Etiology and Treatment of the Nutritional Fatty Liver in Fish: A Review

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INTRODUCTION
T he aquaculture sector has been growing for a century.
Aquatic goods are becoming more and more in demand as a component of human food as the world's population rises.The ecological environment of fish in aquaculture and wild ecology are very different from one another.Yet, fish require a very long period to evolve in an adapted way (Naiel et al., 2022).Fish feed manufacturers have the propensity to use more high-energy raw materials, such as carbohydrates and fats (Beamish and Thomas, 1984;Boujard et al., 2004;Fang et al., 2021;Wang et al., 2019), to boost yield and lower production costs due to the scarcity of protein raw resources and the high market demand (NRC, 2011).However, these high-energy meals frequently result in decreased growth performance (Bright et al., 2005;Fang et al., 2021;Li et al., 2016), liver diseases (Jia et al., 2020b;Li et al., 2021;Zhao et al., 2022), and reduced antioxidant and immunological capability (Jia et al., 2020a;Zhao et al., 2022;Zhou et al., 2020).The liver is typically where lipids and glucose metabolism takes place.In aquaculture, the liver health of farmed fish is very important.Many recent pieces of research have found that liver lipid buildup can cause nutritional fatty liver in fish, which can then result in oxidative stress and liver injury in fish (Du et al., 2008;Lu et al., 2014).Similar to this, lipid buildup in the liver can result from fish feed manufacturing lacking vital nutrients like choline and carnitine.Aquaculture's potential is greatly hampered by the nutritional fatty liver (Gao et al., 2014;Khosravi et al., 2015;Pan et al., 2017).Nutritional fatty liver has received more and more attention in the industry, so this paper mainly discusses the etiology and treatments the nutritional fatty liver in fish.

High-energy diets
High levels of carbohydrates Raising the percentage of carbohydrates in the meal, though can save feed costs and play a specialized function in protein sparing, it can also have many drawbacks, including a drop in fish development performance that causes liver lipid buildup and even fatty liver (Table I).
The protein, energy, and carbohydrate digestibility of rainbow trout (Oncorhynchus mykiss) rapidly reduced as the carbohydrate ratio in the meal rose (Tekİnay and Davies, 2001).It was discovered that adding 19.11% starch to the largemouth bass (Micropterus salmoides) diets significantly elevated the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in blood and that the liver cells were abnormally organized, inflated, and badly

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vacuolated (Zhao et al., 2022).Likewise, snakeheads (Channa argus) fed a diet containing 19% carbohydrates acquired high blood lipid levels and their liver's crude fat and glycogen levels rose (Ding et al., 2020).The plasma triglyceride (TG) content and liver vacuolization area of 45% corn starch-supplemented Nile tilapia (Oreochromis niloticus) were greater than those of the control group (30% corn starch).Also, a 45% corn starch diet caused liver injury in Nile tilapia; plasma AST and ALT activities were higher than in the control group (Li et al., 2021).Another study on Nile tilapia discovered that the expression of the lipid synthesis genes such as diacyl transferase (DGAT), sterol regulatory element binding protein 1 (SREBP1), and fatty acid synthase (FAS)) was significantly enhanced and that 45% of maize starch levels tended to create fatty livers (Zhu et al., 2020).According to study by Fang et al. (2021) early feeding of grass carp (Ctenopharyngodon Idella) with 60% maltodextrin improved the fish's growth performance.Nevertheless, sustained feeding of grass carp with 60% maltodextrin led to the development of clear signs of hyperglycemia, an increase in the expression of FAS mRNA, and the deposition of fat.Studies have revealed that eating a high-carbohydrate diet can make blunt snout breams (Megalobrama amblycephala) hyperlipemia, suppress the expression of genes involved in lipid oxidation, and increase the expression of genes involved in lipid synthesis (Ge et al., 2022;He et al., 2021).

High levels of lipids
Studies conducted as early as 1978 discovered that raising the quantity of fat in feed might have a specific impact on saving protein (Watanabe et al., 1978).Fish have a limited ability to use lipids in their meal.A range of issues, including lipid metabolic disorder, growth limitation, inappropriate lipid buildup in the liver, and even fatty liver, will result from overly blindly raising the level of lipids in feed (Zhou et al., 2020).
For largemouth bass, it was shown that a diet containing 10% fat had the highest growth results (Table II).Plasma ALT and AST levels rose and liver antioxidant capacity fell when dietary fat levels reached 20% (Zhou et al., 2020).Furthermore, research on largemouth bass found that when dietary fat levels exceed 16%, liver damage already develops (Bright et al., 2005).In the Nile tilapia study, it was discovered that the hepatosomatic index (HSI) and visceral somatic indices (VSI) of fish were greatly enhanced when the fat content in the food was 12%, and the concentration of TG in the liver and plasma was also dramatically elevated (Zhang et al., 2020).Hyperlipidemia in tilapia, increased lipid buildup in the liver, reduced antioxidant capability, and increased apoptosis are all linked to lipid levels of 21% or higher (Jia et al., 2020a, b).Studies on the large yellow

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Table II.Physiological effects of high-lipid diets in fish.

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croaker (Larimichthys crocea) have revealed that 17% of lipid levels can cause hyperlipidemia and hepatic lipid buildup (Zhang et al., 2023).Dai et al. (2019) discovered that when the fat content of blunt snout bream feed reached 11%, the antioxidant capacity of the liver reduced compared to the control group (6% lipid).At 11% lipid level, the expression of apoptosis-related genes (cysteineaspartic proteases 3/9 (Caspase 3/9)) in the liver of blunt snout bream was active.In grass carp, the expression of the lipid synthesis genes [acetyl-CoA carboxylase (ACC), FAS, and DGAT)] increased when the lipid content reached 8% (Sun et al., 2021).Nevertheless, other researchers have demonstrated that in grass carp, lipid level reaches 10.7%, lipid synthesis genes (ACC and FAS) are down-regulated and lipid deconstruction genes (peroxisome proliferatorsactivated receptor α (PPARα) and carnitine palmitoyl transferase 1 (CPT1)) are up-regulated (Li et al., 2016).

Essential nutrients deficiency
Choline: In addition to being a component of phospholipids, lecithin, and other tanglesome lipids, choline is a nutrient that functions similarly to vitamins by creating unstable methyl in living things (Zeisel, 1981).
It was discovered in research on parrot fish (Oplegnathus fasciatus) that a choline deficiency would cause a greater buildup of liver lipids and a reduction in the feed efficiency (FE) of parrot fish (Khosravi et al., 2015).By adding the choline synthesis inhibitor 2-amino-2-methyl-1-propanol (AMP) and various levels of choline to the diet of yellowtail kingfish (Seriola lalandi), Liu et al. (2021a) created a low choline model of yellowtail kingfish.According to the study, high levels of choline can reduce the pathological changes in the liver of yellowtail kingfish, which can lack choline and cause liver inflammation and damage.In the study of yellow catfish (Pelteobagrus fulvidraco), it was shown that adding choline to the diet was effective in reducing lipid accumulation in the liver, with the maximum choline addition level of 2273.6 mg/ kg decreasing lipid accumulation in the liver (Luo et al., 2016).Thus, the lack of choline can cause fish liver lipid accumulation and hepatic damage problems.
Vitamin: Vitamins are vital nutrients for all organisms.Fish cannot produce the majority of vitamins which can only be received from the diet.In a study on grass carp, Pan et al. (2017) found that the fish's growth performance, oxidation resistance, and immunology would all suffer from a lack of vitamin E in their diet.Insufficient vitamin E would make grass carp's inflammatory response worse by controlling the expression of nuclear factor kappa-B (NF-κB) and nuclear facter E2-related factor 2 (Nrf2) mRNA.
Nevertheless, the study did not examine how the grass carp's liver would respond to a vitamin deficiency.According to a study on Japanese flounder (Paralichthys olivaceus), adding small amounts of vitamins E and C to oxidized fish oil will help the fish maintain its usual development rate.Nevertheless, excessive vitamin E and C intake might result in lipid peroxidation, which was characterized by a rise in thiobarbituric acid reactive substances (TBARs) in liver tissue (Gao et al., 2014).Similar results were reported in studies on black sea bream (Acanthopagrus schlegeli), which showed that oxidized fish oil can impair growth and significantly raise HSI.Black sea bream's growth performance can be enhanced by adding vitamin E to oxidized fish oil (Peng et al., 2009).

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X. Zhao et al.
Table III displays the physiological impacts of essential nutrients on fish.Figure 1 shows an overview of the etiology and physiological effects of nutritional fatty liver disease.

Carnitine
A key element of lipid metabolism is carnitine.It is a synthetic precursor of the enzymes carnitine acyltransferase (CACT), CPT1, and carnitine palmitoyl transferase 11 (CPT11), and it facilitates the transportation of long-carbon chain fatty acids to the mitochondria for β-oxidation (Li et al., 2019b).
The activity of glutathione peroxidase (GPx) and malondialdehyde (MDA) levels in the liver of largemouth bass were considerably raised and lowered, respectively, by 0.02% L-carnitine supplementation (Chen et al., 2020).In studies on black sea bream, it was shown that supplementing the high-fat diet with 300 mg/kg of L-carnitine dramatically increased the activity of catalase (CAT) and lysozyme (LZM) in the fish's liver.Also, it can ameliorate the histological abnormalities in the liver brought on by a high-fat diet and promote the expression of genes for β-oxidation in the liver of black sea bream (Jin et al., 2019a).
Carnitine has left-handed and right-handed configurations.D-carnitine was discovered to enhance fat buildup in the liver of Nile tilapia and to trigger liver inflammation, oxidative stress, and apoptosis in Nile tilapia.D-carnitine, unlike L-carnitine, does not promote fish development (Li et al., 2019a).

Choline
As previously stated, a shortage of choline, an essential nutrient, induces fat buildup in the liver of parrot fish (Khosravi et al., 2015), and in yellowtail kingfish, it causes inflammation and liver damage (Liu et al., 2021a).Choline shortage is hypothesized to alter lipoprotein translocation in the liver, leading to the buildup of TG and total cholesterol (TC) in the liver and the development of fatty liver (Mai et al., 2009).
The liver lipid of cobia is reduced with increasing dietary choline levels, according to a cobia research (Mai et al., 2009).Choline supplementation in the diet of yellow catfish lowers fat buildup in the liver (Luo et al., 2016).Adding choline to a high-fat diet can minimize the inflammatory response induced by high-fat stress and lower cholesterol levels in the whole fish body and liver.
Additional research has revealed that choline can limit the expression of lipogenesis genes, boost the expression of lipolysis genes, and reduce the production of NF-κB and pro-inflammatory cytokines in the liver and gut of black seabream (Jin et al., 2019b).Different species and even the same species have different needs for choline at different growth stages.Studying the demand for choline in fish is a means to combat fatty liver.

Unsaturated fatty acid
A high-fat diet suppresses β-oxidation and PPARα expression, resulting in steatosis and liver injury (Kang-Le et al., 2014).A study in common carp (Cyprinus carpio) discovered that dietary supplementation with eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) rich algae boosted the expression of PPARα genes in the liver (Eljasik et al., 2020).A study on Atlantic salmon (Salmo salar) discovered that EPA can stimulate mitochondrial proliferation, boost lipid metabolism, and decrease intracellular lipid concentration (Kjaer et al., 2008).Highly unsaturated fatty acid (HUFA) was also discovered to suppress FAS gene activity in the liver, limit fatty acid synthesis, and increase fatty acid catabolism in Atlantic salmon, resulting in hypolipidemia (Morais et al., 2011).
Early research has revealed that saturated fatty acids (SFAs) are more readily deposited in tissues than monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) in rats (Clarke et al., 1990).According to research on giant yellow croakers, employing fish oil or soybean oil rich in unsaturated fatty acids as a source of fat will significantly lower blood ALT and AST levels (Qiu et al., 2017).Studies on silvery black porgy (Sparidentex hasta) has revealed that when the n-3 longchain polyunsaturated fatty acid (n-3 LC-PUFA) content in the diet is too low (0.1%), the range of HSI, plasma TG, and TC of silvery black porgy increases.The rise of n-3 LC-PUFAs will boost the growth performance and immunity of silvery-black porgy, while the plasma TG and TC contents will decrease (Mozanzadeh et al., 2015).We can observe that eating unsaturated fatty acids helps to reduce lipid buildup.If the kinds and need for unsaturated fatty acids in various fish are thoroughly examined, it may be utilized as a strategy to combat nutritional fatty liver.

Plant extract
Plant extracts are classified as alkaloids, polysaccharides, flavonoids, organic acids, and so on.A

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lot of studies have been done on plant extracts as dietary supplements in the battle against fatty liver.
According to a study on black seabream, adding 10g/ kg or 20g/kg of betaine to meals with 17% fat content will reduce steatosis and inflammatory reactions in the liver produced by high-fat diets via SIRT1/SREBP1/PPARα (Jin et al., 2021).It was shown that blunt-nosed seabream's liver apoptosis and oxidative stress were considerably decreased when berberine at doses of 50 mg/kg or 100 mg/kg was added to diets with 15% fat content (Lu et al., 2017).When hybrid grouper (Epinephelus lanceolatus ♂ × Epinephelus fuscoguttatus ♀) are exposed to the stress of a high-fat diet, it has been discovered that the extract of ginkgo biloba leaves lowers their blood lipid levels, increase their antioxidant capacity, and increases the expression of immune-related genes.In hybrid grouper, fewer apoptosis-related genes were expressed (Tan et al., 2018).Forskolin, a type of herbal medicine extract from China, increased the expression of genes for lipolysis and β-oxidation in Nile tilapia diets with 15.31% fat content, which reduced the buildup of lipids in the liver (Zhang et al., 2019).
A lot has been written about polysaccharides as well.Fucoidan, for instance, was discovered to suppress nonalcoholic fatty liver disease through the ASGR/STAT 3 / HNF 4 A signaling pathway in research in zebrafish (Wu et al., 2020).At a dose of 8000 mg/kg, oxidized konjac glucomannan (OKGM), an oxidized polysaccharide, was shown to dramatically lower plasma levels of Yafish (Schizothorax prenanti) TG and TC and to boost the activity of hepatic lipase HL and PPARα (Zhang et al., 2017).In the tilapia investigation, it was discovered that glycyrrhiza total flavones might reduce liver damage brought on by excessive fat through Nrf2 and toll-like receptor (TLR) signaling pathways and boost the fish's antioxidant abilities (Du et al., 2022).The buildup of lipid droplets in the liver caused by excessive fat was also shown to be improved by tea polyphenol in another study on tilapia, and the decrease of lipid droplets was dosedependent.Also, tilapia's immune system and antioxidant capacities can be boosted by tea polyphenols (Qian et al., 2021).Numerous plants contain chlorogenic acid, a kind of polyphenolic acid.Chlorogenic acid can reduce the levels of TG and TC in the plasma of largemouth bass, encourage the expression of genes linked to lipolysis, and enhance the largemouth bass's antioxidant capacities (Yin et al., 2021).
Plant extracts will continue to get attention in intensive aquaculture because of their green color and diversity, but more study is still needed on plant extracts' ability to treat fatty livers.

Short-chain organic acid salt
Some studies show that short-chain organic acids salt can improve the nutritional fatty liver in fish.Calcium pyruvate was found to enhance the growth performance and liver antioxidant capacity of juvenile golden pompano (Trachinotus ovatus) fed a high-fat diet.Moreover, it was shown that calcium pyruvate might up-regulate the expression of genes involved in lipolysis (PPARα, CPT1, and fatty acid binding protein 1 (FABP1)) and down-regulate the expression of genes involved in lipid synthesis (SREBP1, FAS, and ACC), hence decreasing lipid buildup (Shao et al., 2022).Similarly, calcium pyruvate was shown to up-regulate gene expression related to lipolysis and to down-regulate gene expression linked to lipid synthesis in large yellow croaker (Zhang et al., 2023).
In the study of largemouth bass, it found that sodium butyrate could up-regulate the expression of genes of lipolysis (CPTI and PPARα) and improve the growing health and intestinal flora composition of largemouth bass under high-fat stress (Chen et al., 2023).Related investigations on grass carp discovered that sodium butyrate can lower high-fat diet stress plasma AST and ALT levels, lower liver apoptosis and inflammatory factors, and increase grass carp immunity (Gao et al., 2022).Zhou et al. (2019) also proved that sodium butyrate has the effect of reducing liver lipid accumulation.

Selenium element
The antioxidant enzyme GPx active center contains selenoprotein.Liver GPx activity may rise with increased dietary selenium (Chen et al., 2013).A related study found that feeding grass carp 0.3 or 0.6 mg/kg of selenium together with a high-fat diet significantly reduced the amount of MDA in their blood and boosted the activity of superoxide dismutase (SOD) and GPx (Yu et al., 2020;Liu et al., 2021b).Comparable improvements were made in the innate immune system, antioxidant capacity, and growth performance in juvenile Atlantic white croakers (Argyrosomus regius) by food supplementation with 3.98mg of organic selenium (Mansour et al., 2017).Yu et al. (2020) have revealed that nano-selenium may boost the mRNA expression of GPx and CAT as well as Nrf2 in grass carp.
Table IV displays the fish nutritional fatty liver therapy measures.Figure 2 shows an overview of treatment options and physiological effects of nutritional fatty liver disease.

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X. Zhao et al.

CONCLUSION
It is difficult for fish to adjust to the rigorous living circumstances of modern intensive aquaculture.We still can't produce fish in simulated natural settings despite the enormous market demand and economic advantages.We should research new environmentally friendly, secure, and effective anti-nutritional fatty liver chemicals for today's heavy fish production.The majority of recent papers only look at one pathway or one crucial gene when examining the causes of nutritional fatty liver in fish.To study feed formula, future studies should concentrate on how to explore the whole metabolic and regulation network of lipid metabolism and nutritional fatty liver, as well as how to develop a pertinent database for each farmed fish.In short, the problem of nutritional fatty liver in cultured fish will not be solved overnight, which requires us to continue to invest in slow research.

Fig. 1 .
Fig. 1.General overview of the etiology and physiological effects of the nutritional fatty liver.

Fig. 2 .
Fig. 2. General overview of treatment options and physiological effects of the nutritional fatty liver.