LncRNA GAS6-AS1 affects doxorubicin‑induced cardiomyocyte injury in H9c2 cells by regulating miR‑152‑3p: a cellular model relevant to heart failure

Abstract

Objective

To explore the diagnostic significance of LncRNA GAS6-AS1 in chronic heart failure (CHF) and its molecular mechanism of influencing cardiomyocyte function through the regulation of miR-152-3p.

Methods

This study incorporated 126 patients diagnosed with CHF and 124 healthy control subjects. The serum level of GAS6-AS1 was determined via RT-qPCR, and its diagnostic efficacy was assessed through ROC curve. A cell injury model was established by stimulating H9c2 cells with DOX. CCK-8, flow cytometry and RT-qPCR were used to analyze the effects of these on cell viability, apoptosis and inflammatory factors (TNF-α, IL-6, IL-1β). The targeting relationship between GAS6-AS1 and miR-152-3p was verified by DLR.

Results

The expression of GAS6-AS1 in the serum of CHF patients was significantly downregulated. ROC analysis showed that the sensitivity of GAS6-AS1 in the diagnosis of CHF was 84.90%, the specificity was 77.40%, and the AUC was 0.888 (95% CI: 0.849–0.928). Overexpression of GAS6-AS1 could mitigate the reduction in cell viability and the elevation in apoptosis induced by DOX in H9c2 cells, and inhibit the expression of pro-inflammatory factors. GAS6-AS1 could directly bind to miR-152-3p. Overexpression of miR-152-3p could reverse the inhibitory effect of GAS6-AS1 on cell apoptosis and the release of inflammatory factors.

Conclusion

GAS6-AS1 can act as a potential diagnostic biomarker for CHF. It might accomplish this by targeting and suppressing miR-152-3p, thus decreasing cardiomyocyte apoptosis and inflammatory reactions.

Supplementary Information

The online version contains supplementary material available at https://doi.org/10.1186/s13019-026-04524-9.

Introduction

Heart failure (HF) is caused by cardiac injury. This injury triggers structural and functional changes in the myocardium, which in turn impair ventricular systolic or diastolic function [1–3]. Doxorubicin (DOX), a commonly used chemotherapy drug, causes dose-dependent cardiotoxicity that ultimately leads to heart failure. Epidemiological data shows that more than 64 million people worldwide are affected by HF [1].

Chronic heart failure (CHF) is a persistent condition characterized by impaired cardiac function. It disrupts patients’ normal daily activities and quality of life, and places a heavy public and economic burden [4–6] on healthcare systems and families. Therefore, timely diagnosis and therapeutic management are essential to slow or prevent pathological cardiac remodeling, reduce hospitalization rates, and improve long-term survival outcomes for heart failure patients.

Although long non-coding RNAs (LncRNAs) are not directly engaged in protein coding, they play a crucial role in the regulation of gene expression via transcriptional regulation and miRNA sponge action [7]. DOX-induced cardiomyocyte injury in H9c2 cells has been extensively employed as an in vitro model for researching heart failure, as it replicates phenomena such as oxidative stress, apoptosis, and contractile dysfunction observed in the failing heart [8, 9]. Meanwhile, the lncRNA GAS6-AS1 has been proven to play a significant part in apoptosis, metabolic regulation, and inflammatory responses [10–14]. Previous research has indicated that GAS6-AS1 is significantly under-expressed in patients with acute myocardial infarction [15], and is also significantly down-regulated in myocardial fibrosis subsequent to acute myocardial infarction [16]. Myocardial fibrosis after acute myocardial infarction ultimately results in heart failure as well. This implies that GAS6-AS1 has a potential regulatory function in the development of heart failure, yet the specific regulatory mechanisms require further exploration.

The miR-152-3p is involved in multiple biological processes, such as tumorigenesis [17, 18], metabolic regulation [19, 20], and immune responses [21]. According to previous reports, certain studies have been conducted on the influence of miR-152-3p on heart failure. For instance, research shows that miR-152-3p is significantly over-expressed in individuals with advanced CHF [22]. Its expression is elevated in the cardiomyocytes and myocardial tissue of infarcted mice [23]. Thus, miR-152-3p exert have potential pathological effects in heart failure, yet the specific mechanism of action requires further validation.

Based on the aforementioned research background, it can be inferred that both GAS6-AS1 and miR-152-3p may be involved in the pathogenesis of CHF. A targeting relationship between the two was identified in the bioinformatics database. Building upon the above research, it is hypothesized that GAS6-AS1 may regulate the expression of miR-152-3p, thereby influencing cardiomyocyte function and impacting the pathological process of CHF. To validate this hypothesis, the present study was conducted.

Materials and methods

Inclusion of patients and sample collection

In this study, a case-control design was employed. A total of 126 patients with CHF who received treatment at The First Hospital of Lanzhou University from June 2022 to July 2023 were included as the observation group. Simultaneously, 124 age-matched healthy individuals from concurrent medical examinations were selected as the control group. Eligibility for CHF inclusion required a left ventricular ejection fraction of less than 50% and concurrent classification as New York Heart Association (NYHA) functional class II-IV. The exclusion criteria encompassed the presence of malignant tumors, active infections, major surgical operations within the past 2 months, installation of cardiac assist devices, and severe liver and kidney dysfunction. Venous blood samples were collected from all study participants in the morning under fasting conditions. Immediately after collection, the samples were subjected to low-temperature centrifugation (4 °C, 3000×g, 10 min) to achieve serum separation. The ultimately obtained serum components were aliquoted and frozen. They were stored in-20℃ biological sample bank for subsequent experimental analysis. The institutional review board of The First Hospital of Lanzhou University granted ethical approval for this investigation, with written informed consent obtained from all enrolled subjects.

Cell culture and induction

Rat cardiomyocytes H9c2 cells were obtained from ATCC (USA) and cultured in DMEM supplemented with 10% fetal bovine serum (Gibco) under standard conditions (37 °C, 5% CO₂/95% air, 95% humidity), with the culture medium replaced every 48 h.

Considering that the cardiotoxicity of DOX mimics numerous pathological characteristics of CHF, including cardiomyocyte apoptosis, hypertrophy, and disruptions in calcium ion metabolism [24, 25], and based on existing literature [26–28], this study employed 5 µmol/L DOX to treat H9c2 cells for 24 h to simulate cardiomyocyte damage induced by CHF. It is important to note that this in vitro model mainly reflects the direct damage to cardiomyocytes caused by drugs and does not fully represent the complex hemodynamic and neurohumoral changes associated with chronic heart failure. Nevertheless, since DOX-induced myocardial damage shares key pathways, such as apoptosis and inflammation, with CHF, this model is still extensively utilized for preliminary investigations of CHF-related mechanisms.

Cell transfected

H9c2 cells were seeded into 6-well plates and cultured until they reached 80% confluence. Subsequently, in accordance with the experimental design, the following recombinant plasmids and small molecule-RNAs were transfected: pcDNA3.1 negative control (NC), pcDNA3.1 GAS6-AS1, mimic negative control (miR-NC), and miR-152-3p mimic (miR-mimic). The transfection procedure was performed in a proportional manner using the jetPRIME^® transfection reagent (Polyplus). Cells were harvested 48 h post-transfection for subsequent functional assays.

Real-time quantitative reverse transcription PCR (RT-qPCR)

Total RNA was extracted from serum and cells via the TRIzol^® method (Thermo). Regarding serum samples, 750 µL of TRIzol LS reagent was added to 250 µL of serum. The mixture was vortexed for 10–20 s and allowed to stand at room temperature for 10 min. Subsequently, 150 µL of chloroform was added, followed by vortexing and a 5-minute standing period. After centrifugation, the upper aqueous phase was collected, and an equal volume of isopropanol was added. The mixture was gently mixed, and the precipitate was allowed to form overnight. The precipitate, which was the total RNA, was obtained through centrifugation.

For cell samples, 1 mL of TRIzol was directly added to a 6-well plate to lyse the cells. The lysate was then transferred to a 1.5 mL RNAse-free centrifuge tube. After adding 200 µL of chloroform, the same procedure as described above was followed. The purity and concentration of the extracted RNA were measured using a NanoDrop2000 spectrophotometer.

Total RNA served as a template for cDNA synthesis via reverse transcription in accordance with the PrimeScriptRT kit (Takara) protocol. Gene expression analysis was carried out on an AppliedBiosystems7500 real-time quantitative PCR system, with SYBR Green I premix as the fluorescent marker. The reaction system was constructed in strict accordance with the reagent instructions. GAPDH and U6 were employed as endogenous controls respectively. Relative expression was calculated using the 2^−∆∆Ct method.

Western blot

Total cellular protein was extracted from H9c2 cells, and the protein concentration was quantified using a BCA protein assay kit (Thermo Fisher Scientific, USA). Equivalent quantities of protein (30 µg per lane) were resolved by 10–12% SDS-PAGE and then transferred onto a PVDF membrane (Millipore, USA). The membranes were blocked with 5% non-fat milk in Tris-buffered saline supplemented with 0.1% Tween-20 (TBST) for 1 h at ambient temperature, followed by an overnight incubation at 4 °C with primary antibodies directed against: Bax (1:1000, #2772, Cell Signaling Technology), Bcl-2 (1:1000, #4223, Cell Signaling Technology), Caspase-3 (1:1000, #9662, CST), and GAPDH (1:5000, #5174, Cell Signaling Technology) serving as a loading control. After three washes with TBST, the membranes were incubated at room temperature for 1 h with an HRP-conjugated goat anti-rabbit secondary antibody (1:5000, #7074, Cell Signaling Technology). Protein bands were visualized using an ECL substrate (Millipore) and imaged using a ChemiDoc™ imaging system (Bio-Rad, USA). The expression levels of the target proteins were normalized relative to GAPDH as an internal control.

CCK-8

H9c2 cell suspensions (5 × 10³ cells/well) were inoculated in 96-well plates with 100 µL of complete medium per well. After a 48-hour incubation at 37 °C in an atmosphere of 5% CO₂, 10 µL of a cell viability assay reagent (CCK-8; GLPBio) was introduced into each well, and the incubation was prolonged for 2 h under identical conditions. Ultimately, the optical density at 450 nm (OD₄₅₀) was determined using a microplate reader.

Flow cytometry

Collected cells (approximately 1 × 10⁶ cells) were subjected to two rounds of washing with ice-cold phosphate-buffered saline (PBS; Thermo), followed by centrifugation at 300×g for 5 min. The cells were then resuspended in 100 µL of Binding Buffer. Subsequently, 5 µL each of Annexin V-fluorescein isothiocyanate (FITC; BD) and propidium iodide (PI; Sigma) were added, and the mixture was incubated for 15 min at room temperature in the dark. Finally, the sample was diluted with 400 µL of the combined buffer, and the FITC and PI fluorescence signals were detected using a flow cytometer (Beckman Coulter CytoFLEX) within 1 h. Viable cells (Annexin V⁻/PI⁻), early apoptotic cells (Annexin V⁺/PI⁻), and late apoptotic/necrotic cells (Annexin V⁺/PI⁺) were distinguished via a two-parameter scatter plot. The total apoptosis rate was calculated as the percentage of early and late apoptotic cells.

Dual-luciferase reporter assay

The sequence of miR-152-3p binding sites was predicted via bioinformatics (situated in the 3’-UTR region of the GAS6-AS1 gene), and the fragment was directionally cloned into the pmirGLO vector (Promega, USA) to construct the wild-type recombinant plasmid GAS6-AS1-WT. The mutant plasmid GAS6-AS1-MUT was acquired through base substitution in the miR-152-3p seed region (2-8nt) using a targeted mutagenesis kit (Agilent). GAS6-AS1-WT or GAS6-AS1-MUT were co-transfected into H9c2 cells with miR-152-3p mimics, miR-152-3p inhibitor, and miR-NC, respectively. Lipofectamine3000 (ThermoFisher) was employed for transfection, and the cell density was optimized to reach 70%-80% confluence. At the 48-hour post-transfection time point, luminescent output was measured using an assay kit (Promega, USA).

Statistical analysis

The data analysis of this study was accomplished using GraphPad Prism 9.0 and SPSS 23.0. Inter-group statistical comparisons were carried out via the independent samples t-test and one-way ANOVA. The continuous variables in the experimental outcomes were presented as mean ± standard deviation, and the categorical variables were analyzed by means of the chi-square test. The threshold for statistical significance was set at P < 0.05.

Results

Comparison of clinical data between healthy control group and patients with CHF

A total of 126 patients with CHF and 124 healthy controls were included in this study. The comparison of baseline data indicated that there were no statistically significant differences (P > 0.05) between the two groups in the following variables: age (≥ 70 years old: 68% in the CHF group versus 64% in the healthy group), gender, smoking status, alcohol consumption, hypertension, and diabetes. Nevertheless, the B-type natriuretic peptide (BNP) level in the CHF group was significantly elevated, and the left ventricular ejection fraction (LVEF) level was significantly reduced (P < < 0.001, Table 1).

Table 1.

Comparison of clinical data between healthy medical examiners and patients with CHF

Testing Indicators	Health (n = 124)	CHF (n = 126)	P
Age, years	70.70 ± 7.56	70.90 ± 6.52	0.827
Sex
Male	56	71	0.077
Female	68	55
Smoking
NO	61	57	0.531
YES	63	69
Drinking
NO	50	57	0.432
YES	74	69
Hypertensive
NO	53	67	0.099
YES	71	59
Diabetes
NO	62	66	0.707
YES	62	60
BNP, pg/mL	53.84 ± 3.85	892.63 ± 380.48	0.000
LVEF (%)	57.65 ± 2.23	32.81 ± 5.90	0.000

CHF, Chronic heart failure; BNP, Type B natriuretic peptide; LVEF, Left ventricular ejection fraction

Expression and diagnostic value of GAS6-AS1

In comparison with healthy controls, patients with CHF exhibited a substantial down-regulation of GAS6-AS1 gene expression (Fig. 1A). ROC curve analysis further demonstrated that GAS6-AS1 had high sensitivity (84.90%) and specificity (77.40%) in differentiating CHF patients from healthy populations (AUC = 0.888, 95%CI = 0.849–0.928, P < 0.0001, Fig. 1B). The aforementioned results indicate that GAS6-AS1 has the potential as a diagnostic biomarker for CHF.

Fig. 1.

Expression and Diagnostic Significance of GAS6-AS1 in Chronic Heart Failure (CHF). A. Expression of GAS6-AS1 in the Serum of CHF Patients. B. Diagnostic Value of GAS6-AS1 in CHF. **** P < 0.0001

The influence of GAS6-AS1 level on cell activity and apoptosis

To validate the function of lncRNA GAS6-AS1 in DOX-induced cellular damage, GAS6-AS1 was overexpressed in vitro. The RT-QPCR results demonstrated that DOX treatment significantly downregulated the expression of GAS6-AS1, whereas overexpression of GAS6-AS1 notably elevated its expression level (P < 0.01, Fig. 2A). Simultaneously, DOX treatment significantly reduced cell viability (P < 0.01, Fig. 2B) and promoted cell apoptosis (P < 0.01, Fig. 2C-D). Overexpression of GAS6-AS1 effectively reversed the adverse effects induced by DOX, significantly restoring cell viability and suppressing cell apoptosis (P < 0.01, Fig. 2B-D). Further investigations revealed that the protective effect of GAS6-AS1 was associated with the regulation of the expression of apoptosis-related genes. Overexpression of GAS6-AS1 significantly upregulated the expression of the anti-apoptotic gene Bcl-2, while downregulating the expression of the pro-apoptotic genes Bax and Caspase-3 (P < 0.01, Fig. 2D). In summary, these findings suggest that lncRNA GAS6-AS1 can mitigate DOX-induced cellular damage.

Fig. 2.

Influence of GAS6-AS1 on Cell Viability and Apoptosis. A. Expression Levels of GAS6-AS1 in H9c2 Cells under Different Conditions. B. Effect of GAS6-AS1 on the Proliferation of H9c2 Cells. C. Effect of GAS6-AS1 on the Apoptosis of H9c2 Cells. D. Flow Cytometry Apoptosis Scatter Plot Demonstrating the Same Apoptosis Trend. E. Influence of GAS6-AS1 on the Expression Levels of Apoptotic Markers. ** P < 0.01, *** P < 0.001

Overexpression of GAS6-AS1 alleviates the inflammatory response induced by DOX

We conducted a further investigation into the impacts of lncRNA GAS6-AS1 on the expression of inflammatory factors and myocardial injury markers. As depicted in Fig. 3, the overexpression of GAS6-AS1 significantly suppressed the elevated expression of inflammatory factors TNF-α, IL-6, and IL-1β induced by DOX (P < < 0.0001, Fig. 3A-C). These findings suggest that GAS6-AS1 can mitigate the inflammatory response induced by DOX.

Fig. 3.

Overexpression of GAS6-AS1 mitigates the inflammatory response induced by DOX. (A) The impact of GAS6-AS1 on the expression levels of the inflammatory factor TNF-α. (B) The impact of GAS6-AS1 on the expression levels of the inflammatory factor IL-6. (C) The influence of GAS6-AS1 on the expression levels of inflammatory factors IL-1β. **** P < 0.0001

Targeted binding of GAS6-AS1 to miR-152-3p

We identified GAS6-AS1 as a downstream binding target of miR-152-3p from the ENCORI database and predicted the binding sequence of GAS6-AS1 to miR-152-3p (Fig. 4A). The dual-luciferase reporter assay confirmed that co-transfection of wild-type GAS6-AS1 and miR-152-3p mimics significantly suppressed luciferase activity, whereas no significant difference was noted upon transfection of the mutant GAS6-AS1 gene. After co-transfection of wild-type GAS6-AS1 and the miR-152-3p inhibitor, luciferase activity was significantly elevated, while no significant difference was observed when the mutant GAS6-AS1 gene was transfected (Fig. 4B). The expression level of miR-152-3p was significantly lower in patients CHF compared to that in healthy individuals (Fig. 4C). In CHF patients, the expressions of GAS6-AS1 and miR-152-3p showed a significant inverse correlation (Fig. 4D). These results suggest that targeted binding may occur between GAS6-AS1 and miR-152-3p.

Fig. 4.

Binding relationship between GAS6-AS1 and miR-152-3p. (A) The binding site between GAS6-AS1 and miR-152-3p predicted in the ENCORI database. (B) The dual-luciferase reporter assay validates the targeted binding between GAS6-AS1 and miR-152-3p. (C) Expression level of miR-152-3p in CHF patient serum. (D) Correlation between GAS6-AS1 and miR-152-3p expression levels. *** P < 0.001, **** P < 0.0001

GAS6-AS1 regulates the expression of genes related to cell viability and apoptosis by modulating miR-152-3p.

As depicted in Fig. 5, the overexpression of GAS6-AS1 notably suppressed the up-regulation of miR-152-3p induced by DOX (P < 0.0001, Fig. 5A). Subsequently, transfection with the miR-152-3p mimic significantly elevated the expression level of miR-152-3p. The miR-152-3p mimic counteracted the protective effect conferred by the overexpression of GAS6-AS1. This resulted in a decline in cell viability, an augmentation in the apoptosis rate, an increase in the expression of pro-apoptotic proteins Bax and cleaved Caspase-3, and a decrease in the expression of the anti-apoptotic protein Bcl-2 (P < 0.01, Fig. 5B-F). These results were also verified at the protein level. The overexpression of GAS6-AS1 decreased the levels of Caspase-3 and Bax while increasing the levels of Bcl-2. The miR-152-3p mimetic reversed this protective effect (P < 0.01, Fig. 5G-J). These findings confirm that the inhibition of miR-152-3p is the core mechanism of the anti-apoptotic effect of GAS6-AS1.

Fig. 5.

GAS6-AS1 modulates miR-152-3p to impact cell viability and apoptosis. (A) The expression levels of miR-152-3p under diverse treatment conditions. (B) The influence of miR-152-3p on cell proliferation. (C) The effect of miR-152-3p on cell apoptosis. D-F. The influence of miR-152-3p on the expression levels of apoptosis-related genes. G-J. The effect of miR-152-3p on the levels of apoptosis-related proteins. ** P < 0.01, *** P < 0.001 **** P < 0.0001

GAS6-AS1 regulates the expression of inflammatory factors by modulating miR-152-3p

To conduct a more in-depth exploration of the role of miR-152-3p in the inflammatory response, the expression of relevant genes was examined. The findings of RT-qPCR demonstrated that the miR-152-3p mimic could notably counterbalance the inhibitory impact of GAS6-AS1 on inflammatory factors (P < 0.05, Fig. 6A-C). These results suggest that GAS6-AS1 negatively modulates the inflammatory response through targeting and inhibiting miR-152-3p.

Fig. 6.

GAS6-AS1 regulates the expression of inflammatory factors through the modulation of miR-152-3p. A-C. The influence of miR-152-3p on the expression levels of inflammatory factors. * P < 0.05, ** P < 0.01, *** P < 0.001 **** P < 0.0001

Discussion

The morbidity and mortality of CHF show a continuous upward trend. Clinical data indicate that the readmission rate of patients with this disease within six months after discharge reaches as high as 20% or above [29]. Recent research has proposed that epigenetic regulatory molecules, typified by lncRNAs and miRNAs, have emerged as highly promising novel intervention targets in the realm of cardiovascular disease diagnosis and treatment [30, 31]. These findings disclose the diagnostic significance of GAS6-AS1 in CHF, as well as its molecular mechanism through which it regulates cardiomyocyte function by means of targeted regulation of miR-152-3p. It offers a new target for the treatment of CHF.

Long non-coding RNAs (lncRNAs) modulate pathological processes in CHFvia multiple pathways, characterized by tissue specificity and functional diversity. For instance, SNHG5 and ZFAS1 exhibit differential expression in CHF patients and have been employed as biomarkers indicative of the prevalence of malignant diseases in CHF patients [32]. The expression of TUG1 is correlated with the severity of CHF, serving as a dual diagnostic and prognostic biomarker, and it may be involved in the progression of CHF through inflammatory regulation [33]. SRA1 promotes the activation of cardiac muscle fibroblasts by negatively regulating miR-148b and participates in the process of myocardial fibrosis [34]. This study revealed that the expression of GAS6-AS1 in the serum of CHF patients was significantly reduced. Its diagnostic efficacy (AUC = 0.930) implies its potential as a novel non-invasive biomarker. This offers direct evidence for the application of GAS6-AS1 in the clinical auxiliary diagnosis of CHF. Functionally, we used DOX to induce myocardial cell damage to simulate the myocardial cell injury caused by CHF. Further experiments showed that overexpression of GAS6-AS1 could effectively reverse the cell damage induced by DOX. Specifically, GAS6-AS1 promoted cell proliferation, inhibited cell apoptosis, alleviated inflammatory responses. These results clearly demonstrated the positive role of GAS6-AS1 in anti-myocardial injury.

MiRNA modulates the progression of CHF by regulating cardiomyocyte apoptosis, fibrosis, inflammatory response, and metabolism [35, 36]. For instance, inhibiting the expression of miR-21-3p can alter the polarization state of macrophages and reduce excessive mitochondrial autophagy, thereby exerting a protective effect on the myocardium [37]. miR-214-3p may further contribute to the progression of CHF by participating in the rostral ventrolateral medulla (RVLM) inflammatory response [38]; abnormal increase in miR-320a-3p in CHF may aggravate the severity of CHF [39]. In this study, we initially demonstrated that GAS6-AS1 can directly bind to miR-152-3p and negatively regulate its expression. In CHF patients, the expression of miR-152-3p was significantly up-regulated. Further rescue experiments indicated that overexpression of miR-152-3p could reverse the protective effect of GAS6-AS1 on myocardial cell damage. These research findings confirmed that miR-152-3p is the core downstream mediator for the function of GAS6-AS1.

Nevertheless, it is necessary to recognize certain limitations of this study. The research was solely carried out in an in-vitro doxorubicin-induced injury model. Although this model is of great value for clarifying cell-autonomous mechanisms, it fails to comprehensively reflect the hemodynamic, neurohumoral, and systemic inflammatory features of chronic heart failure. Specifically, the clinical serum samples in this study were obtained from patients with chronic heart failure (CHF) of undetermined etiology, while the in-vitro model employed doxorubicin-treated cardiomyocytes, indicating a disparity in etiology. It is acknowledged that the optimal study population for this research would be patients who have undergone doxorubicin treatment and developed CHF, as this would guarantee the consistency between the etiology of the in-vitro model and that of the clinical samples. Consequently, the findings of this study necessitate further validation in animal models of heart failure (such as transverse aortic constriction and myocardial infarction models) and in clinical samples from patients with doxorubicin-associated CHF.

In conclusion, this study not only uncovered the potential of GAS6-AS1 as a diagnostic biomarker for CHF, but also elucidated the crucial role of the GAS6-AS1/miR-152-3p axis in regulating cardiomyocyte function at the experimental level. Our findings offer a novel perspective for comprehending the molecular mechanism of CHF and imply that targeting the GAS6-AS1/miR-152-3p pathway may emerge as a potential new strategy for the treatment of CHF in the future. Given that this study employed a doxorubicin (DOX)-induced cellular injury model, the aforementioned conclusions necessitate validation in patients with doxorubicin-induced CHF and other heart failure models.

Author contributions

Author Contributions: Study concept and design: M. L., N. L., L.X. Y., and J.P. W.; analysis and interpretation of data: M. L., and N. L.; drafting of the manuscript: N. L.; critical revision of the manuscript for important intellectual content: J.P. W.; statistical analysis: M. L., and N. L.

Funding

This study was funded by Uygur Medicine Hospital of Xinjiang Uygur Autonomous Region “Analysis of the Efficacy of Uighur Medicine Fennel Extract Combined with Sacubitril-Valsartan in the Treatment of Heart Failure in Patients with Coronary Heart Disease”(TSYC202301B118).

Data availability

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

This research was conducted following the Declaration of Helsinki. The institutional review board at The First Hospital of Lanzhou University granted ethical approval for this investigation, with written informed consent obtained from all enrolled subjects.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.