In this study, we have shown that elastin haploinsufficiency causes aortic valve malformation and disease in a mouse model, identifying elastin as a crucial factor in the progressive nature of aortic valve disease pathogenesis. These studies distinguish important regional differences in valve malformation and implicate destabilization of the valve annulus as a contributing factor to valve disease. Multiple transgenic mouse models have demonstrated severe cardiac malformation including valve defects;30–32
however, they are typically embryonic lethal and therefore present limited insight into postnatal valve disease pathogenesis. The postnatal valve malformation in the Eln+/−
mouse can be characterized as a “late” valvulogenesis defect (post-cushion formation, post-endothelial mesenchymal transformation), and in this way bears a similarity to human valve disease. The progressive malformation and latent disease manifestation parallels the natural history of human valve disease. Finally, the valve cusp histopathology in elastin haploinsufficiency mimics the findings of human degenerative aortic valve disease, suggesting common mechanisms of elastin dysregulation underlie valve disease pathology. These studies are clinically relevant because they improve our understanding of valve disease pathogenesis and have the potential to contribute to the development of new therapeutics. Taken together, these findings demonstrate a central role for elastin in postnatal valve disease, supporting evidence that aberrant developmental programs cause disease phenotypes that manifest later in life.
The findings of the current study demonstrate that elastin haploinsufficiency results in VIC dysregulation and over time maladaptive ECM remodeling providing an underlying mechanism for disease manifestation. The important functional role of VICs has been emphasized, however VIC phenotyping remains controversial and incompletely understood in part due to the plasticity of these cells.23,25
In this study, we show that elastin haploinsufficiency results in different types of VIC activation that varies spatially and temporally. At the neonatal stage, there is increased expression of both SMA and SMemb concomitant with altered cell proliferation and increased ECM synthesis. At the juvenile stage, the increased expression of SMemb only is spatially restricted and involves the valve annulus-aortic root complex, without changes in either proliferation or apoptosis. Further, SMemb expression is localized to the annulus in adult mice but not aged adult mice, suggesting other factors contribute to valve maintenance and continued ECM remodeling after disease onset. VIC activation is temporally related to aberrant ECM remodeling that results in valve malformation and ultimately valve disease. Since SMA is normally present in the valve, increased expression may reflect maladaptive compensation or uncoordinated VIC differentiation. On the other hand, since SMemb is not typically expressed in the valve, ectopic expression may reflect a different explicitly pathologic response, eg. reactivated endothelial mesenchymal transformation leading to a maladaptive fibrogenesis response.33–35
Since a subset of VICs share a number of attributes with vascular smooth muscle cells, it may be that regulatory phenotypic modulation is also shared.36,37
Future studies exploring VIC activation after disease onset may provide valuable insight into VIC phenotyping, ECM remodeling and the progressive nature of aortic valve disease.
ECM proteins, signaling molecules and transcription factors have been associated with valve disease.6
Gene expression profiling in the current study identified specific genes and pathways involved in the development of aortic valve malformation and disease. Subanalysis of the Fibrogenesis pathway identified patterns of differential gene expression implicating VIC dysregulation and maladaptive ECM remodeling due to fibrogenesis abnormalities. Unexpectedly, we found that elastin
insufficient mice demonstrate decreased TGF-β. Given the central role of elastin in elastic fiber formation, we expected TGF-β to be increased as has been found in the elastic fiber fibrillin1
In the context of valve disease, fibrogenesis abnormalities may represent a spectrum of pathology with fibrosis (too much collagen) on one end and myxomatous change (too much proteoglycan) on the other end. Further, in the Eln+/−
was significantly reduced in aortic valve tissue, consistent with the finding of increased cell proliferation. Jumonji
has been shown to negatively regulate normal proliferation and tissue growth,39
and in the context of elastin haploinsufficiency is downregulated with a concomitant maladaptive increase in cell proliferation that may contribute to early ECM abnormalities and ultimately disease. A limitation of the current study is that the “aortic valve” group contained valve cusp, valve annulus and aortic root. However, the aortic root is known to have different developmental origins than the aorta,40
and therefore from a mechanistic standpoint, the aortic root and valve annulus may represent a functional unit of transitional tissue. Importantly, the microarray was performed prior to the onset of valve disease and therefore is not the result of secondary changes due to hemodynamic perturbation or end stage disease. This type of subanalysis may direct the mechanistic evaluation of inter-related pathways and networks or specific disease-causing mutations, including crosses to other mice with connective tissue phenotypes and conditional transgenic mouse models controlling spatial and temporal gene expression.
These studies clearly demonstrate histopathology intrinsic to the valve cusp, but histochemistry, immunohistochemistry, gene expression and echocardiography findings suggest the region of the valve implicated in disease manifestation is the valve annulus within the aortic root. These observations add significant new implications to the analysis of valve structure and function in the Eln+/−
mouse and may be clinically relevant in the broader study of human valve disease associated with aortopathy. In light of the clinical association between aortic valve disease and aortopathy,41,42
elastic fiber dysregulation in the valve annulus may be particularly relevant in elucidating the pathogenesis of these common and related disorders. These findings challenge the clinical taxonomy and suggest further investigation of the aortic valve annulus region is warranted. The complex anatomy of the aortic valve annulus as it relates to the aortic root has been well-described.21,22
In humans, disease causing gene mutations may be classified as those resulting in both aortic valve disease and aortopathy (ACTA2, MYH11, FBN1, ELN
), valve disease but not aortopathy (FLNA, NOTCH1
), or aortopathy without valve disease (TGFBR1, COL3
). Interestingly, many of these genes were downregulated in elastin insufficient mouse valve tissue (but not aorta tissue), suggesting the primary pathologic tissue is the valve annulus-aortic root complex. These observations raise fundamental questions about both the origin and functional capacity of the valve annulus-aortic root complex and aortopathy. Taken together, these findings confirm that elastin dysregulation plays a central role in human valve disease ultimately leading to variations of aortopathy and aortic valve disease.
Combining functional, molecular and engineering approaches to the evaluation of regional valve tissue may significantly inform our understanding of valve disease natural history and pathogenesis. Synthesizing these approaches will provide important opportunities for early clinical recognition43,44
and potentially refine approaches to valve bioprosthesis development.4,45
For example, one explanation for “acquired” aortic valve disease in these mutant mice may be the combination of a predisposing genotype that results in subtle but maladaptive ECM remodeling and physiologic stresses, including diastolic shear stress at the valve hinge and lateral compressive forces on the valve annulus-aortic root complex. The findings of the current study have significant clinical implications. Longitudinal studies of patients with elastin haploinsufficiency and valve disease may reveal an increased incidence of latent aortic root dilation and aortic regurgitation providing a clinical population with a well defined genotype to study mechanisms of valve disease and aortopathy. Further, elucidating ECM and VIC characteristics will be particularly important in the context of bioprosthetic valve development. Because this area of research is increasingly interested in “living” valve tissue grafts composed of matrix scaffolds seeded with cells, these cell-matrix relationships may inform durability issues.45,46
Finally, different valve sparing root replacement surgery approaches demonstrate different long-term valve disease outcomes, suggesting the integrity of the valve annulus within the aortic root is paramount. Specifically, preservation of the aortic root (reimplantation approach) provides annular stabilization while disruption of the aortic root (remodeling approach) predisposes to annular dilation, significant aortic regurgitation and an increased incidence of subsequent aortic valve replacement.47
The long-standing association between a “dilated aortic root” and aortic regurgitation may actually represent primary valve annulus disease, which questions the premise that valve disease is a secondary complication of aortopathy. This is an important distinction when considering etiology and potential pharmacologic therapies.
Novelty and Significance
What is known?
- Valve disease is characterized by extracellular matrix (ECM) and valve interstitial cell (VIC) abnormalities.
- Animal models of valve disease are limited.
What new information does this article contribute?
- Provides a genetic mouse model of latent valve disease that has origins in development.
- Identifies elastin as a crucial factor in aortic valve disease pathogenesis.
- Implicates the valve annulus region in disease manifestation.
Valve disease is an important public health problem. Elastin, an ECM protein, is a primary component of valve cusp tissue. Despite a long-standing focus on collagen abnormalities in valve disease, elastin dysregulation is a universal if underappreciated finding in human valve disease. Accordingly, we examined the effects of elastin insufficiency on valve structure and function. We identified progressive aortic valve malformation and latent valve disease in elastin insufficient mice. Pathogenesis in this model is due in part to VIC activation resulting in hyperproliferation and maladaptive ECM remodeling in the valve cusp, but surprisingly moreso in the valve annulus. This study identifies important concepts, including the primary role of elastin in valve pathogenesis and the central role of the valve annulus in disease manifestation. These findings provide a new focus for translational research efforts to develop novel therapeutic targets for valve disease.