were similar in size to control hearts (cTnt-Cre, n = 9) (Mann-Whitney U, p > 0.05). I. NipblFlox/+;Sox17-2A-iCre mutant hearts (n = 5), which also display a high frequency of heart defects, were also similar in size to control hearts (Sox17-2A-iCre, n = 5) (Mann-Whitney U, p > 0.05).

Mentions: The possibility that an additional lineage protects against heart defects when Nipbl-deficient led us to consider ways in which essentially non-cardiac developmental events might affect the heart indirectly. One of the most penetrant phenotypes of Nipbl-deficiency is reduced body size (by ~20% at birth; [22] and S3 Fig, Fig 7 below). Not surprisingly, the determinants of body size lie outside the heart. As shown in Fig 7A–7E, in the NipblFlox/+ and NipblFLEX/+ crosses described above, body size is determined by Nipbl-status outside of the cTnt and Sox17 lineages (i.e., all carriers of the NipblFLEX allele are small, and all carriers of the NipblFlox allele are normal in size). Yet it is also observed that heart size (measured as total ventricular volume) correlates strongly with body size (Fig 7F–7I). Thus, lineages outside the heart and endoderm apparently determine the size of the embryonic heart, just as heart and body size are known to scale together in adults [67]. Below (see Discussion), we raise the possibility that the results in Table 1 might be explained by an influence of heart size on ASD risk, with large hearts being at greater risk for defects than small ones.


Conditional Creation and Rescue of Nipbl -Deficiency in Mice Reveals Multiple Determinants of Risk for Congenital Heart Defects
Relationships between Nipbl genotype, embryo size, heart size, and ASDs.A. Table summarizing genotypes, heart size, body size and incidence of ASDs in different crosses. B. Rescued NipblFLEX/+;cTnt-Cre embryos (n = 14) resembled their NipblFLEX/+ littermates (n = 22) in body size and were smaller than control littermates (cTnt-Cre n = 18, wildtype n = 31). C. Similar results were observed in rescued NipblFLEX/+;Sox17-2A-iCre embryos (n = 22; Sox17-2A-iCre n = 16; NipblFLEX/+n = 18; wildtype n = 25). D. NipblFlox/+;cTnt-Cre (n = 15) were similar in overall body size to littermate controls (NipblFlox/+n = 14, cTnt-Cre n = 30, wildtype n = 21). E. Similar results were observed in NipblFlox/+;Sox17-2A-iCre (n = 13) when compared to littermate controls (Sox17-2A-iCre, n = 20; NipblFlox/+n = 10; wildtype n = 19). Note that individual weights for each cross in B–E were normalized to the mean weight of cTnt-Cre controls (B, D), or Sox17-2A-iCre controls (C, E); black bars indicate normalized mean weight for each genotype. F. Ventricular volume analyses (graphed as box plots) show that the overall heart size of rescued NipblFLEX/+;cTnt-Cre embryos (n = 7) were similar in size to NipblFLEX/+ heart size (n = 9) (Mann-Whitney U, p > 0.05). Control hearts (cTnt-Cre, n = 6) were significantly larger than the hearts of their NipblFLEX/+ and NipblFLEX/+;cTnt-Cre littermates (asterisks: Mann-Whitney U, p < 0.05). G. Rescued NipblFLEX/+;Sox17-2A-iCre embryo hearts (n = 5) were also similar in size to NipblFLEX/+ littermate heart size (n = 6) (Mann-Whitney U, p > 0.05) and significantly smaller than control hearts (Sox17-2A-iCre, n = 5) (asterisk: Mann-Whitney U, p < 0.05). H. Ventricular volume analysis show that the ventricle size of NipblFlox/+;cTnt-Cre embryos (n = 9), which display a high frequency of heart defects, were similar in size to control hearts (cTnt-Cre, n = 9) (Mann-Whitney U, p > 0.05). I. NipblFlox/+;Sox17-2A-iCre mutant hearts (n = 5), which also display a high frequency of heart defects, were also similar in size to control hearts (Sox17-2A-iCre, n = 5) (Mann-Whitney U, p > 0.05).
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pbio.2000197.g007: Relationships between Nipbl genotype, embryo size, heart size, and ASDs.A. Table summarizing genotypes, heart size, body size and incidence of ASDs in different crosses. B. Rescued NipblFLEX/+;cTnt-Cre embryos (n = 14) resembled their NipblFLEX/+ littermates (n = 22) in body size and were smaller than control littermates (cTnt-Cre n = 18, wildtype n = 31). C. Similar results were observed in rescued NipblFLEX/+;Sox17-2A-iCre embryos (n = 22; Sox17-2A-iCre n = 16; NipblFLEX/+n = 18; wildtype n = 25). D. NipblFlox/+;cTnt-Cre (n = 15) were similar in overall body size to littermate controls (NipblFlox/+n = 14, cTnt-Cre n = 30, wildtype n = 21). E. Similar results were observed in NipblFlox/+;Sox17-2A-iCre (n = 13) when compared to littermate controls (Sox17-2A-iCre, n = 20; NipblFlox/+n = 10; wildtype n = 19). Note that individual weights for each cross in B–E were normalized to the mean weight of cTnt-Cre controls (B, D), or Sox17-2A-iCre controls (C, E); black bars indicate normalized mean weight for each genotype. F. Ventricular volume analyses (graphed as box plots) show that the overall heart size of rescued NipblFLEX/+;cTnt-Cre embryos (n = 7) were similar in size to NipblFLEX/+ heart size (n = 9) (Mann-Whitney U, p > 0.05). Control hearts (cTnt-Cre, n = 6) were significantly larger than the hearts of their NipblFLEX/+ and NipblFLEX/+;cTnt-Cre littermates (asterisks: Mann-Whitney U, p < 0.05). G. Rescued NipblFLEX/+;Sox17-2A-iCre embryo hearts (n = 5) were also similar in size to NipblFLEX/+ littermate heart size (n = 6) (Mann-Whitney U, p > 0.05) and significantly smaller than control hearts (Sox17-2A-iCre, n = 5) (asterisk: Mann-Whitney U, p < 0.05). H. Ventricular volume analysis show that the ventricle size of NipblFlox/+;cTnt-Cre embryos (n = 9), which display a high frequency of heart defects, were similar in size to control hearts (cTnt-Cre, n = 9) (Mann-Whitney U, p > 0.05). I. NipblFlox/+;Sox17-2A-iCre mutant hearts (n = 5), which also display a high frequency of heart defects, were also similar in size to control hearts (Sox17-2A-iCre, n = 5) (Mann-Whitney U, p > 0.05).
Mentions: The possibility that an additional lineage protects against heart defects when Nipbl-deficient led us to consider ways in which essentially non-cardiac developmental events might affect the heart indirectly. One of the most penetrant phenotypes of Nipbl-deficiency is reduced body size (by ~20% at birth; [22] and S3 Fig, Fig 7 below). Not surprisingly, the determinants of body size lie outside the heart. As shown in Fig 7A–7E, in the NipblFlox/+ and NipblFLEX/+ crosses described above, body size is determined by Nipbl-status outside of the cTnt and Sox17 lineages (i.e., all carriers of the NipblFLEX allele are small, and all carriers of the NipblFlox allele are normal in size). Yet it is also observed that heart size (measured as total ventricular volume) correlates strongly with body size (Fig 7F–7I). Thus, lineages outside the heart and endoderm apparently determine the size of the embryonic heart, just as heart and body size are known to scale together in adults [67]. Below (see Discussion), we raise the possibility that the results in Table 1 might be explained by an influence of heart size on ASD risk, with large hearts being at greater risk for defects than small ones.

View Article: PubMed Central - PubMed

ABSTRACT

Elucidating the causes of congenital heart defects is made difficult by the complex morphogenesis of the mammalian heart, which takes place early in development, involves contributions from multiple germ layers, and is controlled by many genes. Here, we use a conditional/invertible genetic strategy to identify the cell lineage(s) responsible for the development of heart defects in a Nipbl-deficient mouse model of Cornelia de Lange Syndrome, in which global yet subtle transcriptional dysregulation leads to development of atrial septal defects (ASDs) at high frequency. Using an approach that allows for recombinase-mediated creation or rescue of Nipbl deficiency in different lineages, we uncover complex interactions between the cardiac mesoderm, endoderm, and the rest of the embryo, whereby the risk conferred by genetic abnormality in any one lineage is modified, in a surprisingly non-additive way, by the status of others. We argue that these results are best understood in the context of a model in which the risk of heart defects is associated with the adequacy of early progenitor cell populations relative to the sizes of the structures they must eventually form.

No MeSH data available.


Related in: MedlinePlus