Could the Phenotypic Outcomes of Genetic Variability in Cells Operating in Mechanically Dynamic Environments be Influenced by a Disrupted “Cell‐ECM” Relationship? Using Cystic Fibrosis and Marfan Syndrome as an Example

ABSTRACT

Diseases due to mutations in essential molecules can involve tissues functioning in very different environments, with some in mechanically active environments. Diseases arising from mutations in a single molecule, such as the CFTR in cystic fibrosis exhibit varied clinical phenotypes. The lung cells expressing mutations in CFTR are functioning in the mechanically active environment of the lung, but these mutations may also play an adverse role in the cardiovascular system. Similarly, Marfan syndrome arises from mutations in an extracellular matrix (ECM) molecule, fibrillin‐1 and this molecule is also involved in tissues operating in very mechanically active environments. Thus, there is the potential for genetic variants with or without clinical symptoms individually to interact in the same individual to exhibit a unique interdependent phenotype involving disruption of the “Cell‐ECM” relationship. Although the clinical phenotypes for the CFTR and fibrillin‐1 individually are rare, both molecules are known to each have >500 mutations. This may be one example of a molecular pair that could uniquely interact, influencing cell function. This article will discuss this premise and address the potential basis for complementarity using CFTR and fibrillin‐1 as examples.

The “Cell‐Extracellular Matrix Unit” in a mechanically active environment (Centre). Mutations in ECM molecules such as fibrillin‐1 can lead to Marfan Syndrome (A), while mutations in the cellular CFTR can lead to Cystic Fibrosis (B). A combination of such mutations without a clinical phenotype alone could also lead to disease (C).

1. Introduction and Background

1.1. Matrix “Units”

As connective tissues develop and undergo growth and maturation in mechanically active environments, the cells and the extracellular matrix (ECM) develop relationships and function as “units” (Figure 1). Such “units” are dependent on the cells in each tissue interacting with the ECM via integrins, and the cells sensing the mechanical loading via their cytoskeleton mechanosensing molecules (Figure 1 and discussed in more detail below). Thus, the functioning of such “units” requires operating within a physiological mechanical window, as well as having an optimal ECM for that specific environment. An inability to generate effective functional “units” could lead to compromised tissue function and development of disease.

FIGURE 1.

The “Cell‐ECM” unit in mechanically active environments. Cells in connective tissues develop functional relationships with the ECM to regulate cell activity. These relationships are likely tissue specific and dependent on both the cells and the specific composition and structure of the ECM.

1.2. Examples of Genetic Conditions Resulting From an Inability to Generate Functional Cell‐ECM “Units”

Several diseases are believed to have a genetic component that either contributes to disease development directly or indirectly. In this group are a wide range of conditions and diseases resulting from mutations in essential molecules. These include those that are mainly organ‐specific conditions, as well as several others that are more systemic conditions. However, for other complex conditions such as osteoarthritis [1], genome‐wide scans have not revealed a defined gene or set of genes that are affected [2, 3, 4]. Similarly, attempts to uncover the molecular basis for the hypermobility form of Ehlers–Danlos syndrome have not yet been successful [5, 6, 7].

For some conditions/diseases, mutations in a single molecule are believed to be responsible for the disease. Examples of this class of diseases are cystic fibrosis (CF) and Marfan syndrome (MFS), with the CF transmembrane conductance regulator (CFTR) and fibrillin‐1 mutations being the target molecule, respectively. Although MFS and CF are usually considered very different conditions, primarily affecting different tissues, there is the potential for these two conditions and their respective mutations and genetic variants to interact and contribute to the heterogeneity that is observed in the characteristics of CF and MFS, respectively. This Perspective article will primarily use MFS and CF as examples to develop the premise that some mutations in their respective molecules, perhaps even mutations or genetic variants without a detectable clinical phenotype could interact to yield a clinical phenotype via their complementary interactions. Of particular interest would be molecules influencing mechanotransduction and mechanosensing via the extracellular matrix (ECM) and cytoskeleton interacting with other mutations in molecules on cells operating in mechanically active environments. Thus, for example mutations such as in the CFTR are functioning on cells operating in a mechanically active environment, and additional variation in the integrity of the ECM of that environment could affect the functioning of the CFTR and impact the disease phenotype. Conversely, mechanically active tissues known to be adversely affected by fibrillin‐1 mutations such as the mitral valve of the heart, a tissue known to also express CFTR can lose function due to mucin accumulation, could be impacted by genetic variants of the CFTR.

1.3. Cystic Fibrosis (CF)

CF is believed to be the result of mutations in the CFTR. This was reported in 1989 [8] and reviewed in refs. [9, 10]. Since then numerous 17 studies have characterized the role of the membrane localized regulator of chloride exchange and the relationship with mucin [11, 12, 13].

The CFTR is a large plasma membrane spanning molecule with 27 exons and localized to the long arm of human chromosome 7 [14]. Over 750 mutations in the CFTR have been reported across the molecule, with many of them clustered around specific sites [15]. The mutations in the CFTR can vary in type and location [16, 17]. The consequences of mutations in the CFTR are mainly associated with pulmonary, intestinal, and pancreatic pathophysiology leading to loss of lung function and dysfunction of the gastrointestinal tract [15], but can also contribute to eye conditions [18, 19, 20]. The most common mutation is the delta F508 mutation [21], but the frequency and mutation distribution can vary with different ethnic groups [22]. Thus, the involvement of the CFTR in CF is complex, with many mutations associated with the different conditions, including CF of varying severity, and many of the mutations are not associated strongly with particular phenotypes.

Thus, the disease association (i.e., CF) with various mutations is quite heterogeneous, with some mutations leading to very progressive disease while other patients have a less progressive disease. There is also heterogeneity in response to treatment. Innovation in treatment regimens has greatly increased the lifespan of patients with CF. Recently, a complex drug treatment targeting the CFTR has been successful in the treatment of some CF patients (i.e., cocktail of CF modulators; reviewed in [23, 24]), but others are resistant to the intervention [25]. Why there is such significant heterogeneity in CF patients with similar or different mutations in the CFTR has not been explained thus far and as such poses a challenge to treating CF >30 years after the initial findings. Relevant to the present discussion is that some CF patients with severe lung disease, the mutated CFTR can lead to modulation of mechanosensitive channels such as TRPV4 and PIEZO1 [26].

Alterations to the functioning of the CFTR initially leads indirectly to the retention of mucin expressed and a failure to clear such mucin‐based material. This failure to clear the mucins then contributes to the risk for infection by organisms such as Pseudomonas with development of chronic biofilm infection, chronic inflammation, and fibrosis. Until the advent of the CF modulator interventions, the main focus of CF treatment was reactive and designed to control mucin accumulation, chronic inflammation, and infection [23, 24].

In the lung, goblet cells function in a mechanically active environment, being subjected to tensile loading with every breath. Such cells also express the mechanosensitive membrane molecule PIEZO1 [26, 27] and thus, are likely regulated by mechanical loading in conjunction with biological mediators. Mucin secretion is reported to be increased by mechanical loading of goblet cells in vitro [27], but the influence of loading in vivo on mucin secretion is not well characterized. However, since the loading environment would be changed with establishment of a fibrotic environment in CF and the “Cell‐Extracellular Matrix” relationship discussed in [7] also altered, it might be expected that mucin secretion would be impacted as CF progressed.

In endothelial cells, CFTR is also reported to modulate PIEZO1 and TRPV4, and both molecules are mechanosensitive molecules [26, 28]. Therefore, defects in CFTR can also impact the functioning of cells of the cardiovascular system.

Although CF is often considered primarily a lung and gastrointestinal disease, it has also been reported to contribute to heart diseases [29], as well as eye diseases [18, 19, 20]. This has also been shown in mouse models [30] and porcine models [31] of CF. Thus, CFTR mutations expressed in a variety of tissues and organs may contribute to several disease states.

The CFTR has been reported to appear in the rabbit heart after birth and exhibits a gradient of increasing expression in the heart that occurs within 4–5 weeks after birth [32]. Thus, the CFTR appears to be up regulated by the increasing mechanical demands occurring after birth and during early postnatal life.

Thus, in the case of CF where cells are operating in a mechanically active environment, the function of the CFTR can potentially be influenced by the integrity of the ECM (depicted in Figure 2). How the CFTR activity would be influenced by the ECM and the mechanical loading would likely depend on the nature of the mutation occurring in the CFTR (i.e., point mutations in specific domains, loss of ability to interact with the cytoskeleton, internal or external to the cell plasma membrane).

FIGURE 2.

Influence of ECM on cell function with a mutated cell membrane molecule in a mechanically active environment. Cells with a mutated membrane protein such as the CFTR would be compromised in an environment with a normal ECM. The altered function would be restricted to mutations which led to a clinical phenotype under such conditions.

2. Marfan Syndrome (MFS)

MFS is also a rare condition, effecting approximately 1 in 10 000 individuals, and one that develops due to mutations in the fibrillin‐1 gene, a gene on human chromosome 15 [33]. Fibrillin‐1 is a large glycoprotein with 65 exons in the gene [34]. The molecule is involved in elastic fibers in a variety of tissues, including the lung and the heart where it plays a prominent role in the mechanical characteristics of the tissues. Fibrillin‐1 can also interact with cells in such tissues via plasma membrane‐associated integrins [35] and thus, participate in mechanosensing and mechanotransduction [7].

Interestingly, a peptide cleaved from fibrillin‐1 during processing, asprosin, also has biological activity [36, 37, 38]. Asprosin is released from the C‐terminal propeptide of fibrillin‐1 by furin [39]. Therefore, some mutations in fibrillin‐1 that impact the availability of asprosin would lead to both direct effects on ECM integrity in tissues such as lung and cardiovascular tissues, as well as possible indirect effects associated with loss of asprosin.

A large number of mutations (>500) in the fibrillin‐1 gene have been noted [40, 41], and analogous to the CFTR gene some of the mutations are clustered along the protein [34], with some associated with specific characteristics [42, 43, 44]. Why there are so many mutations reported for this gene remains unknown presently. In addition, it is not clear whether additional mutations or genetic variants in fibrillin‐1 exist in the population but are undetected due to no known association with clinical pathology.

Clinically, mutations in the fibrillin‐1 gene lead to increased risk for aortic dissections and mitral valve dysfunctions, conditions that can lead to death [45]. Such mutations can also lead to joint hypermobility [46] and increased skin laxity [47], as well as lung disease discussed in [48, 49, 50, 51, 52], ocular conditions [53, 54], scoliosis [33, 48, 55] and other spinal deformities [56]. The mutations lead to increased risk for disease, but the presentation is quite heterogeneous [34, 57], possibly indicating that perhaps background genes and genetic variants, as well as sex variables may be influencing both risk for heart disease as well as involvement of other tissues [58]. For example, in one family, a 40‐year‐old female died of mitral valve disease while two male siblings developed inguinal aneurisms in exactly the same location and one male sibling also developed other organ involvement [Hart, unpublished observations].

Development of ocular manifestations in Marfan syndrome may also involve mucin accumulation in specific areas of the eye [59]. In the context of the eye, mucins may serve lubrication functions in response to shear loading of the tissue. Thus, in this tissue, mucin expression also appears to be a mechano‐responsive protein. In corneal epithelial cells, this responsiveness is reported to involve TRPV4 [60], a well‐documented plasma membrane localized ion channel that is mechanically gated [61, 62].

In some clinical settings, development of neonatal Marfan syndrome involves severe mitral valve dysfunction, and surgery is required. However, whether such cases are associated with a specific subset of mutations in fibrillin‐1 or other additional factors could not be gleaned from the literature. However, this rapid dysfunction of the mitral valve via a myxomatous process is in contrast to the relatively slow (or later development) development of an analogous process leading to death at age 40 discussed above. Whether this difference is due to background gene variation, the site of the fibrillin mutation, or some other factor, remains to be better understood. However, cardiovascular dysfunction in the presence of a fibrillin‐1 mutations in MFS does appear to involve abnormal involvement of mucins [63].

Although some patients with Marfan syndrome develop severe neonatal valve disease, others develop mitral valve dysfunction as adults [64]. This latter scenario implies that development of cardiac valve disease in the context of Marfan syndrome can be fairly acute (i.e., neonatally) or chronic, taking decades to become symptomatic as an adult [65]. Whether these different scenarios are associated with specific fibrillin‐1 mutations or possibly background gene contributions could not be determined from the available literature.

Further to the above point, some patients with Marfan syndrome do not develop symptomatic cardiac valve disease, but the incidence of aortic dissections is higher than the normal population but certainly not close to 100% [65]. Thus, there is incomplete penetrance of the risk, again possibly due to background gene variability, or the influence of other molecular enhancers of the development of the overt changes in the affected tissues.

Heart valves such as the mitral valve function in a high mechanical stress environment discussed in refs [66, 67], and there is accumulation of mucin in conditions such as floppy mitral valve syndrome associated with Marfan syndrome [68, 69]. There is fibrillin‐1 in mitral valves [70], and there may be molecular interactions in the valves that contribute to an environment not totally dissimilar to the lung and CF. Some of such interactions have been reported in mouse models of Marfan syndrome [71]. In addition, some aspects of Marfan syndrome may relate to the interactions of fibrillin‐1 with the growth factor TGF‐beta [72, 73, 74], and some of this dysregulation may involve integrin‐mediated alterations [75]. Furthermore, DNA methylation is altered in tissues of the cardiovascular system in Marfan patients [76], indicating yet another level of alterations contributing to tissue dysfunction. Thus, there are likely a number of primary and secondary sequelae to fibrillin‐1 mutations in addition to alterations in the ECM.

Thus, some mutations in the fibrillin‐1 gene lead to an altered ECM that is not well suited to maintain a functionally adequate tissue that is needed, such as in the high stress mitral valve environment. This may also lead to an altered “Cell‐ECM” relationship where the cells are functionally compromised regarding the molecules they express (Figure 3), either unmutated ECM or other molecules such as mucins that then may accumulate, and their clearance influenced by other factors.

FIGURE 3.

Impact of mutations in the ECM on Cell‐ECM unit function. Mutations in an ECM molecule that led to overt clinical pathology risk could contribute to loss of the integrity of normal cell‐associated membrane molecules. Under such conditions, mutations in a Marfan syndrome‐causing molecule such as fibrillin‐1 could lead to compromised function of molecules such as the CFTR.

2.1. A Hypothetical Interaction of CFTR and Fibrillin‐1 Mutations in CF and MFS

As discussed above, there is overlap regarding the tissues affected by CFTR and fibrillin‐1 mutations, and the involvement of mucins in both CF and some forms of heart disease (i.e., valve disease), as well as ocular disease. Given the relative commonality of mutations in both CFTR and fibrillin‐1, the overlap in target tissues, the heterogeneity in both CF and associated with Marfan syndrome, the resistance of some forms of CF to modern disease modulators, and the wide‐range of mutations along the length of both CFTR and fibrillin‐1, leads to the concept that mutations or gene variants of both genes may coexist in some forms of both CF and Marfan‐associated conditions, potentiating or inhibiting the risk for developing overt pathology (i.e., positive or negative regulation).

In this scenario, in the lung with a range of CFTR mutations and risk for CF development, the coinheritance of genetic variants in fibrillin‐1, possibly even if not sufficient to result in elevated risk for overt Marfan syndrome‐associated diseases of the heart, eye, or lung, could impact the accumulation of mucin in the lungs at risk for CF (outlined in Figure 4). Such interactions may not correlate with expectations of CF development by CFTR mutations alone.

FIGURE 4.

Subclinical cell and ECM mutations complement to yield a clinical phenotype. The hypothetical situation where subclinical mutations in cell‐associated molecules such as the CFTR are complemented by subclinical mutations in an ECM component such as fibrillin‐1 to yield a clinical phenotype. The severity of the clinical phenotype may be variable and dependent on how the function of the cell‐ECM “unit” is compromised.

Additional insights into how mutations and gene variants in fibrillin‐1 could potentiate the effects of CFTR on CF development may also relate to the fact that individuals with Marfan syndrome are at risk to develop pulmonary disease including pulmonary fibrosis [77, 78] with loss of lung function [52], as well as pneumothorax [79].Thus, genetic variants associated with Marfan syndrome could potentiate the rate and extent of the development and progression of the fibrosis initiated by the inflammation and mucin accumulation that occurs in CF due to CFTR mutations. In such a scenario, the interaction between CFTR and fibrillin‐1 mutations would be indirect with the fibrillin‐1 mutations impacting the consequences of the CFTR mutations and not directly related to the mucin accumulation events. Conversely, in the presence of Marfan syndrome‐associated mutations in the fibrillin‐1 gene, coinheritance of a mutated CFTR could impact mucin turnover and development of either the neonatal or adult forms of mitral valve dysfunction. In this scenario, the fibrillin‐1 mediated altered ECM of the mitral valve would perhaps lead to an enhanced accumulation of mucin and the mutations in the CFTR molecule inhibits the effective clearance of the mucin leading to loss of mitral valve function.

Of note, patients with either CF [80, 81, 82] or MFS [49, 83, 84] have a high prevalence of sleep disorders such as sleep apnea. Furthermore, individuals with joint hypermobility disorders such as Ehlers–Danlos syndromes also have an increased risk for obstructive sleep apnea reviewed in [84]. How these similarities are related mechanistically remains to be determined in detail, but this is another condition that could potentially result from combinations of ECM related mutation events and cell membrane mutation events that individually do not result in a clinical phenotype.

If gene sequencing of both CFTR and fibrillin‐1 genes support this concept, then this information could lead to new interventions. For example, in those with evidence for the development of the chronic form of adult mitral valve dysfunction, one may entertain the use of CFTR modulators that are used in the treatment of CF. Their use in the neonatal form may be tried as well but the rapid progression of this phenotype may or may not benefit from such interventions. Such interventions could also be investigated during development of ocular disease in Marfan syndrome patients using a topical presentation of the drugs which may avoid any complications of intravenous administration.

Conversely, as there are no treatments available for fibrillin‐1 mutations, it is unlikely that at the present time one could negate the influence of a fibrillin ‐1 mutations on CF. However, the sequencing results when both CFTR and fibrillin‐1 undertaken could potentially explain aspects of the heterogeneity observed in CF regarding progression and possibly responsiveness or resistance to treatment protocols. It may also point to the fact that mechanical loading considerations and the “Cell‐ECM” relationship should be entertained when analyzing various CF populations.

How the interactions between CFTR and fibrillin would integrate in light of opposing sex differences that have been reported for CF and Marfan syndrome is not clear but would have to be considered when characterizing outcomes. In Marfan syndrome, aortic events are reported to be more prevalent in males than females [85]. In contrast, CF outcomes are reported to be worse in females than males [86]. The latter potentially may be related to the inflammation associated with a chronic lung infection as females are reported to mount more vigorous immune responses than males [87, 88]. Therefore, the molecular interactions discussed may need to be analyzed in the context of sex.

In summary, while some diseases may be considered caused by a single gene (i.e., CFTR or fibrillin‐1), there may be interactions with those gene variants that influence disease progression. This may be manifested by normal background gene variables, or perhaps the involvement of specific alterations in other molecules that impact the environment that the original mutation operates in, such as the lung, the eye, or the heart. An example of the latter could be mutations in CFTR and fibrillin‐1 where one is the primary effector and the other is an indirect modulator of the consequences of the primary mutations due to the environment.

2.2. Could Other Syndromes Resulting From Mutations in ECM Components and ECM Regulation Also Interact With CFTR Mutations Contributing to CF Development and Progression?

This article has focused on the potential interaction of CFTR and fibrillin‐1 mutations and gene variants on CF and Marfan‐related cardiac diseases to advance the concept that there can be interaction between mutations in diverse molecules that contribute to disease phenotypes. In addition, the concept that there could be correlations between specific mutations in large molecules such as CFTR and fibrillin‐1 that may exert effects on disease progression was advanced.

However, there are also other syndromes such as Ehlers–Danlos syndromes [EDS] (13 subtypes defined by genetic mutations and one as yet not defined)[5, 6, 7, 89], and Loeys–Dietz syndrome where the mutations are also in ECM molecules or in molecules that regulate ECM structure and function [7, 89, 90, 91]. Individuals with some forms of EDS are reported to have compromised lung function [79, 92, 93] or overt lung disease [94]. For those with the hypermobile form of EDS (hEDS) this compromise may be related to inspiratory muscle weakness that can be influenced by exercise programs [95, 96]. Although the chance of having both EDS and CF is likely very rare as the mutations are on different chromosomes, it has been reported that siblings of Turkish consanguineous parents had both EDS type VI and CF [97], so it is possible.

Two additional syndromes involving mutations in genes located on the X chromosome are also relevant to this discussion. The first of these involves loss of function mutations in the biglycan gene, leading to what has been labeled Meester–Loeys syndrome [98, 99]. Biglycan is a small leucine‐rich proteoglycan which can bind to collagens and has a number of other biological functions reviewed in [7, 89]. Individuals with biglycan mutations have increased risk for cardiovascular events, skeletal alterations and joint hypermobility [98, 99]. The second set of mutations are in the filamin A gene (FLNA), leading to variable phenotypes involving the cardiovascular, skeletal and pulmonary systems [100, 101, 102, 103]. Filamin A functions in the cell as a link between plasma membrane located integrins and the cytoskeleton. As such, it is likely involved in mechanosensing and mechanotransduction. Relevant to the present discussion are reports that filamin A interacts with the CF‐associated CFTR and regulates membrane levels of the molecule [104, 105, 106]. Filamin A was also reported to be a biomarker of CF in studies of CF sweat proteomics [107]. Based on the X‐chromosome location of these two molecules, variation in their integrity could impact the clinical phenotype of both CF and Marfan syndrome and contribute to the sex differences in some of the clinical features of those syndromes. Thus, the concept advanced regarding CFTR and fibrillin‐1 may also be expanded to include other molecules with gene variations that affect lung ECM quality and interact directly or indirectly to impact outcomes in CF.

2.3. Feasibility of the Hypothesis

Given the incidence of mutations in genes such as the CFTR and Fibrillin‐1 that lead to the development of clinical pathology are common but still somewhat rare (discussed above), the chances of having mutations in both genes that lead to a clinical phenotype in an individual may be very rare. However, there may be additional mutations in these molecules that do not result in a clinical phenotype on their own but may result in a clinical phenotype when paired with another mutation or genetic variant in a complementary molecule, particularly when one of them is in the ECM of a mechanically active tissue (i.e. fibrillin‐1) or in the mechanical response apparatus (i.e. filamin A). Why there are so many mutations in either the CFTR (>750 reported) or fibrillin‐1 (>500) remains to be determined, but the numbers do not preclude the presence of additional genetic variants that have not yet been identified. The bias would be to identify those mutations associated with a clinical phenotype and not those in the general population.

However, in the literature, there are reports of other circumstances where two separate gene mutations have been implicated in clinical disease [108, 109, 110, 111, 112, 113]. Interestingly, in the reports by Mustacich et al. [110] and Evans et al. [113] one of the mutations involved a mechanosensing molecule, PIEZO1 and TRPV4, respectively. Therefore, one might expect that such mutations would exert some influence via mechanical loading of tissues and mechanotransduction reviewed in [89]. Relevant to the present discussion, in the case report by Phokaew et al. [108], a female‐inherited mutations in both fibrillin‐1 and fibrillin‐2 and had very serious cardiovascular manifestations.

A limitation of the above studies and their rarity is that the patients had to have had a clinically detected and serious phenotype to be selected for sequencing. Furthermore, they likely would have been selected for sequencing of either CFTR or fibrillin‐1, but not both, depending on the clinical presentation. Some mutations in CFTR have been noted but they do not have an identifiable clinical phenotype discussed in [15], so such gene variants could develop a phenotype if paired with perhaps a mutation in fibrillin‐1, as a speculation.

Therefore, to approach any association between CFTR and fibrillin‐1 would likely require sequencing both genes. Although this perhaps would be expensive, the costs for such analysis are decreasing with technological advancements. In addition, a study of multimutation associations with clinical phenotypes may also be bolstered by whole genome sequencing and interrogating the databases. This would likely require using advanced DNA sequencing methods [114]. Projects such as the UK 100,000 Genomes Project [115, 116, 117, 118] are an approach that can be used to detect some gene variant relationships and some results regarding CFTR variants using such databases have already provided relevant information on >4000 variants [119]. However, as whole genome sequencing expands, new ways to interrogate the information may reveal additional complexities hypothesized for CFTR and fibrillin‐1. Thus, testing the proposed linkage between mutations in CF and Marfan syndrome, as an example, may be readily testable in the near future.

2.4. Could this Hypothesis be Extended to Other Diseases of Mechanically Active Tissues?

Although the examples discussed were of the single genetic mutation variety (i.e., CFTR and fibrillin‐1), such genetic diseases are likely a minority with many multigenic conditions more prevalent. For conditions or diseases arising in mechanically active tissues such as osteoarthritis (OA) and intervertebral disc (IVD) degeneration, the diseases are fairly common in the population. For OA, some aspects attributed to genetics, and other aspects related to other variables such as obesity, trauma, sex‐related hormonal factors and use/overuse [1; 120, 121, 122, 123, 124, 125]. Based on twin study outcomes, the genetic contribution can vary up to >50% depending on the joint (i.e., hip, knee, shoulder, hand) involved [120, 123]. However, most studies do not factor in the role of ECM variation and biomechanics in the interpretation of the findings and likely should as the biomechanical environments do vary between joints (i.e. ball and socket for the hip and articulating surfaces for the knee]. Thus, integrity of function is again based on integration of cell responsiveness, ECM and biomechanics for these joint tissues.

Similarly, development of IVD degeneration is also a very common disorder that is complex with many of the same factors as for OA have been implied [126, 127, 128, 129, 130, 131]. Although some variation in the association of gene variants for MMPs, ECM components, and cytokines/immune factors have been noted [122, 123, 125], much of the genetic association (up to 74% depending on the IVD location and degeneration phenotype [127, 129] remains uncharacterized. However, in IVD degeneration some associations with variants of ECM molecules have been detected. How this is translated into alterations in the Cell‐ECM relationship in a mechanically active environment also remains to be determined.

Although some of the variants in ECM regarding OA and IVD degeneration may affect ECM composition or integrity due to proteinase expression, there has not been much evidence generated regarding mutations in ECM molecules impacting cell functionality. This may be a fruitful area to explore via whole genome sequencing.

2.5. Summary, Conclusions, and Future Directions

Considerable progress has been made regarding the link between specific mutations and clinical phenotypes of disease. However, even in conditions such as CF and MFS where mutations in a single gene have been labeled as causing the disease (i.e., CFTR and fibrillin‐1, respectively), there is considerable variation in disease phenotypes and the risk for disease promoted by the mutations. Some of the variation may arise from where in the molecule the mutation is located, but other factors such as involvement of other gene variants and sex‐related factors, as well as the mechanical environment may also play a role.

However, interactions between multiple mutations in interacting molecules has not been very well investigated. In particular, interacting mutations in molecules that exist intracellularly and extracellularly may not come to mind as being related. In addition, mutations in ECM molecules or related to mechanosensing or mechanotransduction also may not come to mind even though the primary mutation is in cells operating in such mechanically active environments. For example, cells in the lung with mutations in the CFTR are interacting with the ECM and functioning in a very mechanically active environment. Therefore, gene variants in the ECM molecules (i.e. as in fibrillin‐1) or the mechanotransduction system (i.e., filamin A) could modulate the consequences of the CFTR mutation on development of CF phenotypes. Conversely, in mitral values, the increased risk for mucin‐related disease development conferred by fibrillin‐1 mutations in Marfan syndrome could be impacted by association with CFTR gene variants that influence mucin turnover and clearance. Thus, the integrity of the “Cell‐ECM” relationship operating in a dynamic mechanical environment may be essential for function and the evolution of a clinical phenotype (Figures 2, 3, 4).

Going forward, greater emphasis on association studies with whole genome sequencing or targeting different genes associated with unique tissue environment may lead to new understanding of disease complexity. Furthermore, in vitro studies related to understanding the effect of mutations in molecules such as the CFTR should likely be performed in mechanically active environments in 3D constructs with relevant ECM components. Such studies with CFTR mutants, and additional molecules related to fibrillin‐1 and filamin A may contribute to improved understanding.

Future directions may also employ assessment of epigenetic variation in outcomes, as well as the use of additional tools such as single‐cell sequencing, spatial transcriptomics, and computational system biology assessments to further elucidate details regarding variations in the regulation of disease activity and progression.

Author Contributions

This article was prepared and finalized solely by David A. Hart and declares that AI was not used in the preparation of the article.

Conflicts of Interest

The author declares no conflicts of interest.

Data Availability Statement

As no new data were presented in this manuscript, a statement is not relevant.