Non-alcoholic fatty liver disease (NAFLD) is a chronic liver disease characterized by excessive fat deposition in hepatocytes, which is not caused by alcohol or other clear liver-damaging factors (Cobbina and Akhlaghi, 2017). The global incidence rate of NAFLD is approximately 25%, particularly in patients with diabetes and obesity (Mundi et al., 2020). NAFLD is the most common chronic liver disease worldwide and is expected to be the main cause of liver transplantation in the future (Younossi et al., 2016b). NAFLD encompasses a wide range of liver disorders, including simple fat accumulation in the liver cells, non-alcoholic steatohepatitis (NASH), fibrosis through the final stages of cirrhosis, and NASH-HCC (Cobbina and Akhlaghi, 2017). The incidence of NAFLD and NASH is related to sedentary lifestyle and excess dietary energy (Farrell et al., 2013). To date, the Food and Drug Administration has not approved any drugs for the treatment of NASH (Eduardo et al., 2015). Currently, NAFLD can be effectively alleviated only through non-drug management approaches, such as healthy lifestyle, diet, and moderate physical activity (Guillaume et al., 2015). Given the limited clinical treatment for NAFLD, the development of drugs that can effectively alleviate NAFLD is of great significance.

The pathogenesis of NAFLD remains unclear so far. However, recent studies have suggested a bidirectional association between NAFLD and metabolic syndrome, with type 2 diabetes increasing the risk of cirrhosis and related complications (Powell et al., 2021). Insulin resistance, diabetes mellitus, and genetic variations in transmembrane 6 superfamily member 2 (TM6SF2) and patatin-like phospholipase domain containing 3 (PNPLA3) play important roles in NAFLD progression (Cobbina and Akhlaghi, 2017). NAFLD is characterized by excessive fatty accumulation in the liver, while simple steatosis is considered pathologically benign. NASH generally indicates liver damage that can progress to severe pathology (Zhang et al., 2018).

The “two-hit” pathogenesis of NAFLD/NASH was widely accepted in the early stage (Chi, 2017). The “first hit” is characterized by an increase in hepatic fat, especially accumulation of hepatic triglycereides and insulin resistance. Once the accumulation of hepatic fat exceeds 5%, it corresponds to hepatic steatosis (Fang et al., 2018). The most direct cause of NAFLD is abnormal liver lipid metabolism, and a large quantity of free fatty acids and triglycerides that accumulate in liver cells (Xiaxia et al., 2019). The “second hit” is that reactive oxygen species (ROS) triggers an inflammatory cascade of liver parenchymal cells and fibrosis (Xiaxia et al., 2019). These effects include high levels of inflammatory cytokines, mitochondrial dysfunction, and oxidative stress. Necrotizing inflammation and fibrosis can progress and eventually lead to cirrhosis (Chi, 2017). However, the widely accepted theory is the “multiple-hit” pathogenesis (Ayonrinde et al., 2015). Changes due to the interaction of genetic and environmental factors, as well as the interactions between different organs and tissues, pancreas, gut, and liver, and broader metabolic dysfunction, are involved (Berardis and Sokal, 2014; Chi, 2017; Vlad et al., 2018). Moreover, scholars believe that environmental and genetic factors and the change in gut microbes in the induction of NAFLD in genetic predisposition, as well as intestinal flora changes lead to intestinal fatty acid, further activate the inflammatory pathways and release proinflammatory factors. Inflammatory cytokines increase liver inflammation and lipid accumulation, and the formation of gut-liver axis to a vicious cycle (Buzzetti et al., 2016; Xiaxia et al., 2019).

In recent years, the functional activity of key genes that synthesize proteins has been decisive in NAFLD. The PNPLA3 variant has been identified as the main genetic determinant of NAFLD. Variants with moderate effect sizes in TM6SF2, membrane bound O-acyltransferase domain containing 7 (MBOAT7), and glucokinase regulator (GCKR) were also shown to contribute significantly (Bellentani et al., 2004). PNPLA3, an enzyme that encodes I148M, is involved in the hydrolysis of triglycerides in adipocytes (Romeo et al., 2008). The lipid TM6SF2 is located in the endoplasmic reticulum and encodes E167K (rs58542926C/T), resulting in the loss of protein function, which in turn increases triglyceride deposition in the liver (Dongiovanni et al., 2015). Natural candidate genes are significantly involved in glucose and lipid metabolism during NAFLD development. Among the single nucleotide polymorphisms (SNPs) that lead to coding region mutations, such as PNPLA3 and TM6SF2, it is reasonable to infer that these defective proteins may be involved. For example, TM6SF2 mutants reduce liver production of very low-density lipoprotein (VLDL), thereby increasing the triglyceride (TG) content in the liver (Bonora et al., 2010).

Some studies have suggested that NAFLD progression follows the process of steatosis, lipotoxicity, and inflammation (Jou et al., 2008). The development of steatosis involves the interaction of many factors, such as dietary habits, gut flora, and genetic factors (Romeo et al., 2008; Jiang et al., 2015; Kirpich et al., 2015). Fat regenesis occurs through upregulation of adipogenic transcription factors, including sterol regulatory binding protein-1c (SREBP1c), carbohydrate-responsive element-binding protein (chREBP), and peroxisome proliferator-activated receptor gamma (PPAR-γ) (Anderson and Borlak, 2008). Fatty acids are mainly stored in the adipose tissue in the form of triacylglycerol. A previousstudy found that fatty acids in obese volunteers seemed to migrate from normal storage organs to the bone and liver tissue. Notably, FAT/CD36 (fatty acid translocation enzymes) promote fatty acid uptake by bone and liver tissues, which are significantly elevated in patients with obesity and NAFLD (Greco et al., 2008; Fabbrini et al., 2009). The accumulation of fat in the liver can lead to lipotoxicity and dysfunction of organelles, such as the mitochondria and endoplasmic reticulum (Browning and Horton, 2004; Bell et al., 2008). Steatosis further leads to the activation of IKKβ, which leads to increased signaling of the transcription factor nuclear factor kappa β (NF-κβ). Activation of NF-κβ induces the production of pro-inflammatory factors. These include tumour necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and interleukin-1beta (IL-1β) levels. These inflammatory factors can promote aggregation and activation of resident hepatic macrophages to further promote NASH inflammation (Ramadori and Armbrust, 2001; Fabbrini et al., 2009).

Oxidative stress may play an important role in NAFLD progression, and under normal physiological conditions, mitochondrial oxidation is the main oxidation pathway of fatty acid deposition. When ROS are overproduced during fatty acid oxidation, hydrogen polyunsaturated fatty acids are extracted from the liver, resulting in mass production of malondialdehyde (MDA) (Esterbauer et al., 1991). MDA can spread from its original site to other cells both inside and outside the cell, causing damage (Esterbauer et al., 1991). Catalase and glutathione levels decrease when ROS levels are elevated, and oxidative stress is exacerbated (Hongming et al., 2018). Lipid peroxidation increases collagen synthesis and cell death, which promotes steatosis and fibrosis (Huang et al., 2018).

Fatty acid outflow from the diet increases, and new fat formation releases free fatty acids from adipose tissue, contributing to TG accumulation in the liver, although to varying degrees (Yeh and Brunt, 2014). However, TG accumulation in the liver itself is not pathological, and may be protective in some cases. Hepatic diacylglycerol acyltransferase 2 (DGAT2) inactivation catalyzes TG synthase and reduces hepatic TG content but increases hepatitis and balloon-like changes (Brunt et al., 1999). This may seem paradoxical, but highlights the importance of liver fat in metabolic function. One possible mechanism for NASH-associated dysfunction involves a shift from minimal to substantial edema. This increase can be achieved by reducing the phosphatidylcholine (PC) levels (Machado et al., 2006) or lipid droplets coated with proteins (Soderberg et al., 2010; Angulo et al., 2013). Total PC levels were reduced in patients with both NAFLD and NASH (Ekstedt et al., 2006), which may be attributable to choline intake associated with NASH rather than choline deficiency (Richardson et al., 2007). In summary, NAFLD is a multifactorial disease with a complex pathogenesis. The prevention and treatment of NAFLD require further clinical and basic research.

Some studies have confirmed that flavonoid intake is inversely related to the risk of NAFLD (Mm et al., 2019). The mechanisms by which flavonoids exert anti-NAFLD effects are mainly through ameliorating inflammation, oxidative stress, and lipid metabolism, and regulating intestinal microbiota imbalance and the related gut liver axis. Flavonoids are natural polyphenol compounds that exist widely in all types of natural plants. Now, more than 9,000 kinds of flavonoids have been identified with a structure of a two phenolic hydroxyl benzene ring (A- and B-loops) interconnected through the central three carbon atoms. The basic parent nucleus is called a 2- phenylchromone (Tsuji et al.), biosynthesis from acetic acid and phenylalanine in plants (Weston and Mathesius, 2013). Flavonoids can be divided into flavonoids, flavonols, orange ketones, isoflavones, anthocyanins, chalcones, and dihydrogen derivatives according to the difference in the three-carbon atomic structure of the linked A and B rings, such as whether the ring is formed, oxidized, or replaced (Tsuchiya, 2010). The types of flavonoids from different sources and their anti-NAFLD mechanisms of action are listed in Table 1.

Flavonoids have a variety of pharmacological effects, including antitumor, antioxidant, antibacterial, antiviral, anti-inflammatory, and analgesic effects (Maleki et al., 2019; Makunga, 2020). Interestingly, flavonoids have positive effects on various NAFLD pathways, such as regulating lipid metabolism, insulin resistance, inflammation, and oxidative stress (Wier et al., 2015). Based on the above advantages, finding new anti-NAFLD drugs derived from plant flavonoids is a hot topic in current research (Figure 1).

The incidence of NAFLD increases each year, similar to clinical stress. Currently, NAFLD has an estimated annual medical and social cost of $292 billion (Younossi et al., 2016a). The different manifestations of NAFLD complicate the diagnosis, which ignores the true condition. The medical system is facing a severe challenge incombating this growing liver disease. Flavonoids have been proven to have very strong pharmacological activity and have excellent alleviating effects on NAFLD and NASH. Flavonoids may ameliorate NAFLD by regulating lipid metabolism, intestinal flora, and autophagy. Therefore, natural flavonoids have huge potential for the clinical development of NAFLD drugs in the future.