Curcumin alleviates arsenic-induced injury in duck skeletal muscle via regulating the PINK1/Parkin pathway and protecting mitochondrial function.

Arsenic is a well-known environmental pollutant due to its toxicity, which can do harm to animals and human. Curcumin is a polyphenolic compound derived from turmeric, commonly accepted to have antioxidant properties. However, whether curcumin can ameliorate the damage caused by arsenic trioxide (ATO) in duck skeletal muscle remains largely unknown. Therefore, the present study aims to investigate the potential molecular mechanism of curcumin against ATO-induced skeletal muscle injury. The results showed that treating with curcumin could attenuate body weight loss induced by ATO and reduced arsenic content accumulation in the skeletal muscle of duck. Curcumin was also able to alleviated the oxidative stress triggered by ATO, which was manifested by the increase of T-AOC and SOD, and MDA decrease. Moreover, we observed that curcumin could ease mitochondrial damage and vacuolate degeneration of nucleus. Our further investigation found that ATO disrupted normal mitochondrial fission/fusion (Drp1, OPA1, Mfn1/2) and restrained mitochondrial biogenesis (PGC-1α, Nrf1/2, TFAM), while curcumin could promote mitochondrial fusion and activated PGC-1α pathway. Furthermore, curcumin was found that it could not only reduce the mRNA and protein levels of mitophagy (PINK1, Parkin, LC3, p62) and pro-apoptotic genes (p53, Bax, Caspase-3, Cytc), but also increased the levels of anti-apoptotic genes (Bcl-2). In conclusion, curcumin was able to alleviate ATO-induced skeletal muscle damage by improving mitophagy and preserving mitochondrial function, which can serve as a novel strategy to take precautions against ATO toxicity.

Arsenic and antimony co-induced nephrotoxicity via autophagy and pyroptosis through ROS-mediated pathway in vivo and in vitro.

Arsenic (As) and antimony (Sb) are commonly accumulated environmental pollutants that often coexist in nature and cause serious widespread biological toxicity. To investigate the nephrotoxicity induced by As and Sb in detail, we explored the mechanism by which As and Sb cotreatment induced autophagy and pyroptosis in vivo and in vitro. In this study, mice were treated with 4 mg/kg arsenic trioxide (ATO) or/and 15 mg/kg antimony trichloride (SbCl3) by intragastric intubation for 60 days. TCMK-1 cells were treated with ATO (12.5 µM), SbCl3 (25 µM) or a combination of As and Sb for 24 h. The results of the in vivo experiment demonstrated that As or/and Sb exposure could induce histopathological changes in the kidneys, and increase the levels of biochemical indicators of nephrotoxicity. In addition, As and Sb can co-induce oxidative stress, which further activate autophagy and pyroptosis. In an in vitro experiment, As and/or Sb coexposure increased ROS generation and decreased MMP. Moreover, the results of related molecular experiments further confirmed that As and Sb coactivated autophagy and pyroptosis. In conclusion, our results indicated that As and Sb co-exposure could cause autophagy and pyroptosis via the ROS pathway, and these two metals might have a synergistic effect on nephrotoxicity.

Protective effects of curcumin on ATO-induced nephrotoxicity in ducks in relation to suppressed autophagy, apoptosis and dyslipidemia by regulating oxidative stress.

Arsenic trioxide (ATO) has been known as common environmental pollution, and is deemed to a threat to global public health. Curcumin (Cur) is a phytoconstituent, which has been demonstrated to have antioxidant effects. In the current experiment, we investigated the efficacy of Cur against ATO-induced kidney injury and explored the potential molecular mechanisms that have not yet been fully elucidated in ducks. The results showed that treatment with Cur attenuated ATO-induced body weight loss, reduced the content of ATO in the kidney, and improved ATO-induced kidney pathological damage. Cur also remarkably alleviated the ascent of ATO-induced MDA level and activated the Nrf2 pathway. Using the TEM, we found Cur relieved mitochondrial swelling, autolysosomes generating and nuclear damage. Simultaneously, Cur was found that it not only significantly reduced autophagy-related mRNA and protein levels (mTOR, LC3-Ⅰ, LC3-Ⅱ, Atg-5, Beclin1, Pink1 and Parkin) and but also decreased apoptosis-related mRNA and protein expression levels (cleaved caspase-3, Cytc, p53 and Bax). Furthermore, through nontargeted metabolomics analysis, we observed that lipid metabolism balance was disordered by ATO exposure, while Cur administration alleviated the disturbance of lipid metabolism. These results showed ATO could induce autophagy and apoptosis by overproducing ROS in the kidney of ducks, and Cur might relieve excessive autophagy, apoptosis and disturbance of lipid metabolism by regulating oxidative stress. Collectively, our findings explicate the potential therapeutic value of Cur as a new strategy to a variety of disorders caused by ATO exposure.

Arsenic (III) and/or Antimony (III) induced disruption of calcium homeostasis and endoplasmic reticulum stress resulting in apoptosis in mice heart.

Arsenic (As) and antimony (Sb) are known as an environmental contaminant with cardiotoxicity properties. The endoplasmic reticulum (ER) is the largest calcium reservoir in the cell, and its calcium homeostasis disorder plays a vital role in endoplasmic reticulum stress (ERS) and apoptosis. The objective of this study was to investigate whether As and Sb induced apoptosis via endoplasmic reticulum stress (ERS) linked to calcium homeostasis disturbance. In this study, thirty-two adult mice were gavage-fed daily with As2O3 (4 mg/kg), SbCl3 (15 mg/kg) and co-treat with SbCl3 (15 mg/kg) and As2O3 (4 mg/kg) daily for 60 days. It was observed that As or/and Sb caused histopathological lesions and ER expansion of the heart. Meanwhile, the gene expression of ER Ca2+ release channels (RyR2 and IP3R) and calmodulin-dependent protein kinase II (CaMKII) increased while the levels of mRNA and protein of ER Ca2+ uptake channel (SERCA2) downregulated significantly compared to the controls. Then, As or/and Sb induced ERS and triggered the ER apoptotic pathway by activating unfolded protein response (UPR)-associated genes ((PERK, ATF6, IRE1, XBP1, JNK, GRP78), and apoptosis-related genes (Caspase12, Caspase3, p53, CHOP). Above indicators in As + Sb group became more severe than that of As group and Sb group. Overall, our results proved that the cardiotoxicity caused by As or/and Sb might be concerning disturbing calcium homeostasis, which induced apoptosis through the ERS pathway.

Multi-omics reveals the protective effects of curcumin against AFB1-induced oxidative stress and inflammatory damage in duckling intestines.

Aflatoxin B1 (AFB1) is the most prevalent and toxic class of aflatoxins, which is considered a significant risk factor for food safety. Curcumin, a phytoconstituent with anti-inflammatory and antioxidant properties, has potential therapeutic value for intestinal inflammatory diseases. In this study, the duckling model susceptible to AFB1 was selected for toxicity testing, aiming to explore the effect of curcumin on AFB1 enterotoxicity and its possible mechanism of action. The results showed that curcumin promoted the growth and development of ducklings and mitigated the changes in morphology and permeability serological index (DAO and D-LA) after AFB1 exposure. Curcumin also mitigated AFB1-induced oxidative stress by activating the Nrf2 pathway, and ameliorated intestinal inflammation by inhibiting the NF-κB/IκB signaling pathway and boosting intestinal autophagy. In terms of gut flora and their metabolites, we found that curcumin supplementation significantly increased the intestinal flora's abundance index and diversity index compared to the AFB1 group, mitigating the decline in the abundance of Actinobacteria and the rise in that of harmful bacteria Clostridia. Furthermore, untargeted metabolomic analysis revealed that the protective effect of curcumin on the intestine was mainly through the regulation of AFB1-induced disorders of lipid metabolism, involving linoleic acid metabolism, α-linolenic acid metabolism, and glycerolipid metabolism. Overall, the enteroprotective effects of curcumin may be of significant value in the future for treating chronic AFB1 poisoning and also provide new therapeutic ideas for other mycotoxicosis.

Curcumin attenuates aflatoxin B1-induced ileum injury in ducks by inhibiting NLRP3 inflammasome and regulating TLR4/NF-κB signaling pathway.

Aflatoxin B1 (AFB1) is a widespread toxic contamination in feed for animals. The primary active component of turmeric, curcumin (Cur), is an antioxidant and an anti-inflammatory. However, it is yet unknown how AFB1 affects the intestinal epithelial barrier and whether Cur acts as a protective mechanism when exposed to AFB1. Here, we explored the mechanism of AFB1-induced intestinal injury from intestinal epithelial barrier, inflammation, pyroptosis, and intestinal flora, and evaluated the protective role of Cur. We found that AFB1 caused weight loss and intestinal morphological damage that is mainly characterized by shortened intestinal villi, deepened crypts, and damaged intestinal epithelium. Exposure to AFB1 decreased the expression of Claudin-1, MUC2, ZO-1, and Occludin and increased the expression of pyroptosis-related factors (NLRP3, GSDMD, Caspase-1, IL-1ß, and IL-18) and inflammation-related factors (TLR4, NF-κB, IκB, IFN-γ, and TNF-α). Furthermore, ileal gut microbiota was altered, and simultaneously, the Lactobacillus abundance was decreased. The gut microbiota interacts with a wide range of physiologic functions and disease development in the host through its metabolites, and disturbances in gut microbial metabolism can cause functional impairment of the ileum. Meanwhile, Cur can ameliorate histological ileum injuries and intestinal flora disturbance caused by AFB1. We found that Cur reversed the effects of AFB1 through modulating both NLRP3 inflammasome and the TLR4/NF-κB signaling pathway. In conclusion, AFB1 can induce inflammatory damage and pyroptosis in duck ileum, while Cur has obviously protective effects on all the above damages.

Curcumin antagonizes inflammation and autophagy induced by arsenic trioxide through immune protection in duck spleen.

Arsenic is a toxic heavy metal widely found in the natural environment and has adverse effects on the health of waterfowl and human. Curcumin (CUR), a natural pigment of the golden spice turmeric, exhibits excellent anti-tumor, anti-inflammatory and anti-oxidant activities. But the effects of CUR on duck spleen exposed to arsenic remain largely unknown. In this study, 75 ducks were divided randomly into Control, L-ATO, M-ATO, H-ATO and CUR + H-ATO groups to systematically analyze the underlying role of CUR. The results showed that arsenic trioxide (ATO) led to growth retardation of ducks, hyaline degeneration and sparse cell arrangement on their spleen. And in the ATO-exposed ducks, the levels of immunoglobulins (Ig; IgA, IgG, IgM) in the serum and the expression of autophagy-related genes (Atg5, P62, LC3I, LC3II, LC3II/I, Beclin-1) were significantly upregulated compared with the control ducks. Moreover, ATO also activated NF-κB signal pathway and upregulated the expression of pro-inflammatory cytokines (TNF-α, IFN-γ, IL-1ß, IL-2, IL-18). Meanwhile, application of CUR alleviated the ATO toxicity with the release of growth inhibition, and the reduced hyaline degeneration and distortion of the spleen capsule. CUR also suppressed ATO-induced NF-κB activation, pro-inflammatory cytokine addition and expression of autophagy-related genes. Overall, these results suggested that CUR might exert a protective effect against ATO-induced immunosuppression in ducks via anti-inflammation and autophagy restoring.

Curcumin activates the Nrf2 Pathway to alleviate AFB1-induced immunosuppression in the spleen of ducklings.

Ducklings is one of the most susceptible poultry to Aflatoxin B1 (AFB1) which widely existed in duckling products will also in turn affect human health. Curcumin (CUR) has significant effects on immune regulation and anti-oxidation. But whether CUR alleviates toxic effects on duckling spleen induced by AFB1 remains largely unknown. In this study we treated duckings with AFB1 and CUR for 21 days before harvesting serum and spleen tissue for analyses. The results showed that AFB1 damaged the spleen tissue of ducklings by activating the NF-κB signaling pathway. And the addition of CUR not only promoted the growth of ducklings, but also enhanced the immune function of the spleen and reduced the damage of AFB1 to the spleen tissue. At the same time, CUR activated the Nrf2 signaling pathway, upregulated the expression of related antioxidant enzymes, inhibited the NF-kB signaling pathway, and ultimately reducing the inflammation of the duckling spleen induced by AFB1. It has been suggested from these results that Nrf2 pathway might be a potential therapeutic target for CUR to treat AFB1-induced immunosuppression in ducklings.

Arsenic trioxide-induced autophagy affected the antioxidant capacity and apoptosis rate of chicken hepatocytes.

Arsenic has recently received widespread attention due to its high toxicological effects on multiple animals; however, the mechanism underlying this toxicity is unclear. We investigated the damaging effects of arsenic trioxide (ATO) on hepatocytes and the effects of regulating autophagy on the hepatocyte damage induced by ATO exposure. First, we investigated the effects of ATO exposure (0, 0.6, 1.2, 2.4, and 4.8 µM) on the biochemical function and autophagy of chicken hepatocytes. The findings showed that as the concentration of ATO increased, the lactate dehydrogenase (LDH) concentration increased, more autophagosomes were observed via transmission electron microscopy (TEM), and the gene and protein expression levels of P62, LC3Ⅱ, and Beclin1 increased. Adding N-acetyl-l-cystine (NAC, 1 mM) attenuated autophagy and the hepatocyte damage induced by ATO. Then, we used rapamycin (Rapa) and 3-methylpurine (3-MA) to regulate the autophagy induced by exposure to 4.8 µM ATO and observed changes in the antioxidant capacity and apoptosis rate of chicken hepatocytes. Induction of autophagy reduced ATO-induced hepatocyte apoptosis but caused no significant effect on oxidative stress in chicken hepatocytes. Inhibition of autophagy exacerbated ATO-induced hepatocyte oxidative stress and apoptosis. These findings demonstrate that autophagy plays an important role in ATO-induced cell damage.

Effects of Long-Term Exposure to Copper on the Keap1/Nrf2 Signaling Pathway and Msr-Related Redox Status in the Kidneys of Rats.

The objective of the present study was to examine the effects of long-term exposure on oxidative damage, Keap1/Nrf2 signaling pathway, and Msr-related redox status in the kidneys of rats. Therefore, in this experimental study, a total of 32 CD-1 rats were randomized into 4 groups and treated with 30-, 60-, and 120-mg/kg Cu for 24 weeks. Different serum biomarkers suggestive of renal functions, pathological changes, and oxidative stress were analyzed in kidney tissues. Moreover, the levels of the Keap1/Nrf2 signaling pathway and redox status-related gene mRNA and proteins were also detected. The results indicated that Cu exposure dramatically increased the contents of creatinine and carbamide. Furthermore, histopathological alterations and mitochondrial damage in kidneys of rats of different Cu-treated groups were obviously observed. In addition, Cu exposure markedly changed the levels of glutathione, catalase, and total antioxidant capacity, and upregulated the contents of protein carbonyl, nitric oxide, and malondialdehyde. Moreover, higher levels of Cu treatments significantly increased the expression of Keap1/Nrf2 signaling pathway and redox status-related genes (NQO1, SOD-1, TRX, MsrA, MsrB1, MsrB2, MsrB3). Simultaneously, the mRNA expression levels of Nrf2, HO-1, and CAT were upregulated in rats exposed to 30- and 60-mg/kg Cu, but downregulated in the 120-mg/kg Cu group compared with the control group. Moreover, the Keap1/Nrf2 signaling pathway and redox status-related protein expression levels (HO-1, SOD-1, TRX, MsrA, MsrB1, MsrB2) were significantly increased in treated rats. In summary, it is suggested that the Keap1/Nrf2 signaling pathway and activation of Msr prevent Cu-induced nephrotoxicity and attenuate oxidative damage.

The protective role of autophagy against arsenic trioxide-induced cytotoxicity and ROS-dependent pyroptosis in NCTC-1469 cells.

Arsenic trioxide (As2O3) is widely used in traditional Chinese medicine to treat tumors. This study investigated the effect of As(III) on pyroptosis in murine hepatocytes in vitro and how this relates to autophagy. NCTC1469-cells were treated with As(III) alone (6, 12 and 18 µM) or in combination with N-acetylcysteine (NAC,1 mM), 3-methyladenine (3-MA, 5 mM) or rapamycin (Rapa,100 nM) for 24 h. The results showed that As(III)-treatment reduced cell viability in a dose-dependent manner, but induced lactic dehydrogenase (LDH) activity. As(III)-treatment also resulted in increased intracellular reactive oxygen species (ROS) levels and decreased mitochondrial membrane potential (MMP), therefore promoting pyroptosis. Moreover, As(III)-treatment upregulated the expression of autophagy and pyroptosis-related genes (LC3-A, LC3-B, P62, Beclin-1, Atg5, Caspase-1, Gasdermin D, IL-18, IL-1ß) and downregulated the expression of m-TOR, NLRP3, ASC genes. Meanwhile the accumulation of light chain 3-B/A (LC3B/LC3A), autophagy-related gene 5 (Atg-5), Bcl-2-interacting protein (Beclin-1), Caspase-1, Gasdermin D, interleukin-1ß (IL-1ß), IL-18 and poptosis-associated speck-like protein (ASC) proteins were upregulated while nucleotide binding and oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) was downregulated in all As(III)-treatment groups. Furthermore, the inhibition of autophagy by 3-MA aggravated AsIII-induced pyroptosis and cytotoxicity. However, NAC or Rapa markedly alleviated the abovementioned phenomenon under As(III) stress. In addition, we speculate that the protective mechanism of NAC on As(III)-induced pyroptosis in hepatocytes mainly include the elimination of ROS because of the chelation of As(III) in the culture medium. In conclusion, these results provide new insight into the mechanisms underlying AsIII-induced cytotoxicity and pyroptosis in hepatocytes in vitro.

Arsenic exposure induces intestinal barrier damage and consequent activation of gut-liver axis leading to inflammation and pyroptosis of liver in ducks.

Arsenic is an important hazardous metalloid commonly found in polluted soil, rivers and groundwater. However, few studies exist regarding the effect of arsenic trioxide (ATO) on the gut-liver axis and consequent hepatotoxicity in waterfowl. Here, we investigated the influence of ATO on duck intestines and livers, and explored the role of the gut-liver axis in ATO-induced hepatotoxicity and intestinal toxicity. Our results demonstrated that ATO-exposure induced intestinal damage, liver inflammatory cell infiltration and vesicle steatosis. Additionally, the intestinal microbiota community in ATO-exposed ducks displayed significantly decreased α-diversity and an altered bacterial composition. Moreover, ATO-exposure markedly reduced the expression of intestinal barrier-related proteins (Claudin-1, MUC2, ZO-1 and Occludin), resulting in increased intestinal permeability and elevated lipopolysaccharide levels. Simultaneously, ATO-exposure also upregulated pyroptosis-related index levels in the liver and jejunum, and increased pro-inflammatory cytokine production (IFN-γ, TNF-α, IL-18, and IL-1ß). Our further mechanistic studies showed that ATO-induced liver and jejunum inflammation were provoked by the activation of the LPS/TLR4/NF-κB signaling pathway and NLRP3 inflammasome. In summary, these results manifested that ATO exposure can cause liver and jejunal inflammation and pyroptosis, and the indirect gut-liver axis pathway may play an essential role in the potential mechanism of ATO-induced hepatotoxicity.

Arsenic or/and antimony induced mitophagy and apoptosis associated with metabolic abnormalities and oxidative stress in the liver of mice.

Arsenic and antimony are coexisting cumulative environmental pollutants that cause severe and extensive biological toxicity. However, their interactions and toxic mechanisms in the liver remain to be fully elucidated. In this study, a total of sixty 4-week-old mice were divided into four groups and treated with 4 mg/kg arsenic trioxide (ATO) or/and 15 mg/kg antimony (Sb) for 60 days. The results demonstrated that biochemical indicators of hepatotoxicity (ALT, AST, ALP) were upregulated in all treated groups. Additionally, the oxidative burden of the liver was increased in the cotreated groups compared with the individual toxicant-treated groups. Meanwhile, mitochondrial injury, autophagosomes, hepatic-congestion and karyopyknosis were obviously observed in cotreated groups. Additionally, coupled with serum biochemical index (TG, TC), histopathology examination and metabolomics results, we found that cotreatment with ATO and Sb resulted in lipid metabolism disorder and steatosis of liver tissues. Our further investigation found that the levels of pro-apoptotic (Caspase-3, Caspase-9, Bax, P53, Cytc) and mitophagy (LC3-B, P62, PINK1, Parkin) indexes in the cotreated groups were markedly increased, whereas the levels of anti-apoptosis index (Bcl-2) were decreased. Collectively, these results show that co-exposure to ATO and Sb can cause abnormal liver energy metabolism and oxidative stress. Moreover, mitophagy and apoptosis play important roles in the mechanisms of arsenic/antimony cytotoxicity to mouse livers.

Evaluation of toxic effects induced by arsenic trioxide or/and antimony on autophagy and apoptosis in testis of adult mice.

Arsenic trioxide (ATO) and antimony (Sb) are well-known ubiquitous environmental contaminants and cause unpromising male reproductive effects in target and non-target exposed organisms. The main objective of this study was to investigate the effects of ATO or/and Sb on process of autophagy, apoptosis, and reproductive organ in adult mice. For this reason, a total of 32 adult mice were randomly divided into different groups like control group, ATO-treated group, Sb-treated group, and combined group. The duration of current experimental trial was 2 months. Various adverse effects of ATO or/and Sb on sperm parameters, oxidative stress, autophagy, and apoptosis were determined in testis of mice. Results indicated that parameters of sperm quality for organ coefficient, sperm count, ratio of sperm survival, testosterone level, and germ cells were significantly decreased, while malformation rate and vacuolization significantly increased in mice exposed to different treatments. Furthermore, the status of antioxidant index of T-AOC, SOD, and MsrB1 levels was reduced, while MDA increased significantly in ATO + Sb group. Results on TEM investigation determined that the autophagosomes, autolysosome, nuclear pyknosis, and chromatin condensation were prominent ailments, and the levels of autophagy and pro-apoptosis indictors including Beclin1, Atg-5, LC3B/LC3A, caspase-8, cytc, cleaved caspase-3, p53, and Bax were up-regulated in treated group, while the content of an anti-apoptosis maker (Bcl-2) was down-regulated. In conclusion, the results of our experiment suggested that abnormal process of autophagy and apoptosis was triggered by arsenic and antimony, and intensity of toxic effects increased in combined treatments of ATO and Sb.