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Evidence from several rodent models has suggested that a reduction of either atrial natriuretic peptide or its receptor in the heart affects cardiac remodeling by promoting the onset of cardiac hypertrophy. The atrial natriuretic peptide receptor mediates signaling at least in part via the generation of intracellular cyclic GMP. To directly test whether accumulation of intracellular cyclic GMP conveys protection against cardiac hypertrophy, we engineered transgenic mice that overexpress a catalytic fragment of constitutively active guanylate cyclase domain of the atrial natriuretic peptide receptor in a cardiomyocyte-specific manner. Expression of the transgene increased the intracellular concentration of cyclic GMP specifically within cardiomyocytes and had no detectable effect on cardiac performance under basal conditions. However, expression of the transgene attenuated the effects of the pharmacologic hypertrophic agent isoproterenol on cardiac wall thickness and prevented the onset of the fetal gene expression program normally associated with cardiac hypertrophy. Likewise, expression of the transgene inhibited the hypertrophic effects of abdominal aortic constriction, since it abolished its effects on ventricular wall thickness and greatly attenuated its effects on cardiomyocyte size. Altogether, our results suggest that cyclic GMP is a cardioprotective agent against hypertrophy that acts via a direct local effect on cardiomyocytes.

In vivo, the normal heart obtains at least 60% of its energy from lipids and the remainder from glucose. Several lines of evidence indicate that an increase in the utilization of glucose (at the expense of fatty acids (FA)) may play a role in the genesis of hypertrophy. Primary cultures of neonatal car- diomyocytes have been used extensively to study the phenotype of these cells as well as their responses to hormonal hyper- trophic agents. Unfortunately, such cultures are most typically cultured in glucose-rich FA-free media, and thus might be hy- pertrophied to start with. We therefore tested the effects of FA- albumin complexes on three different surrogate end points of hypertrophy of cardiomyocytes. Oleate-albumin complexes de- creased the baseline values of all three variables, and increased the relative response of these variables to administration of norepinephrine. Oleate:palmitate-albumin complexes also af- fected all three variables and their responses to norepineph- rine, but the effects differed somewhat from that of oleate- albumin complexes. Our results suggest that addition of long-chain FA, by providing conditions that more closely resem- ble physiological situations, may optimize the expression of hy- pertrophic responses in such cells. However, the differences between the effects of oleate and oleate:palmitate also suggest that the precise composition of FA may affect the phenotype of cardiomyocytes and how these cells respond to hypertrophic agents. —Zahabi, A., and C. F. Deschepper. Long-chain fatty acids modify hypertrophic responses of cultured primary neo- natal cardiomyocytes. J. Lipid Res. 2001. 42: 1325-1330.

Abstract —Cardiac left ventricular hypertrophy (LVH) is commonly associated with hypertension, but its variance is determined for more than 50% by blood pressure–independent genetic factors. Because it constitutes one of the most important risk factors for cardiovascular mortality, we have performed a genome-wide scan of the F2 progeny of crosses between inbred WKY and WKHA rats to detect quantitative trait loci (QTL) linked to cardiac mass. In addition to left ventricular mass (LVM), we also measured left ventricle (LV) concentration of atrial natriuretic factor (ANF), because we have previously established that there was a genetic link between these 2 traits in the same animal cross. We found 2 contiguous QTL on chromosome 5 that were linked to either LVM (logarithm of odds [LOD]=3.5) or log n (LV ANF) (LOD=12). The 1-LOD support intervals of both QTL shared a region overlapping the locus of natriuretic peptide precursor A ( Nppa ) (ie, the ANF-coding gene). We found by sequencing 2 single nucleotide polymorphisms (SNPs) within the first 650 bp of the Nppa minimal promoters of the genes from both strains. One of these SNPs increased the transcriptional activity of the Nppa minimal promoter in transfected neonatal cardiomyocytes in keeping with the higher LV concentration of ANF observed in WKY versus WKHA rats. Taken together with the previous reports showing that ANF may protect cardiomyocytes against hypertrophy, our genetic data single out Nppa as a strong candidate gene for the determination of LVM.

Most cardiovascular traits of interest can be defined as “complex traits,” with the first step in the identification of genetic factors affecting such traits being the detection of quantitative trait loci (QTLs). Animal models have proven particularly useful in this regard. However, only very few of the QTLs identified to date have led to the identification of candidate genes. We describe an example of our own work where the combination of anatomical and a biochemical intermediate phenotypes have led to the identification of the natriuretic peptide precursor A ( Nppa ) gene as a candidate gene for left ventricular hypertrophy (LVH). Combined with the power of comparative genetics, these strategies will continue to improve the chances of finding candidate genes for cardiovascular traits such as susceptibility to heart diseases, hypertension, and hypertension-induced end-organ damage.

55 The cardiac ventricular mass of WKHA inbred rats is higher than that of WKY rats, yet both strains exhibit normal and identical values of blood pressure (as verified by telemetry). We have shown previously in the F2 male progeny of WKY/WKHA crosses that low ventricular concentration of atrial natriuretic factor (ANF) co-segregated significantly with high cardiac ventricular mass (Circ. Res. 84:1453-1458, 1999). To extend these findings, we have performed a scan of the whole genome of 158 rats originating from the male F2 progeny of the WKY/WKHA cross. By linkage analysis, we have identified on chromosome 5 one QTL that was linked to left ventricular ANF concentration (QTL ANF ) with a LOD score of 17, and one QTL that was was linked to left ventricular mass (QTL LVM ) with a LOD score of 3.5. The 2-LOD confidence interval for the location of QTL ANF ranged from positions 94 to 104 cM, which overlaps partly with the 2-LOD confidence interval for the location of QTL LVM between positions 80 to 103 cM. The locus of the natriuretic peptide precursor A ( Nppa ) gene (at about 100 cM) is located within an interval covered by both QTL’s. Sequencing of a 600 bp fragment of the promoter regions of the Nppa genes from WKY and WKHA revealed that there were single nucleotide polymorphisms within two different regions of the promoters. These mutations lie within regions known to contain response elements to transcription factors, but the results of ongoing experiments are needed to determine whether they can affect the ability of the respective promoters to drive the expression of reporter genes in transfected cardiomyocytes. Altogether, these data suggest that the Nppa gene may be considered as a candidate gene to explain, at least in part, the variance of left ventricular ANF concentration and/or left ventricular mass in WKY and WKHA rats. They also define the region that will be studied in congenic lines to obtain a better spatial resolution of the QTL’s, and in particular to determine whether QTL ANF and QTL LVM are identical or adjacent.