Long-term copper exposure caused hepatocytes autophagy in broiler via miR-455-3p-OXSR1 axis.

Copper (Cu) is a common environmental pollutant which has been identified to cause toxic effects on animal bodies. MicroRNAs (miRNAs) are a type of non-coding RNAs involved in the regulation of various cellular activities including autophagy, but the potential regulatory mechanisms after excess Cu intake are still uncertain. Our previous study has prompted that Cu exposure reduced liver miR-455-3p levels. Herein, miR-455-3p was found to be an important molecule in the regulation of Cu-induced autophagy in vivo and in vitro. Histopathology observation of liver tissue indicated that Cu-induced severe hepatic damage including cellular swelling and vacuolization. Meanwhile, excessive Cu exposure not only heighten the mRNA and protein expression levels of Beclin1, Atg5, LC3Ⅰ and LC3Ⅱ, but also decreased miR-455-3p levels. In vitro experiment, Cu-induced autophagy can be attenuated by miR-455-3p overexpression. Additionally, oxidative stress-responsive 1 (OXSR1) was identified as a direct downstream target of miR-455-3p by dual luciferase reporter assays. Moreover, knockdown of OXSR1 can attenuate the autophagy induced by Cu treatment and the miR-455-3p inhibitor. Overall, the miR-455-3p-OXSR1 axis works as a regulator of autophagy under Cu stress, which provides a basis for further revealing the mechanism of chronic Cu poisoning.

Exposure to copper activates mitophagy and endoplasmic reticulum stress-mediated apoptosis in chicken (Gallus gallus) cerebrum.

A large amount of copper (Cu) used in production activities can lead to the enrichment of Cu in the environment, which can cause toxicity to animals. However, the toxicity mechanism of Cu on the cerebrum is still uncertain. Hence, a total of 240 chickens were separated into four groups in this study to reveal the potential connection between mitophagy and endoplasmic reticulum (ER) stress-mediated apoptosis in the chicken cerebrum in the case of excess Cu exposure. The cu exposure situation was simulated by diets containing various levels of copper (11 mg/kg, control group; 110 mg/kg, group I; 220 mg/kg, group II and 330 mg/kg, group III) for 49 days. The results of histology showed that vacuolar degeneration was observed in the treated groups, and the mitochondria swell and autophagosomes formation were found under excess Cu treatment. Additionally, the expression of mitophagy (PINK1, Parkin, LC3I, LC3II and p62) and ER stress (GRP78, PERK, ATF6, IRE1α, XBP1, CHOP, and JNK) indexes were significantly upregulated under excess Cu exposure. Furthermore, the mRNA and protein expression of Bcl-2 were decreased, while Bak1, Bax, Caspase12, and Caspase3 were increased compared to the control group. In summary, this study demonstrated that an overdose of Cu could induce mitophagy and ER stress-mediated apoptosis in the chicken cerebrum. These findings revealed an important potential connection between Cu toxicity and cerebrum damage, which provided a new insight into Cu neurotoxicity.

Effects of Long-Term Exposure to Copper on Mitochondria-Mediated Apoptosis in Pig Liver.

Copper (Cu) is listed as one of the main heavy metal pollutants, which poses potential health risks to humans. Excessive intake of Cu has shown toxic effects on the organs of many animals, and the liver is one of the most important organs to metabolize it. In this study, pigs, the mammal with similar metabolic characteristics to humans, were selected to assess the effects of long-term exposure to Cu on mitochondria-mediated apoptosis, which are of great significance for studying the toxicity of Cu to humans. Pigs were fed a diet with different contents of Cu (10, 125, and 250 mg/kg) for 80 days. Samples of blood and liver tissue were collected on days 40 and 80. Experimental results demonstrated that the accumulation of Cu in the liver was increased in a dose-dependent and time-dependent manner. Meanwhile, the curve of pig's body weight showed that a 125 mg/kg Cu diet promoted the growth of pigs during the first 40 days and then inhibited it from 40 to 80 days, while the 250 mg/kg Cu diet inhibited the growth of pigs during 80 days of feeding. Additionally, the genes and protein expression levels of Caspase-3, p53, Bax, Bak1, Bid, Bad, CytC, and Drp1 in the treatment group were higher than that in the control group, while Bcl-2, Bcl-xL, Opa1, Mfn1, and Mfn2 were decreased. In conclusion, these results indicated that long-term excessive intake of Cu could inhibit the growth of pigs and induced mitochondria-mediated apoptosis by breaking the mitochondrial dynamic balance. Synopsis: Long-term exposure to high doses of Cu could lead to mitochondrial dysfunction by breaking the mitochondrial dynamic balance, which ultimately induced mitochondria-mediated apoptosis in the liver of pigs. This might be closely related to the growth inhibition and liver damage in pigs.

Toxic effects of copper on the jejunum and colon of pigs: mechanisms related to gut barrier dysfunction and inflammation influenced by the gut microbiota.

Copper (Cu) is an essential trace mineral, but its excessive intake can lead to potentially toxic effects on host physiology. The mammalian intestine harbors various microorganisms that are associated with intestinal barrier function and inflammation. In this study, the influences of Cu on barrier function, microbiota, and its metabolites were examined in the jejunum and colon of pigs. Here, we identified that the physical and chemical barrier functions were impaired both in the jejunum and colon, as evidenced by the decreased expression of tight junction proteins (ZO-1, Occludin, Claudin-1, and JAM-1) and mucous secretion-related genes, positive rate of Muc2, and secretion of SIgA and SIgG. Additionally, inflammatory cytokines were overexpressed in the jejunum and colon. Furthermore, Cu might increase the abundances of Mycoplasma, Actinobacillus and unidentified_Enterobacteriaceae in the jejunum, which significantly affected pentose and glucoronate interconversions, histidine metabolism, folate biosynthesis, porphyrin metabolism, and purine metabolism. Meanwhile, the abundances of Lactobacillus and Methanobrevibacter were remarkably decreased and Streptococcus, unidentified_Enterobacteriaceae, and unidentified_Muribaculaceae were significantly increased in the colon, with an evident impact on glycerophospholipid metabolism, retinol metabolism, and steroid hormone biosynthesis. These findings revealed that excess Cu had significant effects on the microbiota and metabolites in the jejunum and colon, which were involved in intestinal barrier dysfunction and inflammation.

Copper induces mitochondria-mediated apoptosis via AMPK-mTOR pathway in hypothalamus of Pigs.

Copper (Cu), one of the heavy metals, is far beyond the carrying capacity of the environment with Cu mining, industrial wastewater discharging and the use of Cu-containing pesticides. Intaking excess Cu can cause toxic effects on liver, kidney, heart, but few studies report Cu toxicity on brain tissue. It is noteworthy that most toxicity tests are based on rodent models, but large mammals chosen as animal models has no reported. To explore the relationship of the Cu toxicity and mitochondria-mediated apoptosis on hypothalamus in pigs, the content of Cu, histomorphology, mitochondrial related indicators, apoptosis, and AMPK-mTOR signaling pathway were detected. Results showed that Cu could accumulate in hypothalamus and lead to mitochondrial dysfunction, evidenced by the decrease of ATP production, activities of respiratory chain complex I-IV, and mitochondrial respiratory function in Cu-treated groups. Additionally, the genes and proteins expression of Bax, Caspase-3, Cytc in treatment group were higher than control group. Furthermore, the protein level of p-AMPK was enhanced significantly and p-mTOR was declined, which manifested that AMPK-mTOR signaling pathway was activated in Cu-treated groups. In conclusion, this study illuminated that the accumulation of Cu could cause mitochondrial dysfunction, induce mitochondria-mediated apoptosis and activate AMPK-mTOR pathway in hypothalamus.

Metabolomics analysis reveals the effect of copper on autophagy in myocardia of pigs.

Among different synthetic compounds copper (Cu) is persistently and frequently used as growth promoter, antibacterial, antifungal and antiparasitic agent and has become common environmental pollutant. Therefore, this study explores the cardio-toxic effects of control group (10 mg/kg bw Cu) and treatment group (125 and 250 mg/kg bw Cu), and it association with process of autophagy and metabolomics in myocardium of pigs kept in three different experimental treatments for a period of 80 days. The results of serum biochemical parameters showed a significantly increase in creatinine kinase (CK), creatine kinase-MB (CK-MB), high density lipoprotein-cholesterol (HDL-C), low density lipoprotein-cholesterol (LDL-C) and aspartate aminotransferase (AST) in pigs exposed to 125 mg/kg bw and 250 mg/kg bw Cu. Meanwhile, the severe structural abnormalities in cardiomyocytes were found when exposed to 250 mg/kg Cu at day 80. In addition, the mRNA and proteins (Beclin1, ATG5 and LC3II) expression levels were significantly increased and p62 was significantly decreased in cardiomyocytes exposed to 250 mg/kg Cu at day 80 of the trial. Further, UPLC-QTOF/MS technique showed that 7 metabolites were up-regulated and 37 metabolites were down-regulated in cardiomyocytes after 250 mg/kg Cu treatment, with a principal impact on the metabolic pathways including glycerophospholipid metabolism, one carbon pool by folate, fatty acid elongation and fatty acid degradation, which were related to autophagy. Overall, our study identified the autophagy processes and metabolites in metabolic pathways in Cu-induced myocardium injury, which provided useful evidence of myocardium toxicity caused by Cu exposure via metabolomics and multiple bioanalytic methods.