The Significance of the Mediterranean Diet in the Management of Non-Alcoholic Fatty Liver Disease: A Systematic Review

Non-alcoholic fatty liver disease (NAFLD) is the accumulation of intrahepatic fat occurring in the absence of alcohol abuse. The fatty changes in the liver are often the beginning of sequelae of complications, potentially causing steatohepatitis, liver cirrhosis, and hepatocellular carcinoma. The Mediterranean diet is not only a way of eating but is considered a lifestyle and primarily consists of a plant-based diet, with olive oil as the primary lipid. In this study, we reviewed the effectiveness of the Mediterranean diet on NAFLD and its efficacy in disease management. This systematic review follows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocol (PRISMA-P) 2009 guidelines. The PubMed database was used to gather articles, using the following terms individually and in combination, “Mediterranean diet,” “non-alcoholic fatty liver disease,” “insulin resistance,” “metabolic syndrome,” “omega-3-fatty acids.” A quality appraisal was completed to include 14 articles in this systematic review. The studies discuss the correlation between the Mediterranean diet and its role in preventing and treating NAFLD. Olive oil is the major monounsaturated fatty acid, whereas nuts, seeds, and fish consist largely of polyunsaturated fatty acids, both of which are essential components of the Mediterranean diet. The plant-based diet, having sufficient amounts of vegetables, legumes, and fruits, provides its anti-oxidant and anti-inflammatory effects, playing a fundamental role in preventing oxidative stress damage. Olive oil polyphenols increase apoptosis and cell cycle arrest. They also decrease proliferation and angiogenesis, all of which prevent neoplasia. Adapting the Mediterranean lifestyle has shown promising effects in NAFLD, reducing overall mortality and morbidity.


Introduction And Background
The diagnosis of non-alcoholic fatty liver disease (NAFLD) is considered when intrahepatic fat accumulation exceeds 5% in the absence of alcohol abuse [1]. The disease prevalence is estimated to be a quarter of the world population, with a continuing rise [2]. Currently, it is the most common cause of abnormal liver tests in developed countries, with a prevalence of 46% in the United States [3,4]. The occurrence of fatty changes is only the beginning of the potentially life-threatening complications occurring over time, including steatohepatitis, liver cirrhosis, and hepatocellular carcinoma [1]. NAFLD is the leading cause of liver transplantation in men and the second leading cause in women [5]. The manifestations from NAFLD cause sustained liver damage and negatively affected the cardiovascular system, increasing overall mortality, and morbidity [5].
The Mediterranean diet dates back to the traditional dietary patterns from the Mediterranean regions, which are considered a virtual geographic location consisting of numerous cultures [6,7]. The Mediterranean diet has many variations, but its main emphasis has been a plant-based diet, with significant fiber, anti-oxidants, vegetable proteins, polyunsaturated, and monounsaturated fat [8]. It is known as a high-fat diet with olive oil as the principal source, and the fat content ranging from 35% to 45% of the total energy intake [6]. The accumulation of fat occurring due to an imbalance in hepatic lipid metabolism is due to insulin resistance [3]. Hence, insulin resistance is known as the initial step in NAFLD. It is responsible for forming reactive oxygen species (ROS), causing oxidative stress, lipotoxicity, subsequent inflammation, and hepatic damage [3]. The high fruit and vegetable content in the Mediterranean diet has sufficient anti-oxidant and antiinflammatory properties, counteracting the effects of ROS and the pathogenesis of NAFLD [3]. A similar effect has been observed with olive oil, decreasing inflammation by inhibiting transcription genes [6]. The anti-oxidant effects of the Mediterranean diet seem to share some form of inverse correlation with the underlying pathology of NAFLD. Research shows that several diets have been tested for NAFLD with various effects. The question remains, is the Mediterranean diet a potential treatment and preventive measure for NAFLD in the near future? It is also the weight loss in response to the diet that significantly affects the disease progression, or is it the ingredients that make the Mediterranean diet that play an essential role [3]? The current treatment recommendations for patients with NAFLD are explored in this article, focusing on how effective the Mediterranean diet is in disease management. Also, we analyze whether adopting the Mediterranean diet early in the steatosis stage of the disease results in any alteration in disease outcome. The fundamental aim of this review is to report the current evidence about the effects of the Mediterranean diet in NAFLD patients and translate this main dietary recommendation to guidelines for clinical practice for disease prevention and treatment.

Review
This section will discuss how articles were gathered and reviewed for this systematic review. The review analyzes the significance of monosaturated and polyunsaturated fats, the anti-inflammatory effects of the Mediterranean diet, the benefits of limiting sodium, and how these factors alter the outcome of NAFLD.

Methods
The protocol of this systematic review follows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocol (PRISMA-P) 2009 guideline. The electronic database PubMed was searched for articles. The keywords and medical subject headings (MeSH) terms were used individually and in combination to identify relevant articles. The keywords and MeSH terms included in the search strategy were "Mediterranean diet," "non-alcoholic fatty liver disease," "insulin resistance," "metabolic syndrome," and "omega-3-fatty acids." A total of 1108 publications were checked for duplications and assessed for relevance to the research topic. The articles were then screened using the inclusion and exclusion criteria to identify the eligibility of the studies. Table 1 and Table 2 summarize the search strategy using MeSH terms and keywords.

Inclusion and Exclusion Criteria
The studies conducted in the last five years (2016)(2017)(2018)(2019)(2020)(2021), studies based on human subjects, English literature only, studies comprised a mixture of systematic reviews, observational (case-control, cohort, and crosssectional studies), meta-analysis, and literature review were included. The animal studies and articles with a publication date more than five years ago were excluded.

Results
A total of 1108 publications were identified from PubMed, from which 526 duplicates were removed. Filters were applied based on inclusion criteria (full-text studies in English, last five years, on humans, all types of reviews), yielding 274 articles. Each article was manually reviewed for its content, and articles irrelevant to the study topic were excluded. The eligibility of the remaining articles was checked using the PRISMA checklist, Joanna Briggs Institute (JBI) checklist, New Castle Ottawa Tool (developed as a collaboration between the University of Newcastle, Australia, and the University of Ottawa, Canada), and a measurement tool to assess systematic reviews (AMSTAR) checklist. A total of 14 studies were ultimately selected and included in the qualitative synthesis. Figure 1 below shows the PRISMA flowchart diagram of literature retrieval. Figure 2 below shows the Mediterranean diet food pyramid with daily, weekly, and monthly consumptions [5].

Daily
The daily servings consist of high consumption of non-refined cereals and grain products, and vegetables and fruits, along with pulses, legumes, and nuts [9]. Olive oil is the main lipid and considered an essential part of the Mediterranean diet. Also, salt intake is limited by replacing herbs and spices at the table [7]. Besides, adequate water intake, approximately six glasses, is ideal, along with wine in moderation. The consumption of red wine with meals dates back to the traditional Mediterranean diet of the European region [7].

Weekly
Eggs, poultry, and fish are limited to weekly intake, with fish having the highest recommended servings of five or six. Sweets are often limited to a maximum of two servings in the week.

Monthly
The Mediterranean diet also focuses on reducing red meat consumption, with only four servings per month [10]. As mentioned above, the main focus of fat is switched over to olive oil. Hence, red meat intake is significantly decreased.

Olive oil and the role of monounsaturated fat
Olive oil is the most representative group of the Mediterranean diet, and oleic acid, a monounsaturated fatty acid (MUFA), is a major component of this essential dietary fat [5]. MUFAs have numerous beneficial effects, including healthy blood lipid profiles, lowering blood pressure, and improving insulin sensitivity which is an essential and first step in preventing NAFLD [3]. Monounsaturated and polyunsaturated fatty acid (PUFA) diets reduce fasting plasma triacylglycerol and very-low-density lipoprotein (VLDL)-cholesterol concentration on average 19% and 22%, respectively [11]. They also cause an increase in high-density lipoprotein (HDL)-cholesterol levels, which is highly beneficial as current research emphasizes higher HDL levels as preventive measures for long-term diseases [12]. The studies indicate a cardiovascular risk reduction and an all-case mortality reduction linked to the benefits of olive oil [13]. The long-standing conditions show a close relationship to NAFLD prevalence, for example, with over 50% of cases occurring in type 2 diabetes mellitus (T2DM) [14]. The large intake of MUFAs as a substitute for trans fatty acids is believed to reduce the risk of T2DM in high-risk patients by up to 83% over a median of 4.4 years [14]. NAFLD is limited to insulin resistance and includes a spectrum of liver conditions associated with intrahepatic fat accumulation and further complications. For example, a quarter of NAFLD patients progress to non-alcoholic steatohepatitis, which is also associated with a higher risk of cardiovascular disease and mortality [14]. In conclusion, a healthy lipid panel is essential in eliminating countless long-standing health concerns. Monounsaturated fatty acids, driven from olive oil, seem to play an outstanding role in preventive medicine.

Nuts and seeds and the role of polyunsaturated fatty acid
Asides from oil olive as the main source of MUFA in the Mediterranean diet, high quantities of PUFAs in nuts, seeds, and fish also play a valuable role [15]. The most important PUFA for NAFLD is long-chain n-3 fatty acids, which include eicosapentaenoic, EPA, and docosahexaenoic, DHA [15]. These prevent liver adipogenesis by modifying transcription and suppressing peroxisome proliferator-activated receptor α and sterol regulatory element-binding protein-1 genes [16]. The main role of these genes is the regulation of hepatic fatty acid oxidation and synthesis, respectively [16]. Additionally, n-3 fatty acids suppress the direct uptake of fat into hepatocytes and modulate the expression of particular miRNAs [16,17]. The wellestablished outcome of PUFA and NAFLD by research is the prevention of hepatic steatosis, resulting in a beneficial role in disease management [18].

Effects of olive oil and a plant-based diet on inflammatory-related mechanisms
A bulk of the Mediterranean diet comprises plant-based foods, with a focus on plant-based protein sources. This includes legumes, moderate consumption of fish, and a limited amount of red meat [13]. Vegetables, fruits, and legumes are high in anti-oxidants and anti-inflammatory, playing a fundamental role in preventing oxidative stress damage and cytokine release [13]. From these food sources, legumes have been identified as the most predictive component for lowering mortality [13]. Vegetables and fruits are equally important and consist of polyphenols, which are important bioactive compounds that increase fatty acid oxidation [7]. Polyphenols promote fatty acid beta-oxidation and exert their anti-inflammatory role by inhibiting NF-κB directly or via peroxisome proliferator-activated receptors (PPARs), and chronic inflammatory pathways such as prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2) [15]. The recent literature has also shown a relationship between the ability of polyphenols to module the immune system by altering the role of WBCs and inflammatory cytokines that participate in the immunological defense [19]. Hence, the Mediterranean diet has been shown to positively influence inflammatory biomarkers, including adhesion molecules and cytokines, in presenting hepatic fat accumulation and ensuring limited blood levels of transaminase, triglycerides, cholesterol, and insulin resistance [20]. In conclusion, olive oil and plantbased diets, which are rich in polyphenols, have shown to have a positive impact on the human body's defense mechanism.

Anti-cancer mechanism from olive oil polyphenols
NAFLD is associated with an increased risk of numerous cancers, with a well-established relationship between NAFLD and hepatocellular cancer [21]. However, research has shown that NAFLD is associated with an increased risk for prostate and colon cancer [22]. When comparing the Mediterranean region with other populations, cancer incidence is drastically lower [22]. The role of olive oil and its associated polyphenols, especially oleuropein hydroxytyrosol, is an important component of the underlying anti-cancer mechanism [23]. Polyphenols increase apoptosis and cell cycle arrest while decreasing proliferation and angiogenesis [22]. The role of apoptosis is indicative of the increase in cell death domains, such as caspase-3,-7, p53, and Bax and Bak [22]. At the same time, suppression of neovascularization by olive oil polyphenols has resulted in a decrease in growth factors such as VEGF and TGFalpha-induced migration of hepatocellular carcinoma cells [21]. These mechanisms drastically inhibited the underlying pathology and worked in preventing malignancy.

Limiting sodium
The Mediterranean diet aims to replace salt at the table with herbs and spices, ensuring favor yet adequate blood pressure control. The relationship between high sodium levels and blood pressure is well established, but the direct effect on NAFLD is controversial. The current World Health Organization (WHO) guidelines indicate a sodium intake of <2000 mg/dl [24]. Many epidemiological studies have proven to show a positive correlation between sodium intake and body weight, independent of energy or sugar intake [24]. The literature has established a relationship between increase sodium intake and triglyceride levels, indicating a potential risk factor for NAFLD [24]. The recommendation of a low-sodium diet has been beneficial in lowering overall mortality and morbidity, with hypertension being an underlying cause of many longstanding conditions [25]. Table 3 summarizes experimental and observational studies that were reviewed, and the study group analyzed along with the findings. These studies involved smaller populations but illustrated a positive correlation between NAFLD and the Mediterranean diet in numerous ways. This includes the reduction of intrahepatic fat accumulation and steatosis in relation to consuming the Mediterranean diet and an overall positive impact on NAFLD score.   Table 4 summarises the current literature reviews that were used in this study to conclude further and establish the effects of the Mediterranean diet on NAFLD. These studies analyzed the risk factors that were influenced by the Mediterranean diet, lowering the outcomes of NAFLD.

Conclusions
This systematic review outlines the significance of the Mediterranean diet in the management of NAFLD and identifies what has been currently studied, and essentially what needs to be focused on in the future to strengthen this relationship. In this review, observational studies, cross-sectional, crossover, and literature reviews were analyzed, indicating a visible link between the two variables. The Mediterranean diet is a regime and a type of lifestyle that has been shown to positively enhancing well-being. It provides a significant number of advantages, being complex in MUFAs, PUFAs, and polyphenols, indicating its antiinflammatory and anti-cancer mechanisms. After reviewing the literature, the Mediterranean diet can be considered a major step forward in NAFLD management. It promotes weight loss by replacing saturated fats and carbohydrates and plays a direct role in eliminating NAFLD pathology. Adapting the Mediterranean diet in the early stages may result in alterations in disease outcome, but more studies are required to confirm this correlation. In the near future, along with more extensive studies, we may potentially be able to conclude the Mediterranean diet as the first-line preventive and treatment option for NAFLD.

Conflicts of interest:
In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work.