Advancements, Gaps, Directions for Exosomes and miRNAs in Pulmonary Valve Diseases and Transcatheter Pulmonary Valve Replacement

1. Introduction

Cells secrete extracellular vesicles (EVs) that contain membrane structures [1,2]. The three major subtypes of EVs are exosomes, microvesicles, and apoptotic bodies, based on their biogenesis, biological characteristics, and receptor composition [3]. Studies indicate that exosomes have a diameter of 30 to 150 nm and possess high stability, low immunogenicity, high biocompatibility, and strong membrane penetration capability[4,5]. As early as 1981, Trams et al. [6] discovered these small membrane-bound vesicles in the supernatant of sheep red blood cells and termed them exosomes. Exosomes are widely present in blood and lymph and can carry proteins, lipids, mRNA, lncRNA, and microRNAs (miRNAs) [7]. Therefore, they can regulate the pathological and physiological states of both local and distant cells [1,2].

Current research indicates that miRNA can serve as diagnostic biomarkers for a number of cardiovascular diseases (CVDs), such as aortic valve stenosis (AVS) [8], pulmonary artery hypertension (PAH) [9], heart failure (HF) [10], and acute myocardial infarction (AMI) [11]. Additionally, they can function as drug delivery vehicles, therapeutic agents, and drug targets [4,5,12]. Moreover, miRNAs contribute to the the occurrence and development [13,14], prognosis [15,16], as well as right ventricular (RV) remodeling of congenital heart disease (CHD) [17,18]. CHD affects approximately one-fifth of patients with right ventricular outflow tract (RVOT) obstruction [19]. Most patients experience pulmonary regurgitation (PR) several years after RVOT reconstruction [20]. Currently, transcatheter pulmonary valve replacement (TPVR), also known as percutaneous pulmonary valve implantation (PPVI), is the primary treatment for patients with RVOT dysfunction and severe PR. As early as 2000, Bonhoeffer et al. completed the world’s first TPVR, successfully treating a child who had CHD and was experiencing valve dysfunction after surgery [21,22]. Studies demonstrate that the survival rates of patients undergoing TPVR are comparable to those of patients undergoing surgical pulmonary valve replacement (SPVR) [23]. However, compared to SPVR, TPVR has the advantages of minimal invasion, rapid recovery, and high safety. This review aims to summarize the research progress of exosomes and miRNAs in pulmonary valve diseases (PVDs) and explores their potential association with TPVR.

2. Physiological Characteristics of Exosomes

2.1. The Biogenesis of Exosomes

The biogenesis of exosomes begins with the invagination of the cell membrane, where the cell internalizes external substances or components from its surface through the process of endocytosis, resulting in early sorting endosome (ESE) formation (Figure 1A) [24,25]. The initiation and maintenance of endocytosis depend on the dynamic changes of the cell membrane and the precise regulation of various endocytosis-related proteins. The membrane of the early endosome buds inward, forming intraluminal vesicles (ILVs) (Figure 1B) [26]. As the number of intraluminal vesicles increases, the ESE gradually transforms into a late sorting endosome (LSE), which subsequently evolves into a multivesicular body (MVB) containing multiple vesicles (Figure 1B, C) [27]. This process is regulated by key proteins such as the endosomal sorting complexes required for transport (ESCRT) complexes (Figure 1C) [28]. During this process, the cell also recruits a variety of molecular components, such as membrane proteins and RNAs, into the vesicles (Figure 1A). A fusion of the MVB with the cell membrane releases the ILVs into the extracellular space, which in turn forms exosomes (Figure 1D). In general, exosomes range from 30 to 150 nm in diameter [29,30,31]. They facilitate communication between cells and material exchange, playing a crucial role both physiologically and pathologically [1].

2.2. The Molecular Composition of EVs

As important mediators of intercellular communication, exosomes possess complex and diverse contents that form the basis for their biological functional diversity. Exosomes contain a variety of proteins, including membrane proteins, cytoplasmic proteins, nuclear proteins, and extracellular matrix proteins (Figure 1D). They are also rich in various RNA components, including mRNA, miRNA, and lncRNA, and may even contain DNA fragments (Figure 1D) [32,33,34,35]. The surface markers of exosomes primarily include CD9, CD63, and CD81, which belong to the tetraspanin family of transmembrane proteins (Figure 1D) [26,36,37,38]. There is significant heterogeneity in their expression across different cell types and environments [26,36,37,38]. Recent studies have shown that circulating miRNAs and tissue miRNAs exhibit significant potential in the diagnosis, prognosis, and pathogenesis of CHD [39,40,41,42]. Furthermore, they contribute significantly to the occurrence and development of the tetralogy of Fallot (TOF) [43,44]. Additionally, when patients with CHD experience fluctuations in their condition or develop complications such as HF or other serious illnesses, the expression levels of specific circulating miRNAs can show significant changes [45,46,47]. These dynamic changes can not only assist clinicians in assessing disease progression, predicting treatment outcomes, and formulating personalized treatment but also enable better judgment regarding the timing for interventional treatments.

3. The Role of miRNA in PVD

In recent years, significant advancements have been made regarding miRNAs in aortic valve, mitral valve, and tricuspid valve diseases. Aortic valve calcification is a prevalent valvular disease, impacting approximately 25% of the adult population [48]. Yanagawa et al. [49] discovered that the expression of miRNA-141 is decreased in patients with bicuspid aortic valve-related aortic stenosis, and it is associated with an increase in the level of bone morphogenetic protein-2 (BMP-2), a key regulator of aortic valve calcification (Table 1). Zhang et al. [50] found that miRNA-30b can negatively regulate Runx2, Smad1, and caspase-3, thereby regulating aortic valve calcification (Table 1). Research has shown that miR-125b is significantly upregulated in aortic valves with severe stenosis compared with controls [51] (Table 1). Hosen et al. [52] identified a correlation between elevated levels of miRNA-122-5p and the absence of improvement in left ventricular function after transcatheter aortic valve replacement (TAVR) (Table 1). Additionally, this miRNA can be transported to cardiomyocytes via EVs and regulates cell apoptosis. Previous research has demonstrated a negative correlation between miRNA-206 levels and left ventricular ejection fraction (LVEF) in patients after TAVR [53] (Table 1). It may thus be possible to assess risk in patients with calcifications of the aortic valve and predict cardiac function following TAVR by evaluating expression levels of these miRNAs [54].

FTR, functional tricuspid regurgitation; HF, heart failure; LV, left ventricular; LVEF, left ventricular ejection fraction; MR, mitral regurgitation; RV, right ventricular TAVR, transcatheter aortic valve replacement; TGA, transposition of the great arteries; TOF, tetralogy of Fallot

In terms of mitral and tricuspid valves, Chen et al. [66] first discovered that the analysis of miRNA in valve tissue can distinguish between two typical degenerative mitral valve diseases, myxomatous mitral valve prolapse (MMVP) and fibroelastic deficiency (FED). Chen et al. [55] found that the significant downregulation of miR-148b-3p and miR-409-3p is associated with the development of HF in patients with mitral regurgitation (MR), providing a new perspective for clinical prediction and intervention of HF (Table 1). In addition, miRNAs in exosomes can predict the recovery of cardiac function in patients with severe MR undergoing MitraClip repair [56,67] (Table 1). Hinojar et al. [57] found that, in comparison to the control group, the expression levels of miR-186-5p, miR-30e-5p, and miR-152-3p were significantly reduced in patients diagnosed with severe functional tricuspid regurgitation (FTR), suggesting their potential as diagnostic biomarkers for FTR (Table 1).

Despite significant advancements in the exploration of miRNAs related to aortic, mitral, and tricuspid valves, research focusing on miRNAs in PVD remains relatively scarce. Firstly, the dynamic changes of exosomes and their miRNAs in the pathogenesis and progression of PVDs await further exploration. It is currently unclear which miRNAs in exosomes are associated with RV function, and it is also uncertain whether they can serve as new PVD treatment targets. Secondly, research on miRNAs in the field of TPVR is nearly absent. We do not yet know which specific miRNAs are closely associated with the success rate, postoperative complications, and long-term prognosis of TPVR. Thirdly, the specific molecular mechanisms of exosomes and their miRNAs in PVDs have not been fully elucidated, leading to a lack of in-depth understanding of their roles in PVDs.

Future research should focus on the following directions. To begin with, it is imperative to conduct additional fundamental research to explore the roles of exosomes and miRNAs in PVDs and to elucidate the associated molecular mechanisms. Secondly, by conducting preclinical trials using animal models of PR, we can more accurately assess the therapeutic effects of exosomes and miRNAs in PVDs. Thirdly, it is necessary to explore the expression patterns of miRNA biomarkers before and after TPVR. Furthermore, to substantiate the biomarker potential of miRNAs, it is imperative that extensive clinical trials be undertaken through multicenter collaborations. In addition, we should integrate miRNAs with traditional cardiac function biomarkers, including troponin, BNP, and NT-proBNP, to comprehensively assess the prognosis of patients [68].

4. miRNA Aids in Determining the Optimal Timing for TPVR Intervention

Patients with RVOT dysfunction, such as TOF or transposition of the great arteries (TGA), may experience RV enlargement, increased volume overload, and PR several years after surgery [69]. As a result, the risk of right HF significantly increases. The timing of intervention for PR should be precisely determined. If the intervention is delayed, the patient’s condition may become too severe, resulting in a poor prognosis. Conversely, if the intervention is performed too early, it may lead to a waste of resources. The timing of intervention should consider both the protection of right heart function and the reduction of intervention frequency. However, the optimal timing for performing TPVR is currently unclear.

According to the guidelines published by the European Society of Cardiology (ESC) in 2020, for symptomatic patients with severe PR and/or at least moderate RVOT obstruction, the implementation of SPVR or TPVR is recommended [70]. Research indicates that the circulating miRNA expression profiles in patients who have undergone surgery for TOF or TGA differ from those in healthy individuals [58,59,60,71,72] (Table 1). Liang et al. [43] demonstrated that the downregulation of miRNA-940 facilitates the proliferation and migration of secondary heart field progenitor cells (Table 1). This process may influence the occurrence and development of TOF, revealing its potential molecular mechanisms. Weldy et al. [46] found that TOF patients had RV enlargement and systolic dysfunction, and increased expression of miR-28-3p, miR-433-3p, and miR-371b-3p (Table 1). These features help monitor RV remodeling and disease progression in patients. Clouthier et al. [73] highlighted the potential of miRNAs as non-invasive biomarkers. They can reflect the systemic response to changes in right heart pressure in children with TOF and associated major aortopulmonary collaterals (MAPCAs), providing important insights for tracking disease progression. Abu-Halima et al. [47] found that miRNA-183-3p can serve as an independent predictor of worsening HF in adult patients diagnosed with TGA and a systemic RV (Table 1). Lai et al. [59] found significant differences in serum miRNA expression levels between patients who underwent atrial switch operation for complete TGA and the control population (Table 1). Notably, serum levels of miR-18a and miR-486-5p were associated with systemic RV function. Other studies indicate that circulating miR-423-5p cannot be considered a biomarker for systemic ventricular function in adults following atrial repair for TGA [74]. In summary, specific miRNA expression profiles can reflect changes in right heart function in patients with CHD after surgery, particularly those with RVOT dysfunction. This helps determine the timing of TPVR. However, not all circulating miRNAs can serve as effective indicators for monitoring cardiac function, and their specific applications require further validation.

5. Exosomes and RV Remodeling Following TPVR

Due to their complex anatomical structures, most CHDs often lead to RV myocyte hypertrophy, apoptosis, and fibrosis, ultimately potentially causing RV dysfunction [75]. Current research indicates that patients with severe PR after CHD corrective surgery experience improvements in RV function and favorable remodeling following TPVR [76,77]. Additionally, TPVR can alleviate RVOT obstruction and promote functional recovery of the ventricle as well as the process of cardiac remodeling [78,79,80].

In recent years, the discovery of molecules such as miRNAs and exosomes has offered new insights into elucidating the pathological mechanisms of RV remodeling and developing intervention strategies. Shi et al. [61] found that mice subjected to hypoxia and pulmonary artery banding developed right HF, accompanied by downregulation of miR-223 expression in vivo (Table 1). Conversely, increasing the expression of miR-223 can alleviate the symptoms of RV dysfunction [61]. This provides new therapeutic targets for the treatment of right HF caused by hypoxia and ischemia. Ma et al. [62] found that upregulation of miR-335-5p can lead to RV remodeling, providing a new strategy for the treatment of RVF caused by pulmonary hypertension (Table 1). Yuan et al. [63] found that the heart secretes miR-378 in response to mechanical stress and inhibits excessive cardiac fibrosis through a paracrine mechanism (Table 1). DiLorenzo et al. [18] discovered that MMP1 in exosomes derived from human amniotic epithelial cells can serve as a biomarker for RV remodeling after TOF repair, providing new tools for clinical monitoring and prognostic assessment. However, the role of miRNA in the process of RV remodeling after TPVR and the related molecular mechanisms remain unclear. In the future, it is essential to further elucidate the specific molecular mechanisms of these exosomes and miRNAs in RV remodeling following TPVR, as well as to develop targeted therapeutic strategies based on these molecules.

6. Exosomes and Postoperative Treatment After TPVR

6.1. The Impact of TPVR on RV Function

TOF is the most common cyanotic CHD, accounting for 20% of all CHDs [81,82]. In addition, CHDs such as complete transposition of the great arteries (CTGA), double outlet right ventricle (DORV), and pulmonary valve stenosis, which involve abnormalities in the RVOT, also require reconstruction of the RVOT and repair of the pulmonary valve [83]. However, these patients often experience PR following reconstruction of RVOT, leading to increased preload on the RV, and RV hypertrophy, which can severely result in right HF [84,85]. TPVR can alleviate the afterload on the RV, reduce pulmonary artery pressure, and improve the long-term prognosis of patients.

Long-term follow-up indicates that the Melody and SAPIEN valves used for TPVR can effectively address RVOT dysfunction [86]. Additionally, patients with residual obstruction or infective endocarditis (IE) after the TPVR have a higher probability of re-valve replacement. Rużyłło et al. [87] conducted a follow-up study on 100 patients who underwent TPVR and found that only 14% of patients experienced serious adverse events during an average observation period of 5.5 years, with IE being the primary cause for reintervention. McElhinney et al. [88] conducted a large multicenter cohort study to assess the mid-term and long-term outcomes of 2,476 patients who underwent TPVR. They found that the cumulative mortality rate 8 years after TPVR was 8.9%, with the primary cause of death being HF [88]. Additionally, the cumulative incidence of any transcatheter pulmonary valve reintervention by the 8th postoperative year was 25.1% [88]. The risk factors for surgical reintervention include age, previous IE, implanted stented bioprosthetic valves, and the postimplant gradient [88].

However, various complications following TPVR, including valve migration or embolism and coronary artery compression, may directly or indirectly damage myocardial tissue [79,89,90]. Compression of the coronary artery can lead to MI, which may then cause necrosis or apoptosis of myocardial cells, subsequently releasing multiple myocardial injury markers. Additionally, IE, as one of the serious complications following TPVR, can elicit an inflammatory response that is not limited to local myocardial necrosis. It may also exacerbate widespread myocardial injury through the activation of the immune system [91]. McElhinney et al. [92] conducted a follow-up study on 2,476 patients from 15 centers between 2005 and 2020 and found that the incidence of IE after TPVR was 9.6% at 5 years and 20% at 10 years. Moreover, a history of previous endocarditis, younger age, and high residual pressure gradients were identified as important risk factors for IE.

6.2. Exosomes as Drug Carriers to Improve Right Heart Function

Exosomes, as an emerging acellular therapeutic strategy, have the potential to treat various CVDs [93,94,95,96]. Currently, exosomes as drug delivery carriers offer several advantages, including low immunogenicity, good safety, ease of storage and transportation, wide availability, multifunctionality, targeting capability, potential for mass production, and minimal ethical restrictions.

Beltrami et al. [64] found that exosomes in pericardial effusion promote cardiac angiogenesis by delivering miRNA-let-7b-5p to endothelial cells (Table 1). Research has shown that cardiac progenitor cells (CPCs) improve myocardial function through paracrine secretion of exosomes, which contain numerous miRNAs that can promote angiogenesis and inhibit apoptosis [65] (Table 1). Barile et al. [97] discovered that the surface protein pregnancy-associated plasma protein A (PAPP-A) from exosomes derived from CPCs protects cardiomyocytes by mediating the release of insulin-like growth factor-1 (IGF-1). In addition, microRNA-210 can not only inhibit apoptosis and promote angiogenesis, but also improve the function of cardiomyocytes by controlling mitochondrial metabolism [98,99]. Therefore, microRNA-210 is considered a novel drug candidate for enhancing myocardial function. Bittle et al. [100] found that exosomes derived from human cardiosphere-derived cells (CDCs) can effectively protect right heart contractile function under conditions of of overloading pressure. Gong et al. [101] found that exosomes derived from mesenchymal stem cells can promote cardiac repair after MI. Subsequently, they found that exosomes promote macrophage M2 polarization by targeting the miR-125a-5p/TRAF6/IRF5 signaling pathway, thereby facilitating the repair process. Additionally, Li et al. [102] were the first to develop a stem cell-derived exosome nebulization therapy (SCENT) for cardiac repair, confirming its efficacy in a mouse model of MI.

The drug delivery methods for exosomes include three strategies, exogenous drug loading, endogenous drug loading, and transgenic approaches [26,103,104] (Figure 2). Exogenous drug loading involves first isolating the exosomes and then introducing the drug into the exosomes through transfection or electroporation (Figure 2A). Endogenous drug loading involves first introducing the drug into the cells (Figure 2B). Once the drug enters the exosomes and is released from the donor cells, the drug-loaded exosomes are then isolated and purified (Figure 2B). The transgenic approach involves genetically modifying the cells to express the desired drug, which is ultimately secreted outside the cells along with the exosomes (Figure 2C). However, the application of exosomes in the treatment following transcatheter right heart valve repair and replacement is still in the early stages. In the future, it may play a significant role in promoting myocardial angiogenesis, inhibiting apoptosis, and protecting the myocardium, providing patients with safer and more effective treatment options (Figure 2D).

7. Conclusion and Perspective

This review summarizes the advances in the application of miRNA within exosomes in PVDs and analyzes the potential relationship between exosomes and TPVR. As important carriers of intercellular communication, exosomes play a significant role in the pathological mechanisms, biomarkers, and therapeutic efficacy assessment of aortic valve, mitral valve, and tricuspid valve diseases. Additionally, miRNA is of significant importance in RV remodeling, cardiac function monitoring, and the timing of TPVR intervention. Exosomes, as drug carriers, demonstrate broad application in improving RV function and reducing postoperative complications. However, the specific miRNA biomarkers related to PVDs and RV function have not yet been identified, and the detailed molecular mechanisms of exosomes and their miRNAs in PVDs still await further elucidation. Additionally, the role of miRNAs in the success rate, postoperative complications, and long-term prognosis of TPVR also needs further exploration. Future research should focus on deepening fundamental studies, validating miRNA biomarkers, and promoting clinical application, with the aim of achieving breakthroughs in the field of PVDs.