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Inhibition of adenosine/A2A receptor signaling suppresses dermal fibrosis by enhancing fatty acid oxidation

Cell Communication and Signaling volume 23, Article number: 206 (2025) Cite this article

Abstract

Background

Skin fibrosis presents a major challenge for clinicians treating conditions like systemic sclerosis (SSc) due to its progressive course and limited treatment options. While the role of metabolism in fibrosis has gained increasing attention, the crucial alterations of metabolic pathway and the underlying signaling of metabolic interconnections in regulating SSc-related skin fibrosis remain largely elusive.

Methods

Metabolomic analysis was performed on plasma samples from 35 SSc patients to identify metabolic alterations. In bleomycin (BLM)- and hypochlorous acid (HOCL)-induced skin fibrosis mouse models, we assessed the impact of global A2a receptor knockout on skin fibrosis. Single-cell RNA sequencing of mouse skin was utilized to investigate the role of A2A in fibroblasts during fibrotic challenge. Human dermal fibroblasts were used in in vitro experiments, employing RNA sequencing and Seahorse assays, to assess the relationship between A2A signaling and fatty acid oxidation (FAO). Finally, fibroblast-specific conditional A2a knockout mice were used to test the effects of specifically targeting A2A in dermal fibroblasts.

Results

Adenosine-centered nucleotide metabolism was elevated in the plasma of SSc patients. Mechanistically, by stimulating dermal fibroblasts with key pathogenic cytokines associated with SSc, we observed significant changes in adenosine receptor A2A expression in response to IL-1β. Immunofluorescence revealed upregulation of A2A expression in dermal fibroblasts of SSc patients. Further, global A2a knockout significantly attenuated skin fibrosis in both BLM- and HOCL-induced skin fibrosis mouse models. Single-cell RNA sequencing of mouse skin revealed significant alterations in fatty acid metabolism in fibroblasts from A2a-deficient mice following fibrotic challenge. RNA sequencing, Seahorse assays and in vitro experiments showed that A2A inhibition promotes FAO by upregulating CPT1A expression via suppressing CREB phosphorylation, alleviating fibrosis in human primary dermal fibroblasts. Furthermore, targeted intervention of A2a specifically in fibroblasts improves outcomes and increases CPT1A expression in BLM-induced skin fibrosis mouse model.

Conclusion

Our study highlights the crucial interplay between adenosine metabolism-A2A receptor axis and FAO in SSc-associated skin fibrosis, suggesting that targeting the adenosine receptor A2A-FAO metabolic axis offers a promising therapeutic strategy for skin fibrosis.

Introduction

Skin fibrosis involves the progressive thickening and stiffening of skin due to excessive extracellular matrix (ECM) deposition, which can lead to restricted joint movement and skin atrophy, seriously affecting patients’ quality of life. Skin fibrosis is a hallmark of fibrotic diseases such as systemic sclerosis (SSc). Despite advances in our understanding of SSc, effective therapies targeting skin fibrosis remain limited [1]. The lack of treatments capable of controlling or reversing skin fibrosis represents a major challenge in managing skin manifestations of SSc and highlights the urgent need for interventions that can effectively address skin fibrosis.

Recent studies have uncovered significant metabolic alterations in SSc, highlighting the role of metabolism in disease progression [2,3,4]. While much attention has been given to metabolic reprogramming in the immune system, it is now clear that metabolic disruptions also play a critical role in fibrosis [5, 6]. Emerging research has identified abnormal metabolic pathways in fibroblasts proliferating and differentiating into myofibroblasts, such as altered glycolysis, fatty acid oxidation (FAO), and glutaminolysis [7,8,9,10,11]. Moreover, metabolism operates as a complex, interconnected network, with different metabolic pathways influencing each other through enzymes, metabolites, and signaling molecules [12,13,14]. This potential crosstalk between pathways has profound implications for disease progression. However, whether these interactions occur in SSc-associated skin fibrosis and the precise role of metabolic changes in driving skin fibrosis remain largely unexplored in SSc.

In this study, we identified that adenosine-centered nucleotide metabolism is elevated in the plasma of SSc patients and revealed that adenosine receptor A2A signaling regulates changes in fatty acid metabolism and energy shifts in fibroblasts involved in SSc-related skin fibrosis. Mechanistically, A2a deficiency significantly reduced skin fibrosis and altered fatty acid metabolism in fibroblasts in both BLM- and HOCL-induced skin fibrosis mouse models. Further in vitro functional studies revealed that A2A inhibition promotes FAO by upregulating CPT1A via suppressing phosphorylation of CREB. Targeting A2A specifically in fibroblasts improves outcomes and upregulates CPT1A expression in the BLM-induced skin fibrosis mouse model. This study highlights the crucial interplay between the adenosine metabolism-A2A axis and FAO in fibroblasts, suggesting that targeting the A2A/p-CREB/CPT1A signaling pathway offers a promising therapeutic strategy for skin fibrosis.

Materials and methods

Human samples

Blood samples were collected from 35 patients with systemic sclerosis (SSc) and 10 healthy controls from the Department of Rheumatology and Immunology at Xiangya Hospital between August 2022 and September 2023. All SSc patients met the 2013 American College of Rheumatology (ACR)/European League Against Rheumatism (EULAR) classification criteria. Exclusion criteria included other systemic autoimmune diseases, malignant tumors, or severe infections.

For blood preparation, samples were drawn into EDTA-coated collection tubes, then centrifuged at 3000 × g for 10 min at 4 °C within 1 h. Plasma samples were aliquoted and stored at -80 °C until further analysis.

The study was conducted in accordance with the principles of the Declaration of Helsinki. All participants provided written informed consent, and the study received approval from the ethics review committee of Xiangya Hospital, Central South University (No. 201212074).

Metabolomic analysis of plasma samples

Metabolomic analysis of plasma samples was conducted using 600MRM analysis (Biotree, Shanghai, China) with UHPLC-QTRAP-MS/MS. For metabolite extraction, 50 μL of plasma was mixed with 250 μL of ddH2O, followed by adding 1200 μL of a methanol (1:1) solution. The mixture was thoroughly vortexed, ultrasonicated in an ice-water bath for 15 min, and incubated at -4 °C for 2 h. After centrifugation at 12,000 rpm and 4 °C for 15 min, the supernatant was transferred to a new tube and concentrated by evaporation for 8 h until dry. The dried powder was then reconstituted in 120 μL of acetonitrile (6:4, v: v) and ultrasonicated in an ice-water bath for 30 s. After a final centrifugation at 12,000 rpm and 4 °C for 15 min, the supernatant was carefully collected for analysis. A mixture of standard solutions served as quality control (QC) samples, and a series of calibration standards were prepared by stepwise dilution.

Chromatographic separation of 600 target metabolites was achieved using an ACQUITY UPLC H-Class system (Waters) on a Waters Atlantis Premier BEH ZHILIC Column (1.7 μm, 2.1 mm x 150 mm). The mobile phase A consisted of ultrapure water and acetonitrile (8:2) with 10 mmol/L ammonium formate, while mobile phase B comprised acetonitrile and ultrapure water (9:1) with 10 mmol/L ammonium formate. Both phases were adjusted to pH 9 with ammonium hydroxide. The sample tray temperature was maintained at 8 °C, and the injection volume was set to 1 μL.

Mass spectrometric analysis was performed using a SCIEX 6500 QTRAP + triple quadrupole mass spectrometer equipped with an IonDrive Turbo V ESI ion source in multiple reaction monitoring (MRM) mode. The ion source parameters were as follows: Curtain Gas = 35 psi, IonSpray Voltage = + 5000 V/-4500 V, Temperature = 500 °C, Ion Source Gas 1 = 50 psi, Ion Source Gas 2 = 50 psi. All data acquisition and quantitative analysis of the target compounds were conducted using SCIEX Analyst Work Station Software (1.7.2) and Data Driven Flow (v-1.0.1).

Metabolites quantification

For adenosine quantification

The concentration of human and mouse plasma adenosine was measured using HPLC/MS/MS. In brief, plasma was aliquoted to tubes containing 10 μM pentostatin (2033/10; Tocris) and 10 μM dipyridamole (0691/500; Tocris). Subsequently, 100 μL of plasma was utilized to extract nucleotides by adding 0.6 M cold perchloric acid. After centrifugation at 14,000 × g for 10 min, 142 μL of the supernatant was sequentially neutralized by 3 M KHCO3/3.6 N KOH. Subsequently, 1.425 μL of 1.8 M ammonium dihydrogen phosphate (pH 5.1) and 3.3 μL of phosphoric acid (30%) were added. The sample was then centrifuged at 14,000 × g for 10 min, and the supernatant was used for HPLC/MS/MS analysis (6470 triple-quadrupole mass spectrometer; Agilent Technologies).

Chromatographic analyses were conducted using high-performance liquid chromatography (HPLC) equipment, consisting of Agilent 1290 infinity UHPLC high-pressure binary gradient pump. Separations were performed using a 4.6*50 mm BEH C18 column with a particle size of 2.5 μm (Waters, XBridge Premier, MA, USA). The mobile phase consisted of (X) water with 0.3% formic acid and (Y) acetonitrile. The binary gradient elution (X: Y proportion, v/v), at a flow rate of 0.3 mL/min, comprised 97:3 from 0 to 5 min; switching to 50:50 from 5 to 7 min; maintained for 10 min. This system was coupled to a mass spectrometer with triple-quadrupole mass analyzers and an electrospray interface, operating in positive mode (ESI +). The temperatures of the source block were set at 100 °C, and desolvation gas was set at 350 °C. Ion detection was performed in the multiple reaction monitoring (MRM) mode, monitoring the transitions of the m/z 268 precursor ion to the m/z 136 product for adenosine (268.1 -> 119.0). The standard curve was constructed using a serial dilution of adenosine (3624/50; Tocris Bioscience).

For acylcarnitine quantification

The concentration of Acylcarnitine in plasma and cells were measured using HPLC/MS/MS. Briefly, 100 μL of plasma or 1 mL of cell culture supernatant was deproteinized with three volumes of cold acetonitrile. Precipitated proteins were removed by centrifugation at 14,000 × g for 15 min at 4 °C. For intracellular acylcarnitine extraction, cells were resuspended in 400 μL of cold methanol after counting, followed by vortexing for 30 s and sonicated on ice. The supernatant containing intracellular metabolites was collected by centrifugation at 14,000 × g for 15 min at 4 °C. The combined plasma and cell-derived supernatants from each sample were evaporated to dryness under vacuum. Samples were reconstituted in 50% (v/v) methanol for analysis. The mobile phase consisted of (X) water with 0.1% formic acid and (Y) acetonitrile. The binary gradient elution (X: Y proportion, v/v) was run at a flow rate of 0.5 mL/min, with the following gradient profile: 5% Y from 0 to 1 min and linear gradient from 5 to 100% Y (1–7 min), 100% Y (7–7.5 min), 100 to 5% Y (7.5–8 min), 5% Y (8–10 min). Ion detection was performed in the multiple reaction monitoring (MRM) mode, monitoring the transitions of the m/z 204 precursor ion to the m/z 85 product ion for acetylcarnitine (204 -> 85) and m/z 400 precursor ion to the m/z 85 product ion for palmitoylcarnitine (400 -> 85). Raw data files generated by LC-MS/MS were processed with MassHunter profile software (Agilent Technologies). The standard curve was constructed using serial dilutions of acetylcarnitine (A275415; Aladdin) and palmitoylcarnitine (T12356; Targetmol).

Dermal fibrosis mouse models

To induce skin fibrosis, six- to eight-week-old mice were shaved in a 1 cm² area on the upper back. For BLM-induced dermal fibrosis, mice received subcutaneous injections of 100 μL of bleomycin (MCE, HY17565A) at a concentration of 1 mg/mL every other day for 21 days. For hypochlorous acid (HOCL)-induced dermal fibrosis, mice received subcutaneous injections of 200 μL of freshly prepared HOCL every day for 6 weeks as previously described [15]. Mice injected subcutaneously with equal volumes of saline were served as controls. Mice were sacrificed and skin tissue was collected for further analysis at the corresponding time point.

Pulmonary fibrosis mouse model

Lung fibrosis was induced by a single intratracheal injection of bleomycin at a dose of 2 mg/kg body weight in six- to eight-week-old mice. Mice injected intratracheally with equal volume of saline were served as controls. Mice were sacrificed on day 21 post-bleomycin challenge.

Sample preparation for 10× genomics

Skin tissues from 5 mice per group were washed with cold PBS, cut into small pieces, and digested at 37°C with constant rotation for 1.5 hours using Dispase II (2 mg/mL, D4693, Sigma-Aldrich) and Collagenase IV (Sigma, #V900893). Following digestion, the cell supernatants were collected, pooled by group, and passed through a 70-μm cell strainer to obtain a single-cell suspension. Single-cell 5′ gene expression libraries were prepared using the Chromium Next GEM Single Cell 5’ Kit v2 protocol (10× Genomics). The resulting libraries were sequenced on the illumina novaseq 6000 platform (Novogene, Beijing, China).

Primary human dermal fibroblast isolation

Human dermal fibroblasts were isolated from normal foreskin tissues. The foreskin was incubated overnight at 4 °C in 2 mg/mL Dispase II (D4693, Sigma-Aldrich). The epidermis and dermis were then gently separated and washed with cold DPBS. The dermis was further cut into small pieces and digested in 3 mg/mL Collagenase IV (Sigma, #V900893) at 37 °C for 2 h, followed by filtration through 40 μm cell strainers (BD Falcon, #352340). After washing twice with DPBS, the cells were resuspended in DMEM/F-12 (1:1) (CellMax, CGM105.05) containing 10% FBS and 25 mM HEPES buffer (Gibco, 15630080). Cells from the third to eighth passages were used for subsequent experiments.

Cell treatment

For primary dermal fibroblasts treatment, cells were first starved in serum-free medium containing 0.1% FBS for 16 h, then treated with the following reagents as specified in each experiment, unless stated otherwise: TGF-β (10ng/ml, Peprotech, 100 −21), IL-1β (20 ng/ml, Peprotech, 200-01B), IL-4 (20 ng/ml, Peprotech, 200-04), IL-6 (20 ng/ml, Peprotech, 200-06), IL-12 (20 ng/ml, Peprotech, 200-12), CGS21680 (10μM, MCE, HY13201A), SCH442416 (10μM, Tocris, 2463), etomoxir (5 μM, Selleck, S8244), forskolin (30μM, Sigma, 344270), IVA337 (20 μM, Selleck, S8770) and 666 −15 (0.1 μM, MCE, HY101120).

Seahorse metabolic assay

Dermal fibroblasts (1.5 × 10^4 cells) were seeded into XFe96 cell culture microplates (Agilent Technologies, Santa Clara, CA, USA) and pretreated for 48 h with or without TGF-β (10 ng/ml) and SCH442416 (10 μM). On the day of the assay, cells were rinsed and pre-incubated in XF RPMI medium supplemented with 1 mM sodium pyruvate, 2 mM L-glutamine, and 10 mM glucose, without CO2. For the fatty acid oxidation metabolism assay, 40 μM etomoxir or control medium was added to port A. Subsequent additions included 15 μM oligomycin to port B, 10 μM FCCP to port C, and 5 μM Rot/AA to port D, all prepared at the recommended 10x concentrations. Automated stimulation was performed according to the manufacturer’s instructions using the Seahorse XF Substrate Oxidation Stress Test-Standard template on the Seahorse XFe Analyzer. Data analysis was conducted using the Seahorse XFp Wave software, and real-time oxygen consumption rates (OCR) were normalized to cell counts, which were quantified using the BioTek Cytation C10 system.

Cell transfection

Human dermal fibroblasts were transfected with siCPT1A (5′-GAAGCUCUUAGACAAAUCTT-3′) using Lipo3000 (L3000015; ThermoFisher Scientific) according to the manufacturer’s instructions for 24 h. The scramble siRNA (siNC) served as controls.

Statistical analyses

Statistical analyses were conducted using Prism 9 software (GraphPad). For metabolomics data, the two-tailed Student′s t test was used for comparisons between two groups. In other in vivo and in vitro studies, where the sample size was insufficient for normality testing, a one-way ANOVA followed by Tukey’s multiple comparisons test was used for experiments with one variable and 3 or more groups, two-way ANOVA followed by Tukey’s multiple comparisons test was used to analyze experiments with 2 variables and 4 groups, and an unpaired two-sided t-test was applied for two-group comparisons. Statistical details (e.g., number of samples/group, number of independent experiments) can be found in the figure legends. A p-value of less than 0.05 was considered statistically significant for all experiments.

A full description of the materials and methods is provided in the Supplementary Material.

Results

Nucleotide metabolism centered around adenosine is significantly elevated in SSc

To elucidate the key metabolic pathways involved in skin fibrosis, such as in conditions like systemic sclerosis (SSc), we performed metabolomics analysis on plasma samples from our SSc cohort, including 35 SSc patients and 14 paired healthy controls (HC), with detailed patient clinical information distribution shown in Fig. 1A. A total of 457 metabolites were identified in plasma. OPLS-DA showed significant variance between HC and SSc groups (Fig. 1B). Compared to healthy controls, we identified 23 significantly elevated metabolites and 8 significantly decreased metabolites in SSc patients (Fig. 1C-D). Using KEGG pathway enrichment analysis, we found that the differential metabolites elevated in SSc plasma were mainly enriched in nucleotide metabolism and purine metabolism pathways (Fig. 1E). Therefore, we further analyzed the differential metabolites in nucleotide metabolic pathway, with the most significant metabolites being Adenosine monophosphate (AMP), 2’-Deoxyguanosine-5’-diphosphate, Adenosine, Adenosine-5’-diphosphate (ADP), and Inosine. (Fig. 1F). Interestingly, we found that most of these differential metabolites were enriched in the ATP to adenosine hydrolysis pathway, including downstream hydrolysis products such as ADP, AMP, adenosine, and inosine, with adenosine showing the highest VIP score (Fig. 1F-G). Given these findings, we further collected SSc patients as a validation cohort to confirm changes in adenosine levels in SSc. As expected, plasma adenosine levels in SSc patients were significantly higher than healthy controls (Fig. 1H). Therefore, we speculate that nucleotide metabolism, centered around adenosine metabolism, may play an important role in the pathogenesis of SSc-associated skin fibrosis.

Fig. 1
figure 1

Nucleotide metabolism centered around adenosine is significantly elevated in SSc. (A) Clinical characteristics of SSc patients (n = 35). (B-D) Orthogonal projections to latent structures-discriminant analysis (OPLS-DA) showing group distribution (B), volcano plot representing differentially expressed metabolites (C) and Z-score plot showing significantly upregulated and downregulated metabolites (D) of plasma samples from SSc patients (n = 35) and HC (n = 14) groups. (E) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis based on significantly upregulated metabolites of SSc patients. (F) Differential metabolites in the nucleotide metabolism pathway of SSc patients compared to HC. (G) Schematic representation of ATP to adenosine hydrolysis pathway. Arrowheads indicate significantly upregulated metabolites in SSc patients. (H) Plasma adenosine concentrations in SSc patients (n = 20) and HC (n = 15). Error bar represents mean +/- SD range. *P < 0.05; HC, healthy control

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Adenosine receptor A2a deficiency reduces fibrosis in both BLM- and HOCL-induced skin fibrosis mouse models

Adenosine is not merely a metabolic byproduct; it functions as a potent GPCR agonist, influencing key pathophysiological processes such as inflammation, neurotransmission, immune response, and fibrosis through its four receptors: A1, A2A, A2B, and A3(Allard et al., 2020, Linden et al., 2019, Meng F. et al., 2019, Zhang et al., 2020). Given the central role of fibroblasts in the pathogenesis of fibrosis, we focused our research on the effects of adenosine on dermal fibroblast biology during the fibrotic process. Previous studies have shown that the baseline expression levels of adenosine receptors in dermal fibroblasts are relatively low. Therefore, we first investigated whether the immune microenvironment associated with SSc could induce abnormal expression of adenosine receptors in dermal fibroblasts. By stimulating dermal fibroblasts with key pathogenic cytokines associated with SSc (Raja and Denton, 2015), we found that only A2A showed significant changes in response to IL-1β stimulation, with its expression increasing by approximately five-fold (Fig. 2A). Consequently, we further analyzed A2A expression in skin samples from SSc patients. Immunofluorescence analysis revealed a significant upregulation of A2A expression in dermal fibroblasts from SSc patients (Fig. 2B). Therefore, our study focuses on the role of adenosine metabolism through dermal fibroblast A2A in SSc and its regulatory mechanisms. To determine whether the elevated levels of adenosine in SSc patients contribute to fibrotic progression via the A2A receptor, we used A2a-deficient mice to establish subcutaneous bleomycin (BLM)-induced classical localized skin fibrosis model and hypochlorous acid (HOCL)-induced diffuse cutaneous lesions model, respectively (Meng M. et al., 2019). First, we examined whether BLM injection would induce adenosine elevation similar to that observed in SSc. Results showed that adenosine levels were significantly elevated in the BLM-induced skin fibrosis model of wild-type (WT) mice, while no changes were observed in A2a−/− mice (Fig. 2C). Furthermore, A2a knockout markedly improved BLM-induced skin fibrosis, evidenced by reduced dermal thickness and collagen deposition, as shown by hematoxylin and eosin (H&E) and Masson’s trichrome staining (Fig. 2D-F). Additionally, hydroxyproline content in skin tissues (Fig. 2G), and the number of α-SMA-positive myofibroblasts were significantly reduced in A2a−/− mice (Fig. 2H). Moreover, the transcriptional levels of fibrotic markers (Acta2, Col1a1, Col1a2, and Ctgf) and inflammation-related cytokines (Il-1b) were notably decreased in A2a−/− mice after BLM challenge (Fig. 2I).

Fig. 2
figure 2

Adenosine receptor A2a deficiency reduces fibrosis in both BLM- and HOCL-induced skin fibrosis mouse models. (A) RT-qPCR analysis of adenosine receptors expression in human dermal fibroblasts treated with TGF-β, IL-1β, IL-4, IL-6 or IL-12 for 48 h. Statistical significance was determined by comparison with the NC group. (B) Representative immunofluorescent staining of A2A (green) and VIMENTIN (red), and quantification of VIMENTIN+ A2A+ cells in the skin sections from HC and SSc patients (n = 5, Scale bar: 100 μm). (C-H) Quantification of plasma adenosine concentrations (C); representative H&E staining (D) and Masson’s trichrome staining (E), quantification of dermal thickness (μm) (F) and hydroxyproline content (G), and representative image of immunohistochemistry staining of α-SMA, along with quantification of α-SMA-positive myofibroblasts in the skin tissues from WT and A2a-/- mice treated with PBS (n = 4) or BLM (n = 8). (I) RT-qPCR analysis of Acta2, Col1a1, Col1a2 and Il1b in the skin from WT and A2a-/- mice treated with PBS or BLM (n = 6). (J-M) Representative H&E staining (J), Masson’s trichrome staining (K), quantification of dermal thickness (μm) (L), and representative image of immunohistochemistry staining of α-SMA, along with quantification of α-SMA-positive myofibroblasts (M) in the skin tissues from WT and A2a-/- mice treated with PBS (n = 4) or HOCL (n = 8) (Scale bar: 200 μm). RT-qPCR of human dermal fibroblasts was normalized against GAPDH, and RT-qPCR results of murine skin were normalized against βactin. n = 4–8/group. Error bars represent mean +/- SD range. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; ns, not significant, HC, healthy control

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Similarly, in the HOCL-induced skin fibrosis model, A2a deficiency resulted in decreased dermal thickness and collagen deposition (Fig. 2J-L), as well as marked reductions in the number of α-SMA-positive myofibroblasts (Fig. 2M). Together, these data suggest that adenosine/A2A signaling plays an important role in the regulating skin fibrosis.

Single-cell RNA-seq revealed altered fatty acid metabolism in fibroblasts after BLM and HOCL challenge in A2a−/− mice

To further investigate the role of A2A in fibroblasts during BLM-induced and HOCL-induced skin fibrosis, we performed single-cell RNA-sequencing (scRNA-seq) on skin samples following fibrotic challenge. By annotating marker gene expression signatures, we identified 15 cell types in murine skin, including keratinocytes, adipocytes, fibroblasts, smooth muscle cells, endothelial cells, lymphatic endothelial cells, Schwann cells, melanocytes, macrophages, neutrophils, dendritic cells, mast cells, T cells, NK cells and NKT cells (Fig. 3A-C).

Fig. 3
figure 3

Single-cell RNA-seq revealed altered fatty acid metabolism in fibroblasts after BLM and HOCL challenge in A2a−/− mice. (A) UMAP plot of main cell types in single-cell RNA-seq data from skin tissue of WT and A2a−/− mice treated with PBS or BLM or HOCL. (B) Dot plot illustrating marker gene expression levels for the different cell types defined in (A). (C) The proportion of different cell types in skin. (D) Number of differentially expressed genes (DEGs) in different cell types of WT and A2a−/− mice treated with BLM (upper) or HOCL (lower). Red bars indicate upregulated genes, and blue bars indicate downregulated genes. (E-F) Gene Ontology (GO) enrichment analysis of DEGs in skin fibroblasts between WT mice treated with PBS or BLM (E), and between WT and A2a−/− mice treated with BLM (F). (G-H) GO enrichment analysis of DEGs in skin fibroblasts between WT mice treated with PBS or HOCL (G), and between WT and A2a−/− mice treated with HOCL (H)

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To identify the A2a-driven transcriptomic changes among different cell types during the fibrotic process, we compared differentially expressed genes (DEGs) among different cell types between WT and A2a-deficient mice following BLM and HOCL challenge, respectively. In both of skin fibrosis preclinical models, fibroblast cluster was one of the cell types with the abundant altered numbers of DEGs in A2a-deficient mice compared to the WT (Fig. 3D).

We further conducted Gene Ontology (GO) analysis on the fibroblast cluster. The results revealed that both BLM- and HOCL-induced wild-type dermal fibroblasts were enriched in fibrosis-related pathways (extracellular structure organization, extracellular matrix assembly, and TGFB-related signaling, etc.) and inflammatory-related pathways (leukocyte chemotaxis, monocyte chemotaxis, regulation of inflammatory response, etc.). Interestingly, fatty acid metabolism, which has been recently reported to be involved in fibrosis, was also significantly enriched in both of our skin fibrosis mouse models [16]. Notably, A2a knockout not only disrupted fibrosis-related pathways but also significantly impacted fatty acid metabolism, mirroring the changes observed in BLM- and HOCL-challenged WT mice (Fig. 3E-H). These findings, revealed by single-cell RNA sequencing, suggest that fibroblast A2A might mediate skin fibrosis by modulating fatty acid metabolism.

A2A mediates fibrotic response by regulating FAO in human dermal fibroblasts

We further investigated the role and mechanisms of A2A in fibroblasts in vitro. Primary human dermal fibroblasts were treated with TGF-β in the presence or absence of the A2A inhibitor SCH442416 or the agonist CGS21680. Stimulation of human dermal fibroblasts with TGF-β promoted the expression of fibronectin and α-SMA at the protein level and increased the mRNA levels of ACTA2 (encoding α-smooth muscle actin), COL1A2, and FN1(encoding fibronectin). This effect was suppressed by targeted inhibition of A2A by SCH442416 in a dose-dependent manner, with significant suppression observed starting at 10 μM (Fig. 4A-B). Accordingly, the concentration was selected for subsequence experiments [17]. By contrast, using A2A activator, CGS21680, led to significant upregulation of fibrosis markers at the protein and mRNA levels in the presence of TGF-β (Supplementary Fig. S1A-B).

Fig. 4
figure 4

A2A mediates fibrotic response by regulating FAO in human dermal fibroblasts. (A-B) Immunoblot analysis of fibronectin and α-SMA (A) and RT-qPCR analysis of ACTA2, COL1A1 and FN1 mRNA (B) in dermal fibroblasts treated with TGF-β, with or without 1–20 μM SCH442416, for 48 h. Statistical significance was determined by comparison with the TGF-β group. (C-D) Volcano plot of the significant DEGs (C) and GO enrichment analysis of DEGs (D) in dermal fibroblasts treated with or without TGF-β for 48 h (left panel), and between the TGF-β-treated dermal fibroblasts treated with or without SCH442416 (10μM) for 48 h (right panel). (E) Heatmap showing the expression of FAO-related genes in the dermal fibroblasts treated with TGF-β, with or without SCH442416, for 48 h. (F) Gene set enrichment analysis (GSEA) of fatty acid oxidation pathway in dermal fibroblasts treated with or without TGF-β (left panel) for 48 h, and in TGF-β-treated dermal fibroblasts treated with or without SCH442416 (right panel) for 48 h. (G) Oxygen-consumption rate (OCR) of dermal fibroblasts pretreated with TGF-β, with or without SCH442416 (10μM), for 48 h. (H-J) Quantification of basal OCR, maximal OCR and ATP production from (G). (K-L) Quantification of acetylcarnitine and palmitoylcarnitine in cell lysate (K) and cell culture supernatant (L) from dermal fibroblasts treated with TGF-β, with or without SCH442416, for 48 h. (M) Quantification of plasma acetylcarnitine and palmitoylcarnitine concentrations from SSc patients (n = 20) and healthy controls (n = 15). (N-O) Immunoblot analysis of fibronectin and α-SMA (N), and RT-qPCR analysis of ACTA2, COL1A1 and FN1 mRNA (O), in dermal fibroblasts treated with TGF-β in combination with different concentrations of IVA337 for 48 h. (P-Q) Immunoblot analysis of fibronectin and α-SMA (P), and RT-qPCR analysis of COL1A1, COL1A2 and FN1 mRNA (Q) in dermal fibroblasts treated with TGF-β, CGS21680 (10μM), and IVA337 (20μM) for 48 h. RT-qPCR results were normalized against GAPDH. Error bars represent mean +/- SD range. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; Oligo, oligomycin, FCCP, Carbonyl cyanide-4 (trifluoromethoxy) phenylhydrazone, Rot/AA, Rotenone/antimycin, Eto, etomoxir, ns, not significant; HC, healthy control

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To understand how A2A inhibition alleviates fibrosis, we performed bulk RNA sequencing (RNA-seq) and compared the TGF-β-treated group with the NC and TGF-β + SCH442416 groups, respectively. We identified 8298 differentially expressed genes (DEGs) in the TGF-β group compared with NC, whereas A2A inhibition affected a total of 1465 genes, with 681 upregulated and 784 downregulated compared to TGF-β treatment alone (Fig. 4C). GO enrichment analysis revealed that, similar to the pathway alterations observed in skin fibrosis mouse models, TGF-β treatment significantly altered fibrosis-related pathways in dermal fibroblasts (collagen fibril organization, extracellular matrix organization and extracellular structure organization, etc.). Meanwhile, fatty acid oxidation (FAO) pathways were markedly downregulated following TGF-β treatment. In contrast, A2A inhibition resulted in a significant upregulation of FAO-related pathways compared to TGF-β alone (Fig. 4D). To further explore the regulation of FAO by A2A, we examined key FAO-regulating genes. The data revealed that the key FAO-related gene carnitine palmitoyltransferase 1 (CPT1A) was most significantly downregulated in fibroblasts after TGF-β stimulation, but their expression was markedly upregulated following A2A inhibition (Fig. 4E). Gene set enrichment analysis (GSEA) further confirmed that FAO pathways were significantly suppressed in fibroblasts following TGF-β treatment, while A2A inhibition reversed this suppression (Fig. 4F).

Recent studies have shown that the downregulation of FAO is a key metabolic pathway driving ECM deposition in fibroblasts [10, 16, 18]. Building on insights from our in vivo scRNA-seq and in vitro RNA-seq data, we employed the Seahorse XF Analyzer to examine the effect of A2A inhibition on FAO during fibrosis. Before adding etomoxir, an inhibitor of FAO enzyme CPT1A, the oxygen consumption rate (OCR) reflected total mitochondrial respiration, encompassing both FAO and other metabolic pathways. TGF-β stimulation did not significantly affect mitochondrial respiration in fibroblasts. However, A2A inhibition markedly increased mitochondrial respiration compared to TGF-β treatment alone. In the etomoxir-treated group, OCR remained largely unchanged in the TGF-β-only group, indicating minimal reliance on FAO during TGF-β stimulation. By contrast, the A2A inhibitor-treated group showed a significant reduction in OCR after etomoxir treatment (Fig. 4G-J), suggesting that the elevated mitochondrial respiration induced by A2A inhibition was largely driven by enhanced FAO activity.

To further investigate the impact of altered FAO activity in fibroblasts, we measured two key downstream metabolites of fatty acid oxidation, acetylcarnitine and palmitoylcarnitine, which are fatty acid intermediates and serve as reliable indicators of FAO activity [19, 20]. Consistent with our findings from gene expression analysis and Seahorse assays, TGF-β treatment significantly reduced intracellular and extracellular levels of both metabolites, whereas inhibition of A2A signaling restored these levels (Fig. 4K-L). To validate these observations in a clinical context, we measured the levels of these metabolites in plasma samples from SSc patients and healthy controls. Our results showed a significant reduction in both acetylcarnitine and palmitoylcarnitine in the plasma of SSc patients (Fig. 4M), further supporting our cellular and murine model data and suggesting that the FAO alterations observed in fibroblasts are also reflected in systemic metabolic changes in SSc.

To directly demonstrate that A2A regulates skin fibrosis through FAO modulation. We further treated dermal fibroblasts with different concentrations of IVA337 (an FAO activator via PPAR receptor activation) in the presence of TGF-β [21]. We observed that IVA337 suppressed fibrosis in a dose-dependent manner, with 20μM effectively limiting the expression of fibrotic markers (Fig. 4N-O). Meanwhile, treatment with the FAO activator IVA337 blocked the profibrotic effects induced by CGS21680 (Fig. 4P-Q). Together, these findings demonstrate that A2A mediates fibrotic phenotype in dermal fibroblasts, potentially through regulating FAO.

A2A mediates FAO through regulating CPT1A expression

Building on our RNA-seq findings, we next investigated whether A2A mediates FAO through CPT1A, a key rate-limiting enzyme of FAO. First, we found that TGF-β downregulated CPT1A at both the protein and RNA levels in a dose-dependent manner, with a significant inhibitory effect observed at the previously recognized dose of 10 ng/mL (Supplementary Fig. S2A-B). Meanwhile, the addition of the A2A inhibitor, SCH442416, dose-dependently upregulated CPT1A expression, with 10 μM demonstrating optimal effects (Fig. 5A-B). We further extended these findings to SSc patients. Immunofluorescence analysis confirmed that CPT1A levels were markedly lower in fibroblasts from SSc patients compared to healthy controls (Fig. 5C-D).

Fig. 5
figure 5

A2A mediates FAO through regulating CPT1A expression. (A-B) Immunoblot analysis of CPT1A (A) and RT-qPCR analysis of CPT1A (B) in dermal fibroblasts treated with TGF-β, with or without 1–20 μM SCH442416, for 48 h. (C-D) Representative immunofluorescent staining of CPT1A (green) and VIMENTIN (red) (C), and quantification of VIMENTIN+ CPT1A+ cells in the skin tissues from HC and SSc patients (D) (n = 5, scale bar: 20 μm). (E-F) Immunoblot analysis of fibronectin and α-SMA (E), and RT-qPCR analysis of COL1A1, COL1A2 and FN1 mRNA (F), in dermal fibroblasts treated with TGF-β, SCH442416 (10μM), and etomoxir (5μM) for 48 h. (G) RT-qPCR analysis of CPT1A mRNA in dermal fibroblasts after silencing of CPT1A. (H-I) Immunoblot analysis of fibronectin, α-SMA and CPT1A (H), and RT-qPCR analysis of ACTA2 and FN1 mRNA (I), in dermal fibroblasts after silencing of CPT1A and incubation with TGF-β for 48 h. (J-K) Immunoblot analysis of fibronectin and α-SMA (J), and RT-qPCR analysis of ACTA2 and FN1 mRNA (K), in dermal fibroblasts after silencing of CPT1A and incubation with TGF-β and SCH442416 (10μM) for 48 h. RT-qPCR results were normalized against GAPDH. Error bars represent mean +/- SD range. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; HC, healthy control

Full size image

To further investigate the role of CPT1A in A2A-regulated fibrosis, we first pharmacologically inhibited CPT1A using etomoxir in the presence of TGF-β. The concentration gradient experiment showed that etomoxir at a concentration of 5 μM significantly enhanced the stimulatory effects of TGF-β on dermal fibroblasts (Supplementary Fig. S2C-D). However, higher concentrations of etomoxir exhibited relatively weak effects on fibrosis, likely due to its off-target activity at high concentrations [22]. Furthermore, inhibition of CPT1A with etomoxir significantly reversed the antifibrotic effects of SCH442416, restoring the levels of fibrotic markers that were previously suppressed by SCH442416 (Fig. 5E-F). In parallel, we performed siRNA to specifically target CPT1A in dermal fibroblasts to further confirm this effect (Fig. 5G). Compared to the scramble siRNA control, siCPT1A significantly promoted the expression of fibronectin and α-SMA at both the protein and mRNA levels under TGF-β stimulation (Fig. 5H-I). Additionally, siCPT1A significantly restored the reduction of fibrotic markers caused by A2A inhibition (Fig. 5J-K). These results demonstrate that A2A mediates FAO by regulating CPT1A expression during fibrosis.

A2A regulates CPT1A expression via p-CREB/CREB pathway

As a GPCR, adenosine receptor A2A can induce phosphorylation of cAMP element regulatory binding protein (p-CREB), which further regulates downstream signaling. Given that p-CREB are reported to be elevated in the skin of SSc patients, we then asked whether p-CREB/CREB pathway is involved in the A2A-regulated fibrotic process [23]. Firstly, incubation with TGF-β promoted phosphorylation of CREB, which was prevented by A2A inhibitor (Fig. 6A). To further confirm the role of CREB phosphorylation in promoting fibrosis, we first used forskolin, an inducer of phosphorylated CREB. Forskolin was shown to upregulate the protein expression of p-CREB and fibronectin, along with mRNA expression of fibrosis markers in the presence of TGF-β (Fig. 6B-C). Notably, forskolin also exacerbated TGF-β-induced CPT1A downregulation (Fig. 6B-C). We next investigated whether A2A-regulated FAO metabolic reprogramming mediated fibrosis through CREB phosphorylation. The addition of forskolin in the presence of an A2A inhibitor reactivated CREB phosphorylation and restored the decline in fibrotic markers caused by A2A inhibition. Furthermore, the increase in CPT1A protein and mRNA expression levels mediated by A2A inhibition was also reversed (Fig. 6D-E). Meanwhile, we used 666 − 15, a highly specific inhibitor of CREB phosphorylation, to further verify the effect. As expected, inhibition of p-CREB markedly reversed the TGF-β-induced downregulation of CPT1A and significantly reduced the expression of fibrotic markers at both the protein and mRNA levels in the presence of TGF-β (Fig. 6F-G). Notably, the reduction in CPT1A expression at both the protein and mRNA levels, which was induced by A2A activation, was also partially reversed by 666 − 15. Additionally, the increase in fibrotic markers caused by A2A activation was also attenuated with 666 − 15 treatment (Fig. 6F-G), indicating that A2A regulates CPT1A expression through CREB phosphorylation to mediate fibrosis.

Fig. 6
figure 6

A2A regulates CPT1A expression via p-CREB/CREB pathway. (A) Immunoblot analysis of p-CREB and CREB in dermal fibroblasts treated with TGF-β, with or without SCH442416 (10μM), for 48 h. (B-C) Immunoblot analysis of p-CREB, CREB, fibronectin and CPT1A (B); and RT-qPCR analysis of COL1A1, FN1 and CPT1A (C) in dermal fibroblasts pretreated with or without forskolin (30μM, 30 min), followed by stimulation with or without TGF-β for 48 h. (D-E) Immunoblot analysis of p-CREB, CREB, fibronectin and CPT1A (D); and RT-qPCR analysis of COL1A1, FN1 and CPT1A (E) in dermal fibroblasts pretreated with or without forskolin, followed by stimulation with or without TGF-β and SCH442416 (10μM) for 48 h. (F-G) Immunoblot analysis of p-CREB, CREB, fibronectin and CPT1A (F), and RT-qPCR analysis of COL1A1, FN1 and CPT1A (G) in dermal fibroblasts treated with TGF-β, CGS21680 (10μM), and 666 − 15 (0.1μM) for 48 h. RT-qPCR results were normalized against GAPDH. Error bars represent mean +/- SD range. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; HC, healthy control

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Fibroblast-specifically knockout of A2a ameliorates BLM-induced skin and lung fibrosis

Finally, we used fibroblast-specific A2a knockout mice to demonstrate the role and mechanism of A2A in skin fibrosis. In the BLM-induced skin fibrosis model, fibroblast-specific A2a knockout mice showed reduced skin thickening, decreased collagen deposition, and lower myofibroblast counts, upon challenge with BLM (Fig. 7A-E). Moreover, fibroblast-specific deletion of A2a resulted in reduced hydroxyproline content in the skin (Fig. 7F). Fibroblast-specific knockout of A2a also significantly suppressed the expression of fibrotic marker genes, including Acta2, Col1a1, and Col1a2, as well as inflammation-related genes like Ifng, Il1b, Il6 and Il11 in the skin tissues (Fig. 7G).

Fig. 7
figure 7

Fibroblast-specifically knockout of A2a ameliorates BLM-induced skin fibrosis. (A-C) Representative H&E staining (A), Masson’s trichrome staining (B) and quantification of dermal thickness (μm) of skin sections (C) from A2af/fCol1a2cre− and A2af/fCol1a2cre+ mice treated with PBS or BLM (Scale bar: 200 μm). (D-E) Representative image of immunohistochemistry staining of α-SMA (D), and quantification of α-SMA-positive myofibroblasts (E) in the skin (Scale bar: 100 μm). (F-G) Quantification of hydroxyproline content and RT-qPCR analysis of fibrotic genes Acta2, Col1a1 and Col1a2, and inflammation-related genes Ifng, Il1b, Il6 and Il11 in the skin. (H) Representative immunofluorescent staining of CPT1A (green) and VIMENTIN (red), and quantification of VIMENTIN+ CPT1A+ cells in the skin (Scale bar: 20 μm). RT-qPCR results were normalized against βactin. n = 5/group; error bars represent mean +/- SD range. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001; ns, not significant

Full size image

Besides, consistent with our observations of A2A’s regulation of CPT1A expression in human dermal fibroblasts, immunofluorescence analysis revealed that CPT1A expression in fibroblasts was reduced in bleomycin-induced skin tissue of wild-type mice, but significantly restored in fibroblast-specific A2a knockout mice (Fig. 7H). These results further validate our in vitro findings, suggesting that fibroblasts A2A regulates dermal fibrosis by modulating FAO through CPT1A.

Given that the lung is an important target organ in SSc, we also employed a BLM-induced pulmonary fibrosis mouse model to investigate the role of A2A in the pulmonary fibrotic process associated with SSc. Consistent with our finding in the BLM-induced skin fibrosis model, fibroblast-specific deletion of A2a significantly alleviated pulmonary fibrosis. The results showed a marked reduction in collagen deposition, as indicated by Masson′s trichrome staining (Supplementary Fig. S3A) and hydroxyproline content (Supplementary Fig. S3B), along with a decrease in the expression of fibrotic genes in the lungs of fibroblast-specific A2a knockout mice following BLM challenge (Supplementary Fig. S3C). Collectively, fibroblast-specifically knockout of A2a ameliorates BLM-induced skin fibrosis, and may potentially improve lung fibrosis.

Discussion

In the current study, we performed metabolomic analysis to uncover metabolic changes in skin fibrosis in diseases like systemic sclerosis (SSc), revealing that nucleotide metabolism centered around adenosine is significantly elevated in SSc patients. Mechanistically, we observed that global knockout of the adenosine A2a receptor markedly reduced skin fibrosis in both BLM- and HOCL-induced scleroderma mouse models. Additionally, single-cell RNA sequencing of mouse skin revealed significant alterations in fatty acid metabolism in fibroblasts from A2a-deficient mice following fibrotic treatments. In vitro experiments further demonstrated that A2A inhibition enhanced fatty acid oxidation (FAO) by upregulating CPT1A expression via suppressing CREB phosphorylation, thereby alleviating the fibrotic response in dermal fibroblasts. Finally, using fibroblast-specific conditional knockout mice, we confirmed that targeting A2A specifically in fibroblasts improves outcomes and increases CPT1A expression in a bleomycin-induced skin fibrosis mouse model. Our findings suggest that specifically targeting A2A-FAO axis in skin fibrosis could be a viable strategy for SSc patients and provide valuable insights into the critical role of energy transformation in skin fibrosis, highlighting a significant interplay between adenosine metabolism and fatty acid oxidation.

Increasing evidence suggests that metabolism plays a central role in the pathogenesis and progression of SSc. In this study, we identified significant enrichment of nucleotide metabolism centered around adenosine in SSc patients through metabolomics analysis. The exact cause of the substantial accumulation of adenosine in these patients remains unknown. A plausible explanation is that the reduced blood flow and vascular dysfunction commonly seen in SSc lead to tissue hypoxia, a strong inducer of adenosine production [24, 25]. Previous studies have suggested that adenosine may play a protective role in acute inflammation and vasodilation by interacting with four G protein-coupled receptors (A1, A2A, A2B, and A3). However, in fibrotic conditions, adenosine has been implicated in promoting fibrosis, as reported in pulmonary, liver, and renal fibrosis [26,27,28,29]. In the context of dermal fibrosis, pharmacological inhibition and global genetic knockout of the A2A receptor has been shown to reduce fibrosis in SSc mouse models [30, 31]. Our study, further combining global and fibroblast-specific A2A receptor knockout mice, provides strong evidence that the fibroblast-specific A2A receptor is a direct and critical mediator of skin fibrosis in SSc. Given adenosine’s potentially opposing roles in disease protection through its downstream receptors in regulating vasodilation, inflammation, and fibrosis, our findings strongly support a targeted therapy strategy focused on the fibroblast A2A receptor for the treatment of SSc-associated skin fibrosis.

Previous studies have demonstrated A2A enhances fibrosis through classic signaling mechanisms, such as the β-catenin and AKT pathways, in dermal fibroblasts [32,33,34,35]. Notably, emerging researches increasingly highlight metabolic alterations as critical pathogenic drivers of fibrosis [8, 10]. Our study uncovered a surprising and critical interaction between adenosine metabolism and fatty acid metabolism, offering a fresh perspective on the complex metabolic interactions that regulate cellular function and influence skin fibrotic outcomes in SSc. Complex interactions exist within the metabolic network, where energy metabolism pathways such as glycolysis, glutaminolysis, oxidative phosphorylation, lipid, and glucose metabolism can undergo shifts during inflammation and cell differentiation through metabolites or metabolic enzymes [7, 36,37,38]. Recent studies have shown that fibrotic conditions, such as renal and peritoneal fibrosis, exhibit enhanced glycolysis and reduced FAO, with FAO inhibition being proposed as a potential strategy to alleviate fibrosis [16, 39,40,41]. In SSc, abnormal regulation of FAO in immune cells has also been reported [42], and FAO agonist has shown promise in improving SSc animal model phenotypes [43, 44]. However, whether FAO plays a role in the fibrosis of dermal fibroblasts and the upstream regulatory mechanisms have remained unclear. Our study advances this understanding by revealing that adenosine metabolism, through its downstream A2A receptor signaling pathway, can mediate the reduction of FAO in dermal fibroblasts by downregulating CPT1A, a key enzyme in FAO, via p-CREB/CREB pathway [45]. This, in turn, alters the energy metabolism of fibroblasts and influences the fibrotic process. This finding not only highlights the metabolic crosstalk between adenosine and FAO in SSc-associated skin fibrosis, but also introduces a potential adenosine-A2A-FAO regulatory mechanism for key physiological and pathological processes involving FAO-related energy alterations.

In summary, our study reveals significant alterations in adenosine-centered nucleotide metabolism in SSc patients. Through genetic mouse models and single-cell sequencing, we uncovered an interplay between adenosine metabolism and FAO that modulates energy shifts during fibrosis. By elucidating the adenosine/A2A/p-CREB/CPT1A signaling pathway in dermal fibroblasts, our research opens new avenues for targeted antifibrotic therapies for skin fibrosis in SSc and enhances our understanding of the metabolic mechanisms underlying skin fibrotic process.

Data availability

No datasets were generated or analysed during the current study.

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Acknowledgements

We acknowledge BioRender.com for providing tools to create schematic illustration.

Funding

This work was supported by National Natural Science Foundation of China (No. 82203931 to Y. T), Key Program of National Natural Science Foundation of China (No. U22A20329 to H. L), China Postdoctoral Science Foundation (No. BX20220355 to Y. T, No. 2023M733957 to Y. T), Science and technology innovation Program of Hunan Province(No. 2022RC3004 to H. L), Central South University Research Program of Advanced Interdisciplinary Studies (No. 2023QYJC004 to H. L), Changsha Municipal Natural Science Foundation (No. kq2202380 to Y. T), Fundamental Research Funds of the Xiangya Hospital (No.2021Q09 to Y. T), The Excellent Youth Project of Hunan Provincial Natural Science Foundation (No.2024JJ4094 to Y.T), as well as The Scientific Research Program of FuRong Laboratory(No. 2023SK2095 to H. L).

Author information

Author notes

  1. Xiaoye Zhang and Jinjian Sun contributed equally to this work.

Authors and Affiliations

  1. Department of Dermatology, Xiangya Hospital, Central South University, Changsha, Hunan, China

    Xiaoye Zhang, Jia Guo, Junyu Zhou, Xiaoyun Xie, Xiang Chen, Hong Liu & Yuzi Tian

  2. Hunan Engineering Research Center of Skin Health and Disease, Central South University, Changsha, Hunan, China

    Xiaoye Zhang, Jia Guo, Junyu Zhou, Xiaoyun Xie, Xiang Chen, Hong Liu & Yuzi Tian

  3. Hunan Key Laboratory of Skin Cancer and Psoriasis, Central South University, Changsha, Hunan, China

    Xiaoye Zhang, Jia Guo, Junyu Zhou, Xiaoyun Xie, Xiang Chen, Hong Liu & Yuzi Tian

  4. National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan, China

    Jinjian Sun, Jia Guo, Xiaoru Hu, Xiaoyun Xie, Xiang Chen, Hui Luo, Hong Liu & Yuzi Tian

  5. Department of General &Vascular Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, China

    Jinjian Sun

  6. Department of Rheumatology and Immunology, Xiangya Hospital, Central South University, Changsha, Hunan, China

    Xiaoru Hu, Xiaoyun Xie, Hui Luo & Yuzi Tian

  7. Provincial Clinical Research Center for Rheumatic and Immunologic Diseases, Xiangya Hospital, Central South University, Changsha, Hunan, China

    Xiaoru Hu, Xiaoyun Xie, Hui Luo & Yuzi Tian

Authors
  1. Xiaoye Zhang
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  2. Jinjian Sun
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  3. Jia Guo
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  4. Xiaoru Hu
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  5. Junyu Zhou
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  6. Xiaoyun Xie
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  7. Xiang Chen
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  8. Hui Luo
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  9. Hong Liu
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  10. Yuzi Tian
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Contributions

Y.T, H.L, X.Z, J.S, H.L, and X.C developed the concepts and discussed experiments. X.Z, Y.T performed the experiments. Y.T, X.Z wrote the manuscript. J.S performed bioinformatics analysis. H.L, X.Z and X.H collected patient samples and provided clinical information. J.G, X.H, J.Z, X.X, analyzed and discussed the data.

Corresponding authors

Correspondence to Hong Liu or Yuzi Tian.

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Ethics approval

The study was approved by the ethics review committee of Xiangya Hospital, Central South University (No. 201212074 and No.202308636). The animal protocols were institutionally approved by the Institutional Animal Care and Use Committee at Xiangya Hospital, Central South University (No.202310043).

Competing interests

The authors declare no competing interests.

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Zhang, X., Sun, J., Guo, J. et al. Inhibition of adenosine/A2A receptor signaling suppresses dermal fibrosis by enhancing fatty acid oxidation. Cell Commun Signal 23, 206 (2025). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12964-025-02210-2

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  • DOI: https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12964-025-02210-2

Cell Communication and Signaling

ISSN: 1478-811X