Lipidomic profiling reveals medium-chain acylcarnitines alterations as metabolic signatures in IgA nephropathy

Abstract

Immunoglobulin A nephropathy (IgAN), the most common primary glomerulonephritis worldwide, exhibits highly variable clinical progression, ranging from asymptomatic urinary abnormalities with potential for spontaneous resolution to progressive kidney failure over 10–20 years. The limited understanding of IgAN pathogenesis continues to constrain the development of effective therapeutic strategies. To elucidate key pathogenic mechanisms, we performed both untargeted and quantitative lipidomic profiling in patients with IgAN. Our results revealed significant disruptions in carnitine metabolism, particularly involving medium-chain acylcarnitines. Notably, medium-chain acylcarnitine levels were strongly correlated with clinical markers of renal function, including serum creatinine and estimated glomerular filtration rate (eGFR). Given their role as key intermediates in mitochondrial fatty acid β-oxidation, we further investigated mitochondrial function. Analysis of transcriptomic data (GSE210098) showed that genes related to the mitochondrial respiratory electron transport chain and oxidative phosphorylation (OXPHOS) system were significantly downregulated in IgAN progressors. Moreover, OXPHOS activity was markedly impaired in renal tubular epithelial cells from IgAN patients (GSE171314). These findings implicate dysregulated medium-chain acylcarnitine metabolism as a potential contributor to IgAN pathogenesis. Importantly, our data suggest a protective role for carnitine in disease development, highlighting a potential novel therapeutic avenue for IgAN.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12882-026-04822-3.

Introduction

Immunoglobulin A nephropathy (IgAN) is the most common primary glomerular disease worldwide, with an estimated incidence of at least 2.5 per 100,000 individuals. Disease progression of IgAN varied greatly, ranging from asymptomatic urinary abnormalities with the potential for spontaneous resolution to gradually progressive kidney failure within 10 to 20 years [1–3]. Although IgAN has been first described more than 50 years ago [4], our understanding of the disease pathophysiology is still deemed insufficient partly result from limited animal models that mimic the human disease. The widely accepted pathogenesis of IgAN is known as the “multi-hit hypothesis”: (1) patients with IgAN have increased circulating levels of galactose-deficient IgA1 (Gd-IgA1); (2) the galactose-deficient hinge region acts as a neoepitope to promote the formation of autoantibodies; (3) circulating immune complexes form containing Gd-IgA1, autoantibodies and complement C3; (4) immune complexes deposit in the mesangium and activate inflammatory and cellular proliferative signaling cascades [1]. Multi-hit hypothesis brought us a preliminary understanding of the pathogenesis of IgA nephropathy, further research on the key nodes of IgA nephropathy is crucial for the development of clinical therapeutic drugs.

Lipids are essential metabolites that serve key cellular functions, including maintenance of cellular barriers, involvement in energy metabolism, and participation in cell signaling [5]. Dyslipidemia is a hallmark of CKD, and its severity not only correlates with advancing CKD stages but is also associated with increased cardiovascular risk and mortality [6]. Research on dyslipidemia in CKD remains an evolving area, and further experimental validation of current findings is needed. Lipidomics, the comprehensive analysis of individual lipids within a biological system, enables the profiling of a wide range of lipid species. Compared to untargeted metabolomics, lipidomics offers greater effectiveness, sensitivity, and specificity. This approach has been widely applied in lipid biochemistry, clinical biomarker discovery, and disease diagnosis [7–9]. In CKD, lipidomic signatures have shown potential in identifying risk factors for disease progression, and the alterations in lipid metabolism are believed to play a significant role in disease pathogenesis. Specifically, research on lipidomic profiling in IgAN has highlighted disruptions in carnitine metabolism, including the accumulation of medium-chain acylcarnitines, which may provide valuable insights into the underlying mechanisms of IgAN. Although studies like those by Deng et al. have demonstrated a prospective association between circulating lipids and treatment outcomes in IgAN [10]. Nevertheless, significant gaps remain in the characterization of the serum lipid profile in patients with IgAN.

Carnitine, first discovered in muscle, is a low molecular weight compound widely distributing in all organs of mammals [11, 12]. In physiological state, carnitine is involved in many important biochemical reactions including but not limited to fatty acid oxidation [13]. In some organs which regard fatty acids as a crucial adjective source of energy, including skeletal and heart, deficiency of carnitine may cause life-threatening consequences [13, 14]. Kidney, as another carnitine rich site, is an important occasion not only for carnitine related metabolism but also for homeostasis of carnitine [11]. It was also reported that circadian clock in the renal tubule strongly affects several key metabolic pathways in the kidney, including NAD⁺ synthesis, β-oxidation of fatty acids, mitochondrial activity and carnitine handling, particularly [15]. Regrettably, what we know about the relationship between carnitine and renal diseases is very limited.

In this study, we performed targeted serum lipidomics and quantitative lipidomic analysis on patients with IgAN to explore the role of lipid metabolism in the disease. Our findings revealed significant alterations in carnitine metabolism in IgAN, particularly in medium-chain acylcarnitines, which showed a strong correlation with clinical parameters such as serum creatinine and estimated glomerular filtration rate (eGFR). These results suggest that changes in medium-chain acylcarnitines may play a pivotal role in the pathogenesis of IgAN. Importantly, we identified a protective role of carnitine in the pathogenesis of IgAN, offering a novel therapeutic strategy for the treatment of the disease.

Materials and methods

Subjects

The research workflow is shown in Fig. 1. This study was reviewed and approved by the Ethics Committee of Xiangya Hospital, Central South University (No. 202004194). All experiments were conducted in accordance with the principles of the Declaration of Helsinki. Informed consent was obtained from all participants in this study. We collected serum samples from 120 individuals, including 85 patients with IgAN and 35 age-, gender-, and BMI-matched healthy controls. Inclusion criteria included (1) aged 18 to 50 years, (2) biopsy-proven IgAN, and (3) proteinuria greater than 1 g/d. Major exclusion criteria included secondary, familial, crescentic IgAN, and presence of other CKD. The demographic characteristics of the participants are summarized in Table 1. The samples were randomly divided into two groups: the discovery cohort, which included 50 IgAN patients and 20 controls, and the validation cohort, which comprised 35 IgAN patients and 15 controls. The demographic characteristics of the IgAN participants in discovery and validation cohort are summarized in Supplementary Table 1.

Fig. 1.

Schematic diagram of the workflow of the present study

Table 1.

Demographics of IgAN patients and healthy control subjects

Parameters	HC (n = 35)	IgAN (n = 85)	P value#
Age (years)	33.46 ± 9.24	31.26 ± 10.01	0.2659
Gender (male, n (%))	16 (45.7%)	39 (45.9%)	1.000
Weight (kg)	64.80 ± 12.80	65.39 ± 13.56	0.8262
Serum creatinine (µmol/L)	68.00 (56.26, 74.66)	87.00(69.40, 121.10)	0.000
eGFR(mL/min/1.73 m2)	116.20 (109.60, 122.70)	92.60 (58.19, 116.30)	0.000
24 h proteinuria (g)	-	2.31 (1.46, 4.07)	-

#Comparisons between the healthy control group and the IgAN group. eGFR, estimated glomerular filtration rate. Bold values indicate P < 0.05

Data collection

The bulk RNA-Seq dataset GSE210098 was accessed from the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/geo/), which includes kidney biopsies from 15 adult IgAN progressors and 21 IgAN patients with clinical remission (non-progressors) [16]. The single-cell RNA-Seq dataset GSE171314 includes kidney biopsies from 4 IgAN patients and 1 control subject [17]. We used the R package “Seurat” (version 4.0.2) to analyze the single-cell data, as described in previous studies [18, 19]. The UMAP algorithm was applied to explore and visualize cluster classifications across cell samples.

Untargeted lipidomic profiling and quantitative lipidomic profiling

Metabolite extraction was primarily performed following previously reported methods [19]. Briefly, 50 µL of each sample was mixed with 1 mL of lipid extraction solvent, followed by vortexing and sonication. Then, 200 µL of water was added, and the mixture was vortexed again. The samples were centrifuged at 12,000 rpm for 10 min at 4 °C. A 200 µL aliquot of the supernatant was transferred to a centrifuge tube and concentrated. The residue was reconstituted in 200 µL of lipid resuspension solution for subsequent LC-MS/MS analysis.

The data acquisition system consisted of Ultra Performance Liquid Chromatography (UPLC) using an ExionLC™ AD and Tandem Mass Spectrometry (MS/MS) using a QTRAP^® 6500+. Chromatographic separation of all samples was performed using a Thermo Accucore™ C30 column. The UPLC system employed a gradient elution program with acetonitrile/water (solvent A) and acetonitrile/isopropanol (solvent B). The gradient was set as follows: 0 min, A/B = 80:20 (v/v); 2 min, A/B = 70:30 (v/v); 4 min, A/B = 40:60 (v/v); 9 min, A/B = 15:85 (v/v); 14 min, A/B = 10:90 (v/v); 15.5 min, A/B = 5:95 (v/v); 17.3 min, A/B = 5:95 (v/v); 17.5 min, A/B = 80:20 (v/v); and 20 min, A/B = 80:20 (v/v). The flow rate was 0.35 mL/min, the column temperature was maintained at 45 °C, and the injection volume was 2 µL. Mass spectrometry analysis was conducted in both positive and negative ion modes, with a mass spectrometer voltage of 5500 V in positive ion mode and − 4500 V in negative ion mode. The ion source settings were as follows: gas 1 (GS1) at 45 psi, gas 2 (GS2) at 55 psi, and curtain gas (CUR) at 35 psi. Each ion pair was scanned and detected in the triple quadrupole using optimized declustering potential (DP) and collision energy (CE).

Qualitative analysis was performed based on the MWDB (Metware Biotechnology Co., Ltd., Wuhan, China) in-house database by matching retention time (RT) and parent/daughter ion pair information. Lipid quantification was completed using the multiple reaction monitoring (MRM) mode of the triple quadrupole mass spectrometer.

Raw data were imported into Analyst 1.6.3 software for automated data processing. The data processing and analysis workflow included retention time correction, experimental design setup, peak selection, normalization, deconvolution, and compound identification.

Differential metabolites selected

Based on the results of OPLS-DA, metabolites that exhibit significant differences between different varieties or tissues can be initially screened from the variable importance in projection (VIP) derived from the multivariate analysis model. Additionally, univariate analysis, including p-value or fold change, can be used to further refine the selection of differential metabolites. Lipids with VIP > 1 and p-value < 0.05 are identified as significantly differential lipids.

Statistical analysis

The statistical analyses were performed using R software (version 4.0.4) or GraphPad Prism (version 8.0). Gene expression levels were compared using Student’s t-test. p < 0.05 was considered statistically significant. *p < 0.05; **p < 0.01; ***p < 0.001.

Results

General lipid composition of the plasma of IgAN patients

The characteristics of all subjects are shown in Table 1 and Supplementary Table 1. Initially, we analyzed the discovery cohort, revealing that serum creatinine levels in IgAN patients were significantly higher than those in the control group, while the eGFR levels were significantly lower in the IgAN group compared to the control group (Fig. 2A). Subsequently, we performed a targeted serum lipidomics analysis in the discovery cohort, identifying 788 lipid metabolites in total. Among them, 361 were glycerophospholipids, 222 were glycerolipids, 90 were sphingolipids, 86 were fatty acyls, 27 were sterolipids, and 2 were prenol lipids. OPLS-DA models based on PCA were used to distinguish significant differences between the control and IgAN groups. As shown in Fig. 2D, samples from each group were clearly separated along different sides of the plot, indicating distinct metabolic profiles. The grouping sample line lies below the random sampling line, and the Q2 values of permutation tests were all < 0.05 (Fig. 2D). Therefore, these OPLS-DA models demonstrate clear reliability in identifying characteristic metabolite biomarkers.

Fig. 2.

General lipid composition of the plasma of IgAN patients. (A) Serum creatinine and (B) eGFR from the discovery cohort were determined and quantified. (C) Classification of identified lipids. (D) OPLS-DA of serum lipids data for IgAN patients and controls from the discovery cohort. (E) Differential lipids in IgAN patients and controls from the discovery cohort. (F) Heatmap of differential lipids quantified in human serum. (G) The relative level of differential acylcarnitine in IgAN patients and controls from the discovery cohort. Results are expressed as mean ± SEM. **P < 0.01; ***P < 0.001

Using VIP > 1 and p-value < 0.05 as thresholds, a total of 63 differential lipid metabolites were identified. These metabolites were categorized as follows: 26 triglycerides (TG), 15 phosphatidylethanolamines (PE), 12 acylcarnitines (CAR), 4 diglycerides (DG), 3 ceramides (Cer), and the remaining were lysophosphatidylethanolamines (LPE), oxidized lipids (Eicosanoids), and hexosylceramides (HexCer) (Fig. 2E). Among these 63 differential lipids, 49 were upregulated, while 14 were downregulated. Interestingly, nearly all of the downregulated lipids were acylcarnitine species (Fig. 2E-F). KEGG pathway enrichment analysis of the differential lipids revealed significant enrichment in carnitine-related pathways (Supplementary Fig. 1). The 11 acylcarnitines that were significantly downregulated in IgAN patients were C8:0-carnitine, C9:0-carnitine, C10:0-carnitine, C12:0-carnitine, C13:0-carnitine, C8:1-OH-carnitine, C10:1-carnitine, C10:1-OH-carnitine, C12:1-carnitine, C12:1-OH-carnitine, and C14:2-carnitine (Fig. 2G). These results suggest that medium-chain acylcarnitines are significantly downregulated in IgAN patients in the discovery cohort.

Medium-chain acylcarnitines associated with the progression of IgAN

Subsequently, we performed a correlation analysis between the levels of these medium-chain acylcarnitines and the clinical information of IgAN patients. The results showed that serum levels of C8:0-carnitine, C9:0-carnitine, C10:0-carnitine, C12:0-carnitine, C13:0-carnitine, C8:1-OH-carnitine, C10:1-carnitine, C10:1-OH-carnitine, C12:1-carnitine, C12:1-OH-carnitine, and C14:2-carnitine were significantly negatively correlated with eGFR levels and significantly positively correlated with serum creatinine levels (Fig. 3). These results suggest that medium-chain acylcarnitines related to clinical indicators could serve as potential biomarkers for the disease.

Fig. 3.

Medium-chain acylcarnitines associated with the progression of IgAN. The correlation of (A) serum creatinine or (B) eGFR and C8:0-carnitine, C9:0-carnitine, C10:0-carnitine, C12:0-carnitine, C13:0-carnitine, C8:1-OH-carnitine, C10:1-carnitine, C10:1-OH-carnitine, C12:1-carnitine, C12:1-OH-carnitine, and C14:2-carnitin

Validation of the association of medium-chain acylcarnitines in IgAN

To further clarify the relationship between medium-chain acylcarnitines and IgAN, we conducted quantitative lipidomics analyses to determine the specific levels of medium-chain acylcarnitines in IgAN. Analysis of the validation cohort revealed that, consistent with the previous results, the majority of medium-chain acylcarnitines were significantly downregulated in IgAN patients compared to the control group (Fig. 4).

Fig. 4.

Heatmap of short, medium, and long chain acylcarnitines quantified in IgAN patients and controls from the validation cohort

Correlation analysis revealed that C9:0-carnitine, C13:0-carnitine, C12-OH-carnitine, C8:1-OH-carnitine, C12:1-OH-carnitine, and C10:2-carnitine were significantly negatively correlated with eGFR levels or significantly positively correlated with serum creatinine levels in patients (Fig. 5A-B). These findings further validate the reliability of medium-chain acylcarnitines as potential clinical biomarkers for IgAN.

Fig. 5.

Validation of the association of medium-chain acylcarnitines in IgAN. The correlation of (A) serum creatinine or (B) eGFR and medium-chain acylcarnitines

Medium-chain acylcarnitines deficiency may be associated with mitochondrial injury

As medium-chain acylcarnitines are key intermediates in the β-oxidation of fatty acids within the mitochondria, we next investigated changes in mitochondrial function in the kidney tissue of IgAN patients. Analysis of the transcriptional dataset (GSE210098) from kidney biopsies of IgAN patients showed that, compared to clinical remission (non-progressors) patients, genes related to respiratory electron transport and the electron transport chain oxidative phosphorylation (OxPhos) system in mitochondria were significantly downregulated in the IgAN progressor group (Fig. 6A-B). Single-cell transcriptomic analysis (GSE171314) of IgAN patient kidney tissue revealed that energy metabolism pathways, including OxPhos, were highly enriched in the proximal and distal tubular epithelial cells (such as CD, DT, PT) of normal kidneys, but these pathways were significantly downregulated in the tubular epithelial cells of IgAN patients (Fig. 6C-D). These results suggest that the deficiency of medium-chain acylcarnitines in IgAN patients may be associated with mitochondrial injury.

Fig. 6.

Medium-chain acylcarnitines deficiency may be associated with mitochondrial injury. (A) The gene set enrichment analysis (GSEA) for the term of “Respiratory Electron Transport” and “Electron Transport Chain Oxphos System In Mitochondria” on the GSE210098 transcriptional dataset. (B) Heatmap of respiratory electron transport genes of GSE210098 transcriptional dataset. (C) Composition and distribution of single cells from GSE171314. (D) Energy metabolism pathways were highly enriched in the proximal and distal tubular epithelial cells of normal kidneys

Discussion

In this study, we performed serum lipidomics analysis to identify significant differences between healthy controls and patients with IgAN. We found that medium-chain acylcarnitine levels were markedly reduced in IgAN patients and were associated with disease progression. Mechanistically, this downregulation may be linked to mitochondrial dysfunction. These findings underscore the potential role of medium-chain acylcarnitines as key regulators of mitochondrial function and highlight their promise as candidate biomarkers for IgAN.

Mitochondria are heterogeneous and highly dynamic organelles essential for adenosine triphosphate (ATP) production, metabolic regulation, reactive oxygen species (ROS) generation, as well as cell differentiation and death [20–24]. Among human organs, the kidney ranks second only to the heart in mitochondrial content and oxygen consumption, reflecting its high metabolic demands. As a result, the fundamental cellular functions of the kidney are heavily reliant on intact mitochondrial activity. Under pathological conditions, mitochondrial dysfunction can result in impaired ATP synthesis, excessive ROS accumulation, and induction of apoptosis. These perturbations collectively contribute to the onset and progression of various renal diseases, including focal segmental glomerulosclerosis [25, 26], acute kidney injury [27, 28], and CKD [29–31].

The immunopathogenesis of IgAN is widely recognized as a multi-hit process involving sequential pathogenic events [1, 2]. Elevated levels of underglycosylated IgA1 trigger the production of autoantibodies and the formation of circulating immune complexes. These complexes activate the complement cascade, leading to mesangial cell activation and proliferation. The ensuing glomerular inflammation results in podocyte injury, proteinuria, tubulointerstitial inflammation, and progressive fibrosis [32–34]. Emerging evidence indicates that mitochondrial dysfunction may be intimately involved in this pathogenic cascade. Mitochondrial DNA (mtDNA), a surrogate marker of mitochondrial integrity, is released into the cytosol and subsequently into systemic circulation following mitochondrial injury [35, 36]. Additionally, renal mitochondrial damage can lead to the excretion of mtDNA fragments into the urine [37, 38]. Elevated urinary mtDNA levels have been reported in IgAN patients and are positively correlated with disease severity [39]. Conversely, Liu et al. demonstrated that lower mtDNA copy numbers in peripheral blood were associated with better renal function and milder histopathological injury in IgAN [40]. Collectively, these findings suggest that systemic mitochondrial dysfunction may not only reflect disease burden but also play an active role in the pathogenesis of IgAN.

Our study identified mitochondrial injury in patients with IgAN, which may be associated with a deficiency of medium-chain acylcarnitines. Acylcarnitines are indispensable for mitochondrial fatty acid β-oxidation and overall energy homeostasis [41–43]. In addition to their central role in fatty acid metabolism, they contribute to the protection of cellular membranes, prevent intracellular lipid accumulation, modulate ketogenesis and gluconeogenesis, and facilitate the elimination of toxic metabolic intermediates. Under physiological conditions, acylcarnitines are essential for transporting long-chain fatty acids (LCFAs) across the inner mitochondrial membrane, which is impermeable to fatty acids with carbon chains longer than 12 carbons. They are the exclusive carriers that enable the import of fatty acids into the mitochondrial matrix for subsequent β-oxidation. Therefore, a deficiency in acylcarnitines suggests impaired fatty acid oxidation.

Such impairment can lead to structural and functional disruptions in mesangial cells, podocytes, and proximal tubule epithelial cells—cell types integral to nephron function [44, 45]. Among these, proximal tubular cells—the most abundant cell type in the kidney—serve as major sites for renal lipid handling [46, 47]. Accumulation of fatty acids in these cells promotes tubular damage and contributes to the progression of interstitial fibrosis. Genome-wide transcriptomic analyses in both murine and human models of renal interstitial fibrosis have demonstrated downregulation of key mitochondrial enzymes and regulators of fatty acid oxidation, including PPARα and PPARγ, resulting in excessive intracellular lipid accumulation [48]. Moreover, the PPAR signaling pathway has been implicated in the pathogenesis of IgAN, potentially promoting disease progression through the induction of ferroptosis in mesangial cells [49]. However, direct evidence linking renal lipid accumulation to IgAN remains limited. Based on our findings, we propose that acylcarnitine deficiency may represent a previously underappreciated contributor to disrupted lipid metabolism in IgAN.

Acylcarnitine deficiency has been reported in a variety of pathological conditions, including insulin resistance [50], endocrine disorders [51], and hepatic [52] as well as renal diseases [53, 54]. In our study, serum levels of acylcarnitines, particularly medium-chain species, were significantly decreased in IgAN patients and were associated with disease progression. Medium-chain acylcarnitines have also been identified as biomarkers in other pathological contexts. For instance, reduced levels of short- and medium-chain acylcarnitines have been observed in HIV-infected individuals and were correlated with increased susceptibility to infection [55]. Similarly, significantly lower blood concentrations of medium-chain acylcarnitines have been reported in low-birth-weight infants of both sexes [56]. Disruption of acylcarnitine homeostasis is also frequently observed in patients with advanced CKD undergoing hemodialysis [53, 54].

Clinically, acylcarnitine deficiency has been associated with a spectrum of symptoms, including muscle weakness, cramps, fatigue, erythropoietin-resistant anemia, and cardiac dysfunction, all of which are closely linked to reduced quality of life in CKD patients [53, 54, 57]. Notably, in our cohort, acylcarnitine deficiency was already detectable in early-stage IgAN patients exhibiting progressive disease trajectories, underscoring its potential utility as an early prognostic biomarker. This metabolic alteration may represent a novel disease hallmark in IgAN. Plasma acylcarnitines has also been reported to be associated with prognosis and treatment responses in patients with IgAN [58]. However, the underlying mechanisms driving acylcarnitine depletion, as well as its broader pathophysiological implications, remain to be fully elucidated. Furthermore, given the relatively short follow-up duration in our study, longitudinal validation in larger, independent cohorts is warranted to confirm the prognostic relevance of serum acylcarnitines in IgAN.

IgAN is the most common biopsy-proven primary glomerulonephritis, though its geographic prevalence varies. In East Asia, the male-to-female ratio of IgAN is about 1:1 or less than 2:1, which differs from the ratio of up to 6:1 reported in Europe and the United States [59–61]. This strongly indicates that environmental and/or genetic factors may have certain roles in the pathogenesis of IgAN. Our study mainly included Chinese IgAN patients. The role of acylcarnitine deficiency in IgAN patients of other ethnicities needs to be supported by more data.

Conclusion

In summary, untargeted and quantitative lipidomic profiling were employed to elucidate key pathogenic mechanisms underlying IgAN. Our analysis revealed significant dysregulation of medium-chain acylcarnitines in IgAN, potentially linked to mitochondrial injury. These findings offer new insights into the role of carnitine metabolism in the pathogenesis of IgAN.

Supplementary Information

Below is the link to the electronic supplementary material.

Author contributions

Jiawei Cheng: Conceptualization, Methodology, Investigation, Resources, Writing - Original Draft. Yuanyuan Han: Validation, Formal analysis, Visualization. Yan Zhang: Methodology, Investigation, Data Curation. Xin He: Methodology, Investigation, Data Curation. Sijue Zou: Methodology, Investigation, Data Curation. Wenzhe She: Methodology, Investigation, Data Curation, Funding acquisition. Zhangzhe Peng: Conceptualization, Project administration. Ling Huang: Supervision, Project administration, Writing - Review & Editing. Hao Huang: Conceptualization, Software, Writing - Review & Editing, Supervision, Project administration, Funding acquisition.

Funding

This study was supported by the Project funded by National Natural Science Foundation of China (82300787); the Natural Science Foundation of Hunan province (2025JJ50519); and the Fundamental Research Funds for the Central Universities of Central South University (2025ZZTS0172).

Data availability

All data generated or analysed during this study are included in this published article.

Declarations

Ethical approval

The Review Board of the Xiangya Hospital of Central South University approved the present study (No. 202004194). Informed consent was obtained from all participants in this study. This study had been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki and its later amendments.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.