Neonatal mouse-derived engineered cardiac tissue: a novel model system for studying genetic heart disease.

RATIONALE: Cardiomyocytes cultured in a mechanically active 3-dimensional configuration can be used for studies that correlate contractile performance to cellular physiology. Current engineered cardiac tissue (ECT) models use cells derived from either rat or chick hearts. Development of a murine ECT would provide access to many existing models of cardiac disease and open the possibility of performing targeted genetic manipulation with the ability to directly assess contractile and molecular variables. OBJECTIVE: To generate, characterize, and validate mouse ECT with a physiologically relevant model of hypertrophic cardiomyopathy. METHODS AND RESULTS: We generated mechanically integrated ECT using isolated neonatal mouse cardiac cells derived from both wild-type and myosin-binding protein C (cMyBP-C)-null mouse hearts. The murine ECTs produced consistent contractile forces that followed the Frank-Starling law and accepted physiological pacing. cMyBP-C-null ECTs showed characteristic acceleration of contraction kinetics. Adenovirus-mediated expression of human cMyBP-C in murine cMyBP-C-null ECT restored contractile properties to levels indistinguishable from those of wild-type ECT. Importantly, the cardiomyocytes used to construct the cMyBP-C(-/-) ECT had yet to undergo the significant hypertrophic remodeling that occurs in vivo. Thus, this murine ECT model reveals a contractile phenotype that is specific to the genetic mutation rather than to secondary remodeling events. CONCLUSIONS: Data presented here show mouse ECT to be an efficient and cost-effective platform to study the primary effects of genetic manipulation on cardiac contractile function. This model provides a previously unavailable tool to study specific sarcomeric protein mutations in an intact mammalian muscle system.

The ROSA26 LacZ-neo(R) insertion confers resistance to mammary tumors in Apc(Min/+) mice.

B6.129S7-Gtrosa26 (ROSA26) mice carry a LacZ-neo(R) insertion on Chromosome (Chr) 6, made by promoter trapping with AB1 129 ES cells. Female C57BL/6J Apc(Min/+) (B6 Min/+) mice are very susceptible to the induction of mammary tumors after treatment with ethylnitrosourea (ENU). However, ENU-treated B6 mice carrying both Apc(Min) and ROSA26 are resistant to mammary tumor formation. Thus, ROSA26 mice carry a modifier of Min-induced mammary tumor susceptibility. We have previously mapped the modifier to a 4-cM interval of 129-derived DNA that also contains the ROSA26 insertion. Here we report additional evidence for the effect of the ROSA26 insertion on mammary tumor formation. To test the hypothesis that the resistance was due to a linked modifier locus, we utilized two approaches. We have derived and tested two lines of mice that are congenic for 129-derived DNA within the minimal modifier interval and show that they are as susceptible to mammary tumors as are B6 mice. Additionally, we analyzed a backcross population segregating for the insertion and show that mice carrying the insertion are more resistant to mammary tumor development than are mice not carrying the insertion. Thus, the resistance is not due to a 129-derived modifier allele, but must be due to the ROSA26 insertion. In addition, the effect of the ROSA26 insertion can be detected in a backcross population segregating for other mammary modifiers.

Genetic background affects susceptibility to mammary hyperplasias and carcinomas in Apc(min)/+ mice.

Treatment of female C57BL/6J (B6) mice carrying the mutant Min allele of the adenomatous polyposis coli (Apc) gene with ethylnitrosourea (ENU) results in approximately 90% of mice developing an average of three mammary tumors within 65 days. As a first step in the identification of loci modifying susceptibility to ENU-induced mammary tumors and hyperplasias, we have tested ENU-treated Apc(Min)/+ (Min/+) mice on several hybrid backgrounds for susceptibility to mammary and intestinal tumors. C57BR/cdJxB6 (BRB6) Min/+ mice were more sensitive to development of mammary squamous cell carcinomas than B6 Min/+ mice. In contrast, Min/+ hybrids between B6 and FVB/NTac (FVB), 129X1/SvJ (129X1), and 129S6/SvEvTac (129S6) were all significantly more resistant to mammary carcinoma development. However, mice from these three crosses developed more focal mammary hyperplasias than did the B6 or BRB6 Min/+ mice. Susceptibility to intestinal tumors was independent of mammary tumor susceptibility in most hybrids. These results indicate that genetic background can affect independently the phenotypes conferred by the Min allele of APC:

ROSA26 mice carry a modifier of Min-induced mammary and intestinal tumor development.

B6.129S7-Gtrosa26 (B6.R26) mice carry a LacZ-neoR insertion on Chromosome (Chr) 6, made by promoter trapping with 129 ES cells. Female C57BL/6J ApcMin/+ (B6Min/+) mice are highly susceptible to intestinal tumors and to the induction of mammary tumors after treatment with ethylnitrosourea (ENU). However, B6.R26/+ Min/+ females develop fewer mammary and intestinal tumors after ENU treatment than do B6 Min/+ mice. B6.R26/+ mice from two independently derived congenic lines show this modifier effect. Each of these congenic lines carries approximately 20 cM of 129-derived DNA flanking the insertion, raising the possibility that the resistance is due to a linked modifier locus. To further map the modifier locus, we have generated several lines of mice carrying different regions of the congenic interval. We have found that resistance to mammary and intestinal tumors in ENU-treated Min/+ mice maps to a minimum 4-cM interval that includes the ROSA26 LacZ-neoR insertion. Therefore, the resistance to tumor development is due to either the ROSA26 insertion or a very tightly linked modifier locus.