Introduction

Dilated Cardiomyopathy (DCM) is a primary myocardial disease characterized by ventricular dilation and impaired myocardial contractility, often leading to congestive heart failure and cardiac death1,2. The etiology of DCM is complex and multifactorial, encompassing genetic predispositions, viral infections, autoimmune reactions, and toxin exposures3,4. Despite extensive research on the pathophysiology of DCM, its precise molecular mechanisms remain incompletely understood. Recently, microRNAs (miRNAs), as pivotal regulators of gene expression, have gained increasing attention for their roles in cardiovascular diseases.

miRNAs regulate the expression of target genes at the post-transcriptional level, playing a crucial role in the development and metabolism of eukaryotes5. Circulating miRNAs (c-miRNAs), which possess key characteristics of effective biomarkers, including high specificity and sensitivity to disease-related changes, as well as a long half-life in samples, can be obtained through non-invasive methods, enabling rapid and accurate early detection6. Given these characteristics and their role in biological regulation, c-miRNAs may emerge as a new class of biomarkers for diagnosing DCM and potentially serve as therapeutic targets. Previous studies have reported varieties in the manifestation of several c-miRNAs in DCM and their potential diagnostic values7,8,9. The role of miR-214 in cardiovascular diseases, such as myocardial ischemia and heart failure, has been extensively investigated, with notable changes in its expression observed10. Studies have shown that miR-214 plays a crucial role in regulating cardiomyocyte apoptosis and energy metabolism by targeting specific genes and signaling pathways11,12. Nevertheless, the precise mechanisms underlying the actions of miR-214 in DCM remain elusive.

In this study, by integrating bioinformatics analysis with experimental verification, we found that miR-214 expression is significantly downregulated in the serum of DCM patients. Further mechanistic studies have shown that miR-214 inhibits the expression of phosphodiesterase 5 A (PDE5A), thereby regulating the cGMP signaling pathway, crucial for controlling intracellular calcium levels and the metabolic state of cardiomyocytes. Based on these findings, we propose that miR-214 may serve as a crucial biomarker in DCM, regulating PDE5A expression and influencing cardiomyocyte energy metabolism. This discovery offers new insights into the pathogenesis of DCM and opens up novel avenues for the early diagnosis and targeted therapy of the disease.

Materials and methods

Data acquisition and analysis

Circulating miRNA sequencing data from 11 DCM patients and 8 healthy individuals (GSE53081) and mRNA sequencing data from heart tissue of 332 DCM patients and healthy controls (GSE141910) were downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/). Raw sequencing data were processed using Trimmomatic for quality control and mapping. Low-quality reads and poly-N-containing reads were removed to obtain clean reads, which were then mapped to the reference genome using HISAT2. Differentially expressed circulating miRNAs (DE c-miRNAs) and genes (DEGs) were identified using the DESeq2 R package, with significance thresholds set at adj-p < 0.05 and |log2FoldChange| > 1.The target genes of the DE c-miRNAs were screened using the Targetscan (https://www.targetscan.org/vert_80/) and the miRDB (http://mirdb.org/) databases13,14. The GO and KEGG enrichment analyses were conducted using the DAVID database (https://david.ncifcrf.gov/)15. Protein-protein interaction (PPI) analysis was performed using the STRING database (https://cn.string-db.org/)16and visualized using Cytoscape software (version 3.7.2, http://www.cytoscape.org/).

Clinical sample collection

Serum samples were collected from patients diagnosed with DCM at the Cardiology Department of Henan Provincial Hospital of Traditional Chinese Medicine, all of whom had provided informed consent. The study was approved by the hospital’s ethics committee and complied with the ethical standards of the Declaration of Helsinki. Initially, peripheral venous blood samples were obtained from the patients. The serum was separated using centrifugation and stored at -80 °C for future use. Concurrently, clinical information for each patient was documented, including age, gender, disease stage, co-morbidities, treatment, left ventricular ejection fraction (LVEF), and left ventricular fractional shortening (LVFS), as shown in Table S1. These measurements were performed by experienced echocardiographers using echocardiography and documented in dedicated case report forms.

Animal ethics statement

The animal experiment protocol for this study was reviewed and approved by the Ethical Committee of Henan Province Hospital of Traditional Chinese Medicine (approval number: PZ-HNSZYY-2023-035). We confirmed that all experiments in this study were performed in accordance with the relevant guidelines and regulations. All the procedures of the study were followed by the ARRIVE guidelines (https://arriveguidelines.org/arrive-guidelines).

Primary myocardial cell culture and modeling

Primary cardiomyocytes were isolated from the myocardium of neonatal C57BL/6 mice (1–3 days old), sourced from the Experimental Animal Center of Henan Provincial Hospital of TCM. The adult mice were procured from Spefor Biotechnology Co., Ltd. (Beijing, China). The neonatal mice were euthanized with an overdose of CO2. Following disinfection, the heart was excised via thoracotomy, placed in pre-cooled D-Hanks buffer, and gently compressed to expel blood. The heart tissue was then cut into 1–2 mm³ fragments. Tissue digestion was performed using 0.2% collagenase II (Cat. # 9001-12-1, Sigma-Aldrich) for 5–10 min per cycle, with digestion halted by adding 10% FBS (Cat. # 04-007-1 A, BI) in DMEM complete medium (Cat. # 2112078, BI). This process was repeated for 4–5 cycles until complete dissociation of the myocardial tissue was achieved. The resulting cell suspension was filtered through a 70 μm cell strainer to remove undigested tissue. After centrifugation at 1000 rpm for 5 min, the cells were plated in polylysine-coated culture dishes. Differential adhesion was employed after 1 h to eliminate fibroblasts, thereby enriching the cardiomyocyte population. Cardiomyocytes were identified through α-Actin staining. The cells were then seeded into culture plates and maintained in DMEM containing 10% FBS. When they reached 70–80% confluence, angiotensin II (Ang II) (Cat. # 4474-91-3, Sigma-Aldrich) was added to a final concentration of 1 µM for 48 h. Post-treatment, morphological changes in the cardiomyocytes, such as increased cell size and disordered arrangement, were observed under a microscope. Quantitative reverse transcription PCR (RT-qPCR) was used to detect ANP and BNP expression, simulating the pathological characteristics of DCM.

RT-qPCR

Total cardiomyocytes RNA was extracted using TRIzol reagent (Cat. # 15596018, Thermo Fisher Scientific), and subsequently reverse transcribed to cDNA. RT-qPCR was employed to detect and analyze the expression levels of DE c-miRNAs and mRNAs in primary cardiomyocytes. Primer sequences for RT-qPCR are listed in Table S2. Each sample was run in triplicate technical replicates, and relative expression levels were calculated using the 2−ΔΔCt method. For miRNA detection, a synthetic external reference (Cel-miR-39) was used for normalization, while GAPDH served as the internal control gene for mRNA detection.

Western blot

Total cardiomyocytes cell lysates were extracted using RIPA buffer and the protein concentration was determined using a standard BCA assay kit. The lysates were separated by SDS-PAGE and transferred onto polyvinylidene fluoride (PVDF) membranes (0.45 μm, Millipore). The membranes were incubated overnight at 4 °C with primary antibodies against PDE5A (Cat. # 22624-1-A, Proteintech, 1:1000 dilution) or β-Actin (Cat. # 66009-1-Ig, Proteintech,1:2000 dilution), followed by incubation with secondary antibodies. After washing, chemiluminescence ECL was added to the imaging. The band intensity was quantified using ImageJ software (version 1.8.0, https://imagej.net/ij/).

Luciferase assay

Predicted miRNA binding sites were inserted into the 3’UTR region of a luciferase reporter gene vector to create the wild-type and mutant reporter gene vectors (Cat. # E2920, Promega). Forty-eight hours after transfection, 293T cells were harvested, lysed, and then treated with luciferase substrate to measure luciferase activity by a luminometer (GloMax-Multi, Promage). Firefly luciferase activity was normalized to Renilla luciferase activity to account for differences in transfection efficiency.

qPCR array

Total RNA was extracted from cardiomyocytes samples according to the qPCR array manual (Cat. # wc-mRNA0215-M, Wcgene® biotech). miRNAs were reverse-transcribed into cDNA using a miRNA reverse transcription kit (Cat. # RR047A, Takara). The cDNA was then added to the qPCR reaction mix and dispensed into the pre-designed miRNA qPCR array plates. Amplification was performed using a real-time fluorescence quantitative PCR instrument (7500FAST, ABI), and the cycle threshold values for each miRNA were recorded. Primer sequences for qPCR Array are listed in Table S3.

cGMP level detection

The intracellular cGMP levels were detected using GloSensor™ technology, developed by Promega. This technique utilizes the plasmid pGloSensor™ 40 F, which encodes a fusion of the cGMP-binding domain with a mutant form of Photinus pyralis luciferase. Initially, the pGloSensor™ 40 F plasmid was transfected into mouse cardiomyocytes. Subsequently, cells were cultured for 48 h to ensure the expression of the reporter gene. Transfected cells were harvested and seeded in a 96-well plate. The GloSensor reagent (Cat. # E1291, Promega) was added to the culture medium 30 min prior to measurement. Test samples or treatments were added to the cells, and following appropriate incubation periods, luciferase activity was measured using a luminometer (GloMax-Multi, Promage) to record the light output.

Statistical analysis

Statistical analyses were conducted using IBM SPSS 19.0. Data are reported as mean ± standard deviation (Mean ± SD). An independent samples t-test was employed for comparisons between two groups. For comparisons among multiple groups, a One-Way analysis of variance (ANOVA) followed by Tukey’s HSD post-hoc test was utilized. Pearson correlation analysis was utilized to assess the relationship between miRNA expression levels and LVEF or LVFS. Receiver operating characteristic (ROC) curve analysis was conducted to evaluate the diagnostic efficacy of miRNA-214 in DCM, with the area under the curve (AUC) being calculated. Statistical significance was established at P < 0.05.

Results

Identification of DE c-miRNAs and DEGs in DCM

Based on differential expression analysis of the GSE53080 dataset, with significance thresholds set at adj-p < 0.05 and |log2FoldChange| > 1, a total of 125 DE c-miRNAs associated with DCM were identified, with 86 miRNAs upregulated and 39 miRNAs downregulated (Fig. 1A, B). Subsequently, using the TargetScan and miRDB databases, we screened the target genes for 50 DE c-miRNAs with |Log2FC| > 2, including 44 upregulated and 6 downregulated miRNAs, identifying 4463 target genes.

Analysis of the GSE141910 dataset, with significance thresholds set at adj-p < 0.05 and |log2FoldChange| > 1, identified 784 DEGs in human cardiac tissue, including 227 upregulated and 507 downregulated genes (Fig. 1C). Then, we performed an intersection analysis of 4463 target genes of DE c-miRNAs and 784 DEGs, identifying 124 common genes (Fig. 1D).

Fig. 1
figure 1

Prediction of circulating miRNAs and genes associated with DCM. (A) Clustering analysis of circulating miRNAs associated with DCM. (B) Volcano plot of DE c-miRNAs in DCM, identified with thresholds of adj-p < 0.05 and |log2FoldChange| > 1 (C) Volcano plot of DEGs in DCM, identified with thresholds of adj-p < 0.05 and |log2FoldChange| > 1 (D) Venn diagram of common genes identified between DE c-miRNA target genes and DEGs in DCM.

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Enrichment and PPI analysis

To further elucidate the functions of these common genes, we conducted KEGG and GO enrichment analyses using the DAVID database. The results revealed that these common genes were significantly enriched in several key pathways and biological processes, including the PI3K-Akt signaling pathway, ECM-receptor interaction, Spinocerebellar ataxia, Human papillomavirus infection, cell adhesion, and inflammatory response (Fig. 2A, B).

Additionally, we carried out PPI network analysis of the 124 common genes using the STRING platform. This analysis identified several hub genes, including CCL5, NT5E, THBS1, ASPN, GRIA1, CTLA4, COL9A1, COMP, IL2RB, FGF10, and COL8A2 (degree ≥ 5) (Fig. 2C). These hub genes play crucial regulatory roles within the network and may be key contributors to the pathophysiological processes in cardiac tissue. Notably, CCL5, NT5E, and THBS1 play significant roles in cardiovascular pathology. CCL5 contributes to the progression of atherosclerosis by recruiting immune cells during inflammation17,18. NT5E, encoding CD73, is associated with vascular calcification, and mutations in this gene can lead to arterial calcification19. THBS1 is involved in cardiac remodeling and fibrosis, affecting cell survival and tissue repair, especially in conditions such as heart failure and DCM20,21. These genes are integral to the inflammatory and fibrotic processes affecting cardiac tissue.

Fig. 2
figure 2

Enrichment and PPI analysis of common genes (A) KEGG pathway22 enrichment analysis of common genes. (B) GO enrichment analysis of common genes. (C) PPI network analysis of common genes with identified hub genes highlighted in red.

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Validation of DE c-miRNAs in DCM

To validate the results of our bioinformatics analysis, we selected 12 DE c-miRNAs (seven upregulated and five downregulated) for experimental verification. Serum samples were collected from 47 healthy individuals and 29 patients with DCM. The expression levels of these miRNAs were quantified using RT-qPCR. The results showed that the expression trends of these 12 miRNAs in the serum of healthy individuals and DCM patients were consistent with the expected trends from our analysis (Fig. 3A-L). This validation further supports our bioinformatics findings, suggesting that these differentially expressed miRNAs may play important roles in the pathophysiology of DCM and provide compelling evidence for their potential application as diagnostic and therapeutic biomarkers.

Fig. 3
figure 3

Validation of DE c-miRNAs in DCM. (A-L) Expression levels of selected miRNAs in serum samples from healthy individuals and DCM patients, as determined by RT-qPCR. Statistical comparisons between two groups were performed using an unpaired Student t-test. ****P < 0.0001.

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Correlation of miRNA-214 with cardiac function indicators and diagnostic efficacy analysis

In this study, we conducted an in-depth analysis of the correlation between the expression levels of DE c-miRNAs and cardiac function indicators, specifically LVEF and LVFS. The results demonstrated significant correlations between the expression levels of various DE c-miRNAs and both LVEF and LVFS. Notably, the expression level of miRNA-214 was significantly positively correlated with both LVEF (R2 = 0.5519, P < 0.01) and LVFS (R2 = 0.5866, P < 0.01), indicating that as LVEF and LVFS significantly decrease, miRNA-214 expression decreases (Fig. 4A, B).

To evaluate the diagnostic efficacy of miRNA-214 in DCM, we performed ROC curve analysis. The results indicated that miRNA-214 has high diagnostic efficacy in assessing DCM, with an AUC of 0.7275 (95% CI: 0.65–0.81, p < 0.01), suggesting good sensitivity and specificity in distinguishing DCM patients from healthy individuals (Fig. 4C).

Fig. 4
figure 4

Correlation and Diagnostic Efficacy of miRNA-214 in DCM. (A) Correlation between miRNA-214 expression and LVEF. (B) Correlation between miRNA-214 expression and LVFS. (C) ROC curve analysis of miRNA-214 for diagnosing DCM.

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Validation of miRNA-214’s Regulatory Role in DCM

To further validate the relationship between miRNA-214 and DCM, we established a DCM cell model by treating neonatal mouse cardiomyocytes with Ang II. The model’s validity was confirmed by measuring cardiomyocyte area using ImageJ and assessing the mRNA levels of key DCM markers, ANP and BNP, via RT-qPCR. In the Ang II-treated group, we observed a significant increase in cell area, accompanied by elevated ANP and BNP mRNA levels (Fig. 5A-C), confirming the successful establishment of the DCM cell model. Additionally, miRNA-214 expression was notably decreased in the Ang II-treated cardiomyocytes (Fig. 5D).

To further explore the role of miRNA-214, we inhibited its expression using a miRNA-214 inhibitor and examined changes in DCM-related genes through a qPCR array. With significance thresholds set at adj-p < 0.05 and |log2FoldChange| > 1, we identified 11 downregulated and 12 upregulated genes in the Ang II-treated DCM model. Upon miRNA-214 inhibition, 15 genes were downregulated and 7 were upregulated (Fig. 5E, F). Venn diagram analysis revealed that 9 genes (MYH6, LMNA, ATP2A2, MYBPC3, TNNI3, TNNT2, DES, ITGA7, CACNA1C) were commonly downregulated, and 3 genes (ITGA5, PLN, TGFB1) were commonly upregulated in both conditions (Fig. 5G). These findings further substantiate miRNA-214’s critical regulatory role in the pathophysiology of DCM, illustrating that changes in its expression significantly affect the expression patterns of genes related to DCM. Specifically, miRNA-214 appears to affect myocardial contractile function (MYH6, MYBPC3, TNNI3, TNNT2, and DES)23,24,25,26, calcium homeostasis (ATP2A2, CACNA1C, and PLN)27,28,29, and extracellular matrix remodeling and myocardial fibrosis (ITGA7, ITGA5, and TGFB1)30,31,32. Consequently, PDE5A, critical for regulating the cGMP signaling pathway—which impacts cardiomyocyte contraction, relaxation, and myocardial fiber and calcium ion homeostasis33,34—may be a downstream target influenced by miRNA-214’s modulation of gene expression.

Fig. 5
figure 5

Validation of miRNA-214’s regulatory role in DCM. Neonatal mouse cardiomyocytes were treated with Ang II, and the following parameters were analyzed across different experimental groups: cell area (A), ANP (B), BNP (C) mRNA expression levels, and miR-214 expression levels (D). (E) Expression changes of DCM-related genes in the Ang II-treated DCM cell model. DEGs were identified using thresholds of adj-p < 0.05 and |log2FoldChange| > 1. (F) Expression changes of DCM-related genes following miRNA-214 inhibition. DEGs were identified using the same thresholds. (G) Venn diagram showing the common DEGs affected by both Ang II treatment and miRNA-214 inhibition. Data are presented as Mean ± SD. *P < 0.05, **P < 0.01, ****P < 0.0001.

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miRNA-214 regulates cGMP metabolism via PDE5A and its role in DCM pathogenesis

Integrated with the PPI network analysis in Fig. 2C, examination of common differentially expressed genes in the Ang II treatment model and the miRNA-214 inhibition model suggests that PDE5A may be a direct target of miRNA-214 in DCM regulation. To validate this finding, we assessed the expression levels of miRNA-214 and its impact on PDE5A gene and protein levels in neonatal mouse cardiomyocytes using RT-qPCR and western blot. The results showed that miRNA-214 inhibitor significantly reduced the level of miRNA-214 in neonatal mouse cardiomyocytes and significantly increased both the mRNA and protein levels of PDE5A. RT-qPCR results indicated that PDE5A mRNA levels were significantly elevated following miRNA-214 inhibition, and western blot results confirmed the increase in PDE5A protein levels. These findings demonstrate that miRNA-214 negatively regulates PDE5A expression (Fig. 6A-C).

To further confirm the direct binding relationship between miRNA-214 and PDE5A, we performed bioinformatics prediction using TargetScan, which identified a 6-mer binding site in the 3’UTR of PDE5A for miRNA-214 (Fig. 6D). This prediction was experimentally validated using a dual-luciferase reporter assay. The results showed that miRNA-214 inhibited the wild-type PDE5A 3’UTR by 32.4%, while it had no inhibitory effect on the mutant PDE5A 3’UTR (Fig. 6E). This result demonstrates that miRNA-214 directly binds to the 3’UTR of PDE5A to regulate its expression.

Given the function of PDE5A, we hypothesized that miRNA-214 might participate in the pathogenesis of DCM by regulating cGMP metabolism via PDE5A. To test this hypothesis, we used the Promega cGMP GloSensor®luciferase detection system to measure the effect of miRNA-214 on cGMP levels in neonatal mouse cardiomyocytes and evaluated the role of miRNA-214 in regulating cGMP levels using the PDE5A inhibitor sildenafil35. The results showed that inhibition of miRNA-214 significantly reduced cGMP levels in neonatal mouse cardiomyocytes, while sildenafil, a specific inhibitor of PDE5A, significantly increased cGMP levels and mitigated the reduction in cGMP levels caused by miRNA-214 inhibition (Fig. 6F). These results indicate that miRNA-214 may regulate DCM by modulating PDE5A and its mediated cGMP metabolism. Our study elucidates the molecular mechanism of miRNA-214 in DCM, providing new insights and potential targets for the development of therapeutic strategies based on miRNA-214 and PDE5A.

Fig. 6
figure 6

miRNA-214 regulates PDE5A and affects cGMP metabolism in DCM. (A) Expression of miRNA-214 in different groups of cells. (B) Expression of PDE5A mRNA in different groups of cells. (C) Immunoblot analysis of PDE5A expression in various cell groups (cropped images; full blots are available in Supplementary Fig. 2). (D) Dual-luciferase reporter assay showing the targeting relationship between miRNA-214 and PDE5A. (E) Validation of miRNA-214 binding to PDE5A 3’UTR. (F) Effect of miRNA-214 inhibition and sildenafil treatment on cGMP levels in neonatal mouse cardiomyocytes. Data are expressed as Mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001.

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Discussion

DCM, a myocardial disease, is a major cause of heart failure and death, characterized by high morbidity and mortality1,36. In the last decades, miRNAs have garnered significant attention for their roles in cardiovascular diseases as crucial regulators of gene expression. Our bioinformatics analysis and experimental validation have revealed that miR-214 is significantly downregulated in the serum of DCM patients and negatively correlates with cardiac function indicators LVEF and LVFS. Further analysis has indicated that miR-214 negatively regulates PDE5A gene expression by directly targeting it.

miR-214 is encoded by the Dnm3os gene located on chromosome 137. It is widely expressed in the myocardium, playing critical roles in cardiomyocyte differentiation and apoptosis38,39. In the normal heart, miR-214 expression is maintained at a stable level. However, in DCM patients, miR-214 expression typically decreases40. Studies have demonstrated that decreased miR-214 expression is closely associated with cardiomyocyte fibrosis and apoptosis41,42. Our findings suggest that miR-214 levels are significantly decreased in the serum of DCM patients and are positively related to LVEF and LVFS. This provides a valuable reference for the early non-invasive diagnosis of DCM.

PDE5A is primarily responsible for degrading cGMP, playing a critical regulatory role in the cardiovascular system. As an important intracellular signaling molecule, cGMP plays a crucial role in cardiomyocyte proliferation, apoptosis, and vasodilation43,44,45. Increased expression or activity of PDE5A leads to reduced cGMP levels, thereby inhibiting PKG activity and increasing the risk of cardiomyocyte apoptosis, which plays a significant role in the pathological progression of DCM46. Conversely, inhibiting PDE5A expression can enhance the NO-cGMP signaling pathway, improving mitochondrial function, reducing oxidative stress, and alleviating cardiomyocyte hypertrophy47,48,49. Our study demonstrated that miR-214 significantly reduces cGMP levels by inhibiting PDE5A expression. Moreover, we found that the PDE5A inhibitor sildenafil partially reversed the effect of miR-214 inhibition on cGMP levels, further supporting the conclusion that miR-214 regulates cGMP through PDE5A.

In our study investigating the role of miR-214 in DCM, we found that the combination of miR-214 inhibition and sildenafil led to a more pronounced effect on cGMP levels in neonatal mouse cardiomyocytes compared to either treatment alone. This suggests that additional mechanisms, beyond the direct targeting of PDE5A by miR-214, may be involved. miR-214 is a key regulator of cardiac remodeling, apoptosis, and fibrosis50,51, all of which are central to the pathogenesis of DCM. Our dual-luciferase reporter assays confirmed that miR-214 directly binds to the 3’UTR of PDE5A, thereby modulating its expression and influencing cGMP levels, a critical secondary messenger in cardiac function. However, the enhanced effect observed with the combined treatment suggests the possibility of compensatory upregulation of other PDEs, such as PDE2 and PDE3, which also degrade cGMP and may be upregulated when PDE5A activity is inhibited52,53. Additionally, the cross-talk between cAMP and cGMP signaling pathways in the heart could be influenced by miR-214, further affecting cGMP metabolism. miR-214’s regulatory role may extend beyond PDE5A, potentially targeting other proteins involved in critical cardiac processes, thus amplifying the effects of sildenafil. Off-target effects of sildenafil, particularly at higher concentrations, might also contribute to the observed synergistic effect. These findings highlight the complexity of miR-214’s role in DCM and underscore the need for further research to elucidate these additional mechanisms. Such investigations could reveal novel therapeutic targets for DCM, offering more effective treatment strategies, especially in cases where PDE5A regulation is crucial. The interplay between miR-214, PDE5A, and other signaling pathways in the heart represents a promising avenue for future research, with potential implications for developing therapies that target the underlying molecular mechanisms of DCM.

Our study elucidates the molecular mechanism by which miR-214 influences cGMP metabolism through the regulation of PDE5A in DCM. This finding not only deepens our understanding of DCM pathogenesis but also identifies new potential targets for therapeutic strategies based on miR-214 and PDE5A. However, several limitations persist. First, the sample size of our study was relatively small, necessitating further validation with larger clinical samples in future research. Second, our study primarily focused on the regulatory relationship between miR-214 and PDE5A; thus, future research should explore other miRNAs and target genes potentially involved in the pathological processes of DCM. Additionally, further studies employing animal models and clinical trials are essential to evaluate the practical application potential of therapeutic strategies based on miR-214 and PDE5A in DCM.

Conclusion

In conclusion, our study reveals a new mechanism by which miR-214 influences the progression of DCM by regulating cGMP metabolism through PDE5A. This provides new potential targets for the diagnosis and treatment of DCM. Future research based on this finding holds the promise of bringing new therapeutic hope to patients with DCM.

Data availability

The datasets extracted and analyzed during the current study are available in the GEO database under accession numbers GSE53081 and GSE141910.