Introduction
Cirrhotic cardiomyopathy (CCM) is a cardiac disorder that occurs in patients with cirrhosis, independent of other heart diseases. It is characterized by an impaired contractile response to stress, diastolic dysfunction, and a prolonged QT interval.1 Reported prevalence ranges from 26% to 81%,2 but the mechanisms underlying cirrhosis-induced cardiac injury remain unclear, and no specific preventive treatments for CCM have yet been developed.
The 2005 World Congress of Gastroenterology established diagnostic criteria for CCM, which were later revised in 2020 by the Cirrhotic Cardiomyopathy Consortium.3 Since then, numerous studies have examined its pathophysiology and clinical course before transplantation, although research using animal models remains limited. Existing animal studies have produced inconsistent findings regarding the timing and mechanisms of injury, with conflicting results on cardiac hypertrophy, atrophy, and impaired contractile function at rest.4–7
Uhlig et al. conducted a detailed investigation of temporal dynamics and cardiac fragility in a rat model, but the absence of gallbladders in rats may limit the applicability of their findings to CCM.8 In contrast, Matyas et al. evaluated heart function in mice with bile duct ligation (BDL)-induced cirrhosis and showed that mice reproduce the pathophysiological features of CCM.7 Nevertheless, the two-week study period was insufficient to capture the dynamic progression of the disease.
This study assessed cardiac function in mice with BDL-induced cirrhosis at weeks 2, 4, and 8, while also examining dynamic changes in inflammatory and mitochondrial markers. In addition, echocardiographic data were collected from patients with CCM to analyze the correlation between cirrhosis severity and cardiac dysfunction.
Methods
Animal model
All animal care and experimental procedures were approved by the Ethics Committee of Anhui Medical University (LLSC20210241) and conducted in accordance with the Declaration of Helsinki (2024). Seventy male C57BL/6J mice (seven weeks old, 20–25 g) were purchased from Jiangsu Jicui Pharmaceutical Biotechnology Co., Ltd. The mice were housed under controlled conditions with an ambient temperature of 24 ± 1°C, a 12-h light/dark cycle, and a relative humidity of 55% ± 5%. Before the study, they were acclimated to the laboratory for one week and then randomly divided into five groups using a randomized block design: Control, Sham, two weeks, four weeks, and eight weeks. There were 14 mice in each group. Since the survival rate of BDL surgery is only about 50%, 14 mice were included in each group (a total of 70) to ensure a final sample size of n = 6 per group. BDL surgery was performed in all groups except the Sham group, which underwent the same procedure without ligation. BDL was conducted as previously described.9 At the endpoint, mice were fasted for 8 h, anesthetized with isoflurane (R510-22-10; RWD, China), and euthanized by cervical dislocation following orbital blood sampling (∼0.2 mL). Death was confirmed by an audible click from cervical vertebral dislocation, followed by the absence of heartbeat and spontaneous respiration. The heart and liver were then rapidly excised. Potential confounding by cirrhosis severity and comorbidities was considered, and efforts were made to collect standardized echocardiographic and biochemical data.
Patient cases of liver cirrhosis
Data were collected from 65 patients with liver cirrhosis treated at the First Affiliated Hospital of Anhui Medical University between 2021 and 2025. The inclusion criteria required a diagnosis of liver cirrhosis with no history of cardiovascular disease. The control group consisted of 39 healthy individuals who underwent routine physical examinations at the same hospital. Informed consent was obtained from all patients and healthy controls. This study was approved by the Biomedical Ethics Committee of Anhui Medical University (Ethics Number: PJ2024-05-76), and all procedures complied with the ethical principles of the Declaration of Helsinki and the International Ethical Guidelines for Biomedical Research.
Echocardiography
Mice were anesthetized with isoflurane (R510-22-10; RWD, China), and two-dimensional M-mode echocardiography was performed at two, four, and eight weeks using a VINNO 6LAB ultrasound system (VINNO, China). The E/A ratio was assessed by pulsed Doppler echocardiography, and the E/e′ ratio was measured by tissue Doppler echocardiography.
Electrocardiography
Mice were anesthetized with isoflurane (R510-22-10; RWD, China) and placed in the supine position on the table, secured with tape. Subcutaneous needle electrodes from a commercial electrocardiography system (BL-420F; Chengdu Tailian Technology Co., Ltd., China) were then attached to record the electrocardiogram.
Treadmill exercise performance test
Mice were first acclimated to the treadmill for 5 min at an initial speed of 2 m/min. The speed was then increased by 2 m/min every 2 min up to a maximum of 16 m/min. The test was terminated when a mouse remained on the active electric grid at the rear of the treadmill at least 18 times for more than 10 s each.
Blood pressure measurement
Systolic blood pressure and diastolic blood pressure were measured at the tail using a Softron BP-2010 animal sphygmomanometer. Before the formal test, all mice were trained to adapt to the measurement procedure. Measurements were obtained while the mice were calm in a warm (37°C), dark environment. Blood pressure was measured at least three times, and the average value was recorded.
Histopathology
Fixed tissues were sectioned at a thickness of 5 µm for histological analysis. Sections were stained with hematoxylin and eosin (B006 & B005; ebiogo, China), Masson’s trichrome (B022; ebiogo, China), wheat germ agglutinin (L4895; Sigma-Aldrich, USA), and 0.1% Sirius Red (BP-DL030; Sbjbio, China) following the manufacturers’ protocols. Stained sections were examined and imaged using a 3D HISTECH Panoramic MIDI system, and whole-slide scans were analyzed with CaseViewer software.
Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining
TUNEL staining was performed as previously described.10 After deparaffinization, cardiac tissue sections were incubated with 20 µg/mL DNase-free protease, followed by TUNEL detection solution. The reaction was terminated with labeled reaction stop solution, after which streptavidin–horseradish peroxidase working solution was applied. After washing, 3,3′-diaminobenzidine tetrahydrochloride color development solution was added, and the sections were counterstained with hematoxylin. Finally, the sections were differentiated with 1% hydrochloric acid–ethanol solution. Stained sections were examined and imaged using a 3D HISTECH Panoramic MIDI system, and whole-slide scans were analyzed with CaseViewer software.
Immunohistochemistry
Immunohistochemistry was performed as described previously.11 Fixed tissues were sectioned at a thickness of 5 µm for immunohistochemical analysis. The expression of interleukin (IL)-1β, IL-6, p65, and tumor necrosis factor (TNF) was detected using microwave-assisted antigen retrieval. Sections were incubated overnight at 4°C with primary antibodies: mouse anti-IL-1β (1:50), rabbit anti-IL-6 (1:50), mouse anti-p65 (1:50), and rabbit anti-TNF (1:50). After incubation with the appropriate secondary antibodies, staining was visualized using 3,3′-diaminobenzidine tetrahydrochloride, and the slides were counterstained with hematoxylin. Stained sections were examined and imaged using a 3D HISTECH Panoramic MIDI system, and whole-slide scans were analyzed with CaseViewer software.
Western blotting
Western blotting was performed as previously described.12 Total protein was extracted from treated heart tissue using radioimmunoprecipitation assay lysis buffer (P0013B; Beyotime, China) supplemented with protease inhibitors (P1005; Beyotime, China) and phosphatase inhibitors (ST506-2; Beyotime, China). Protein concentrations were determined using the BCA Protein Assay Kit (P0010; Beyotime, China). Equal amounts of protein were separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (8%–12%) and transferred onto polyvinylidene fluoride membranes (P2120; Millipore, USA). Membranes were blocked with 5% skim milk for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: B-cell lymphoma 2 (Bcl-2; 1:1,000, 60178-1-Ig; Proteintech, China), Bcl-2-associated X protein (Bax; 1:1,000, 60267-1-Ig; Proteintech, China), caspase-3 (1:1,000, 19677-1-AP; Proteintech, China), lactate dehydrogenase A (LDHA; 1:1,000, 19987-1-AP; Proteintech, China), pyruvate kinase M2 (PKM2; 1:1,000, 60268-1-Ig; Proteintech, China), carnitine palmitoyltransferase 1A (CPT1A; 1:1,000, 15184-1-AP; Proteintech, China), phosphorylated p65 (1:1,000, PAB53260; Bio-Swamp, China), p65 (1:1,000, MAB51128; Bio-Swamp, China), and β-actin (1:10,000, 81115-1-RR; Proteintech, China). After washing, membranes were incubated for 1 h at room temperature with either anti-rabbit (1:10,000, ZB-2301; Origene, China) or anti-mouse (1:10,000, SA00001-1; Proteintech, China) secondary antibodies, followed by additional washing. Protein bands were visualized using an enhanced chemiluminescence reagent (BL161A; Biosharp, China), and densitometric analysis was performed with ImageJ software.
Reverse transcription quantitative polymerase chain reaction (RT-qPCR)
Total RNA was extracted from snap-frozen heart tissue using TRIzol reagent (Invitrogen, Grand Island, NY, USA). RT-qPCR was performed with SYBR® Green and mouse-specific primer sets. Relative gene expression was calculated using the 2–ΔΔCt method, with normalization to glyceraldehyde-3-phosphate dehydrogenase.13 The primer sequences used in this study are listed in Supplementary Table 1.
Electron microscopy
Animals were euthanized with an overdose of isoflurane (RWD, China). A 1 mm3 piece of cardiac tissue was excised with a surgical blade and immediately fixed in 2.5% glutaraldehyde for 24 h. The fixed tissue was embedded in pure epoxy resin and sectioned at a thickness of 70 nm for histological analysis. Sections were stained with Grade B stain (22400; Electron Microscopy Sciences, USA) and examined using a transmission electron microscope (JEM1400; JEOL, Japan).
Statistical analysis
Data are expressed as mean ± standard error. Group differences in normally distributed data were analyzed using one-way analysis of variance followed by Tukey’s post hoc test, performed with GraphPad Prism version 9.5 (GraphPad Software, San Diego, CA, USA). Non-normally distributed data were analyzed using the Kruskal–Wallis test in SPSS version 17 (IBM Corp., Armonk, NY, USA). Categorical variables were compared using the chi-squared test. Each experiment was repeated three times, and statistical significance was defined as p < 0.05.
Results
BDL-induced liver injury and cirrhosis in mice
Pathological examination of the liver was performed across the five groups of mice. After BDL, mice developed severe cholestasis by two weeks, which progressively worsened and led to reduced liver weight (Fig. 1A). Serum analysis showed significant increases in alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, and total bilirubin at two weeks, indicating liver dysfunction. Aspartate aminotransferase levels rose at two weeks, then declined at four to eight weeks but remained higher than those in the control group (Fig. 1B). Histological assessment revealed extensive structural changes. Hematoxylin and eosin staining showed distorted liver architecture in mice after BDL, with marked ductal proliferation, enlarged hepatocyte nuclei, hepatocyte swelling, pseudolobule formation, and inflammatory cell infiltration. By two weeks, partial confluent necrosis appeared in the central region around the lesion, and the necrotic areas expanded progressively (Fig. 1C). Masson’s trichrome staining demonstrated intense collagen deposition around the central hepatic vein, with fibrosis evident in the hepatic bile ducts. Morphological analysis revealed dark-stained masses with pronounced radial adhesion of collagen fibers in hepatocytes, and the degree of fibrosis increased steadily over time (Fig. 1D). Together, these pathological findings confirmed that mice subjected to BDL developed severe liver injury, progressing to cirrhosis by four weeks.
BDL-induced cardiac dysfunction in mice
Echocardiography was performed to evaluate cardiac function. Mice subjected to BDL showed impaired cardiac performance, with significant reductions in end-diastolic interventricular septal thickness, end-diastolic left ventricular internal diameter, end-systolic left ventricular internal diameter, stroke volume (SV), left ventricular end-diastolic volume, cardiac output, and heart rate. Notably, the decreases in end-systolic left ventricular internal diameter, SV, left ventricular end-diastolic volume, and cardiac output at two weeks were more pronounced than those observed at four weeks. These changes were accompanied by bradycardia and thickening of the left ventricular posterior wall at the end of systole. Although left ventricular ejection fraction (EF) and fractional shortening (FS) were slightly increased, cardiac output was significantly reduced (Fig. 2A and D). Pulsed Doppler and tissue Doppler echocardiography further demonstrated diastolic dysfunction, characterized by a decreased E/A ratio and an increased E/e′ ratio (Fig. 2B). Mice after BDL also exhibited significant reductions in both distance and time on the treadmill exercise test (Fig. 2C). Severe hemodynamic abnormalities were observed, including reductions in systolic blood pressure, diastolic blood pressure, and mean arterial pressure (Fig. 2D). Furthermore, electrocardiograms showed prolonged corrected QT (QTc) and QRS intervals. QTc increased most prominently at two weeks, then declined at four to eight weeks, but remained significantly elevated compared with controls (Supplementary Fig. 1A). Together, these findings demonstrate that mice subjected to BDL develop significant impairment of cardiac function.
BDL-induced cardiac damage in mice
We next examined the cardiac pathology and physiology of mice subjected to BDL. Serum analysis following BDL indicated the onset of cardiomyopathy. Markers of myocardial injury, including creatine kinase and creatine kinase-MB, gradually decreased from two to eight weeks after BDL. In contrast, brain natriuretic peptide (BNP), its precursor N-terminal pro-B-type natriuretic peptide, and cardiac troponin I increased significantly at two weeks post-surgery, then declined at four and eight weeks but remained elevated compared with controls, indicating persistent myocardial injury (Fig. 3A). Heart weight and heart-to-tibia length ratio were significantly reduced in mice subjected to BDL compared with controls (Fig. 3B), and reduced heart volume was further confirmed by morphological analysis (Fig. 3C). Wheat germ agglutinin staining showed that myocardial cell diameter decreased progressively, with reductions beginning at two weeks and becoming more pronounced at four weeks (Fig. 3D). These findings suggest that CCM induces ventricular remodeling, characterized primarily by myocardial atrophy. Histological evaluation supported these observations. Hematoxylin and eosin staining demonstrated that myocardial cells in controls were tightly and orderly arranged, with evenly stained cytoplasm and clearly defined nuclei. In contrast, myocardial fibers in mice with CCM were hypertrophic, thickened, and loosely and irregularly arranged, with enlarged intercellular spaces, variable nuclear size, and occasional nuclear fusion. Inflammatory cell infiltration was markedly increased, with clusters surrounding blood vessels and extending into adjacent tissues (Fig. 3E).
BDL-induced myocardial apoptosis and fibrosis in mice
Protein expression analysis of Bax, Bcl-2, and caspase-3 demonstrated that cardiomyocytes in mice with CCM underwent apoptosis, with minimal apoptosis at two weeks, significant apoptosis at four weeks, and further increases at eight weeks (Fig. 4A). TUNEL staining confirmed progressive myocardial apoptosis (Fig. 4B). Masson’s trichrome and Sirius Red staining revealed abundant collagen fiber deposition in the interstitial and perivascular regions of the myocardium in mice subjected to BDL. Fibrosis was mild at two to four weeks but became more pronounced by eight weeks (Fig. 4C and D). RT-qPCR analysis showed that the expression of fibrosis markers α-smooth muscle actin, collagen type I alpha 1, and transforming growth factor-β1 progressively increased over time after BDL (Fig. 4E). These findings indicate that myocardial fibrosis is a key contributor to the development of CCM.
BDL-induced myocardial inflammation in mice
Inflammation plays a crucial role in cardiac injury; therefore, we further examined inflammatory factors in mice subjected to BDL. Gene expression analysis showed significantly higher levels of IL-1β, IL-6, p65, and TNF in myocardial tissue, with TNF expression peaking at two weeks and declining slightly at four weeks (Fig. 5A). Consistently, immunohistochemistry revealed markedly elevated levels of IL-1β, IL-6, p65, and TNF in myocardial tissue, indicating increased inflammatory cell infiltration (Fig. 5B). Western blotting further confirmed upregulated expression of phosphorylated p65 and total p65 proteins, supporting the involvement of inflammatory signaling pathways in the pathogenesis of CCM (Fig. 5C).
BDL-induced cardiac mitochondrial dysfunction in mice
Given that the heart is the most energy-demanding organ in the body, we investigated mitochondrial alterations in the hearts of mice subjected to BDL. Morphological examination revealed cardiomyocyte edema, widened nuclear spaces, nuclear condensation, mitochondrial swelling, and cristae rupture at two weeks. By eight weeks, mitochondrial damage had further progressed, with persistent swelling and rupture (Fig. 6A). Western blot analysis demonstrated a progressive decline in peroxisome proliferator-activated receptor alpha, a marker of mitochondrial biogenesis, as well as CPT1A, a key enzyme in fatty acid β-oxidation, from two to eight weeks post-surgery. In contrast, the expression of glycolytic enzymes, including hexokinase 2, PKM2, and LDHA, was significantly increased in the hearts of mice with CCM. These findings indicate that BDL induces severe mitochondrial injury in the heart, characterized by reduced β-oxidation and enhanced glycolysis, suggesting a metabolic shift from fatty acid β-oxidation to glycolysis (Fig. 6B).
Cardiac dysfunction in patients with cirrhosis
Compared with healthy controls, patients with cirrhosis showed significantly elevated liver function markers, including aspartate aminotransferase, alkaline phosphatase, alanine aminotransferase, total bile acid, total bilirubin, and lactate dehydrogenase (Fig. 7A). Echocardiographic analysis demonstrated significant increases in end-diastolic interventricular septal thickness, end-diastolic left ventricular posterior wall thickness, and left atrial diameter, whereas no significant differences were observed in EF or FS, suggesting myocardial hypertrophy or dilation in these patients (Fig. 7B). The 65 patients with cirrhosis were then classified into two groups: compensated cirrhosis and decompensated cirrhosis. Patients with decompensated cirrhosis had significantly higher total bilirubin, total bile acid, and BNP levels compared with those in the compensated cirrhosis group (Fig. 7C). Echocardiographic parameters, including aortic root diameter, left ventricular diameter, and left atrial diameter, were also significantly elevated in the decompensated cirrhosis group, whereas EF and FS did not differ significantly (Fig. 7D). Correlation analysis showed that total bilirubin and total bile acid were positively associated with BNP, QT, QTc, SV, left ventricular diameter, left atrial diameter, and aortic root diameter, and negatively associated with EF and FS. These findings indicate that the extent of cardiac dysfunction in patients with cirrhosis is closely linked to disease severity (Supplementary Tables 2 and 3).
Discussion
Echocardiography is considered the most reliable method for assessing cardiac function in CCM. In this study, echocardiography was first performed to evaluate cardiac performance in mice. The E/A ratio reflects left ventricular inflow velocity during early rapid passive filling (E-wave) and atrial contraction (A-wave); in diastolic dysfunction, the E/A ratio falls below 1. Furthermore, the E/e′ ratio serves as an indicator of left ventricular filling pressure and is less dependent on loading conditions in the presence of diastolic dysfunction. As diastolic function deteriorates, e′ decreases and the E/e′ ratio increases.14,15 Shahvaran et al. summarized echocardiographic data from 615 patients with cirrhosis and reported that the mean E/A ratio and mean left ventricular EF were significantly lower in patients with cirrhosis than in healthy controls.16 In our BDL mouse model, the E/A ratio was consistent with that observed in patients with cirrhosis, although the left ventricular EF in mice subjected to BDL was slightly higher than that in controls. This finding contrasts with the 2020 diagnostic criteria for CCM established by the Cirrhotic Cardiomyopathy Consortium, which define systolic dysfunction as an EF < 50%. EF, a key afterload-dependent parameter, is the ratio of SV to end-diastolic volume. The increase in EF values observed in our study may reflect a smaller reduction in SV relative to the decrease in end-diastolic volume. Indeed, several murine studies have reported preserved or even increased EF in CCM.7,17–19 Likewise, clinical studies suggest that CCM is often characterized by diastolic dysfunction and preserved systolic function at rest, with normal or elevated EF.20–22 Bushyhead et al. further reported that increased EF is associated with worse outcomes in cirrhosis.23 In our clinical cohort, increased end-diastolic left ventricular posterior wall thickness, end-diastolic interventricular septal thickness, and left atrial diameter, together with unchanged EF and FS, indicated impaired diastolic function with preserved systolic function, findings that align with previous research. We hypothesize that this discrepancy may reflect the heterogeneous phenotypes of CCM. CCM develops gradually, progressing from a compensated to a decompensated state, and typically manifests as chronic cardiac dysfunction with myocardial hypertrophy, systolic or diastolic impairment, and electrophysiological abnormalities. Myocardial hypertrophy, induced by increased cardiac load, represents a compensatory mechanism. However, the BDL mouse model is relatively acute, inducing CCM over a short timeframe without allowing for the development of long-term compensatory hypertrophy. In our study, heart weight decreased, myocardial cells underwent atrophy, and end-diastolic left ventricular internal diameter, end-systolic left ventricular internal diameter, and left ventricular end-diastolic volume all declined. Consequently, myocardial atrophy and reduced ventricular volume contributed to an increased rather than decreased EF.
We monitored blood pressure in mice and observed hypotension following BDL surgery. In patients with cirrhosis, alterations in liver structure and metabolic function increase intrahepatic vascular resistance and cause portal hypertension. Portal hypertension,24 together with arterial vasodilation, results in central hypovolemia, which in turn leads to hypotension, activation of a hyperdynamic circulatory state, and stimulation of potent vasoconstrictor systems. These hemodynamic changes contribute to complications such as CCM.25 Our study showed that BDL induces electrocardiographic abnormalities, most notably prolongation of the QTc interval. The QT interval on the electrocardiogram represents the time from the onset of the QRS complex to the end of the T wave, reflecting the total duration of ventricular depolarization and repolarization, which generally corresponds to ventricular contraction during systole. Because the QT interval is influenced by heart rate, the QTc interval is commonly used to determine whether QT prolongation is present. A prolonged QTc interval reflects slowed depolarization and increased action potential duration.26 Notably, 30%–70% of patients with cirrhosis present with prolonged QT intervals,27 highlighting the importance of monitoring the QTc interval in patients with CCM.
In our study, mice with CCM showed increased circulating cardiac biomarkers, including BNP, N-terminal pro-B-type natriuretic peptide, and cardiac troponin I, indicating potential cardiac dysfunction. BNP and N-terminal pro-B-type natriuretic peptide, secreted by the ventricles, are elevated in both compensated and decompensated cirrhosis in response to myocardial damage and hypertrophy.28,29 Cardiac troponin I is a key marker of myocardial damage and is elevated in some patients with cirrhosis, who often exhibit subclinical myocardial injury, such as reduced SV and left ventricular mass index.30 These findings are consistent with our observations.
The results of this study clearly demonstrate that CCM is associated with myocardial fibrosis and apoptosis, both of which play central roles in cardiac dysfunction. Masson’s trichrome staining revealed collagen fiber accumulation in the myocardial interstitium and perivascular regions, indicating structural alterations secondary to chronic liver disease. Fibrosis increases myocardial stiffness, impairing both systolic and diastolic function. RT-qPCR results showed progressive upregulation of fibrotic markers, such as α-smooth muscle actin, collagen type I alpha 1, and transforming growth factor-β1, from two to eight weeks after BDL surgery, reflecting worsening fibrosis over time. This accumulation disrupts normal myocardial architecture and function, leading to reduced contractility and increased cardiac workload in CCM. These findings are consistent with those of Isaak et al., who reported that myocardial fibrosis in cirrhosis correlates with systemic inflammation and the degree of cirrhosis on magnetic resonance imaging.31 Such structural changes exacerbate cardiac dysfunction.
Beyond fibrosis, cardiomyocyte apoptosis is a critical factor in the progression of CCM. In our study, pro-apoptotic markers such as Bax and caspase-3 were upregulated, whereas the anti-apoptotic protein Bcl-2 was downregulated, indicating enhanced cardiomyocyte apoptosis in mice subjected to BDL. The activation of apoptotic pathways in CCM may be driven by the chronic inflammatory state of cirrhosis. As cirrhosis advances, inflammatory cytokines and reactive oxygen species contribute to mitochondrial dysfunction and DNA damage, triggering cardiomyocyte apoptosis. This loss of cardiomyocytes reduces cardiac contractile capacity and further promotes myocardial fibrosis. Nam et al. also reported a link between apoptosis and cardiomyopathy in bile duct-ligated mice, showing that inhibition of apoptosis alleviated myocardial damage.4 The persistence of apoptosis in our CCM model suggests that therapeutic strategies targeting apoptotic pathways may help preserve myocardial function in patients with cirrhosis.
This study also showed that inflammatory cytokines, including IL-1β, IL-6, IL-18, p65, and TNF, were significantly elevated following BDL, consistent with previous reports implicating inflammation in CCM.32 The activation of p65 has been reported to play a major role in reducing cardiac contractility in CCM.33 Moreover, oxidative stress and p65 activation enhance the production of proinflammatory mediators such as TNF and IL-1β. In turn, excessive TNF stimulates p65 transcription and promotes oxidative stress in cardiomyocytes.34 Studies have also demonstrated that TNF reduces cardiac contractility in mice subjected to BDL through the p65–inducible nitric oxide synthase and p38 mitogen-activated protein kinase signaling pathways.35 These findings suggest that inhibiting TNF may improve cardiac contractility during systemic inflammation. In addition, inflammation-induced mitochondrial dysfunction can impair fatty acid oxidation and promote a glycolytic shift, thereby contributing to the progression of CCM.36
Although not traditionally classified as a metabolic organ, the heart is the most energy-demanding organ in the body.37,38 Under normal conditions, 60%–90% of cardiac adenosine triphosphate is generated through fatty acid oxidation, with the remainder derived from pyruvate oxidation.37 However, in pathological states, overt cardiac dysfunction is often associated with reduced fatty acid oxidation.39–42 To assess whether this occurs in CCM, we examined the effects of BDL on fatty acid β-oxidation. CPT1A, the rate-limiting enzyme of β-oxidation in mitochondria, was significantly reduced in cardiac tissue from mice subjected to BDL. This finding suggests a link between impaired fatty acid oxidation and the development of CCM. Alterations in mitochondrial fission dynamics have been shown to disrupt cellular metabolism,43 and mitochondrial fission is associated with a glycolytic metabolic profile.44 Consistent with this, we observed increased levels of glycolytic enzymes, including hexokinase 2 and PKM2, in the hearts of BDL-treated mice, indicating a metabolic shift toward glycolysis and activation of the Warburg effect, which is characterized by enhanced aerobic glycolysis and lactate production.45 Furthermore, PKM2 can translocate to the nucleus, induce the expression of glycolytic enzymes such as LDHA,46 and contribute to pro-apoptotic processes.47 In line with this, our study demonstrated increased expression of hexokinase 2, PKM2, and LDHA, suggesting that similar metabolic reprogramming occurs in the hearts of mice with CCM. These findings raise the possibility that targeted mitochondrial therapies may mitigate CCM-associated cardiac damage by improving myocardial energy metabolism. Vázquez-Abuín et al. reported that stimulation of soluble guanylate cyclase improved cardiac function and mitochondrial activity in a rat model of early-stage heart failure with preserved EF.48 Similarly, Moheimani et al. demonstrated that metformin and cyclosporine A exert cardioprotective effects in patients with liver cirrhosis through modulation of cardiac energy metabolism,49 potentially by enhancing myocardial mitochondrial function.