Liver-spleen axis: Deciphering a crucial crosstalk in non-alcoholic fatty liver disease progression and therapeutic implications

Abstract

Non-alcoholic fatty liver disease (NAFLD), recently renamed metabolic dysfunction-associated steatotic fatty liver disease, is a highly prevalent metabolic liver disorder, closely linked to obesity, insulin resistance, and chronic low-grade inflammation. While traditionally viewed through the liver-adipose-gut axes, emerging evidence identifies the liver-spleen axis as a critical pathway in disease progression. This review examines the anatomical and portal vascular connections that facilitate direct communication between these organs, focusing on how splenic immune reprogramming—specifically involving myeloid-derived suppressor cells, natural killer/natural killer T cells, and pro-inflammatory signaling—drives the transition from simple steatosis to advanced cirrhosis. We further explore the diagnostic utility of splenic volume, metabolic activity, and stiffness as non-invasive markers that correlate with the severity of hepatic fibrosis and portal hypertension. In conclusion, the spleen serves as a vital immunometabolic regulatory hub and a sentinel for hepatic stress. Recognizing this axis offers significant potential for precision diagnostics and novel therapeutic interventions. Future research should focus on longitudinal intervention trials and multi-omics studies to fully decode the signaling crosstalk within the NAFLD spectrum, shifting the management of the disease toward a more systemic, integrated approach.

Key Words: Immune cells; Diagnostic imaging; Spleen longitudinal diameter at ultrasound; Portal circulation; Chronic inflammation; Comorbidities; Central nervous system involvement; Therapeutic approach

Core Tip: Traditional models of non-alcoholic fatty liver disease focus on the “liver-adipose-gut axis,” but emerging evidence establishes the “liver-spleen axis” as a critical immunometabolic rheostat. The spleen acts as an active “priming station” where metabolic stress triggers immune reprogramming. Splenic monocytes and pro-inflammatory cytokines are deployed via the portal circulation to the liver, accelerating steatohepatitis and fibrosis. Clinically, spleen longitudinal diameter measured by ultrasound and splenic metabolic activity assessed via positron emission tomography/computed tomography serve as high-reliability, non-invasive biomarkers of disease severity. Targeting this axis offers a novel frontier for precision diagnostics and interventions to halt metabolic and systemic inflammatory decay.

INTRODUCTION

Non-alcoholic fatty liver disease (NAFLD) represents a spectrum of hepatic steatosis (HS) evolving to fibrosis, cirrhosis, and hepatocellular carcinoma in a subset of patients. The nomenclature for NAFLD was recently updated to metabolic dysfunction-associated steatotic fatty liver disease (MASLD). The revised definition mandates the co-occurrence of HS with the presence of at least one of five specified cardiometabolic criteria, including body mass index (BMI), fasting serum glucose levels, blood pressure, plasma triglycerides, or high-density lipoprotein-cholesterol levels. Although NAFLD and MASLD differ conceptually, their diagnostic overlap is substantial. Given the widespread use of NAFLD in the literature cited here, we retained the NAFLD terminology throughout this manuscript[1]. Consequently, and due to the prevalent use of the NAFLD designation in the majority of literature cited herein, we shall uniformly employ the NAFLD terminology throughout the entirety of this manuscript. The progression of NAFLD has been confirmed to be multifactorial, where distinct, yet interrelated, parallel insults converge to drive disease progression[2]. Insulin resistance (IR) is central to this mechanism, promoting de novo lipogenesis, peripheral lipolysis and increasing the flux of free fatty acids to the liver. The resulting accumulation of ectopic lipids, which includes toxic species such as ceramides and diacylglycerols, defines lipotoxicity; these compounds directly induce hepatocellular injury and mitochondrial dysfunction[3,4]. The impairment of mitochondrial β-oxidation and enhancement of fatty acid metabolism generates excessive reactive oxygen species (ROS), leading to significant oxidative stress[5]. This cellular damage activates pattern recognition receptors and the inflammasome (e.g., NOD-like receptor pyrin domain-containing protein 3 [NLRP3]), culminating in a state of low-grade, sterile chronic inflammation driven by the release of pro-inflammatory cytokines, including tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6)[6]. The intrinsic (mitochondrial) pathway of apoptosis is highly relevant in NAFLD, governed by the B-cell lymphoma 2 (Bcl-2) family of proteins. Anti-apoptotic members of this family, such as Bcl-2 itself and B-cell lymphoma xL, prevent mitochondrial membrane permeabilization, thereby inhibiting the release of pro-apoptotic factors like cytochrome C[7]. Serum Bcl-2 concentrations were significantly higher in the simple steatosis group than in the nonalcoholic steatohepatitis (NASH) group, suggesting that apoptotic activity is inversely correlated with disease severity[8]. In progressive NAFLD, however, the expression and activity of pro-apoptotic members (e.g., Bcl-2-associated X protein and Bcl-2 homologous antagonist/killer) increase relative to their anti-apoptotic counterparts, leading to mitochondrial dysfunction and the initiation of the caspase cascade, culminating in hepatocyte death and subsequent inflammation[9]. Furthermore, gut microbiome dysbiosis plays a pivotal role as a disease modifier. Altered microbial composition compromises the intestinal epithelial barrier, enabling the translocation of bacterial products, such as lipopolysaccharide, to the liver via the portal circulation, thereby amplifying the inflammatory cascade and exacerbating hepatic injury[10,11]. Despite its close anatomical and functional links to the liver, the spleen has long been overlooked in NAFLD pathophysiology[12]. This organ plays an important role in immune system modulation (regulatory B cells [Bregs] and regulatory T cells [Tregs]) and maintaining peripheral tolerance through the clearance of circulating apoptotic cells, T and B cell differentiation and activation, and antibody production in the white pulp[13]. Traditional organ-system axes in NAFLD include the gut-liver axis and adipose-liver axis. However, an emerging concept is the “liver-spleen axis”—a bidirectional interaction between the liver and spleen mediated via vascular, immunologic and metabolic pathways (Figure 1)[14].

Figure 1 Crosstalk between the spleen and liver. The liver-spleen axis is governed by a dynamic bidirectional immune-cell trafficking network in which splenic monocytes, natural killer (NK) cells, and lymphocytes are mobilized via sphingosine-1-phosphate (S1P)/S1P receptor 1-driven egress, guided through the portal circulation by C-X-C motif chemokine ligand 9/10 (CXCL9/10) and CXCL12 chemokine gradients, arrested on liver sinusoidal endothelial cells through selectin-P-selectin glycoprotein ligand-1 and intercellular adhesion molecule-1/Lymphocyte function-associated antigen-1 integrin-mediated firm adhesion, and conditioned within the hepatic microenvironment by reciprocal cytokine signals including interleukin (IL-6)/Janus kinase/signal transducer and activator of transcription 3, interferon gamma, transforming growth factor-beta/SMAD, and tumor necrosis factor alpha, while the liver in turn reprograms splenic immune tone through portal delivery of lipopolysaccharides/Toll-like receptor 4 (TLR4) agonists, bile acids, damage-associated molecular patterns-driven NOD-like receptor pyrin domain-containing protein 3/IL-1β, hepatokines such as fibroblast growth factor 21 and hepatocyte growth factor, and extracellular vesicle-encapsulated microRNA cargo, with the entire axis held in homeostatic balance by regulatory checkpoints including programmed cell death protein 1/programmed death-ligand 1 on liver sinusoidal endothelial cells, T-cell immunoglobulin and mucin domain-containing protein 3/galectin-9-mediated NK and T-cell exhaustion, cytotoxic T-lymphocyte associated protein 4/cluster of differentiation 80 (CD80)/CD86 regulatory T cell (Treg) suppression, and indoleamine-driven tryptophan depletion that collectively sustain hepatic immune tolerance. Cellular players: Kupffer cells, hepatic stellate cells, Tregs, regulatory B cells (Bregs), macrophages, and dendritic cells. Immune signaling: Tumor necrosis factor alpha (TNF-α), IL-6, IL-10, CXCL10, nuclear factor kappa B (NF-κB), TLR4; nonalcoholic steatohepatitis (NASH). Pathological progression: From steatosis to NASH, including fibrosis and hypoxia. Metabolic dysfunction: Lipid disruption, insulin resistance, de novo lipogenesis.

Eosinophils, alongside splenic and hepatic monocytes, are essential components of components of the liver-spleen axis. Data show that splenectomy results in a long-term impairment in the regulation of eosinophils, an effect that becomes obvious during periods of acute stress. This indicates that the spleen is critical for maintaining normal eosinophil control, and that an excessive eosinophil response could be a mechanism behind the adverse outcomes observed in patients without a spleen[15]. To determine cause and effect, researchers applied a two-sample Mendelian randomization method to 91 inflammatory proteins and NAFLD. The analysis revealed that three inflammatory biomarkers (eotaxin, chemokine that features prominently, osteoprotegerin, and TNF receptor superfamily member 9) are genetically associated with an increased risk of NAFLD, strongly suggesting a causal relationship where higher levels of these proteins lead to a higher disease risk[16]. Accordingly, serum eotaxin concentrations were distinctly associated with TNF-α, IL-6, IL-1β, vascular endothelial growth factor (VEGF) and platelet-derived growth factor levels but not with C-reactive protein (CRP), fibrinogen, heat shock protein-70, or spleen size in patients with NAFLD. Among metabolic and anthropometric parameters, eotaxin showed significant predictive power for IR expressed as homeostatic model assessment (HOMA). NAFLD was distinctly associated with HOMA. Interestingly, carotid intima-media thickness was strongly predicted by both eotaxin levels and NAFLD severity on ultrasound (US), although no correlation was observed between eotaxin and NAFLD severity[17].

THE FOUNDATION OF THE AXIS

In healthy adults, the spleen measures approximately 11 cm in length and weighs about 150 g. It is positioned beneath the 9^th to 12^th thoracic ribs and functions as an intraperitoneal organ with a smooth serosal surface. It is attached to the retroperitoneum by fatty ligaments that also contain its vascular supply. The splenic surfaces are described relative to their locations and are termed the diaphragmatic (phrenic) and visceral surfaces, with the visceral surface divided into an anterior or gastric border and a posterior or renal portion, while the splenic hilum is directed anteromedially and the splenic artery and vein emerge from the splenic hilum in the form of six or more branches, with the splenic artery being remarkable for its large size and tortuosity and positioned slightly superior to the vein[18]. The spleen is part of the lymphatic system, with germinal centers supplied by arterioles called penicilliary radicles, and is derived from mesenchymal tissue rather than endodermal tissue like most gut organs[19]. Its microarchitecture is highly sophisticated and little is known about specific processes that are performed in its differentiation, with mesenchymal, hematopoietic and endothelial cells interacting with each other thanks to complex, organized and still undiscovered signals leading to the development of its complex micro-architecture, deepened in[20]. The spleen drains via the splenic vein, which joins the portal vein and thereby delivers blood to the liver. Thus, splenic venous output directly impacts hepatic circulation.

In liver cirrhosis of any etiology, splenomegaly and hypersplenism are well-recognized consequences[21,22]. More recently, altered spleen size have been documented even in earlier stages of liver disease such as NAFLD[23]. In a positron emission tomography/computed tomography (PET/CT) study, splenic glucose uptake rate was higher in patients with HS compared to those without, showing an inverse correlation with hepatic computed tomography (CT) density[24]. Thus, anatomical and functional connectivity support the biological plausibility of a spleen-liver cross-talk in NAFLD.

Several studies have found that spleen volume or metabolic activity correlates with NAFLD severity, particularly as the disease progresses toward its most severe manifestation, liver cirrhosis. Combining splenic diameter with sonographic features such as echo attenuation and focal fat sparing improves differentiation between NASH and simple steatosis[25]. Tarantino et al[26] conducted an US study showing that patients with NASH had higher spleen longitudinal diameter (SLD) values, as well as significantly higher IL-6 and VEGF concentrations than the other patients with simple HS and controls. The optimal cutoff value for SLD that best discriminated NASH from patients with HS was 116 mm (specificity 95% and sensitivity 88%); the sensitivity and specificity of this parameter were better than both IL-6 and VEGF in the same setting. Subsequently, other authors retrospectively analyzed CT images from 84 patients with histologically confirmed NAFLD. To standardize the measurement, they introduced and calculated the spleen-body index based on a defined formula. Findings suggested that spleen enlargement may be a distinctive feature of early NASH[27]. The mean value of the measured splenic volume at CT in 140 subjects was 214.6 cm³; interestingly, splenic volume did not vary significantly with the patient’s height, weight, or BMI[28]. The SLD was identified as a significant, independent predictor of obesity-related HS, a finding particularly pronounced in males of advanced age. Crucially, in assessing the mechanistic link between splenic morphology and liver disease, authors found that circulating concentrations of the pro-inflammatory cytokine IL-6 exerted a strong influence on the prediction of HS by SLD, underscoring the role of systemic inflammation[29]. Hepatic fibrosis severity is directly and positively correlated with spleen volume; specifically, patients reaching advanced stages of fibrosis exhibit marked and evident splenomegaly[30]. While total liver volume assessed by multidetector CT is a very poor predictor of underlying fibrosis, total splenic volume is associated with the degree of hepatic fibrosis[31]. Research aimed to evaluate the diagnostic efficacy of the spleen volume for accurately quantifying the severity of hepatic fibrosis across the spectrum of chronic liver disease showed that in the NAFLD subgroup, the spleen size increased with the progression of liver fibrosis, peaking at F3 (Figure 2)[32].

Figure 2 Line graph showing the trend of the spleen volume vs fibrosis in non-alcoholic fatty liver disease. Splenic volume generally increases in parallel with the progression of liver fibrosis, with the most significant increases often observed in the transition from F3 to F4, although this can be variable due to collateral circulation. NASH: Nonalcoholic steatohepatitis.

Confirming that simply measuring liver volume is not enough to determine the extent of liver damage, total splenic volume at enhanced CT exhibited a linear correlation with hepatic venous pressure gradient assessment[33]. In another study, patients with metabolic syndrome (MS), main driver of NAFLD[34]. had larger spleen size than healthy controls. SLD significantly correlated with waist circumference but not IL-10 concentrations in patients with MS[35]. Studying early atherosclerosis determined by carotid intima-media thickness, the SLD measurements at US of spleen predicted the presence of obesity-related NAFLD, but there was no prediction between the serum levels of eotaxin, chemokine associated to coronary artery disease[36] and the values of SLD[17]. Waist circumference, hip circumference, waist to hip ratio, waist to height ratio and body fat distribution, as well as body composition markers both in 525 men and 570 women, aged between 30 years and 90 years, were significantly associated with spleen volume measured by magnetic resonance imaging (MRI)[37]. Hypothesizing that obesity-induced inflammation might be responsible for larger spleen volume in the study[38]. The aforementioned findings on spleen size are in line with greater threat to national health posed by central obesity than general obesity, although both are independently associated with increased risk of NAFLD[39]. Surprisingly, in 275 patients with NAFLD, an SDL greater than 13 cm was not found, although there was a strong correlation between spleen size and body weight[40]. More recently, a large United Kingdom Biobank-based study found that larger spleen volume, assessed by neck-to-knee body Magnetic Resonance Imaging (MRI), was independently associated with NAFLD, liver fat fraction, liver volume and fibrosis-4 score, but not independently associated with type 2 diabetes mellitus[41]. Another work reviewed the role of mediterranean diet (MD), inflammation and spleen in the “liver-spleen axis” context: Splenomegaly is often observed in patients with NAFLD, reflecting chronic low-grade inflammation and immune activation[42]. The MD has been shown to significantly improve levels of specific inflammation biomarkers. This improvement is particularly notable for high-sensitivity CRP, IL-6, and IL-17. These findings support the potent role of the MD in reducing risks associated with chronic inflammation[42]. Authors conducted three-dimensional reconstruction of the spleen through a deep learning network model using a two-stage coarse-to-fine segmentation approach. Studying 90 NAFLD cases and 47 healthy controls who had received contrast-enhanced CT scan of the abdomen. Results suggested that spleen enlargement and increased metabolic activity may serve as surrogate markers of HS or impending disease progression[43]. As HS severity increases, both the portal vein and splenic diameters show a corresponding rise, resulting in a larger spleen in advanced cases[44]. The measurement of splenic length, which is the most commonly used in clinical practice, correlates well with splenic volume, assessed by CT, particularly when performed with the patient in the right lateral decubitus[45].

SPLEEN AS AN ACTIVE IMMUNOMETABOLIC DRIVER

The spleen serves as a major immunologic organ, reservoir and modulator of myeloid and lymphoid cells. In obesity and NAFLD, splenic immune reprogramming appears to contribute to hepatic inflammation. A recent mouse study found that in obese mice, fatty liver inflammation was associated with enrichment of splenic myeloid-derived suppressor cells (MDSCs) and natural killer T (NKT) cells, and a strong positive correlation between spleen and liver MDSC/NKT distributions[46]. Mechanistically, splenic immune cells may traffic to the liver via the portal circulation or release cytokines/mediators (e.g., transforming growth factor [TGF-β1], TNF-α, IL-6) that stimulate hepatic inflammation and fibrogenesis. Some review evidence suggests that TGF-β1 derived from the spleen contributes to hepatic fibrosis[47]. Recent research pinpoints a specific population of monocytes originating in the spleen—the CD11b⁺CD43hiLy6Clo subset—as key drivers of liver pathology. The specific cell surface markers CD11b⁺CD43hiLy6Clo define the sM-1 subpopulation, which exhibits distinct features even before migrating: These monocytes are defined by a mature F4/80 expression and their high output of pro-inflammatory ROS, which promote tissue damage. The particular cells actively migrate to the liver, transform into pro-fibrotic macrophages, and are directly responsible for exacerbating fibrosis. This mechanism highlights the functional connection within the spleen-liver axis and suggests that targeting the sM-1 precursor cells could be a strategy for treating liver disease[48]. Interesting research demonstrates how macrophages in the spleen worsen liver fibrosis. They signal liver cells to release C-C motif chemokine ligand 2 (CCL2), which attracts inflammatory monocytes and shifts the liver's immune environment toward a damaging M1 state[49]. Migration of classical C-X3-C motif chemokine receptor 1 (CX3CR1) monocytes from the spleen to the liver exacerbates fibrosis by altering hepatic CX3CR-1GFP cell dynamics and increasing cytokine production. These results establish the importance of the spleen-liver axis in fibrotic pathology and identify splenic monocytes as a key target for clinical intervention[50]. Similarly, authors observed a significant increase in double-negative T cells (DNTs) in the peripheral blood of patients with liver fibrosis. Mechanistically, these DNTs secrete TNF-α to activate the TNFR1-NLRP3 axis, driving hepatic stellate cell (HSC) activation and subsequent fibrosis progression[51]. Authors observed DNT cells with enhanced suppressive activity in the spleen and lymph nodes of thymectomized and CD8-deficient mice[52].

Spleen as a central component of the neuroimmune axis

The spleen is recognized as a central component of the neuroimmune axis, where sympathetic nerve fibers modulate immune responses primarily through catecholaminergic signaling. This signaling directly influences cytokine production and inflammatory responses, offering several pathways for immunomodulatory intervention. One key mechanism is the cholinergic anti-inflammatory pathway. Here, norepinephrine released by splenic catecholaminergic fibers binds to β2-adrenergic receptors located on CD4⁺ T cells. This binding prompts the T cells to secrete acetylcholine (ACh). The ACh then acts on macrophages by binding to the α7 nicotinic ACh receptor, which ultimately suppresses the production of inflammatory cytokines. In addition to this pathway, the sympathetic nervous system also engages in direct immunomodulation of splenic immunocytes. This involves regulating key genes and signaling pathways, altering cytokine secretion, and modulating ion flux to influence cellular functions directly. Given the spleen’s pivotal role, therapeutic strategies focused on its modulation have shown great promise. Physical methods, specifically electrical stimulation, is emerging as one of the most viable approaches for clinical applications in managing diseases by modulating splenic immunity[53]. Sympathetic innervation maintains expression of specific chemokines critical to immune cell organization, with chemical sympathectomy significantly reducing chemokine ligand 13, also known as B lymphocyte chemoattractant expression without affecting chemokine ligand 19 or chemokine ligand 21 expression in the spleen, the last two guiding immune cells, especially naive T cells and dendritic cells (DCs) to secondary lymphoid organs[54]. Has the notion of the spleen serving as a monocyte reservoir during acute inflammatory responses withstood scientific scrutiny over time? While it may be too early to provide a definitive answer, initial findings appear promising. Temporal tracking studies revealed that monocytes mobilized to the injured myocardium originate initially from circulating blood pools followed subsequently by splenic sources[55].

The spleen as a bioactive lipid hub: Immunometabolic drivers of hepatic fibrosis

The spleen’s influence stems from its massive population of immune cells, especially macrophages, which are central to lipid homeostasis and inflammation. The processes impacting on lipid metabolism are displayed during its macrophage activity and lipid clearance. The scavenging function is due to splenic macrophages that are highly phagocytic, much like Kupffer cells in the liver. They are crucial for clearing aged or damaged blood cells, including red blood cells, which release lipids (through enzymes like heme oxygenase 1, which breaks down heme into carbon monoxide and biliverdin upon degradation). This clearance process directly involves lipid processing and storage[56]. Another mechanistic level is related to foam cell formation: Under inflammatory or dyslipidemic conditions, splenic macrophages can accumulate lipids, transforming into foam cells. This process involves the excessive uptake of lipids, primarily oxidized low-density lipoproteins, which macrophages then store as lipid droplets in their cytoplasm[57]. This local lipid burden affects the spleen’s overall metabolic state and can influence the systemic circulation of lipids and lipoproteins. Activated immune cells, particularly macrophages and monocytes (M-1) mobilized from the spleen to the liver, significantly impact lipid metabolism by actively producing and releasing specialized lipid mediators. These molecules act as powerful paracrine and autocrine signals that directly govern inflammation, fibrosis, and cellular fate[58]. There are two major classes of bioactive lipids relevant to the spleen-liver: (1) Eicosanoids/sphingolipids and endocannabinoids: Pro-fibrotic/pro-inflammatory eicosanoids (prostaglandins (prostaglandin 2, prostaglandin D2 [PGD-2]) and leukotriene B4 [LTB-4]) are signaling molecules derived primarily from the omega-6 arachidonic acid via enzymes like cyclooxygenase and lipoxygenase and the omega-3 polyunsaturated fatty acids. Splenic immune cells can undergo a “class-switching” of enzyme expression, shifting the balance between pro- and anti-inflammatory outputs. The spleen is noted to contain elevated levels of pro-inflammatory mediators such as LTB-4 and PGD-2 in disease models[59]. When splenic monocytes migrate, they contribute these powerful pro-inflammatory signals to the liver microenvironment, which directly promotes HSC activation, in the sense that they stimulate HSC to transform into fibrogenic myofibroblasts[60]. They further amplify local inflammation and immune cell recruitment. Specialized pro-resolving mediators (SPMs), derived from omega-3/polyunsaturated fatty acid, specifically eicosapentaenoic acid and docosahexaenoic acid. They are resolvins (Rvs), protectins, and maresins (MaRs). These mediators, including Rv series and MaR series, are synthesized by macrophages and are crucial for the active resolution of inflammation. The shift from pro-inflammatory eicosanoids to SPM is a sign of tissue healing. While SPM are beneficial, their deficiency or delayed production by splenic cells can exacerbate chronic inflammation and fibrosis[61]; and (2) Sphingolipids and endocannabinoids: Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid synthesized by sphingosine kinases. It is a critical regulator of immune cell trafficking, and S1P receptor 5 is highly expressed in the spleen[62]. The concentration gradient of S1P between the blood and lymphoid tissues (like the spleen) is vital for controlling immune cell egress. High S1P in blood facilitates the exit of cells from the spleen. Low S1P in lymphoid tissue promotes retention of cells. S1P signaling, particularly through S1PR1 and S1PR3, is known to promote angiogenesis and HSC activation in the liver[63]. Thus, the spleen’s role as a major regulator of circulating S1P levels can indirectly dictate the number of migrating, pro-fibrotic immune cells entering the liver. Endocannabinoids (cannabinoid [CB]) are lipid-signaling molecules derived from arachnid acid, such as anandamide and 2-arachidonoylglycerol[64]. The CB system is highly upregulated in advanced liver disease. CB-1 receptor activation primarily functions as pro-fibrotic, stimulating HSC and increasing inflammation[65]. CB-2 receptor activation has generally anti-fibrotic and anti-inflammatory action[66]. The splenic immune cell reservoir is a likely source of these circulating macrophage-derived CB, linking splenic activation to both hepatic and vascular pathology[67]. In liver fibrosis, the sM-1 cells exacerbate disease by activating HSCs[68]. This activation process is often intertwined with metabolic dysregulation within the liver microenvironment, suggesting the spleen contributes indirectly by dispatching metabolically active immune cells[69]. In essence, the spleen regulates lipid metabolism not by being a primary processing organ like the liver or adipose tissue, but by serving as a major reservoir and processing center for immune cells whose activity (e.g., ROS production, cytokine release) directly impacts the inflammatory state, which is a powerful driver of MS and dyslipidemia[70]. Understanding this axis may provide novel diagnostic and therapeutic opportunities.

Splenic nexus: At the heart of liver decay, muscle wasting, and the estrogen shield

Excess fat mass serves as the primary architect of a persistent inflammatory state, which in turn fosters IR and the accumulation of fat within the liver. This metabolic strain, compounded by aging, disrupts the delicate growth hormone/insulin-like growth factor 1 (IGF-1) axis, where inflammation may further stifle essential IGF-I signaling. In an interesting study of overweight and obese women, authors observed that lower IGF-I availability is closely linked to higher body fat, an enlarged spleen, evaluated as SLD at US and systemic markers of inflammation, reflecting a more severe progression of NAFLD[71]. Beyond its role in managing energy and lipids, the adipocyte hormone leptin plays a critical role in liver disease. Multiple studies have established that long-term elevations in leptin levels promote steatosis, fibrogenesis, and the eventual onset of liver cancer[72]. Evidence indicates that leptin exerts an inhibitory effect on splenic lymphocyte function indirectly via the urocortin-sympathetic nervous system, contrasting with its direct stimulatory properties[73]. Recent data indicate that 17 β-estradiol regulates immune activity by reducing the mouse spleen DCs numbers and stimulation. Concurrently, it promotes an anti-inflammatory profile by increasing endocytosis and upregulating IL-10 mRNA expression[74]. The spleen has been identified as a vital site for monocyte storage and deployment during inflammatory events. This challenges the current paradigm that monocytes are merely circulating precursors that undergo terminal differentiation into DCs or macrophages only upon migrating into peripheral tissue[75]. Bone marrow-derived macrophages may traffic through the spleen on their way to the to the liver, contributing to the increased hepatic macrophage population during NAFLD progression[76]. A previously quoted study explicitly proves that a specific subset of immune cells (derived from the spleen) migrates to the liver to worsen fibrosis[48]. It provides direct evidence for the theory that the spleen acts as a command center for liver inflammation. In contrast with previous data, for some authors immunogenic DCs are closely linked to hepatic lipid accumulation, suggesting a potential pathophysiological pathway for the development and progression of NAFLD[77], even though findings support the widely held notion that liver DC are generally weak activators of immunity[78]. Women of reproductive age are relatively protected against NAFLD compared to men. However, this advantage disappears after menopause, at which point NAFLD rates in women equal or surpass those seen in men of the same age[79]. Is it possible that estrogen exerts its protective effects through spleen-related pathways, thereby driving the sexual dimorphism seen in NAFLD? Myostatin is a key driver of muscle atrophy, fat accumulation, and sarcopenia. In the context of liver disease, animal models of fibrosis show an up-regulation of the myostatin receptor, activin type IIB, on HSCs. This muscle-liver axis is further complicated in postmenopausal women, as declining estrogen levels accelerate muscle deterioration and impair protein synthesis. Together, these factors increase the risk of frailty, falls, and fractures[80-82]. An intriguing study reveals that splenic responses differ markedly between obesity-susceptible and obesity-resistant mice fed high-fat diets (HFDs), with the spleen emerging as a key tissue linking metabolic and immune dysfunction. In obesity-susceptible mice, spleen tissue showed increased myostatin expression along with elevated proinflammatory cytokine production (IL-17, IL-1β, and interferon gamma) from splenocytes, while obesity-resistant mice exhibited decreased splenic myostatin levels. Furthermore, C57BL/6 mice on HFDs demonstrated increased frequencies of activated CD4⁺ and CD8⁺ T cells in the spleen, indicating heightened immune activation in this lymphoid organ during diet-induced obesity. These findings suggest that the spleen plays a critical role in obesity susceptibility through coordinated changes in myostatin regulation and immune cell function, establishing it as an important site where growth factor signaling and inflammatory responses converge to influence metabolic outcomes[83]. For many years myostatin has been established as a principal inhibitor of skeletal muscle growth and development. Emerging evidence now demonstrates that myostatin also influences carbohydrate and lipid metabolism, particularly relevant in the context of sarcopenic obesity[84]. IL-7-deficient mice show dramatic reduction of lymphoid cells, in spleen[85]. However, it is now considered a myokine that may help regulate muscle cell development (Figure 3)[86]. The spleen serves as a vital signal-processing center that complements its role in monocyte mobilization by acting as a major source of dopamine. The splenic environment facilitates dopamine synthesis via noradrenergic fibers and tyrosine hydroxylase-containing cells; this localized neuroendocrine output, produced by both neuronal and non-neuronal cell subsets, is a critical driver of the systemic immune modulation observed in the muscle-spleen-liver loop[87]. Myosteatosis, characterized by pathological fat accumulation within skeletal muscle, engages in a bidirectional crosstalk with the spleen, whereby altered myokine and lipid signals from steatotic muscle activate splenic immune responses, which in turn release pro-inflammatory cytokines that impair muscle insulin signaling and mitochondrial function, exacerbating muscle pathology and systemic metabolic dysfunction. This dynamic interplay drives systemic inflammation, IR, and sarcopenia risk, highlighting the muscle-spleen axis as a critical mediator of metabolic inflexibility and chronic disease progression through integrated immunometabolic mechanisms[88-90]. Intramuscular triglyceride (IMTG) serves as both a vital energy reserve and a dynamic fat storage depot that expands during periods of elevated lipid availability while functioning as a fatty acid source. Observing about the cytokine network regulating the interplay between IMTG and obesity, ordered probit regression analysis revealed that US-detected HS predicted IMTG accumulation[91]. As illustrated in Figure 3, myosteatosis in skeletal muscle is hypothesized to release signals that activate splenic immune cells. These activated cells, in turn, can traffic to the liver and contribute to hepatic IR and fibrogenesis, establishing a detrimental tri-organ loop.

Figure 3 Crosstalk among the muscle, spleen, and liver. The diagram shows a bidirectional crosstalk among the skeletal muscle, spleen, and liver in the context of metabolic dysfunction and chronic inflammation. Skeletal muscle communicates with the liver through myokines, aminoacid fluxes, and insulin sensitivity pathways, influencing hepatic lipid handling and systemic glucose homeostasis. The spleen contributes immunometabolic signals, including cytokines, monocyte trafficking, and modulation of systemic inflammatory tone that shape hepatic inflammation and fibrogenesis. Reciprocal feedback from the liver, via hepatokines and metabolic intermediates, affects both muscle metabolism and splenic immune activity, highlighting a dynamic tri-organ network central to metabolic disease progression[89-91].

Iron-immune bridge

The liver and spleen synergize in iron homeostasis. The spleen’s iron salvage pathway supplies hepatic stores, maintaining systemic iron balance[92]. The spleen plays a vital role in systemic iron metabolism through its red pulp macrophages, which recycle iron from senescent erythrocytes. This process is regulated by cytokines like IL-33, which promote macrophage maturation and efficient iron recovery. During anemia, splenic hematopoietic stem cells increase iron uptake to stimulate erythroid regeneration via Tet methylcytosine dioxygenase 2 signaling. Genetic factors such as Hfe and TfR2 are also crucial for proper iron handling in splenic macrophages, and their absence can disrupt hematopoiesis (Figure 4). Additionally, iron accumulation patterns in the spleen vary with metabolic diseases, suggesting potential diagnostic applications through imaging[93-96]. The relationship between iron metabolism and NAFLD has garnered renewed scientific interest in recent years, largely due to the emergence of the ferroptosis concept—an iron-dependent mechanism of programmed cell death[97]. Because iron-rich splenocytes drive inflammation and aggravate NASH through the portal vein, managing how the spleen processes iron is a vital step in creating novel therapeutic targets for the disease[98].

Figure 4 Interleukin 33 signaling in the spleen. Diagram illustrates interleukin 33 (IL-33) signaling in splenic iron recycling, highlighting its role in red pulp macrophage maturation and erythrocyte clearance via the IL-1 receptor-like 1 (IL1RL1)-suppression of tumorigenicity 2 receptor pathway and downstream transcription factors GATA-binding factor 2 (GATA2) and Spi-C. RPM: Rat promegakaryoblast-like.

Beyond immunity: The spleen’s metabolic secret

During fasting or infection, hepatic gluconeogenesis is modulated by splenic norepinephrine release[99,100]. Research suggests the spleen plays a vital role in keeping blood sugar levels stable. This is because the spleen’s immune cells prioritize glucose uptake for energy. Evidence for this regulatory link is found in surgical history: Healthy individuals who have their spleen removed eventually develop type 2 diabetes mellitus, indicating that losing the spleen significantly disrupts the body’s ability to manage sugar[101,102]. Beyond its immunological functions, the human spleen exhibits a 47% increase in 18F-fluorodeoxyglucose uptake following Bacillus Calmette-Guérin (BCG) vaccine administration. This metabolic shift in splenic lymphocytes and monocytes provides a potential explanation for the systemic lowering of blood glucose to near-normal levels. Ultimately, these results indicate that BCG can trigger the spleen to serve as a pivotal site for glucose regulation, even when pancreatic function is absent[103]. In a classical study dealing with healthy male Fischer 344 rats, aging is associated with a coordinated elevation in the activities of key glucose metabolizing enzymes within splenic cells, reflecting an age dependent remodeling of metabolic pathways that may influence immune cell function and energy homeostasis[104]. While investigating potential immunotherapies for type 1 diabetes, authors identified a population of islet derived stem cells within the spleen capable of reversing diabetes in a murine model. Initial experiments demonstrated that splenic cells, once isolated from donor mice and co-administered into diabetic recipients together with TNF-α-inducing agent (used to selectively eliminate pathogenic T cells), migrated to the pancreas and differentiated into fully functional islet β cells, thereby restoring normoglycemia[105]. The spleen influences IR by regulating immune cells that affect inflammation in fat tissue, with spleen-Bregs producing anti-inflammatory IL-10, which protects against obesity-related IR; however, chronic inflammation associated with MS can also enlarge the spleen, suggesting a complex interplay where spleen function impacts glucose metabolism and inflammation, influencing insulin sensitivity and potentially protecting against or contributing to metabolic dysfunction (Figure 5).

Figure 5 Mechanisms of spleen-mediated insulin resistance regulation. This schematic illustrates six interconnected pathways through which the spleen modulates systemic inflammation and metabolic homeostasis[35]. NAFLD: Non-alcoholic fatty liver disease; TNF-α: Tumor necrosis factor alpha.

The spleen supplies “innate-like” B cells to visceral fat. These cells release anti-inflammatory IL-10, reducing inflammation and improving insulin sensitivity in obesity. Specific splenic immune cells (B and T cells) are crucial for maintaining an anti-inflammatory environment, impacting energy balance and adipocyte function[106]. Splenic participation in glycemic homeostasis in obese and non-obese male rats. Spleen-nerve (vagus nerve) interactions play a role in glucose homeostasis, affecting insulin secretion and pancreas function, according to this study[107]. In conditions like MS or NAFLD, the spleen may enlarge due to chronic inflammation, a hallmark of IR[108]. Recent findings showed that repeated human umbilical-mesenchymal stem cell (MSC) infusions reduced IR in the epicardial adipose tissue of mice with HFD-induced obesity, operating in part through enhanced IL-10 secretion by splenic regulatory T cells. These results offer significant evidence that interactions between MSCs and splenocytes are central to the therapeutic benefits produced by MSC treatment[109]. Exercise reduces in spleen the pro-inflammatory molecule TNF-α through cholinergic pathways, improving insulin sensitivity, according to Huang et al’s study[110]. Some small studies suggest splenectomy might increase diabetes risk, possibly due to loss of stem cells or altered glucoseregulation, while others show improved insulin sensitivity in diabetic patients after splenectomy, indicating a complex role[111].

Splenic-hepatic-central nervous system axis: Mediating lipid-driven systemic inflammation

Authors examined comprehensive gene lists for NAFLD and Alzheimer’s disease (AD)—illnesses potentially sharing mechanisms based on risk factors and previous studies. Network clustering and pathway enrichment revealed 42 pathways, with three major groups identified as key links between both diseases: Carbohydrate metabolism, long-chain fatty acid metabolism, and IL signaling pathways[112]. A recent study provides class II evidence (not definitive proof) demonstrating that nonalcoholic fatty liver disease is independently associated with an increased risk for the development of both vascular and nonvascular forms of dementia[113]. A more recent one analyzed 4582 individuals with NAFLD and 6318 control participants. Cases of AD were more frequent among patients with NAFLD than in the control group. In the fully adjusted Poisson regression model, NAFLD was associated with a higher risk of AD (relative risk = 2.80)[114]. Emerging data demonstrate that immune cells from peripheral organs such as the spleen, including monocytes, NK cells, and T lymphocytes, undergo active recruitment to the central nervous system where they could influence AD pathogenesis through their effects on neuroinflammation[115]. Accumulating evidence increasingly supports the hypothesis that impaired immune regulation, autoimmunity, and inflammatory processes are plausible mechanisms underlying AD pathogenesis. IL-6 is the plausible trigger[116]. Compared to patients with HS, those with NASH had larger spleen volumes along with significantly higher IL-6 and VEGF concentrations[117]. The findings of a systematic review and meta-analysis reveal that patients with AD exhibited elevated serum VEGF concentrations consistent with upregulated VEGF expression, though VEGF levels inversely correlated with disease severity, declining as the condition worsened[118]. Emerging evidence suggests that spleen involvement in NAFLD may contribute to neuroinflammation relevant to AD, via the brain-spleen-liver axis (Figure 6) and systemic immune dysregulation. This connection is increasingly recognized in aging and metabolic disease models[119].

Figure 6 Liver-spleen-brain axis in patients with non-alcoholic fatty liver disease/nonalcoholic steatohepatitis with Alzheimer’s disease comorbidity. Hepatic steatosis and steatohepatitis trigger systemic inflammation through Kupffer cell activation and tumor necrosis factor alpha (TNF-α) signaling, leading to insulin resistance and immune activation. The spleen acts as a central immune hub, where macrophages, dendritic cells, and monocytes are activated and recruited. These peripheral immune cells traffic to the central nervous system, where they drive neuroinflammation through microglial activation, promoting amyloid-β deposition and tau phosphorylation, key pathological hallmarks of Alzheimer’s disease. NAFLD: Non-alcoholic fatty liver disease; NASH: Nonalcoholic steatohepatitis; IL: Interleukin; CXCL10: C-X-C motif chemokine ligand 10; ROS: Reactive oxygen species; TLR4: Toll-like receptor 4; BBB: Blood-brain barrier.

Critical physiological and pathological triangle: Gut-spleen-liver axis

While the gut-liver axis is well-known for transporting microbial products to the liver, the inclusion of the spleen recognizes its role as a “priming station” and reservoir for the systemic immune cells that drive chronic liver disease. The gut-spleen-liver axis constitutes an intricate, multi-layered defense and metabolic coordination network where the integrity of the intestinal epithelial barrier dictates the initial payload entering the system. Specifically, dysbiosis or increased permeability allows microbial products like lipopolysaccharide and other pathogen-associated molecular patterns (PAMPs) to flood the portal vein. This subjects the liver’s resident macrophages, the Kupffer cells, to immediate and continuous inflammatory signaling via TLR4 activation, thereby initiating the hepatic clearance and detoxification processes. Concurrently, disseminated antigens are channeled through the mesenteric lymph nodes to the spleen, the main site for filtering blood-borne threats, where T-cell priming and B-cell differentiation orchestrate a systemic adaptive immune response. This floods the circulation with effector cells and regulatory cytokines, which ultimately feed back into the liver via the systemic circulation to drive the acute phase response by hepatocytes. This demonstrates a continuous, bidirectional regulatory loop essential for balancing intestinal homeostasis, systemic immunity, and metabolic partitioning. The microbiome’s role within this axis is foundational, acting as the primary sculptor of host immunity and metabolism. Microbial-derived signals, notably short-chain fatty acids (SCFAs) such as butyrate, profoundly influence the differentiation and function of DCs within the gut-associated lymphoid tissue. These in turn dictate the repertoire of lymphocytes that seed the systemic circulation and subsequently populate the spleen’s white pulp. This splenic population, educated by gut-derived factors, is crucial for maintaining tolerance vs mounting effective systemic defense against circulating pathogens. A dysbiotic state therefore directly translates to an aberrantly programmed splenic immune response capable of either under-reacting to systemic threats or over-reacting in autoimmune manifestations. The gut-spleen-liver axis is intimately linked to the pathogenesis and progression of NAFLD, as the chronic, low-grade inflammatory state driven by gut-derived PAMPs overwhelms Kupffer cell detoxification capacity[120]. Leading to persistent HSC activation and fibrosis, furthermore, the spleen exacerbates this process by releasing pro-fibrogenic cytokines and monocytes into the circulation during systemic inflammation. Pro-fibrogenic cytokines and monocytes contribute to the inflammatory burden that drives the transition from simple HS to more severe steatohepatitis. The microbiome’s influence on SCFA production further disrupts host lipid metabolism within the liver parenchyma, solidifying the axis’s central role in chronic metabolic liver disease (Figure 7). The impact of the non-bacterial microbiome, encompassing viruses (virome), fungi (mycobiome), and archaea, provides a crucial, yet often underestimated, dimension in the etiology and progression of NAFLD/NASH. Specifically, alterations in the virome, such as increased abundance of bacteriophages that lyse beneficial bacteria[121]. can indirectly contribute to NAFLD by disrupting SCFA production and increasing the influx of bacterial endotoxins; simultaneously, fungal overgrowth (e.g., Candida species) can directly stimulate Toll-like receptor 2 pathways in the gut, leading to heightened systemic inflammation that is ultimately processed by the spleen and then projected onto the liver, where fungal products can exacerbate hepatocyte lipotoxicity and inflammation, thus linking shifts in the entire resident microbial community, beyond just bacteria, directly to the severity of HS and subsequent progression toward advanced NAFLD. The gut-spleen-liver axis operates through a coordinated network of immune mediators that regulate inflammation, tissue repair, and immune surveillance[122,123].

Figure 7 Systemic pathophysiology of the gut-liver-spleen axis. This diagram illustrates the complex inter-organ communication and metabolic signaling that drive systemic inflammation and hepatic fibrosis. CXCL: C-X-C motif chemokine ligand; IL: Interleukin; LPS: Lipopolysaccharide; NAFLD: Non-alcoholic fatty liver disease; TGF-β: Transforming growth factor beta; TNF-α: Tumor necrosis factor alpha.

From sleep to skin to hormones: The liver-spleen axis as a common thread in chronic diseases

The liver-spleen axis represents an emerging concept in integrative medicine that describes the bidirectional communication between the liver and the spleen, with dysregulation in this axis increasingly linked to both chronic inflammation and metabolic dysfunction, characteristics of NAFLD and related, immune-mediated diseases including obstructive sleep apnea syndrome (OSAS), polycystic ovary syndrome (PCOS), and psoriasis. In OSAS, intermittent hypoxia generates oxidative stress and endothelial dysfunction that impair liver function while promoting overactivity in splenic immune cells, with gut permeability changes leading to endotoxin translocation that activates both hepatic Kupffer cells and splenic macrophages, perpetuating systemic inflammation through elevated pro-inflammatory cytokines such as IL-6, TNF-α, and CRP. Therapeutic approaches include continuous positive airway pressure therapy to reduce hypoxia, anti-inflammatory MD[124], probiotics to restore gut barrier function[125] and supplements like berberine[126] and milk thistle that support liver detoxification. In PCOS, IR and hyperinsulinemia overload hepatic metabolic function[127], while disrupting spleen-mediated immune tolerance, assessed by spleen transcriptome sequencing[128] and chronic low-grade inflammation characterized by elevated CRP, IL-6, and TNF-α activates both liver and splenic immune cells, with visceral fat-derived adipokines further disrupting liver-spleen communication[129,130] and gut dysbiosis leading to lipopolysaccharide (LPS) translocation that compounds hepatic and splenic inflammation. Interventions such as metformin and inositol improve insulin sensitivity, as well as high-fiber low-glycemic diets and omega-3 fatty acids reduce systemic inflammation, and probiotics being particularly valuable[131,132]. In psoriasis, T helper 17 (Th17)-driven immune dysregulation with elevated IL-17 and IL-23 activates hepatic Kupffer cells and splenic immune cells while increasing the risk of MS and NAFLD, which further disrupts liver-spleen immune homeostasis[133] and gut dysbiosis[134]. This increases intestinal permeability and allows bacterial metabolites to enter circulation and activate liver-spleen inflammation, making therapeutic strategies including topical and systemic immunosuppressants, gluten-free or MD, probiotics, omega-3 fatty acids useful, and N-acetylcysteine particularly effective[135]. Moreover, vitamin D[136] and magnesium modulate immune responses[137] in patients with psoriasis in the light that low Mg²⁺ diet increases indicators of inflammatory activity in the spleen[138]. Obviously, exercise and weight loss reduce visceral adiposity and improve both hepatic and splenic function, and sleep optimization that mitigates hypoxia-related damage. Patients with psoriasis experience a higher prevalence of sleep disorders that cannot be explained by sleep duration alone[139]. In medical literature, critical convergence of factors between psoriasis, OSAS, and NAFLD refers to a dangerous feedback loop where each condition worsens the others through shared inflammatory and metabolic pathways. Accordingly, a recent meta-analysis found that increased eotaxin levels were linked to a higher risk of OSAS[140].

Hemodynamic and immunological crosstalk: The spleen as a driver of hepatic dysfunction

The spleen is not just a “victim” of liver disease; its hemodynamic reaction (increasing blood volume into a blocked system) actively worsens the liver’s condition. In the context of liver cirrhosis, the spleen shifts from a protective lymphoid organ to a primary driver of hepatic inflammation and fibrogenesis through a complex bidirectional immune crosstalk. As portal hypertension causes splenic congestion, the organ undergoes structural remodeling that activates resident splenic macrophages[141] to secrete high levels of pro-fibrogenic cytokines, most notably TGF-β1, which travels through the portal vein to directly stimulate HSCs. Simultaneously, the spleen acts as a reservoir and “training ground” for inflammatory monocytes and T-cell subsets (e.g., Th17 cells) that migrate to the liver[142], exacerbating the local inflammatory microenvironment and suppressing regenerative signals like IL-6. This immunological axis effectively transforms the spleen into an endocrine-like organ that perpetuates a chronic cycle of injury, systemic inflammation, and immune dysfunction making it an active participant in the progression toward end-stage liver disease rather than a mere bystander. The spleen’s immune dysfunction plays a critical role in disrupting the gut-liver axis in the most advanced form of NAFLD, i.e. liver cirrhosis (Figure 8)[143]. This intimate relationship means the liver is constantly exposed to gut-derived substances including bacterial metabolites, SCFAs, bile acids, hormones, and PAMPs from the gut microbiome, while simultaneously the liver regulates gut homeostasis through bile acid secretion, production of antimicrobial proteins, and immune modulation. The spleen’s altered immune function contributes to cirrhosis-associated immune dysfunction syndrome, a paradoxical state of simultaneous immunodeficiency and systemic inflammation, where inadequate bacterial clearance perpetuates gut dysbiosis and bacterial overgrowth while excessive inflammatory responses accelerate hepatic fibrogenesis, promote complications like spontaneous bacterial peritonitis[144] and hepatorenal syndrome, and increase mortality risk[145], thereby establishing splenic immune involvement as a pivotal component linking gut microbial disturbances, hepatic decompensation, and the overall severity of cirrhotic disease.

Figure 8 Gut-liver axis in cirrhosis: A vicious cycle. Initial events and sub-sequential molecular processes linking gut and liver in cirrhosis. α-SMA: Alpha smooth muscle actin; HDAC: Histone deacetylase; IL: Interleukin; LPS: Lipopolysaccharide; NF-κB: Nuclear factor kappa B; NLRP3: NOD-like receptor pyrin domain-containing protein 3; PAMP: Pathogen-associated molecular pattern; PPAR: Peroxisome proliferator-activated receptor; ROS: Reactive oxygen species; SCFA: Short-chain fatty acid; TGF-β: Transforming growth factor-beta; TJ: Tight junctions; TLR4: Toll-like receptor 4; TMAO: Trimethylamine N-oxide; TNF-α: Tumor necrosis factor alpha.

Splenic sink: Modulating immunotherapy resistance in NAFLD-driven carcinoma

Evidence from clinical trials indicates that immune checkpoint inhibitor (ICI)-based combination therapies, particularly when augmented by anti-angiogenic agents and tyrosine kinase inhibitors, correlate with prolonged survival in a subset of patients with hepatocarcinoma. This pharmacological synergy provides a viable therapeutic alternative for patients failing to maintain a response to first-line agents[146]. Also to address the prevalence of treatment-emergent hepatotoxicity and other immuno-related adverse events, current oncological research emphasizes combination therapies. By leveraging targeted drugs and microbiota modulation, these novel approaches aim to broaden the efficacy-to-safety window in patients with advanced malignancies[147]. Immunotherapy-induced hepatotoxicity typically manifests as panlobular hepatitis with lymphocytic infiltrates rather than drug-induced steatosis. Nevertheless, NAFLD—and its associated steatohepatitis—represents a known risk factor that increases susceptibility to, and may coexist with, immune-related hepatitis in this patient population[148]. In many advanced cancers, the spleen shifts from an immune-activating organ to a reservoir for immunosuppressive cells, particularly MDSCs and certain regulatory immune cells[149].

Tumors release signals that stimulate the spleen to produce and expand these cells, which then enter the bloodstream and migrate to tumors and lymphoid tissues. Once there, they inhibit cytotoxic T-cell activation, reduce NK cell function, and release suppressive molecules such as arginase and inhibitory cytokines[150,151]. This systemic immune suppression weakens the body’s ability to recognize and eliminate cancer cells, allowing tumors to grow more aggressively and resist immune-based therapies. As a result, splenic enlargement and altered immune composition are often associated with greater cancer severity and poorer prognosis[152]. A larger spleen volume at baseline, as well as spleen enlargement during ICI therapy, were associated with poorer prognosis in patients with cancer receiving ICI treatment[153]. Some clinical studies have demonstrated an association between MDSC levels and splenic volume[154]. The future of metabolic and oncological medicine may rest not in the liver itself, but in the therapeutic recalibration of the spleen. By shifting the clinical focus toward this “immunometabolic switchboard,” physicians open a new frontier for precision interventions that could overcome the “splenic sink” of immunotherapy resistance. Specifically, utilizing splenic volume and MDSC concentration as non-invasive biomarkers would allow clinicians to predict a patient’s response to ICI with unprecedented accuracy.

MODULATORS AND AMPLIFICATIONS OF THE AXIS

Exogenous stressors and comorbid conditions have been demonstrated to potentiate the pathophysiological dysregulation of the hepatosplenic axis. Microplastics (MPs), small plastic particles resulting from the degradation of larger plastic items and from primary sources such as textiles, engineered plastic pellets, among others, have become a ubiquitous environmental pollutant. As their prevalence in the natural environment grows, concerns about their potential impacts on human health have escalated[155]. The integrated function of the spleen is vital for sustaining systemic homeostasis, enabling appropriate catabolic waste clearance, beyond supporting immunological competence[156]. Evidence from both in vitro and in vivo studies indicates that exposure to MPs and nanoplastics (NPs) disrupts immune function. This disruption includes impacts on hematopoiesis (blood cell formation), the activation of immune cells, and the production of inflammatory cytokines. While laboratory (in vitro) studies often highlight direct cellular toxicity and altered immune cell behavior, animal (in vivo) studies show more complex outcomes, revealing significant effects on organ systems, such as the spleen[157-160]. In mice spleens, exposure to polystyrene NPs resulted in a cascade of adverse effects: They triggered oxidative stress, activated the mitogen-activated protein kinase signaling pathway, and caused both necroptosis and subsequent inflammation[161]. Spleen toxicity stems from the elevated release of argentum (Ag) ions from the NPs. Due to their specific properties, Ag NPs easily traverse the cell membrane, subsequently converting into toxic ions within the cell. The accumulation of these ions damages cellular DNA and organelles, potentially inducing signaling pathways that result in ROS production and apoptosis and necrosis[162]. A recent study showed that MPs introduced via drinking water severely compromised the murine immune system. Key findings included splenic atrophy, characterized by reduced weight of the spleen, diminished populations of CD8⁺ T cells, and an inversion of the T cell balance, demonstrated by an increased CD4⁺/CD8⁺ ratio[163]. The spleen of six-week-old BALB/c female mouse, showed that the white pulp was abundant and consisted of dense lymphoid tissue. Furthermore, the transcriptome analysis pointed to a broad systemic disturbance, with the most significant impact observed in the body's machinery for processing, breaking down, and absorbing food, alongside widespread changes in fundamental metabolic and endocrine signaling pathways[164]. Evidence suggests that MPs and their chemical additives act as obesogens, potentially driving NAFLD pathogenesis by altering gut microbiota or exacerbating liver fibrosis[165]. Recent literature underscores the prevalence of MPs contamination within the bottled beverage industry; however, the long-term toxicological impacts remain largely uncharacterized[166]. New evidence suggests NPs in bottled water may drive NAFLD development, by disrupting lipid metabolism via inhibition of peroxisome proliferator-activated receptor-gamma, highlighting a burgeoning threat to public health[167]. Very recent findings indicate that prolonged NPs exposure heightens susceptibility to NAFLD progression. This suggests that NPs represent a potential toxicological factor contributing to the initiation and advancement of hepatic pathology[168]. The liver and spleen play vital roles in immune response. Disruption of their function due to MPs/NPs exposure can compromise immune defenses and increase susceptibility to other liver diseases. Environmental toxicants as MPs/NPs can directly prime splenic immune cells, as shown in Table 1, thereby lowering the threshold for their deployment to the liver and worsening NAFLD. Examining the liver-spleen axis in relation to MPs/NPs can foster collaboration across various fields, including toxicology, immunology, and environmental science. Moreover, research findings can lead to innovative solutions to mitigate plastic-related health risks, such as developing materials that are less harmful to human health.

Table 1 Microplastics and nanoplastics deeply permeate the liver and spleen, unsettling their natural functions.

Domain	Organ/system	Mechanism	Cellular alterations	Physiological outcome
Immune dysfunction	Spleen	ROS-MAPK pathway activation; necroptosis; tissue atrophy	Reduced CD8+ T cells; increased CD4+/CD8+ ratio	Immunosuppression; impaired immune surveillance
Metabolic dysfunction	Liver	Disruption of transcriptomic networks governing nutrient handling and endocrine signaling	Altered lipid metabolism and insulin signaling pathways	Promotion of non-alcoholic fatty liver disease
Axis dysregulation	Liver-spleen axis	Impaired coordination of blood filtration and waste clearance	Accumulation of catabolic by-products; dysregulated inflammatory signaling	Increased susceptibility to metabolic and inflammatory disease
Systemic effects	Whole organism	Reprogramming of cellular survival and metabolic homeostasis	Persistent immune-metabolic imbalance	Progression to chronic organ disease

Microplastics (MPs) and nanoplastics (NPs) act as a dual-organ toxin that collapses the body’s metabolic and immune defenses. The synergy of their impact can be synthesized into three core failures: Immune failure (spleen): NPs trigger the reactive oxygen species (ROS)/mitogen-activated protein kinase pathway, leading to necroptosis and physical atrophy. This causes a critical “immune inversion” (reduced CD8⁺ T cells and increased CD4⁺/CD8⁺ ratio), resulting in systemic immunosuppression. Metabolic failure (liver): Chronic exposure serves as a catalyst for non-alcoholic fatty liver disease. MPs/NPs disrupt transcriptomic “machinery” responsible for nutrient absorption and endocrine signaling, rewiring the body’s fundamental metabolism. Systemic collapse (the axis): Because the liver and spleen work together to filter blood and manage waste, their simultaneous disruption creates a “pathological cascade.” The body loses its ability to clear catabolic waste while becoming hyper-susceptible to inflammatory and metabolic pathologies. Bottom line: MPs/NPs fundamentally reprogram cellular survival and metabolic homeostasis, transforming an environmental pollutant into a driver of chronic organ disease.

The literature has disproportionately emphasized respiratory and cardiovascular outcomes associated with particulate matter exposure, neglecting the potential relationship between air pollution and the contemporary epidemic of obesity and associated MS, including NAFLD[169].

Authors observed a positive association between ambient particulate matter 2.5 (PM2.5) exposure and NAFLD odds among hospitalized patients in this nationwide cross-sectional analysis of the NIS database[170]. Furthermore, PM2.5 long-term exposure was associated with major severity of steatotic liver disease[171]. Findings indicated that PM2.5 exposure precipitated hepatic injury marked by steatosis and fibrosis, with alterations in Par3 polarity protein expression and cellular localization serving as a pivotal molecular pathway[172]. Surprisingly, the Par3 polarity protein serves a vital function in splenic and lymphocytic immune responses by maintaining cell polarity at the immune synapse, which is necessary for B-cell activation, antigen presentation, and the assembly of antigen-processing machinery[173].

An intriguing study reveals a critical pathway for PM2.5-induced spleen damage in the sense that PM2.5 (and its water-soluble parts) triggers autophagy and inflammation through the NLRP3 inflammasome in spleen lymphocytes. The fact that blocking autophagy significantly reduces the NLRP3 activation provides new understanding that links autophagy to the inflammasome-driven spleen injury caused by PM2.5[174]. Exposure to PM2.5 negatively impacted the spleen’s metabolic function, notably by upregulating the tricarboxylic acid cycle and changing branched-chain amino acids metabolism. This specific metabolic alteration in the spleen, potentially linked to its lipid regulation role, intensifies the harmful effects of a HFD, resulting in a failure of the spleen to metabolize lipids properly and exacerbate the hepatic fat storage, i.e. NAFLD[175]. PM2.5 exposure and cigarette smoking demonstrated independent associations with elevated MS risk, as well as a joint effect when occurring concurrently[176]. It is worth emphasizing that MS is the predominant underlying cause of NAFLD[177]. Women with PCOS exhibit a moderate elevation in circulating CRP alongside a decline in adiponectin. These alterations are primarily attributable to PCOS itself, independent of obesity, although the presence of obesity further exacerbates them[178]. In premenopausal women with PCOS, an association was evidenced between higher serum bisphenol A concentrations, HS, clinical features (hirsutism) and inflammatory markers, including increased spleen size, thereby indicating the activation of the liver-spleen axis[179]. It should be stressed that women suffering from PCOS face a dramatically elevated risk of developing NAFLD, with research indicating the odds may be higher than in the general population. This amplified risk stems from shared root causes common in PCOS, specifically IR, obesity, chronic inflammation and hyperandrogenism[180]. Hence, the spleen may act not simply as an innocent bystander but as an active participant in the immune-metabolic milieu that drives obesity-related NAFLD progression, due to specific population of monocytes, which suggests that specific chemotactic signaling exists between the spleen and the fibrotic liver. Obesity reduces the spleen's synthesis of the anti-inflammatory cytokine IL-10, creating a systemic deficiency. The spleen’s remaining IL-10 is protective, and its removal (splenectomy) significantly increases inflammation in the liver and white adipose tissue, while also promoting a pathological shift in fat storage toward the liver[181]. Obesity is pathologically characterized by macrophage infiltration of adipose tissue, a source of multipotent adult stem cells. This inflammatory environment drives IR, a central precursor to NAFLD characterized by HS. The cytokine stem cell growth factor (SCGF)—which is known to act on granulocyte/macrophage progenitor cells, especially synergistically with granulocyte-macrophage colony-stimulating factor and macrophage colony-stimulating factor—is strongly implicated in this pathology. High SCGF-β levels were linked to the severity of both IR and HS. This predictive relationship was most pronounced in male patients, where SCGF-β and IL-6 levels successfully predicted HOMA-IR values. Furthermore, in males, the concentrations of inflammatory markers CRP and IL-6 predicted SCGF-β concentrations, which then in turn predicted the presence of HS at US. Intriguingly, SLD was a significant predictor of SCGF-β concentrations. The association between SCGF-β levels and the severity of IR and HS was found to be mediated by CRP. Conversely, the protective anti-inflammatory cytokine IL-10 showed a negative correlation with SCGF-β concentrations, suggesting that IL-10 may mitigate the pathogenic cascade driven by SCGF-β[182]. A previous research aimed to understand the direct contribution of HS, a component of NAFLD, to early atherosclerosis, which was evaluated by measuring carotid intima-media thickness. The authors investigated the behavior of HSP-70 and gamma-glutamyl transferase, two molecules central to the function of both the liver and the vascular endothelium, across obese patients exhibiting varying degrees of liver fat storage. The primary goal was to determine if NAFLD was an independent risk factor for atherogenesis even after accounting for its strong association with MS and its individual components. In their assessment of inflammatory and structural markers related to HS severity, the previously quoted study[117] found a progressive increase in both spleen size, measured as SLD, and, more notably, CRP levels as the severity of HS increased.

DISCUSSION

Bridging the gap between traditional imaging standards and the latest non-invasive breakthroughs necessitates a fundamental reconceptualization of diagnostic frameworks. Detecting spleen size and other spleen-related parameters at US is common practice in every medical setting, mainly in the metabolic one[183]. The consistency of this approach is subject to certain constraints: (1) Standardize measurement protocols across operators and institutions, including consistent patient positioning (supine or right lateral decubitus), standardized scanning planes (maximum longitudinal diameter in the coronal or oblique plane), and precise anatomical landmarks; (2) Establish clear criteria for image quality acceptance to minimize measurement variability; (3) Combine spleen size with other splenic parameters such as spleen stiffness measured by elastography, which may reflect portal hypertension and fibrosis more accurately than size alone. Splenic stiffness has shown promise in differentiating stages of liver disease[184]; (4) Develop composite scoring systems that integrate spleen measurements with other US findings like liver echogenicity, portal vein diameter, other vascular indices[185], and liver stiffness measurements. Multi-parameter approaches typically outperform single measurements; (5) Create population-specific reference ranges adjusted for body habitus, age, sex, and ethnicity, since spleen size varies significantly across demographics. Current cutoffs may not be universally applicable; (6) Conduct prospective studies with histological correlation using liver biopsy as the reference standard to establish optimal spleen size thresholds for NAFLD vs NASH. Determine sensitivity, specificity, and likelihood ratios for different cutoff values; (7) Perform inter-observer and intra-observer assessment to quantify measurement reproducibility and identify factors that contribute to variability; (8) Incorporate artificial intelligence and machine learning algorithms trained on large datasets to identify subtle splenic changes and patterns that may not be apparent to human observers. Artificial intelligence could analyze texture, echotexture heterogeneity, and dimensional relationships; (9) Consider complementary imaging modalities like MRI for splenomegaly confirmation and to assess additional features such as iron deposition or perfusion patterns that may correlate with disease severity[186]; and (10) Combine spleen measurements with clinical data (BMI) and non-invasive biomarkers (fibrosis-4, NAFLD fibrosis score, aspartate aminotransferase/alanine aminotransferase ratio) to create comprehensive risk stratification models[187]. Indeed, the association between spleen volume and spleen length was substantially stronger than that observed with its thickness or width. Conversely, age and BMI demonstrated only modest correlations with spleen volume[188] substantially confirming historic findings[26]. Splenic size or longitudinal diameter on US has been proposed as adjunctive findings in NAFLD[23,189]. The length of the spleen was closely linked to the volume shown on the CT scan[190]. Various authors evaluated the diagnostic accuracy of the Controlled Attenuation Parameter (CAP^TM), a specialized software algorithm implemented in the FibroScan^® (Echosens, Paris, France) system. By using liver biopsy as gold standard, the researchers established that a CAP value of 248 dB/m is the optimal cut-point for identifying the presence of HS[191]. Contextually, measuring spleen stiffness with US is a reliable tool for diagnosing portal hypertension without the need for invasive procedures (Figure 9)[192-194].

Figure 9 Validated metrics for non-alcoholic fatty liver disease/chronic liver disease. Utilizing this suite of metrics enables physicians to longitudinally track disease progression, spanning the spectrum from initial hepatic steatosis to end-stage portal hypertension[193,194]. CAP: Controlled attenuation parameter; F: Fibrosis; LSM: Liver stiffness measurement; S: Steatosis; SSM: Spleen stiffness measurement.

Endoscopic US-guided shear wave elastography is an emerging, non-invasive method for identifying clinically significant portal hypertension—defined as a hepatic venous pressure gradient of 10 mmHg or higher[195]. Based on their findings on PET/CT measures, authors propose that liver and spleen metabolism are interconnected. They suggest that shared insulin sensitivity may be a mechanism underlying this link—a hypothesis warranting further study. The study also reports that in individuals with NAFLD, both the splenic glucose uptake rate (normalized for tracer distribution volume) and spleen size are elevated[23]. Very recently, automated segmentation of liver and spleen on CT to assess liver-spleen ratio has been used in artificial intelligence approaches for NAFLD severity grading[196]. These suggest that combined liver plus spleen imaging metrics may improve sensitivity or specificity for NAFLD/NASH detection or staging, but more validation is needed. Because the spleen participates in multiple inter-organ crosstalk axes, its clinical evaluation can indirectly reflect the health status of several other organs. Yet, its fragile structure and the inherent limitations of conventional pathological approaches, such as biopsy, make direct assessment difficult. Molecular imaging has therefore emerged as a noninvasive and efficient strategy capable of providing highly precise visualization of splenic phenotypes. This technology offers a mesoscopic view of the spleen’s physical characteristics as well as the biological processes occurring at cellular and molecular scales in vivo. With the advent of advanced molecular imaging modalities, the spleen is increasingly recognized as a pivotal organ in the onset and progression of numerous diseases[197]. What are the therapeutic implications and the future of targeting the axis? Given the emerging role of the spleen in NAFLD pathophysiology, therapeutic strategies could consider the axis rather than only the liver. For example, vitamin D deficiency and magnesium status have been discussed in this context. Although direct interventional data on spleen change are limited, the conceptual framework is relevant for integrated organ-axis thinking. It is essential to revisit the central player: Chronic low-grade inflammation, which is a persistent, systemic, and subtle state of immune activation. It is driven by factors like: (1) Obesity and unhealthy diet: Excess fat (especially visceral fat) is not just storage; it is metabolically active. Adipocytes release pro-inflammatory cytokines (e.g., TNF-α, IL-6)[198]; (2) IR: A key feature of MS it further promotes inflammatory pathways; and (3) Gut dysbiosis: An imbalance in gut bacteria can lead to a “leaky gut”, allowing bacterial endotoxins, LPS, to enter the bloodstream, triggering inflammation in the liver[199]. Potential avenues include dietary factors (macro- and micro-nutrients) influencing chronic inflammation, HS and possibly spleen involvement. Modulating splenic immune cell populations (e.g., MDSCs, NKT cells) to reduce hepatic inflammation. One of the key mechanisms for spleen-mediated immune modulation involves neural-immune communication: Nerve fibers in the spleen discharge norepinephrine, which locks specific receptors found on certain T cells. This interaction causes these T cells to produce ACh, a chemical messenger that subsequently blocks macrophages from releasing inflammation-promoting molecules. A further process works by directly influencing immune cells within the spleen, adjusting their genetic activity, molecular communication networks, and chemical signaling patterns, ultimately changing how these cells behave. When comparing different treatment options, non-invasive physical approaches—particularly targeted electrical currents and focused US waves directed at the spleen—have emerged as the most clinically viable methods for controlling splenic immune activity and managing related conditions[200]. Targeting IL-15, as pro-fibrotic cytokine[201] in the light that a retrospective study suggests that, in addition to age, IL-15 levels may be associated with intima-media thickness in obese patients with NAFLD, indicating a possible involvement of this cytokine in the atherosclerotic process[202]. Exosome-mediated delivery: Engineered exosomes can shuttle therapeutic miRNAs between spleen and liver via targeting p53 up-regulated modulator of apoptosis, cyclin E1, and K (lysine) acetyltransferase 5, respectively[203]. The synthesis and application of nanoparticles, liposomes, and colloidal systems. They represent a shift toward site-specific modulation of the inflammatory microenvironment, with a critical emphasis on the spleen as a central immunomodulatory hub. These platforms exploit the distinctive physiological hallmarks of inflammation, such as leaky vasculature and the fenestrated endothelial lining of the splenic sinusoids, which facilitate the passive entrapment of carriers within the splenic red pulp. By encapsulating therapeutic agents, the drug’s pharmacokinetic profile is altered to favor sequestration within the spleen’s marginal zone, effectively diverting the active compounds from healthy systemic circulation toward this primary site of immune cell maturation. Consequently, these systems facilitate a higher therapeutic index by concentrating anti-inflammatory payloads directly within activated splenic macrophages and DCs. This localized accumulation allows for the systemic suppression of pro-inflammatory cytokines at their source of production, thereby mitigating “cytokine storms” and chronic inflammatory states with minimal off-target toxicity to non-lymphoid tissues[204]. The drug disulfiram (DSF), prescribed for alcohol dependence due to its ability to inhibit aldehyde dehydrogenase activity, has an additional effect: It blocks the migration of specific immune cells—monocytes and macrophages—through its action on FROUNT, an internal molecule that regulates the CCR2/CCR5 communication system. DSF has been tested in idiomatic pulmonary fibrosis[205]. Evidence shows that statins reduce the production of key proinflammatory cytokines such as TNF-α and IL-6 in monocytes by interfering with geranylgeranylationdependent Rac1 activation[206]. They also modulate monocyte recruitment by downregulating the CCL2/CCR2 axis, a pathway central to monocyte trafficking in chronic inflammatory diseases[204,205]. Other agents—such as omega-3 fatty acids, GLP1 receptor agonists, and FXR agonists—also influence monocyte activity (Figure 10), but statins remain the most consistently documented in suppressing monocyte activation and shifting macrophage polarization toward anti-inflammatory phenotypes[206]. This makes statins uniquely positioned as dual metabolic and immunomodulating therapies, particularly relevant in conditions like NAFLD where monocyte-Kupffer cell crosstalk drives hepatic inflammation (Figure 10)[207-209]. Splenectomy attenuates liver fibrosis by decreasing the expression of tumor necrosis factor superfamily 14 (also known as LIGHT), though direct application to NAFLD is speculative[210]. Gaps in current knowledge and future research directions remain inevitable. Despite the promising concept of the liver-spleen axis in NAFLD, major gaps remain: Most human data are cross-sectional; longitudinal studies linking spleen changes to NAFLD progression are lacking. Mechanistic human studies linking splenic-derived mediators to hepatic injury in NAFLD are very limited. Interventional trials targeting the spleen (directly or via immune modulation) in NAFLD have not been reported. Integration of the liver-spleen axis with other axes (gut-liver, adipose-liver) and multi-organ network approaches remains to be developed. The traditional view of the spleen as a “bystander”—merely a reservoir that passively congests due to portal pressure—is being replaced by the concept of the spleen as an active “rheostat” of systemic inflammation. This is a central question in modern immunometabolism. In the context of NAFLD and its comorbidities, the spleen acts as a proactive participant through three primary mechanisms: (1) The “sensory” function/monitoring distant stress: The spleen is the body’s largest filter for blood-borne antigens and “danger signals” (damage-associated molecular patterns. When the liver is under metabolic stress (lipotoxicity), it releases inflammatory cytokines and extracellular vesicles into the circulation. The spleen “senses” these signals, triggering a transition from a resting state to a pro-inflammatory programmed state; (2) The “amplifier” function/immune cell trafficking: In lieu of remaining a static anatomical site, the spleen actively modulates the immune response. The spleen houses a massive reservoir of undifferentiated monocytes. During chronic inflammation in NAFLD, the spleen “deploys” these cells, which migrate to the liver and adipose tissue, where they differentiate into aggressive macrophages, worsening the original organ damage; (3) The splenic vein “highway”: This is a direct route. The spleen exports pro-inflammatory mediators directly to the liver via the portal circulation, creating a “feed-forward” loop of injury; and (4) The “metabolic mirror”: Spleen reflecting IR: Data showing a correlation between splenic diameter and surrogate markers of IR suggests the spleen is not just reacting to the liver; it is reacting to the systemic metabolic environment. In obese youth, IR leads to chronic low-grade inflammation. Interestingly enough, understanding this axis is essential for implementing the correct therapy, as addressing the splenic-driven systemic inflammation is key to halting disease progression. Finally, investigating carbohydrate restriction (a cornerstone of metabolic health) should be deepened to include its effects on the spleen, particularly how it modulates splenic immune cell mobilization and metabolic stress. In fact, high carbohydrate-mTORC1-liver-spleen axis as a single, integrated inflammatory circuit. When dietary intake exceeds metabolic capacity, it triggers a systemic shift that transforms the liver and spleen from metabolic and immune filters into sources of chronic inflammation[211]. Recent research demonstrates that spleen-targeted lipid nanoparticles can ameliorate behavioral deficits in Mecp2 transgenic mice by suppressing disease-promoting gene expression within peripheral immune tissues. Selective MeCP2 knockdown in the spleen modulated systemic immune function reducing inflammation, thereby establishing that peripheral immunomodulation can effectively treat centrally-manifested disease[212]. Lately, authors investigated strategies for cell-specific targeting within the spleen, mechanisms to prevent enzymatic degradation of therapeutic cargo, and methodologies to optimize intracellular bioavailability of pharmacological agents and nucleic acid[213]. Due to the organ’s high blood flow and unique microcirculatory characteristics suggest spleen is significantly exposed to new-generation therapeutics that often struggle to penetrate other tissues protected by tight endothelial barriers. Authors examined the spleen’s involvement in the disposition of monoclonal antibodies, nanoparticles, and exosomes, while discussing the resulting implications for their therapeutic efficacy and toxicity[214]. Recent progress in the physical, genetic, and pharmacological modulation of splenic immunity has revealed substantial potential for treating a broad spectrum of conditions, including neurological, inflammatory, cardiovascular, autoimmune, and oncological diseases. A particularly promising frontier in this field is bioelectronic medicine; specifically, splenic nerve stimulation offers a novel approach for achieving precise, targeted immune regulation[215]. To optimize the delivery of personalized neoantigen mRNA vaccines, researchers utilized the L242-20 Lipo nanoparticle, specifically engineered for spleen-targeted uptake. Evaluated within a HCC model, the NeoPol-mL242 mRNA vaccine successfully triggered a robust anti-tumor immune response while demonstrating a high level of safety[216].

Figure 10  Integrated immune and metabolic actions of non-alcoholic fatty liver disease therapies on the spleen-liver axis. Statins and other non-alcoholic fatty liver disease therapies act on monocytes and Kupffer cells, dampening cytokine output and reshaping immune-metabolic signaling across the spleen-liver axis[207-209]. FXR: Farnesoid X receptor; GLP-1RA: Glucagon-like peptide-1 receptor; IL: Interleukin; PPAR: Peroxisome proliferator-activated receptor; TNF-α: Tumor necrosis factor alpha.

CONCLUSION

To remain consistent with the foundational literature, the traditional NAFLD acronym has been used throughout this work. The presence of chronic low-grade inflammation is the engine that drives NAFLD to its more severe form, i.e., NASH. The spleen becomes involved both as a consequence of the resulting liver damage (via portal hypertension) and as an active contributor to the systemic inflammatory state. Therefore, in clinical practice, the health of the spleen can serve as a barometer for the severity of the underlying liver disease in a patient with NAFLD. Emerging evidence highlights the interplay between the liver and spleen, termed the liver-spleen axis, as a critical component in NAFLD progression. This axis reflects the bidirectional relationship where liver dysfunction influences splenic activity and vice versa. In NAFLD, hepatic fat accumulation and inflammation lead to liver stiffness and fibrosis, which disrupt normal liver function. This triggers systemic inflammation and altered cytokine profiles, affecting the spleen. The spleen, a key immune organ, responds by increasing its activity, often leading to splenomegaly in advanced NAFLD particularly in steatohepatitis and cirrhosis. Splenic activation exacerbates systemic inflammation, further compromising liver health. Portal hypertension in NAFLD-related cirrhosis also triggers splenic congestion. The resulting immune dysfunction increases infection rates, posing a major risk for these patients. Interventions aimed at reducing hepatic inflammation and fibrosis may mitigate splenic hyperactivity, while addressing splenic dysfunction could alleviate systemic inflammation, improving outcomes in NAFLD. The cross-talk between liver and spleen represents an emerging dimension in the pathophysiology of NAFLD, linking splenic metabolic and immunologic changes with HS, inflammation and fibrosis. While still in early stages of investigation, evidence of splenic enlargement, increased metabolic activity and altered immune cell populations supports this concept. Integrating spleen assessment into diagnostics and exploring therapeutic strategies that address this axis may enrich future NAFLD management. Robust longitudinal and interventional studies are needed to clarify the clinical utility of the liver-spleen axis.

ACKNOWLEDGEMENTS

We used various large language models, such as Gemini 3 Flash, ChatGPT-5.2, Claude Sonnet 4.6 and Microsoft Copilot to support the refinement of my manuscripts, particularly for professional editing tasks such as polishing the English language, identifying and correcting minor typographical issues, and reducing inadvertent duplication within the reference list. We also employed these tools to assist in generating original schematic figures and to streamline other small but time-consuming editorial adjustments, always ensuring that the scientific content, interpretation, and conclusions remained entirely our own.