Modulation of Autophagy as a Therapeutic Target in the Treatment of Metabolic Dysfunction-Associated Steatotic Liver disease: Evidence from Pharmacological, Nutritional, and Bioactive Compound Interventions

1. Introduction

Metabolic dysfunction-associated steatotic liver disease (MASLD) represents a substantial and growing global health challenge, affecting approximately one quarter of the world’s adult population [1]. Its continued increase is intrinsically linked to obesity and sedentary lifestyle pandemics. The socioeconomic burden of MASLD is substantial, being one of the leading causes of hepatocellular carcinoma and the need for liver transplantation in the United States, with lifetime healthcare costs that reached approximately US$100 billion in 2016 [2].

MASLD is defined by the presence of hepatic steatosis (fat accumulation ≥ 5% in hepatocytes) in individuals with at least one of five metabolic disorders: high body mass index (BMI ≥ 25 kg/m²), elevated fasting glucose (≥ 100 mg/dL), high blood pressure (≥ 130/85 mmHg), high triglyceride levels (TG ≥ 150 mg/dL), or high-density lipoprotein cholesterol (HDL-C). This new nomenclature, adopted in 2023, emphasizes the central contribution of metabolic dysfunction, without requiring the exclusion of alcohol consumption [3,4,5].

Clinically, MASLD is a progressive disease. Its initial stage is simple hepatic steatosis, characterized by fat accumulation without significant inflammation. The most advanced stage is metabolic dysfunction-associated steatohepatitis (MASH), evidenced by hepatocellular inflammation, ballooning, and risk of developing fibrosis, cirrhosis, and hepatocellular carcinoma, with consequent increased mortality [5,6].

Autophagy is a crucial catabolic process that contributes to cellular homeostasis by degrading dysfunctional components and lipids. Lipophagy, a selective autophagic mechanism targeting lipid vesicles, is the selective mechanism by which lipid vesicles are sequestered by autophagosomes, fused to lysosomes, and degraded by lysosomal acid lipase [7]. Evidence indicates that MASLD is positively correlated with autophagy suppression in the liver [8,9]. It is suggested that the abundance of cellular energy, typical of obesity and insulin resistance (IR), inhibits the AMP-activated protein kinase (AMPK)/ mammalian target of rapamycin complex 1 (mTORC1) pathway, which regulates autophagy [10,11].

Inhibition of mTORC1 is essential to activate the autophagic machinery [e.g., Unc-51-like kinase 1 (ULK1), the beclin 1 complex, conversion of microtubule-associated protein 1 light chain 3 form I (LC3-I) to form II (LC3-II)]. Impaired hepatic autophagy leads to increased lipid accumulation, which, in turn, further suppresses autophagy, creating a vicious cycle. This accumulation not only perpetuates steatosis but also contributes to cellular stress and inflammation, driving MASLD progression [12,13].

Some evidence suggests the potential of pharmacological and non-pharmacological agents to induce autophagy, including antidiabetic drugs such as glucagon-like peptide-1 (GLP-1) agonists and metformin, dietary interventions such as calorie restriction and intermittent fasting, and bioactive compounds [14]. Given this scenario, the relevance of autophagy as a mechanism for attenuating the pathogenesis of MASLD stands out, as does the importance of a better understanding of potential therapeutic agents and of modulating this pathway.

Thus, this review aims to compile existing evidence on the effects of pharmacological, nutritional, and bioactive compound interventions on autophagy and to explore their potential role in the pathogenesis of MASLD.

1.1 Pharmacological interventions

1.1.1 GLP-1 analogs

GLP-1 analogs are a class of drugs used to treat type 2 diabetes mellitus (T2DM) and obesity. In general, these drugs exert their effects through the activation of membrane receptors coupled to G proteins, expressed mainly in the lung, kidney, stomach, heart, intestine, alpha and beta cells of the pancreatic islets, and in multiple regions of the central nervous system, promoting effects such as increased satiety, augmented glucose-dependent insulin secretion, decreased glucagon secretion, and attenuated gastric emptying [15,16].

Within this family, liraglutide has demonstrated consistent positive effects on MASLD in experimental and clinical studies. Some evidence suggests that autophagy activation may be a mechanism underlying the improvement in MASLD observed in some experimental models using liraglutide. For example, a study using in vivo and in vitro approaches showed that liraglutide alleviates hepatic steatosis induced by a high-fat diet (HFD) by activating the transcription factor EB (TFEB)-regulated autophagy–lysosomal pathway and promoting autophagy-dependent lipid degradation. This activation occurs via the GLP-1R/ TFEB pathway, highlighting the therapeutic potential of liraglutide in MASLD. In this sense, liraglutide promoted nuclear translocation of TFEB and restored autophagic activity by improving lysosomal function, leading to reduced hepatic expression of LC3-II and sequestosome 1 (p62) [17,18,19,20,21,22].

Interestingly, a study in an HFD-induced MASLD model with Sprague–Dawley rats and HepG2 cells treated with palmitic acid (PA) showed that liraglutide administration for 4 weeks increased beclin 1 and autophagy-related 7 (ATG7) protein expression, as well as the LC3-II/ LC3-I ratio, which were reduced in response to HFD consumption, indicating induction of autophagy. Additionally, an increase in the expression of phospho-AMP-activated protein kinase (p-AMPK) was observed, along with a decrease in the expression of phospho-mammalian target of rapamycin (p-mTOR), suggesting that liraglutide modulates the AMPK/ mammalian target of rapamycin (mTOR) pathway to promote autophagy (Table 1) [14].

Another study investigated the effect of liraglutide on hepatocellular steatosis in C57BL/6 mice and L-O2 cells treated with free fatty acids (FFAs). It was demonstrated that liraglutide administration for 12 weeks increased the expression of LC3-II and beclin 1 (reduced with HFD) and reduced that of p62 (increased with HFD), indicating induction of autophagy. An increase in p-AMPK expression and a decrease in p-mTOR expression were also reported, suggesting that liraglutide modulates the AMPK/ mTOR pathway to promote autophagy (Table 1) [23].

In a study investigating the effects of liraglutide on T2DM-associated hepatic steatosis in an animal model, liraglutide significantly reduced liver damage in mice with MASLD and T2DM, as evidenced by histological and biochemical analyses. The authors also found that liraglutide activated the AMPK/ mTOR signaling pathway, which was suppressed in the MASLD model (Table 1) [24].

Another study evaluated whether liraglutide could attenuate MASLD in mice fed an HFD, exploring the underlying molecular mechanisms, with emphasis on mitochondrial architecture and autophagy mediated by the SIRT1/ SIRT3-FOXO3a pathways. Treatment with liraglutide for 4 weeks resulted in a significant decrease in body mass, improved glucose tolerance, and reduced hepatic triglyceride content in mice with MASLD. Additionally, decreased p62 gene and protein expression, and increased beclin 1 expression and LC3-II/I ratio were observed, indicating that liraglutide promoted hepatic autophagy (Table 1) [25].

1.1.2 Metformin

Metformin is a synthetic biguanide with an antihyperglycemic effect, used in the treatment of T2DM. Its effects are mainly due to increased insulin sensitivity, reduced hepatic glucose synthesis (by inhibiting hepatic gluconeogenesis), and decreased intestinal glucose absorption, with the safe dose for the treatment of T2DM between 250–2550 mg/day [26].

Two studies showed that metformin significantly improves steatosis in patients with MASLD without T2DM, although it has no impact on fibrosis or necroinflammation in these individuals [27,28]. Additionally, patients with MASH presented metformin-improved plasma alanine aminotransferase (ALT) levels and liver histology in 30% of patients [29].

Regarding autophagy, a study reported that this pathway is involved in improving MASLD with metformin use. More specifically, in an HFD-induced MASLD mouse model, metformin treatment for 9 weeks increased LC3-II protein expression, the number of autophagic vesicles and TFEB in the nuclear fraction of the liver, and decreased p62 expression in the liver, simultaneously improving obesity, steatosis, and IR [30].

Furthermore, a study conducted on human carcinoma cells showed that metformin induced autophagy and increased autophagic flux by activating the AMPK/ mTOR pathway, as shown in Table 2. Although these findings do not derive directly from hepatic MASLD models, the identified mechanism is highly biologically relevant, as AMPK/ mTOR signaling dysfunction and reduced autophagic capacity are central to the disease’s pathophysiology [31].

1.2. Calorie restriction

Calorie restriction is a dietary intervention that consists of a significant reduction in calorie intake without compromising the nutritional content of the diet. In animal models and primates, this type of intervention is associated with increased longevity, through adaptations at the metabolic and molecular levels [32].

In metabolic terms, one study showed that a 30% reduction in calorie intake in primates reduced the presence of T2DM and cardiovascular diseases [33,34]. An association between negative energy balance and improved MASLD has also been suggested, possibly through reduced body mass, with improvements in steatosis, liver histology, and plasma ALT levels [35,36].

The activation of autophagy has been studied as a potential mechanism involved in the metabolic effects observed with calorie restriction. A study using in vivo and in vitro methods demonstrated that calorie restriction and its mimetics activate chaperone-mediated autophagy, promoting the selective degradation of damaged proteins and conferring cellular protection against stress and aging. Concerning MASLD, characterized by the accumulation of dysfunctional proteins and impairment of autophagic pathways, this mechanism offers a plausible explanation for the beneficial metabolic effects of calorie restriction and reinforces its potential as a therapeutic intervention (Table 3) [37].

A study aimed at understanding the effects of calorie restriction and resveratrol on hepatic steatosis showed that both reduced hepatic steatosis and hepatocyte ballooning induced by HFD. In this study, calorie restriction provided superior protection compared with resveratrol, associated with lower total calorie intake and reduced body mass. Moreover, animals fed an HFD had decreased SIRT1 levels and autophagy markers LC3-II and beclin 1, as well as p62 accumulation, indicating compromised autophagic flux. Treatment with calorie restriction increased the expression of SIRT1, LC3-II, and beclin 1, in addition to reducing p62 levels, suggesting a restoration of hepatic autophagic flux (Table 3) [38].

1.3. Intermittent fasting

Intermittent fasting is a dietary intervention characterized by alternating periods of fasting and eating. The most commonly used protocols include alternate-day fasting, in which full days of fasting or very reduced calorie intake alternate with days of ad libitum eating, and full-day fasting, which involves 1–3 days of total or partial fasting per week; and time-restricted feeding, limiting eating to a short daily window (usually 4–10 hours) and maintaining fasting during the remaining hours of the day. As a consequence of these cycles of food abstinence, there is greater mobilization of fatty acids and increased production of ketone bodies, a metabolic process associated with improved cardiometabolic parameters, including plasma lipids, insulin sensitivity, and blood pressure [39].

Some studies suggest that certain intermittent fasting protocols are associated with reduced body mass and inhibition of MASLD progression [40,41,42,43]. There is also evidence suggesting a relationship between autophagy, MASLD, and intermittent fasting.

One clinical trial investigated how intermittent fasting influences hepatic autophagy in patients with MASLD, evaluating the effects of the intervention on autophagy markers, including autophagy-related 5 (ATG5) and beclin 1. Intermittent fasting increased serum ATG5, reduced hepatic steatosis, hepatic fibrosis, body mass, BMI, abdominal circumference, ALT, and aspartate aminotransferase (AST). The study suggests that intermittent fasting may be an effective strategy to improve hepatic steatosis and activate autophagy in patients with MASLD. These effects may contribute to enhanced hepatic metabolism and reduced risk of progression to more severe forms of liver disease (Table 4) [44].

Another study demonstrated that a 24-hour intermittent fasting protocol over three non-consecutive days increased LC3 protein and gene expression in the livers of C57BL/6 mice fed both an HFD and a control diet. Increases in lysosomal-associated membrane protein 1 (LAMP1) protein expression and beclin 1 gene expression in the liver of mice fed a control diet were also observed (Table 4) [45].

1.4. Bioactive compounds

1.4.1 Epigallocatechin-3-gallate

Epigallocatechin-3-gallate (EGCG) is the catechin present in the greatest quantity in green tea (Camellia sinensis), contributing to its antioxidant, anti-inflammatory, and cardioprotective properties. Doses between 800 and 1600 mg/day are well tolerated and safe, and some studies have demonstrated hepatoprotective and antifibrotic effects with EGCG, with a reduction in hepatic steatosis [46,47,48,49,50].

A study using C57BL/6 mice showed that 2 weeks of EGCG treatment exerted anti-obesity effects, mainly by activating beclin 1-mediated autophagy in gonadal white adipose tissue, resulting in reduced visceral adiposity and improved glucose tolerance. These findings highlight the therapeutic potential of EGCG in the prevention and treatment of obesity and associated metabolic disorders, such as MASLD (Table 5) [51].

Another study demonstrated that EGCG stimulates autophagy in primary mouse hepatocytes. These effects are partially mediated by activation of the AMPK pathway, leading to increased protein expression. Thus, EGCG may have therapeutic potential for the prevention and treatment of hepatic steatosis and other hepatic conditions associated with autophagy dysfunction [52]. Additionally, it was demonstrated that EGCG increased the protein expression of LC3 and AMPK in differentiated adipocytes [53]. Table 5 presents the effects of EGCG on autophagy in both studies.

Moreover, a study on the effect of EGCG-rich green tea extract showed that supplementation can prevent HFD-induced hepatic steatosis in Swiss rats, possibly through activation of the AMPK pathway, with increased expression of AMPK and SIRT1 in this model, as shown in Table 5 [54].

1.4.2 Quercetin

Quercetin (3,5,7,3′,4′-pentahydroxyflavone) is a polyphenol found mainly in fruits, vegetables, and derivatives, such as onions, apples, and wine, and is attributed with antioxidant, anti-inflammatory, and antitumor effects. In addition, quercetin exhibits cardioprotective effects, including a reduction in blood pressure [55]. Although beneficial effects are attributed to this bioactive compound, its low oral bioavailability makes quercetin poorly usable in the human body, with a maximum of 20 mg/day absorbed by the digestive tract [56,57].

A hepatoprotective effect is attributed to quercetin, although the mechanisms underlying this effect remain poorly elucidated. It has been proposed that this polyphenol attenuates inflammation by improving mitochondrial function, regulating inflammatory cytokine production, and modulating oxidative stress [58,59,60,61,62].

Supplementation of Goto–Kakizaki (GK) rats with quercetin was able to promote hepatic autophagy, increasing the expression of proteins involved in autophagy and related pathways, such as LC3, beclin 1, PTEN-induced putative kinase 1 protein (Pink-1) and Parkin RBR E3 ubiquitin protein ligase (Parkin), and reducing the expression of p62, phosphatidylinositol 3-kinase (PI3K), phospho-phosphatidylinositol 3-kinase (p-PI3K), protein kinase B (AKT), phospho-protein kinase B (p-AKT), mTOR, and p-mTOR proteins, reversing the protein expression scenario observed in GK animals fed an HFD (Table 6) [63].

Another study used ApoE-deficient mice fed an HFD as a model. Quercetin, by promoting autophagy, reduced the hepatic accumulation of oxidized low-density lipoprotein (ox-LDL). In this study, increased LC3-II protein expression and reduced p62 expression were observed, suggesting augmented autophagic degradation (Table 6) [64].

Additionally, in a mouse model of HFD-induced MASLD, 4 weeks of quercetin treatment reduced the NAFLD Activity Score (NAS), a marker of MALD severity, possibly by modulating autophagy-related proteins, as evidenced by increased LC3 protein expression in the liver. However, hepatic p62 expression was also increased by quercetin treatment (Table 6) [65].

2. Conclusions

Autophagy emerges as a central mechanism in maintaining hepatic homeostasis and protecting against MASLD progression. The evidence gathered in this review demonstrates that different pharmacological, nutritional, and bioactive compound interventions converge on activating pathways associated with increased autophagic flux—especially those mediated by AMPK/ mTOR signaling and the regulation of TFEB.

Among the drugs analyzed, GLP-1 agonists, particularly liraglutide, show consistent effects in attenuating steatosis, inflammation, and mitochondrial dysfunction, frequently accompanied by the induction of autophagy in experimental models. Metformin also restores autophagic flux and reduces steatosis, although the magnitude of these effects in humans remains limited. Nutritional interventions, such as calorie restriction, support the notion that energy-deficient states are potent physiological inducers of autophagy, thereby reversing structural and functional alterations associated with MASLD. Bioactive compounds with autophagoregulatory potential offer promising perspectives, but still lack robust evidence.

Collectively, the data indicate that modulating autophagy represents a plausible and strategic therapeutic target in MASLD. However, its clinical translation depends on studies evaluating dose, duration, safety, and the magnitude of long-term benefits. Advancing the understanding of the mechanisms regulating autophagic flux, as well as identifying sensitive and specific markers of its activity in vivo, will be essential for the development of effective and personalized interventions. Thus, integrating pharmacological and nutritional strategies that optimize autophagy may represent a promising approach to halt the progression of MASLD and improve metabolic and hepatic outcomes.