Metabolic dysfunction-associated steatotic liver disease (MASLD) poses a significant challenge to public health, impacting a substantial proportion of the population, with approximately 30% of individuals affected globally. Particularly concerning is its escalating prevalence in Latin America (Arab et al., 2021). Despite its growing significance, there remains a notable absence of targeted pharmacological interventions for this condition.

It is well established that a carotenoid-rich diet benefits liver health (Elvira-Torales et al., 2019). Dietary carotenoids are mainly accumulated in the liver, where they are packaged into different lipoproteins for their release to the blood circulation and other organs and tissues, such as the kidneys, adipose tissue, skin, brain, and retina (Elvira-Torales et al., 2019). The accumulation of these antioxidants and their metabolites in the liver allows to rise carotenoids as a promising therapy for liver diseases. Individuals who consume the highest amounts of carotenoids have the lowest risk of developing MASLD (Ferramosca et al., 2017).

Among the array of carotenoids garnering attention for their hepatoprotective properties, lutein stands out for its antioxidant, anti-inflammatory, and hypolipidemic effects. (Cao et al., 2015). Lutein supplementation in rats fed with a high-fat diet (HFD), a model of MASLD, recovered liver function, improved lipid accumulation, and restored hepatic lipid metabolism and insulin signaling, preventing hepatic dyslipidemia and insulin resistance (Wang et al., 2021). In addition, lutein reduced lipid peroxidation and decreased pro-inflammatory cytokine production in the liver of guinea pigs fed with a high-cholesterol diet (Kim et al., 2012). Moreover, in rats with hepatic injury, lutein lowers ALT and AST concentrations, two markers of liver damage (Sindhu et al., 2010). In summary, there is mounting evidence that lutein has beneficial effects on the liver. At the same time, its anti-oxidative potential has been well-documented, but the precise mechanisms underlying its hypolipidemic effects remain inadequately explored. This review aims to consolidate existing evidence and elucidate the potential mechanism by which lutein may exert its hypolipidemic effects, possibly through the modulation of autophagy via the activation of the master transcription factor TFEB.

TFEB is a crucial regulator of cellular processes, among them lysosomal biogenesis and autophagy, and its signaling pathways influence cellular energy metabolism. Its activity and localization are modulated by mTORC1-mediated phosphorylation, determining its subcellular localization in the cytoplasm or nucleus, thus its function as a transcription factor. TFEB’s involvement includes lipid catabolism and it’s a clear potential therapeutic target mediating cellular clearance in a variety of diseases, including rare lysosomal storage diseases and more prevalent ones such as Parkinson’s and Alzheimer’s (Napolitano and Ballabio, 2016).

Furthermore, we propose a putative intracellular transport pathway for lutein mediated by STARD3, drawing upon insights from studies on the intracellular trafficking of other carotenoids. STARD3 is a cholesterol and lutein-binding protein found in late endosomes, featuring a domain that anchors it to endosome membranes and facilitates interactions with other proteins. Its exact role remains unclear, but it’s known to create contact sites between the endoplasmic reticulum (ER) and endosomes potentially influencing cholesterol and lipid distribution within cells (Alpy et al., 2013).

This investigation holds substantial importance as it offers prospects for therapeutic advancements in hepatic steatosis and promises insights into its underlying pathophysiology. This article delineates the current understanding of lutein’s mechanisms and its therapeutic potential in addressing fatty liver disease.

Between 25 and 30 percent of the general population suffers from Metabolic dysfunction-associated steatotic liver disease (MASLD), formerly known as non-alcoholic fatty liver disease or NAFLD. It has become the most frequent cause of liver disease worldwide (Asrani et al., 2019). Higher prevalence rates are found in particular subpopulations, such as people with obesity (90%) and type 2 diabetes (76%) (Younossi et al., 2016). Furthermore, metabolic disorders frequently coexist with the onset of MASLD syndrome, including increased triglycerides (TGs, neutral fats) in the plasma, insulin resistance, hypertension, and reduced high-density lipoprotein (HDL) cholesterol. Hepatic steatosis, defined as the intracytoplasmic TGs accumulation, representing at least 5% of liver weight, is the defining characteristic of MASLD (Chalasani et al., 2018). Most patients have simple steatosis, but steatohepatitis (liver inflammation) is present in about 25% of MASLD patients. Although this inflammatory injury is reversible, it can progress to an irreversible stage of liver damage such as fibrosis, cirrhosis, and hepatocellular carcinoma (Haas et al., 2016), being crucial to intervene therapeutically at the reversible stages of the disease.

Hepatic free fatty acids (FFAs) can be acquired from the diet, adipose tissue lipolysis, and de novo lipogenesis. FFAs are then oxidized through β-oxidation or esterified into TGs, stored as lipid droplets (LD), or packaged into lipoproteins, such as VLDL, to be secreted into the bloodstream (Nassir et al., 2015). The imbalance among these major molecular pathways and how their dysregulation contributes to the development of MASLD is not the focus of discussion in this review. However, an important pathway that may contribute to MASLD is lipolysis, which will be discussed below.

Lutein is one of the most abundant carotenoids in human blood samples, together with lycopene, β-carotene, β-cryptoxanthin, α-carotene, and zeaxanthin (Torsten, 2008). It is a non-provitamin A carotenoid that belongs to the xanthophyll family and is found in spinach, kale, eggs, corn, and green leafy vegetables (Abdel-Aal el et al., 2013). It is known to accumulate in human and primate retina and brain selectively (Erdman et al., 2015), being one of two major carotenoids found as a color pigment in the human eye (macula and retina). It functions as a light filter, protecting the eye tissues from sunlight and oxidative damage (Buscemi et al., 2018). Recently, lutein has drawn increasing attention to its function in chronic diseases other than oculopathy. It has been reported that lutein has anti-inflammatory, antiapoptotic, and anti-oxidative properties through the blockage of the NF-κB pathway (Li et al., 2012a; Li et al., 2012b) and by decreasing IL-6 (Ozawa et al., 2012; Ahn et al., 2022), among other pathways in different pathological conditions, including atherosclerosis and cerebral ischemia/reperfusion models. Lutein is also being studied as a neuroprotective agent (Sun et al., 2014) and as a cognitive performance enhancer (Johnson, 2012); Clinical trials have supported the beneficial effect of oral supplementation with 10 mg of lutein in Alzheimer’s disease patients, particularly in terms of clinically meaningful improvements in visual function but not in cognitive function (Nolan et al., 2015). Furthermore, ongoing clinical trials are evaluating the effect of lutein in reducing oxidative stress caused by amyloid beta (Aβ) in Alzheimer’s patients, although the results of these trials have not yet been published ¹ . Also in rats Lutein decreased the Aβ effects on learning and memory (Nazari et al., 2022). However, its role in liver pathologies has been little researched, but the evidence suggests that lutein would be a promising therapy for liver disease.

Lutein supplementation in rats increased the expression of critical factors in autophagy signaling, such as phosphatidylinositol 3-kinase, an upstream regulator of TFEB (Qiu et al., 2015). Also, a high dose of lutein increased the expression of PPARα (Qiu et al., 2015), which is upstream of TFEB (Kim et al., 2017b) and is associated with lipid metabolism, as previously described (Ghosh et al., 2015). Also, long-term oral feeding of lutein increases total AMP-activated protein kinase (tAMPK) and phosphorylated AMP-activated protein kinase (pAMPK) contents in muscle (Matsumoto et al., 2014), and there is abundant evidence linking AMPK phosphorylation with TFEB activation (Collodet et al., 2019; Huang et al., 2019; Paquette et al., 2021). Moreover, in a model of rat intestinal epithelial cells, lutein treatment induces the processing of LC3-II and the upregulation of autophagy-related genes, including Atg4A, Atg5, Atg7, Atg12, and Becn1 (Chang et al., 2017). On the other hand, there is evidence that lutein inhibits autophagy in rat müller cells exposed to hypoxia (Fung et al., 2016), suggesting that the lutein effect on autophagy could depend on the cellular type. This compelling evidence suggests that lutein may contribute to the induction of autophagy, mediated by the activation of TFEB, PI3K/Akt, and AMPK pathways. However, there is a lack of relevant studies in this area, warranting further investigation to conclusively establish the role of lutein in autophagy.

Steroidogenic Acute Regulatory (StAR)-related lipid transfer Domain–3 (STARD3), also known as Metastatic axillary Lymph Node clone 64 (MLN64) (Tomasetto et al., 1995), is a late endosome (LE) membrane-associated protein (Alpy et al., 2001), which is evolutionarily conserved throughout the animal kingdom, as well as in unicellular organisms closely related to animals (Voilquin et al., 2019). STARD3 belongs to the START family of proteins, whose members share a conserved StAR-related lipid transfer (START) domain. The START family comprises 15 members, with STARD3 being the only member with a transmembrane anchoring domain (Clark, 2020; Asif et al., 2021).

START proteins have a conserved fold, but their primary sequences differ, allowing selectivity in ligand binding. These proteins are involved in the transport of lipids such as sterols, phospholipids, bile acids, ceramides, and carotenoids (Alpy and Tomasetto, 2014; Clark, 2020; Sluchanko et al., 2022).

STARD3 has been identified as the unique protein with high-affinity binding to lutein (Li et al., 2011). Primarily recognized for its role in cholesterol binding, STARD3 serves as a transporter facilitating cholesterol transfer between the late-endosome/lysosome (LE/Lys) and the endoplasmic reticulum (ER) by forming Membrane Contact Sites (MCSs) (Alpy et al., 2001; Alpy et al., 2013; Wilhelm et al., 2017). Functionally, it has been implicated in cholesterol transport during steroidogenesis in placental tissue (Watari et al., 1997; Bose et al., 2000; Zhang et al., 2002). Despite the established involvement of STARD3 in cholesterol mobilization towards the mitochondria (Hoglinger et al., 2019; Nara et al., 2023), the precise transfer mechanism remains incompletely elucidated. Notably, the detection of STARD3 within confined regions between LE/Lys and mitochondria suggests a potential mechanism for interaction associated with MCS formation (Zhang et al., 2002; Charman et al., 2010)

Structurally, STARD3 comprises the MENTAL domain (MLN64 NH2-terminal) at the N-terminus and the C-terminal START domain. The MENTAL domain anchors STARD3 to the endolysosomal membrane and contains a nonconventional FFAT motif crucial for interactions with vesicle-associated membrane proteins and the formation of MCS (Di Mattia et al., 2020). The START domain in the cytosol facilitates cholesterol positioning and mobilization between LE and other subcellular structures (Tsujishita and Hurley, 2000; Zhang et al., 2002; Charman et al., 2010; Alpy et al., 2013; van der Kant et al., 2013; Wilhelm et al., 2017; Nara et al., 2023).

STARD3 was identified as a lutein-binding protein because of its high homology with the carotenoid-binding protein (CBP) found in silkworms (Li et al., 2011). Binding studies demonstrated that STARD3 binds free lutein with high affinity (KD = 0.45 μM) and specificity; even so far, no other protein has been described to bind lutein with this affinity (Li et al., 2011; Horvath et al., 2016). The lutein-binding function of STARD3 resides within the C-terminal START domain as a tunnel-like hydrophobic cavity (Horvath et al., 2016), which can also accommodate a single cholesterol molecule with 1.5 mM affinity. Recently, Hempelmann et al., 2023 elegantly demonstrated the mobilization of sphingosine by STARD3 at inter-organellar level. Therefore, lutein cholesterol and maybe sphingosine share the same protein binding site, with lutein exhibiting a higher affinity. While STARD3 is primarily described as a lysosomal protein, evidence suggests its presence in the plasma membrane. Studies show STARD3 localization at the plasma membrane, with positive vesicles mediating cholesterol recycling (van der Kant et al., 2013) and the transfer of cholesterol from the endoplasmic reticulum to LE affecting the plasma membrane cholesterol pool (Wilhelm et al., 2017).

In this context, it is reasonable to speculate that STARD3 might prefer specific ligands based on their subcellular localization and the concentrations of available ligands. For example, under elevated lutein concentrations, the affinity of STARD3 for lutein may be heightened compared to cholesterol binding. This scenario could mitigate the adverse effects associated with STARD3-mediated cholesterol transportation to the mitochondria, which include its involvement in tumor progression and its correlation with increased mitochondrial cholesterol levels, thereby inhibiting apoptosis and increasing ROS (Asif et al., 2021). Our studies found that overexpression of STARD3 in mice livers resulted in hepatic damage and apoptosis (Tichauer et al., 2007). Moreover, overexpression of STARD3 in hepatocytes increased mitochondrial dysfunction by reducing membrane potential and increasing mitochondrial ROS levels, correlated with elevated mitochondrial cholesterol content (Balboa et al., 2017). Interestingly, this mechanism could explain the antiapoptotic and antioxidant effect of lutein, which likely occurs by lutein occupying the binding site of STARD3, thereby inhibiting its binding to cholesterol.

As was discussed before, STARD3 would not only reside in endosomes but can also cycle between the endocytic pathway and the plasma membrane (Alpy et al., 2001; Zhang et al., 2002; van der Kant et al., 2013; Vassilev et al., 2015). Therefore, considering this possibility, STARD3, located in the plasma membrane, could contribute to the lutein entry in the hepatocyte. Nevertheless, our prior findings have indicated that STARD3 is predominantly situated at the intracellular level (Balboa et al., 2017). However, under pathological conditions, such as steatosis, the subcellular localization of STARD3 remains understudied.

Regarding STARD3 gene expression, it has been described that its promoter region contains putative binding sites for lipid-responsive transcription factors, such as PPARs, retinoid X receptors (RXR), and SREBPs (Borthwick et al., 2009). Moreover, there is evidence that STARD3 expression is sterol-dependent in human THP-1 macrophages in which the increase in sterol content induces a decrease in STARD3 gene expression, while sterol depletion using methyl beta-cyclodextrin enhances STARD3 gene expression (Borthwick et al., 2009). In this context, it is interesting that lutein favors an increase in LDLR expression (also regulated by SREBP2) and activates PPARα in liver tissue in male Wistar rats (Soundarya et al., 2018), suggesting that lutein could regulate STARD3 expression. In line with this, STARD3 protein levels are correlated with lutein concentrations in the human brain (Tanprasertsuk et al., 2016). On the other hand, in insulinoma cells, it has been observed that the expression of STARD3 is not regulated by lutein (Pinto and Graham, 2016). However, this observation lacks clarity regarding the methodology employed for lutein administration, such as direct addition or incorporation via liposomes, as well as the duration of treatment. Therefore, we cannot guarantee proper cellular uptake of lutein or sufficient time for its intended effects to manifest.

Since in liver diseases such as MASLD, there is lipid accumulation, it is possible to infer that STARD3 levels will decrease, as in obesity models (Soffientini et al., 2014b).

Therefore, considering the hepatoprotective effects of lutein and the potential role of STARD3 as a lutein transporter, it is reasonable to propose that a decline in STARD3 expression could compromise liver health by restricting lutein uptake into cells. Additionally, lutein treatment may potentially restore physiological levels of STARD3 expression, thereby promoting liver health.

Considering the evidence that in hepatocytes, the overexpression of STARD3 facilitates the lipidation of exogenous apoA-I and promotes de novo biosynthetic pathways for neutral lipids, leading to an increase in triacylglycerol accumulation (Soffientini et al., 2014a) and that overexpression of STARD3 resulted in enhanced cholesterol efflux and reduced intracellular oxidized LDL accumulation (Almarhoun et al., 2021), it can be inferred that STARD3 plays a dual role in lipid metabolism depending on the cell type and this dual role could be determined by the binding to its ligands.

The role of STARD3 in the presence of lutein in hepatocytes is unclear. In this context, it remains unclear whether lutein competes against cholesterol for STARD3 binding, reducing TG accumulation, or if lutein has a hepatoprotective role independent of its interaction with STARD3. In this review, we have presented evidence of the hepatoprotective effect of lutein and proposed a mechanism for how lutein may exert this effect. The significance of its transporter, STARD3, is not well-documented, and we believe that further research into the role of STARD3 in the pathogenesis of hepatic steatosis is warranted. We posit that lutein’s effect may depend on its transporter, STARD3, making this protein crucial for the intracellular impact of lutein.

Our proposed model (Figure 2) positions lutein as a promising therapeutic candidate for hepatic steatosis.

Hepatic steatosis arises from an imbalance in various lipid homeostasis processes. Among these, we firmly believe that decreased lipophagy plays a crucial role in the development of steatosis, leading to reduced LD hydrolysis and availability of FA for critical reactions such as beta-oxidation. In this context, lutein, a known hepatoprotective agent, is proposed to exert its effect on the liver by enhancing lipophagy, and TFEB likely mediates this action. Additionally, we propose a model wherein STARD3 participates in the intracellular transport of lutein into hepatocytes, further emphasizing the relevance of this protein in the effects of the carotenoid. The data and evidence reviewed in this article strongly support our proposed model. While we recognize the need for additional in vivo models to validate our hypothesis, we believe that models currently under development, such as liver organoids derived from patients, could help address questions regarding the expression levels of STARD3 in MASLD patients versus controls, whether patients exhibit a different response to lutein compared to controls, and whether differences in TFEB expression, previously documented in MASLD patients, could also influence the response to lutein treatment. Based on the findings presented in this review, the current evidence supports lutein and STARD3 as promising nutraceutical and therapeutic targets, respectively, for treating hepatic steatosis.