Silybin prevented avermectin-induced cardiotoxicity in carp by modulating oxidative stress, inflammation, endoplasmic reticulum stress, mitochondrial apoptosis, and autophagy.

Avermectin is one of the widely used anthelmintics in aquaculture and exhibits substantial toxicity to aquatic organisms. Silybin is extensively used for its anti-inflammatory, antioxidant and anti-apoptotic biological properties. Heart is essential for the survival of fish and plays a vital role in pumping blood oxygen and nutrients. Residual avermectin in water poses harm to carp. However, there is still insufficient research on whether silybin can mitigate the toxicity of avermectin to carp heart tissues. In this research, we established a model involving carp subjected to acute avermectin exposure and administered diets containing silybin to explore the potential protective effects of silybin against avermectin-induced cardiotoxicity. The results revealed that avermectin induced oxidative stress, inflammation, endoplasmic reticulum (ER) stress, mitochondrial pathway apoptosis and autophagy in the cardiac tissues of carp. Compared with the avermectin group, silybin significantly reduced ROS accumulation in cardiac tissues, restored antioxidant enzyme activity, inhibited mRNA transcript levels of pro-inflammatory-related factors, and attenuated ER stress, mitochondrial pathway apoptosis and autophagy. Protein-protein interaction (PPI) analysis demonstrated that silybin mitigated avermectin-induced cardiac oxidative stress, inflammation, ER stress, mitochondrial pathway apoptosis and autophagy. Silybin exerted anti-inflammatory effects through the Nuclear Factor kappa B (NF-κB) pathway, antioxidant effects through the Nuclear factor erythroid 2-related factor 2 (Nrf2) - Kelch-like ECH-associated protein 1 (Keap1) pathway, alleviated cardiac ER stress through the Glucose-regulated protein 78 (GRP78)/Activating Transcription Factor 6 (ATF6)/C/EBP homologous protein (CHOP) axis, suppressed apoptosis through the mitochondrial pathway, and inhibited excessive autophagy initiation through the PTEN-induced putative kinase 1 (PINK1)/Parkin RBR E3 ubiquitin protein ligase (PARKIN) signaling pathway. This study provided evidence supporting the protective effect of silybin against avermectin-induced cardiotoxicity in carp, highlighting its potential as a dietary additive to protect fish from adverse effects caused by avermectin exposure.

Quercetin attenuates avermectin-induced cardiac injury in carp through inflammation, oxidative stress, apoptosis and autophagy.

As an important antibiotic, avermectin (AVM) has been widely used in China, but its unreasonable application has caused serious harm to the water environment. In view of the various pharmacological effects of quercetin (QUE), such as anti-inflammatory and antioxidant, the scientific hypothesis that "QUE may cause carp poisoning by inhibiting AVM" was proposed in this study. However, its protective effect in AVM -induced heart damage has not been reported. QUE reduced the symptoms of AVM toxicity and decreased the levels of creatine kinase, lactate dehydrogenase, and creatine kinase in the serum of carp. By histological observation, QUE was found to significantly reduce cardiac fiber swelling in carp. A DHE fluorescence probe study showed that QUE was able to inhibit AVM -induced accumulation of reactive oxygen species (ROS) in carp myocardium. We found that QUE significantly increased the intracellular antioxidant enzymes CAT, T-AOC and GSH enzyme activity and reduced intracellular MDA content. In addition, QUE significantly increased il-10 and tgf-ß1 expression, and significantly down-regulated tnf-α, il-6, il-1ß and inos expression. Tunel assay showed that QUE attenuated AVM -induced apoptosis, significantly decreased the transcript levels of pro-apoptosis-related genes, and increased the expression of anti-apoptosis-related genes. We also detected the protein expression of LC3 in the AVM group and QUE + AVM group, and found that the expression of LC3 was significantly increased in both groups compared with the Control group, but after adding QUE, the expression of LC3 was significantly decreased compared with the AVM group. In addition, the transcript levels of p62 and atg5 were also detected by qPCR. QUE significantly increased the expression of p62 and decreased the expression of atg5, suggesting that QUE could attenuate AVM -induced cardiac autophagy in carp. This study will provide preliminary evidence of the principle of QUE attenuating AVM -induced myocardial injury in carp from four aspects, including oxidative stress, inflammatory response, apoptosis and autophagy, and provide a theoretical basis for its prevention and treatment.

Protective effect of quercetin on avermectin induced splenic toxicity in carp: Resistance to inflammatory response and oxidative damage.

Avermectin pollution is an important problem that cannot be ignored in aquatic system in recent years. It has brought great trouble to freshwater aquaculture, especially fishery aquaculture. Plant-derived quercetin has anti-inflammatory and antioxidant properties and is widely used as a dietary additive, but its protective effect on immune damage induced by avermectin in freshwater carp remains unclear. This study evaluated the role of dietary additive quercetin supplementation in chronic avermectin exposure of carp spleen. Sixty carp were divided into 4 groups (n = 15/ group), including control group, avermectin treatment group, quercetin treatment group, quercetin and avermectin co-treatment group. Carp were exposed to a 1/10 96 h LC50 dose of avermectin for 30 d and fed a carp diet containing 400 mg/kg quercetin twice a day (3% body weigh/ carp). The results showed that chronic avermectin exposure caused the loose parenchymal structure of carp spleen tissue and the increase of inflammatory cells, accompanied by increased transcription levels of pro-inflammatory il-1ß, il-6, tnf-α and decreased levels of anti-inflammatory factors il-10 and tgf-ß1, ROS accumulation in spleen tissue. MDA content increased and T-AOC, CAT and GSH levels decreased. Quercetin down-regulates the NF-κB pathway by inhibiting the expression of iNOS and activating p38 MAPK, blocking the transcription of inflammatory factors, and alleviating the inflammation of carp spleen caused by chronic avermectin exposure. In addition, quercetin inhibits the over-activation of Nrf2/Keap-1 signaling axis, blocks ROS accumulation, and restores the spleen REDOX homeostasis. In conclusion, quercetin, as a dietary additive for carp feed, can effectively improve the immune damage caused by avermectin pollution in aquatic environment, resist spleen inflammation and oxidative stress, and provide a theoretical basis for clinical development of freshwater carp feed.

Silybin protected from avermectin-induced carp (Cyprinus carpio) nephrotoxicity by regulating PPAR-γ-involved inflammation, oxidative stress, ferroptosis and autophagy.

Avermectin, a widely used deworming drug, poses a significant threat to fisheries. Silybin is recognized for its antioxidant and anti-inflammatory properties. The kidney, being crucial for fish survival, plays a vital role in maintaining ion balance, nitrogen metabolism, and hormone regulation. While residual avermectin in water could pose a risk to carp (Cyprinus carpio), it remains unclear whether silybin can alleviate the renal tissue toxicity induced by avermectin in this species. In current study, we developed a model of long-term exposure of carp to avermectin to investigate the potential protective effect of silybin against avermectin-induced nephrotoxicity. The results indicated that avermectin induced renal inflammation, oxidative stress, ferroptosis, and autophagy in carp. Silybin suppressed the mRNA transcript levels of pro-inflammatory factors, increased catalase (CAT) activity, reduced glutathione (GSH) activity, diminished reactive oxygen species (ROS) accumulation in renal tissues, and promoted the activation of the Nrf2-Keap1 signaling pathway. Furthermore, the transcript levels of ferroptosis-associated proteins, including gpx4 and slc7a11, were significantly reduced, while those of cox2, ftl, and ncoa4 were elevated. The transcript levels of autophagy-related genes, including p62 and atg5, were also regulated. Network pharmacological analysis revealed that silybin inhibited ROS accumulation and mitigated avermectin-induced renal inflammation, oxidative stress, ferroptosis, and autophagy in carp through the involvement of PPAR-γ. Silybin exerted its anti-inflammatory effect through the NF-κB pathway and antioxidant effect through the Nrf2-Keap1 pathway, induced renal cell iron efflux through the SLC7A11/GSH/GPX4, and suppressed autophagy initiation via the PI3K/AKT pathway. This study provides evidence of the protective effect of silybin against avermectin-induced nephrotoxicity in carp, highlighting its potential as a therapeutic agent to alleviate the adverse effects of avermectin exposure in fish.

Preparation, characterisation and in vitro anti-inflammatory activity of Baicalin microsponges.

Baicalin, a flavonoid extracted from traditional Chinese medicine, Scutellaria baicalensis has significant anti-inflammatory effects. Microsponges are drug delivery systems that improve drug stability and slow the release rate. The combination of baicalin and the microsponges produced a new and stable system for its delivery, resulting in a novel formulation of baicalin. Baicalin microsponges (BM) were prepared using the quasi-emulsion solvent diffusion method. Effects of the mass ratio of the polymer (ethylcellulose) to baicalin, the concentration of the emulsifier polyvinyl alcohol (PVA), the stirring speed on the encapsulation efficiency (EE), and yield of the microsponges were investigated by combining the one-factor test and Box-Behnken design (BBD). The preparation process was standardised using 2.61:1 mass ratio of ethyl cellulose to baicalin, 2.17% concentration of PVA, with stirring at 794 rpm. Optimised BM formulations were evaluated for the parameters of EE (54.06 ± 3.02)% and yield of (70.37 ± 2.41)%, transmission electron microscopy (TEM), and in vitro cell evaluation. Results of the in vitro anti-inflammatory assay showed that baicalin microsponges-pretreated-lipopolysaccharide (LPS)-induced RAW264.7, mouse macrophages showed reduced inflammatory response, similar to that seen in baicalin-treated macrophages.

Quercetin attenuates environmental Avermectin-induced ROS accumulation and alleviates gill damage in carp through activation of the Nrf2 pathway.

Avermectin (AVM) is one of the most often used insecticides which is toxic to aquatic organisms, and cause oxidative-induced damages to the fish respiratory organ, the "gills". To better understand the mechanism by which an antioxidant reduces AVM-induced gill damage, we investigated the effects of Quercetin (Que) on AVM induction of oxidative stress to inhibit damages to the gills using common carp as a model organism. The Que is a fruit and vegetable rich flavonoid with antioxidant activity. In this study, four groups were created: the Control group, the Que group (400 mg/kg), the AVM group (2.404 µg/L), and the Que plus AVM group. The analytical methods were pathological structure examination, qPCR, Reactive Oxygen Species (ROS) and Western blot. The results showed that Que alleviated AVM-induced oxidative stress, inflammatory damage and apoptosis in the carp gills by activating the Nrf2 pathway. The mechanism was that Que alleviated the accumulation of ROS, reduced the balance between oxidation and antioxidant disrupted by AVM exposure, lowered the content of lipid peroxidation produced malondialdehyde (MDA), and increased the content of antioxidant enzymes including glutathione (GSH) and catalase (CAT). Nrf2 pathway was activated. Meanwhile, Que inhibited gill apoptosis in carp by decreasing the levels of Bax, Cytochrome C, Caspase9, Cleaved-Caspase3 and reduced Bcl2. This has important implications for future studies on Que and AVM. New suggestions are provided to reduce the threat of aquatic environmental pollution.