Cardiovascular calcification is an integral part of many cardiovascular diseases, which collectively are the leading cause of death worldwide and contribute to ∼17 million deaths per year (Radvar et al., 2021). One main location of calcification in cardiovascular tissues is the aortic valve, which leads to atherosclerotic vascular microcalcifications, heart failure, and, ultimately, heart attacks (Demer and Tintut, 2008, Hutcheson et al., 2017, Kempf et al., 2021). Various risk factors have been identified to increase the likelihood of cardiovascular calcification including age, gender, diabetes, chronic renal disease, atherosclerosis, hypertension, and smoking (Jondeau et al., 2011, Lavall et al., 2012, Tsamis et al., 2013). However, aging is considered to be the primary risk factor for cardiovascular calcification; it is responsible for changing the structure of collagen and elastin proteins in cardiovascular tissues, such as aorta and mitral valves (Tsamis et al., 2013). The human aorta circulates oxygenated blood from the heart throughout the body (Komutrattananont et al., 2019) and has three layers – the tunica adventitia, tunica media, and tunica intima (Tsamis et al., 2013). Within the microstructure of the aortic wall, elastin and collagen are the main contributors to elasticity and mechanical strength, respectively (Berillis, 2013). Collagen is mainly located in the tunica adventitia (outer layer) (Komutrattananont et al., 2019) and the tunica media (middle layer) (Berillis, 2013), while elastin is primarily located in the tunica media (middle layer). In cardiovascular diseases, inflammatory conditions can affect the compliance of the aortic wall by altering the aorta’s diameter, length, and thickness. Moreover, age-related changes cause an enlargement of, and structural changes in, the tunica media (Berillis, 2013, Komutrattananont et al., 2019). Vascular calcification manifests differently across tissue layers, notably in the intima, media, and heart valve. Intimal calcification primarily affects atherosclerotic plaques especially those caused by apoptotic bodies, contributing to arterial stenosis and thrombosis (Proudfoot et al., 2000). In contrast, medial calcification, prevalent in conditions like diabetes and chronic kidney disease, stiffens arteries and increases cardiovascular risk (Amann, 2008).

Collagen is the most important component of the extracellular matrix (ECM) of heart valves: it provides stiffness, strength, and stability for the valve’s cusps (Kodigepalli et al., 2020). Elastic fibers are mainly composed of elastin, are predominantly arranged as continuous sheets along the radial and circumferential axes (Kodigepalli et al., 2020), facilitate valve motion, and bear a substantial amount of load without deformation (Hinton and Yutzey, 2011). In a diseased environment, changes in the organization of collagen fibers and the emergence of calcification result in thickening of the cusp and leaflet of the valves (Kodigepalli et al., 2020).

While the molecular mechanisms regulating pathological calcification in heart tissue remain unclear, several studies have highlighted events that are similar to some observed in bone formation (Kempf et al., 2021, Tintut et al., 2021). For instance, some cellular processes in vessel wall calcification resemble developmental osteogenesis (Kapustin and Shanahan, 2009), including the competition between mineralization inhibitors and promoters(Herrmann et al., 2020, Kaartinen et al., 2007, Persy and McKee, 2011), osteoblastic differentiation, expression of bone matrix proteins, and formation of hydroxyapatite (Kapustin and Shanahan, 2009, Persy and McKee, 2011, Radvar et al., 2021, Rajamannan et al., 2011). For example, pyrophosphate and matrix gla protein crucially prevent calcification by hindering mineral deposition (Fleisch and Bisaz, 1962, Luo et al., 1997). Beyond inhibitor deficiency, factors like inflammation and oxidative stress also contribute to vascular calcification (Demer and Tintut, 2008). Furthermore, calcified particles have been shown to trigger osteoblastic differentiation of mesenchymal stem cells associated with vascular tissue (Bertazzo et al., 2013), but the source of these calcified particles has not been identified. Several studies have taken a materials science approach to investigate this mysterious process (Bertazzo et al., 2013, Hutcheson et al., 2016). For example, a pioneering study used advanced nano-analytical microscopy techniques to determine that the onset of cardiovascular calcification is not associated with surface precipitation of calcium phosphate, but rather a more complex biomineralization process that occurs within the bulk of the tissue (Bertazzo et al., 2013, Radvar et al., 2021).

The ECM is important for the formation of calcified structures. Even in the absence of cells, tissues can become calcified (Watson et al., 1998). Elastin is the predominant ECM component of elastic fibers in cardiovascular connective tissues. Elastic fibers have a very low turnover rate, so insult to elastic tissue can result in either degradation due to chronic loss(Humphrey et al., 2014) or excess (detrimental) accumulation(Bailey et al., 2014). During the initial stages of cardiovascular calcification, macrophage-derived elastolytic enzymes(Ruiz et al., 2015) and matrix metalloproteinases degrade elastin(Bailey et al., 2004), resulting in the release of soluble elastin-derived peptides that can promote osteogenic differentiation and subsequent calcification(Green et al., 2014). Murshed and co-workers showed that in the absence of a matrix gla protein that is a structural ECM component (Murshed et al., 2004), as well as the elastin content in the arterial walls acts as a critical player for medial calcification (Khavandgar et al., 2014). Sakata et al. reported that modifying the elastin content of the aorta can cause calcification in the aortic media (Sakata et al., 2003). On the other hand, collagen is also believed to play major role in extracellular calcification of cardiovascular tissues, by participating in a slow process that takes years or decades to yield accumulation of collagen, calcification, and disruption of the tissue microarchitecture (Ruiz et al., 2015). However, these studies have, for the most part, analyzed tissues at later stages of calcification and have not addressed the initial events triggering this process. In addition, mineral–associated vesicles (MVs) from cells undergoing osteoblastic differentiation can nucleate and grow hydroxyapatite crystals when bound not only to collagen, but also to other ECM components such glycosaminoglycans (GAGs) (Kapustin et al., 2011). Rodriguez et al. exploited collagen-deficient aortic valvular leaflets via collagenase enzymatic digestion, where the disruption of collagen structure stimulates pathological consequences (Rodriguez et al., 2014).

It has been shown that elastin can be associated with matrix vesicles (Kapustin et al., 2011) and has a greater propensity for calcification because of aging or specific pathologies (Tsamis et al., 2013). The role of elastin in inducing mineralization has been reported in a few in-vitro studies (Parashar et al., 2021). For example, Gourgas et al. reported the deposition of globular calcium phosphate minerals on the fibers and filaments of elastin-like polypeptide (ELP) membranes (Gourgas et al., 2019). Furthermore, we have developed methodologies to engineer ELP-based membranes (Tejeda-Montes et al., 2014, Tejeda-Montes et al., 2012) with tunable ELP conformation (Elsharkawy et al., 2018) and geometrical confinement (Deng et al., 2021) to investigate organic–inorganic interactions within bulk environments.

There is currently no definitive therapy to prevent or treat cardiovascular calcification and the underlying mechanisms triggering this condition are not fully understood. The known risk factors are also not consistently correlated, leaving clinicians uncertain about the optimal therapies for these patients (Nicoll et al., 2015). Surgery is the only effective treatment but can lead to aortic or mitral valve damage, which then requires surgical replacement of the valve. In addition, bio-prosthetic valves still have a high risk of calcification and operative mortality. Given this reality, as well as our studies investigating the role of elastin in mineralization (Deng et al., 2021, Elsharkawy et al., 2018), we hypothesize that cardiovascular tissue compositional damage and elastin degradation play an important, yet not fully understood role in the mineralization of aortic and valve tissues. Validation of this hypothesis would help reshape the current paradigms for pathological mineralization of cardiovascular tissues and pave the way for new therapeutic strategies to prevent, treat, and reverse these conditions. Our findings suggest that elastin interacts favorably to calcium ions, as well as cardiovascular tissues with compositional damage and elastin degradation showed the highest levels of calcifications.