Total ammonia nitrogen (TAN, the sum of NH₃ and NH₄+) is a major water quality concern in aquaculture systems worldwide, where its accumulation can lead to severe fish health deterioration and economic losses. The toxic fraction, unionized ammonia (NH₃), increases with pH and temperature. However, the precise cellular mechanisms underlying ammonia-induced hepatotoxicity remain poorly defined, hindering the development of targeted management strategies. In this study, we tested the hypothesis that ferroptosis, a novel form of regulated cell death, serves as a key execution mechanism. Largemouth bass (Micropterus salmoides), an economically important aquaculture species highly sensitive to ammonia stress, were exposed to environmentally relevant concentrations of total ammonia nitrogen (14.94 and 29.88 mg/L) for 96 h. Based on the measured TAN and water parameters (25 °C, pH 8.0), the calculated NH₃ concentrations were 0.80 mg/L for LA and 1.60 mg/L for HA. An integrated approach combining histopathology, biochemistry, and transcriptomics was employed. Ammonia exposure caused dose-dependent hepatic damage and lipid peroxidation. Mechanistically, ammonia influx, associated with altered expression of ammonia transporter genes RHAG and AQP12, depleted the intracellular glutamate pool, crippling glutathione (GSH) synthesis and suppressing antioxidant enzymes (SOD1, CAT, GPX4). Concurrently, ammonia disrupted iron homeostasis in association with changes in the SLC46A1/HMOX1 pathway, leading to significant hepatic iron accumulation. The coordinated downregulation of the core ferroptosis defense system (SLC7A11 and GPX4) created biochemical conditions consistent with ferroptosis-related lipid peroxidation. These findings support the involvement of ferroptosis-related pathways in ammonia-induced hepatotoxicity, providing a mechanistic basis for improved ammonia management in aquaculture systems.

The effects of extracts of Ginkgo biloba leaves (EGb) on feed quality, hypoxia tolerance, and disease resistance were examined in this study. EGb were prepared by using four different solvents: aqueous, ethanol, ethyl acetate, and petroleum ether. Subsequently, five distinct fish diets were formulated, including one basal diet and four EGb-supplemented diets (1 % EGb addition respectively). A total of 450 juvenile Jian carp weighing 10.55 +/- 0.22 g were randomly assigned and fed these diets for 15 d. The results showed that EGb supplementation decreased the leaching loss ratio and inhibited mold growth and lipid peroxidation in pelleted fish feeds (P < 0.05). The activities of trypsin, chymotrypsin, amylase, superoxide dismutase, lipase, glutathione-S-transferase, and anti-hydroxy radical in fish digestive organs were all improved, while H2O2 levels were decreased by dietary EGb supplementation (P < 0.05). Under hypoxic conditions, dietary EGb supplementation lowered the rate of oxygen consumption while increasing the duration for fish (P < 0.05). Furthermore, dietary EGb supplementation decreased the mortality of Jian carp after challenge with Aeromonas hydrophilia and increased plasma lysozyme activity (P < 0.05). Overall, our results suggest that EGb supplementation improves feed quality and inhibits mold growth. The beneficial effects of EGb on fish growth performance may be associated with improved digestive ability. Dietary inclusion of EGb enhanced the antioxidant status to improve hypoxia tolerance and disease-resistance ability in fish. The aqueous extract (AQE) derived from Ginkgo biloba leaves demonstrated superior beneficial effects compared to other extracts.

Intestinal functional zonation and segment-specific microbiota responses to high-fat diet (HFD) are poorly understood in largemouth bass. In this study, a feeding experiment was conducted using 390 largemouth bass with an initial body weight of 10.87 ± 0.05 g. The impacts of HFDs on intestinal histomorphology and microbial communities across four intestinal segments, compared with a normal diet (ND), using 16S rRNA gene sequencing and histological analysis. The intestine was divided into four segments based on morphological features and length. Results showed that HFDs significantly decreased intestinal fold height and induced inflammatory lesions. Microbial genera exhibited distinct spatial distribution: Klebsiella was concentrated in segments 1 and 2, Cetobacterium in segment 4, and Acinetobacter was abundant in most segments. HFDs significantly increased opportunistic, potentially pathogenic taxa, including Acinetobacter and Plesiomonas, while reducing probiotics such as Paenibacillus and Bacillus. Segmented analysis detected more differential microbes and revealed potentially pathogenic taxa overlooked by whole-intestine analysis. These findings reveal segment-specific microbial distribution and HFD-induced intestinal dysbiosis in largemouth bass, providing a theoretical basis for intestinal functional zonation and the optimization of healthy aquaculture strategies.

Largemouth bass (Micropterus salmoides) is an economically important aquaculture species in China. Aeromonas veronii (A. veronii), a conditional pathogen, causes substantial economic losses in largemouth bass farming. Valine (Val), an essential amino acid for fish, plays a crucial role in the growth of largemouth bass; however, its effects on liver health under A. veronii infection remain unclear. This study investigated the influence of dietary Val on hepatic antioxidant capacity, inflammatory response, apoptosis, and autophagy in largemouth bass following A. veronii challenge. A total of 720 juveniles (37.99 ± 0.01 g) were fed six isonitrogenous diets with graded levels of Val (11.0-26.1 g/kg diet) for 11 weeks, and the fish were subjected to a 7 d challenge trial with A. veronii. Results showed that dietary Val supplementation (20.1 g/kg) effectively mitigated A. veronii-induced histopathological damage, notably attenuating both necrotic changes and inflammatory infiltration. Mechanistically, dietary Val enhanced hepatic antioxidant capacity by activating the Nrf2/Keap1 pathway, significantly increasing antioxidant enzyme activities (SOD1, CAT, GST, GPx, GR, GCLC) and reducing oxidative stress markers (ROS, MDA, PC) (P < 0.05); Dietary Val attenuated apoptosis through downregulation of endoplasmic reticulum (ER) stress markers (GRP78, ATF6, IRE1, PERK, eIF2α, ATF4, CHOP) and modulation of Bcl2/Bax ratio (P < 0.05); Dietary Val suppressed excessive autophagy as evidenced by decreased expression of key autophagy-related genes (ULK1, ATG9, BECN1, ATG7, ATG5, LC3) and proteins (P < 0.05). Dietary Val regulated inflammatory responses by suppressing pro-inflammatory cytokines (TNF-α, IL-1β) while promoting anti-inflammatory factors (TGF-β2, IL-10). These findings demonstrate Val's multifaceted hepatoprotective effects through coordinated regulation of oxidative stress, apoptosis and autophagy pathways in A. veronii-infected largemouth bass.

This review aims to summarize the current knowledge regarding the effects of various dietary additives on osmoregulation in aquaculture. It covers salt supplements, potassium supplements, myo-inositol supplements, metal ion supplements, and monosaccharides, emphasizing their importance in improving osmotic adjustment. The review provides an overview of the underlying mechanisms involved in osmoregulation and highlights the role of some specific dietary additives in promoting osmotic balance. Dietary salt and inorganic potassium directly provide ions for osmotic balance, while dietary metals, inositol, and monosaccharides can help mitigate the effects of osmotic stress and enhance the body's capacity for osmoregulation. Additionally, the addition of exogenous glucose not only provides energy required for osmoregulation but also acts as an osmolyte itself, contributing to osmoregulation. The review underscores the need for further research to better understand the interactions between dietary additives and osmoregulation in aquaculture. It also highlights the importance of considering species-specific requirements, optimal dosage levels, and the potential synergistic effects of combining multiple additives. Overall, carefully selected and properly administered dietary additives have the potential to improve osmoregulatory capacity, mitigate osmotic stress, and enhance the overall performance and welfare of cultured aquatic organisms. Future research should focus on elucidating the underlying mechanisms and optimizing the application of dietary additives to maximize their effectiveness in aquaculture systems.

This experiment was arranged to explore the impacts of dietary MHA on liver lipid metabolism in largemouth bass. A total of 480 fish (14.49 ± 0.13 g) were randomly allocated into four groups, each with three replicates. They were then given four different diets containing graded levels of MHA (0.0, 3.0, 6.0, and 9.0 g/kg) for 84 days. The results showed that dietary MHA increased hepatic lipid vacuoles and lipid content (p < 0.05). Dietary supplementation with MHA 9.0 g/kg diets increased the activities of acetyl-coA carboxylase (ACC), fatty acid synthase (FAS), and stearoyl-coA desaturase 1 (SCD-1). Dietary MHA up-regulated the mRNA expressions of liver lipid synthesis (ACC, FAS, SCD-1 and SREBP-1c) (p < 0.05). Furthermore, compared with the 0.0 g/kg diet group, the group supplemented with 9.0 g/kg MHA in the diet exhibited a significant decrease in the activities of liver lipid-oxidation-related enzymes (acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), and stearoyl-CoA desaturase 1 (SCD-1), as well as HSL and CPT1) and the gene expressions of ATGL, HSLa, HSLb, CPT1a, and PPARα (p < 0.05). Additionally, the mRNA expressions and protein levels of SIRT1 and AMPK in the 9.0 g/kg MHA-supplemented group were significantly lower than those in the 0.0 g/kg diet group (p < 0.05). Overall, the present results suggested that dietary MHA could increase lipid accumulation through regulating SIRT1/AMPK signaling pathways in the livers of largemouth bass.

A 13 weeks feeding assessment was performed with the aim of determining the influence of enzyme-treated soy protein (ETSP) supplementation on growth performance, intestinal barrier function, liver lipid metabolism and related signaling pathways of largemouth bass (Micropterus salmoides). A total of 1000 largemouth bass (initial average weight: 22.3 +/- 0.1 g) were fed five different diets used in the experiment, each with a different amount of ETSP: 0.0 (M1), 20.0 (M2), 40.0 (M3), 60.0 (M4), and 80.0 (M5) g/kg. The results demonstrated that dietary ETSP supplementation significantly improved growth performance, body composition, and nutrient retention (P<0.05). Histological examination revealed significantly greater intestinal villus height in fish fed optimal ETSP levels (P<0.05). The 40.0 g/kg ETSP diet enhanced intestinal barrier function by upregulating tight junction proteins and anti-inflammatory cytokines while downregulating pro-inflammatory factors through modulation of the AMPK-SIRT1-NF-kappa B signaling pathway (P<0.05). Moreover, histological examination revealed optimal ETSP (40.0 g/kg) improved hepatic macrovesicular steatosis and cellular nuclei displacement, restrain lipid vacuoles' relative area, also activities of plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST), thereby lowered liver damage. Dietary ETSP may suppress fatty acid transport and lipid synthesis, and promote lipid catabolism by activating FGF21/AMPK pathway. Finally, quadratic regression analysis of the specific growth rate (SGR) revealed that the optimal dietary ETSP level for largemouth bass was 40.0 g/kg.

This study investigated the effects ofAstragalus membranaceus by-product (AP) on feed quality, mold growth, and fish disease-resistant ability. Seven fish diets were prepared with gradient levels of AP (0, 1.0, 2.0, 4.0, 8.0, 16 and 32 %). A total of 420 juvenile Crucian carp (7.17 +/- 0.05 g) were randomly assigned and fed for 60 days. Results showed that AP supplementation decreased pellet hardness, and increased pellet durability index, as well as inhibited mold growth of pelleted fish feed (P<0.05). Fish weight gain (WG), feed efficiency (FE), content of reduced glutathione, and the activity of anti-hydroxy radical and anti-superoxide anion, as well as enzymes such as trypsin, amylase, alkaline phosphatase, Na+/K+-ATPase, superoxide dismutase, glutathione reductase, lipase, and glutathione-S-transferase in fish digestive organs were all improved (P<0.05), while malonaldehyde levels were decreased by dietary AP supplementation (P<0.05). The optimal AP supplementation levels for Crucian carp were 1.02 % and 1.03 %, as determined by broken-line regression analysis of WG and FE. Furthermore, dietary AP supplementation decreased the mortality of crucian carp after challenge with A. hydrophilia and increased lysozyme activity, and levels of total amino acids and protein in the plasma, and decreased content of ammonia, and activities of glutamate-oxaloacetate transaminase and glutamate-pyruvate transaminase in the plasma (P<0.05). Our study suggested that AP supplementation improves feed quality and inhibites mold growth. The beneficial of AP on fish growth performance may be associated with improved absorption and digestion. Dietary inclusion of AP enhanced non-specific immunity and antioxidant status so as to improve fish disease-resistant ability.

This study aimed to investigate the effects of dietary chromium picolinate (Cr-Pic) on growth performance, glucose (GLU) metabolism, and hypoxia tolerance of juvenile grass carp (Ctenopharyngodon idella). A total of 2160 juvenile grass carp (10.98 ± 0.01 g) were randomly divided into six groups with six cages each, containing 60 fish per cage. They were fed experimental diets with chromium (Cr3+) at 0.00 (basal diet), 0.40, 0.80, 1.20, 1.60, and 2.00 mg/kg (actual contents: 0.25, 0.67, 1.08, 1.63, 2.08, and 2.65 mg/kg, respectively) for 10 weeks. Then, 96 fish were selected and divided into a normoxia group (n = 48) and a hypoxia group (n = 48) for a 96 h hypoxia trial. The growth trial results showed that, compared to the 0.25 mg/kg Cr3+ group, the 0.67 to 2.65 mg/kg Cr3+ groups enhanced the percent weight gain (PWG; P < 0.001) and increased the protein retention value (P < 0.001), and promoted the growth and development of juvenile grass carp. In the hypoxia group, dietary 1.08 mg/kg Cr3+ reduced the activities of serum glutamic pyruvic transaminase (GPT; P < 0.001) and glutamic oxaloacetic transaminase (GOT; P < 0.001), thereby alleviating liver damage. Furthermore, dietary 1.63 mg/kg Cr3+ also increased glycolysis, the tricarboxylic acid cycle, and oxidative phosphorylation in juvenile grass carp under hypoxia stress, as evidenced by the enhanced activities of liver enzymes, such as hexose kinase (P < 0.001), pyruvate kinase (P < 0.001), and malate dehydrogenase (P < 0.001). Additionally, in the liver, dietary 1.08 to 1.63 mg/kg Cr3+ inhibited the expression of proteins related to gluconeogenesis, like glucose-6-phosphatase (P < 0.001) and phosphoenolpyruvate carboxykinase (P < 0.001), and elevated the relative mRNA expression levels of mitochondrially encoded NADH dehydrogenase I (P < 0.001) and adenosine triphosphate (ATP) synthase-coupling factor 6 (P < 0.001), thereby increased ATP production under hypoxia stress. Moreover, in the hypoxia group, dietary 1.08 to 1.63 mg/kg Cr3+ up-regulated the liver relative mRNA expression levels of nuclear respiratory factor 1 (P < 0.001) and mitofusin 1 (P < 0.001), and down-regulated the relative mRNA expression levels of mitochondrial fission protein 1 (P < 0.001) and B-cell lymphoma-2-interacting protein 3 (P < 0.001), which contributed to the maintenance of normal mitochondrial function. Regression analysis was conducted utilizing the parameters of PWG, GPT activity, and ATP content, and the dietary Cr3+ requirements of juvenile grass carp were 1.02, 1.55, and 1.54 mg/kg, respectively. Overall, this study provided new insights into the effects of Cr3+ on GLU metabolism of juvenile grass carp under hypoxia stress, offering a nutritional regulation strategy for improving the ability of fish to resist hypoxia stress.

Vitamin D3 (VD3) has been recognized to improve glycolipid metabolism, whereas the mechanism behind is still unknown. Herein, largemouth bass, Micropterus salmoides (LMB) were fed with six diets with different concentrations of VD3 for 12-weeks. The results demonstrated that VD3 improved growth, feed utilization and proximate composition. In addition, VD3 enhanced liver glycogen content, glycolysis-related genes, glycogen synthase kinase 3(3 a (GSK3(3 a) and insulin receptor a (INSRa) mRNA expression, suppressed plasma glucose content and gluconeogenesisrelated genes expression. VD3 down-regulated forkhead box O1a (FoxO1a) mRNA expression and nucleus-FoxO1 protein level and increased cytoplasm-FoxO1 protein level. VD3 decreased plasma triglyceride (TG), total cholesterol (TC) contents and aspartate aminotransferase (AST), alanine aminotransferase (ALT) activities. Histological analyses revealed that VD3 decreased macrovesicular steatosis of liver. VD3 improved liver lipid metabolism via spursing lipolysis, transportation, and inhibiting lipogenesis. RT-PCR and western blot tests suggested that the potential mechanism may partially be attributed to the VDR/AMPK/SIRT1 pathway mediated glycolipid metabolism. According to the broken line regression analysis of PWG, the dietary VD3 requirement of LMB was estimated to be 1980.86 IU/kg. This study provides theoretical support and a novel mechanism of action VD3 glycolipid metabolism via VDR/AMPK/SIRT1 pathway.

Hypoxia is a prevalent phenomenon in aquaculture, threatening the survival and growth of fish. The present study investigated the effects of dietary glutathione on hepatopancreatic injury in juvenile grass carp under hypoxia stress and its possible mechanisms. We formulated six diets with different levels of glutathione (0.00, 145.95, 291.90, 437.85, 583.80 and 729.75 mg/kg) to feed juvenile grass carp with initial body weights (11.08 +/- 0.01 g) for 70 days, followed by a 96-h hypoxia stress experiment. The results of this study showed that glutathione improved the growth performance of juvenile grass carp and alleviated hypoxia-induced glutathione depletion (GSH and GCL). In addition, glutathione decreased serum GPT and GOT activities and reduced ROS, MDA and PC contents in the hepatopancreas to attenuate tissue oxidative damage. Moreover, glutathione alleviated hypoxia stress-induced hepatopancreatic mitochondrial dysfunction, which may be related to the enhancement of mitochondrial oxidative phosphorylation and improvement of the mitochondrial quality control system. Furthermore, the ability of glutathione to alleviate hypoxia-induced hepatopancreatic autophagy may be associated with the inhibition of autophagy initiation, nucleation, elongation and degradation phases. In conclusion, glutathione attenuated hypoxia stress-induced hepatopancreatic injury in fish, which may be related to the fact that glutathione attenuated mitochondrial dysfunction and autophagy. Using the SGR from growth experiment and serum GPT activity under hypoxia stress as markers, it was determined that the optimum levels of glutathione to be added to juvenile grass carp for growth and against hypoxia stress were 451.50 mg/kg and 437.88 mg/kg, respectively.

The influence of dietary glutamate (Glu) was evaluated in a 56-day feeding trial on the growth performance and flesh quality of largemouth bass (Micropterus salmoides). A total of 1170 fish (average body weight 24.05 ± 0.22 g) were randomly allocated into six groups, with three replicates per group. They were fed diets containing Glu in levels of 11.40% (G1), 11.88% (G2), 12.53% (G3), 13.27% (G4), 14.33% (G5), and 15.62% (G6). We found that, over a 56-day feeding period, the final body weight (FBW) of largemouth bass was about 4–5 times the IBW. The FBW, percent weight gain (PWG), specific growth rate (SGR), feed efficiency (FE), and protein efficiency ratio (PER) initially increased and then decreased with elevating dietary Glu levels. Likewise, protein content, lipid content, apparent digestibility coefficient of dry matter (ADCD), and apparent digestibility coefficient of protein (ADCP) followed a similar pattern. Supplementation with Glu significantly improved the hepatosomatic index (HSI), viscerosomatic index (VSI), and relative gut length (RGL). Moreover, dietary Glu augmentation noticeably enhanced flesh composition such as muscle protein, ash, lipid, amino acid contents, and polyunsaturated fatty acids (PUFAs). Furthermore, dietary Glu supplementation enhanced muscle physicochemical quality (such as drip loss and pH), textural properties (adhesiveness and cohesiveness), and biochemical indices such as total protein (TP) and salt-soluble protein, while decreasing muscle cathepsin B (CtsB) and lactate dehydrogenase (LD) contents, thereby improving flesh quality. In conclusion, these findings suggest that Glu plays a crucial role in enhancing both growth performance and muscle quality in largemouth bass. The optimal dietary requirement for juvenile largemouth bass was estimated to be approximately 125.1 g/kg of diet based on SGR analysis.

Fish meal (FM) is a crucial high-quality protein source in aquafeeds, prized for its excellent palatability, high digestibility, and rich protein content [...]

Hypoxia often induces inflammatory in fish, likely due to disrupted ion homeostasis and DNA damage. Chromium (Cr3+), as an essential trace element for animals, aids DNA repair and regulates ion homeostasis. However, whether Cr3+ could alleviate hypoxia-induced liver inflammation by regulating ion homeostasis or reducing DNA damage is unknown, which is the purpose of this study. A total of 2160 juvenile grass carp (10.98 ± 0.01 g) were randomized to 6 treatments. The fish were fed diets containing Cr3+ at concentrations of 0.25 (basal diet), 0.67, 1.08, 1.63, 2.08, and 2.65 mg/kg for 70 d. Then, 96 fish were selected, randomized to control (n = 48) or hypoxia (n = 48) groups, and subjected to hypoxia stress for 96 h. The results showed that: (1) In the hypoxia group, Cr3+ inhibited the gene expression of pro-inflammatory cytokines and pyroptosis-related factors, while it enhanced the gene expression of anti-inflammatory cytokines. This indicated that Cr3+ alleviated hypoxia-induced liver inflammatory responses. (2) Cr3+ reduced the relative content of mitochondrial DNA (mtDNA) in the cytoplasm of the hypoxia group (P < 0.05), which suggested that Cr3+ had a potential benefit in alleviating DNA damage. (3) In the hypoxia group, Cr3+ suppressed the expression of proteins associated with K+ and Cl- efflux and promoted the expression of proteins related to K+ and Cl- influx, thereby contributing to ion homeostasis. In summary, Cr3+ likely maintained ion homeostasis and preserved mtDNA integrity, thereby suppressing liver inflammatory responses, which may provide a novel nutritional regulation strategy for enhancing hypoxia stress resistance in fish.

Cetobacterium somerae, a symbiotic microorganism resident in various fish intestines, is recognized for its beneficial effects on fish gut health. However, the mechanisms underlying the effects of C. somerae on gut health remain unclear. In this experiment, we investigated the influence of C. somerae (CGMCC No.28843) on the growth performance, intestinal digestive and absorptive capacity, and intestinal structural integrity of juvenile grass carp (Ctenopharyngodon idella) and explored its potential mechanisms. A cohort of 2,160 juvenile grass carp with an initial mean body weight of 11.30 ± 0.01 g were randomly allocated into 6 treatment groups, each comprising 6 replicates (60 fish per replicate). The experimental diets were supplemented with C. somerae at graded levels of 0.00 (control), 0.68 × 10⁹, 1.35 × 10⁹, 2.04 × 10⁹, 2.70 × 10⁹, and 3.40 × 10⁹ cells/kg feed. Following a 10-week experimental period, biological samples were collected for subsequent analyses. Dietary supplementation with C. somerae at 1.35 × 10⁹ cells/kg significantly enhanced growth performance, intestinal development, and nutrient retention rate in juvenile grass carp (P < 0.05). The treatment resulted in increased intestinal acetic acid concentration and enhanced activities of digestive enzymes and brush border enzymes (P < 0.05). Furthermore, it reduced intestinal permeability (P < 0.05), preserved tight junctions (TJ) ultrastructural integrity, and increased the expression of TJ and adherens junctions (AJ) biomarkers at both protein and transcriptional levels (P < 0.05). Mechanistically, these effects may be correlated with enhanced antioxidant capacity and coordinated modulation of the RhoA/ROCK, Sirt1, and PI3K/AKT signaling pathways. The appropriate supplementation levels, based on weight gain rate, feed conversion ratio, the activity of serum diamine oxidase and the content of lipopolysaccharide, were 1.27 × 10⁹, 1.27 × 10⁹, 1.34 × 10⁹ and 1.34 × 10⁹ cells/kg, respectively. C. somerae improved intestinal digestive and absorptive capacity of juvenile grass carp, maintained intestinal structural integrity, and thus promoted their growth and development. This work demonstrates the potential of C. somerae as a probiotic for aquatic animals and provides a theoretical basis for its utilization in aquaculture.

Atherosclerosis (AS) is a prevalent cardiovascular condition, and the growth and phenotypic switch of vascular smooth muscle cells (VSMCs) play a crucial role in its development. Studies have revealed that the activation of certain transcription factors and signaling pathways can trigger these cellular changes. Consequently, targeting these pathways and pivotal molecules has emerged as a promising strategy for AS treatment. Drugs that can reverse the cellular changes in VSMCs may offer new therapeutic options for AS, marking a significant advancement. While previous research has suggested that urolithin B (Uro B) possesses anti-atherosclerotic properties, its exact mechanism remains to be fully understood, especially the effect of Uro B in VSMCs. This study discovered that Uro B can impede the proliferation and migration of VSMCs prompted by PDGF-BB, as well as their phenotypic changes, indicating that Uro B could potentially prevent AS by inhibiting the phenotypic switch of VSMCs.

This study explored the effects of dietary vitamin C on growth performance, flesh quality, and collagen and elastin content in adult grass carp (Ctenopharyngodon idella). A total of 540 fish (initial weight 585.00 ± 0.96 g) were randomly distributed into six groups, with three replicates per group and 30 fish per replicate. Fish were fed six experimental diets containing 4.01, 44.13, 83.47, 124.17, and 167.18 mg/kg L-ascorbyl-2-polyphosphate (C2PP), as well as 82.82 mg/kg ethylcellulose-coated ascorbic acid (EC-AA), for 9 weeks. The results indicated that C2PP supplementation (44.13-167.18 mg/kg) enhanced adult grass carp growth performance. It also increased muscle hardness, chewiness, and springiness, as well as crude protein and crude lipid content, while decreasing cooking loss and moisture content. Vitamin C supplementation increased collagen and elastin synthesis in adult grass carp through the activation of the transforming growth factor-beta (TGF-β)/mothers against decapentaplegic homolog (Smads), phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/protein kinase B (AKT)/target of rapamycin (TOR), La ribonucleoprotein domain family member 6 (LARP6), and Fibulin-5 signaling pathways. Concurrently, vitamin C could inhibit collagen and elastin degradation by upregulating tissue inhibitor of metalloproteinases (TIMPs). Based on percent weight gain (PWG), hardness, and chewiness, the dietary vitamin C requirements for adult grass carp were 107.05, 104.42, and 97.00 mg/kg, respectively. These results indicate that vitamin C can improve the flesh quality of grass carp by promoting collagen and elastin deposition.

Appropriately increasing the level of fat in feed exhibits potential protein-saving effects, but high levels of fat in the feed can cause liver injury in fish of largemouth bass. 18β-Glycyrrhetinic acid (GA), a natural extract from glycyrrhiza, has been shown to enhance growth performance and improve intestinal health in animals. Thus, the present study was to investigate the effects of dietary GA on high-fat-induced liver fibrosis and the potential mechanisms in juvenile largemouth bass. The experiment comprised five different diets, including normal control (NC) diet (NC group), high-fat (HF) diet (HF group), HF diet supplemented with 0.5 mg/kg GA (HFL group), HF diet supplemented with 1.0 mg/kg GA (HFM group), and HF diet supplemented with 1.5 mg/kg GA (HFH group). A total of 750 healthy largemouth bass with an initial average weight of 17.39 ± 0.09 g were selected and randomly divided into five treatment groups, with three replicates per group and 50 fish per replicate. The feeding experiment was conducted for 11 weeks. In the liver, compared to NC, HF diet led to a marked reduction of the mitochondria-associated membranes (MAMs) gap, resulting in mitochondrial Ca2+ overload, augmented liver Ca2+ transport proteins expressions and reactive oxygen species (ROS) content (P < 0.05), decreased mitochondrial membrane potential (MMP) and adenosine 5'-triphosphate (ATP) content in the liver (P < 0.05). Thus, the activation of transforming growth factor-β1 (TGFβ1) - SMAD family member 2 (Smad2/3) signaling pathway resulted in the upregulation of pro-fibrogenic markers (α-Sma, collagen I, fibrontein and Mmp9) mRNA expressions (P < 0.05). However, GA could alleviate this phenomenon. Through virtual docking analysis showed that the docking energy between GA and Sirtuin 3 (SIRT3) was -9.0 kcal/mol, and the binding effect between the two was stable. Such proposed mechanism was further corroborated by the SIRT3 siRNA transfection cell experiment, indicating that GA may potentially influence the mitochondrial calcium uniporter (MCU) expression by targeting SIRT3, thereby reducing mitochondrial Ca2+ content. Together, this study elucidates the essential role of GA in ameliorating liver lipid accumulation and improving liver health, highlighting that GA improves liver fibrosis and mitigates mitochondrial Ca2+ overload-mediated liver injury. Consequently, GA holds promise as a potential health-functional additive or therapeutic agent for liver fibrosis.

The specific miRNA regulation triggered by enzyme-treated soybean protein in response to well-known stressors, such as the prophylactic use of the antimicrobial oxytetracycline, remains unknown. Hence, this study aimed to evaluate the regulatory changes of hepatic miRNAs induced by oxytetracycline and enzyme-treated soybean protein in largemouth bass dietary formulations. The experiment was designed with three groups: the normal control (NC), the oxytetracycline exposure treatment group (OTC), and the pre-treatment with enzyme-treated soybean protein before oxytetracycline exposure group (ETSP). miRNA sequencing was employed to characterize the differences between these groups. In conclusion, the NC group exhibited up-regulation of 13 host miRNAs and down-regulation of 1 miRNA compared to the OTC group, whereas the ETSP group showed an increasing trend of 36 host miRNAs and a decreasing trend of 13 host miRNAs compared to the OTC group. Nine miRNAs were identified as prudential targets for enzyme-treated soy protein, protecting the largemouth bass liver from oxytetracycline. Furthermore, gene ontology analysis revealed nine key miRNAs that mediate signaling pathways with significant differences. The cellular lipid metabolic process was identified as the most important biological process, and the propanoate metabolism pathway was highlighted as significant. These results will facilitate further exploration of the mechanism by which enzyme-treated soy protein alleviates the effects of oxytetracycline on largemouth bass in water environments.