Haidong Yao

Gene Expression of Endoplasmic Reticulum Resident Selenoproteins Correlates with Apoptosis in Various Muscles of Se-Deficient Chicks

Dietary selenium (Se) deficiency causes muscular dystrophy in various species, but the molecular mechanism remains unclear. Our objectives were to investigate: 1) if dietary Se deficiency induced different amounts of oxidative stress, lipid peroxidation, and cell apoptosis in 3 skeletal muscles; and 2) if the distribution and expression of 4 endoplasmic reticulum (ER) resident selenoprotein genes (Sepn1, Selk, Sels, and Selt) were related to oxidative damages in these muscles. Two groups of day-old layer chicks (n = 60/group) were fed a corn-soy basal diet (33 μg Se/kg; produced in the Se-deficient area of Heilongjiang, China) or the diet supplemented with Se (as sodium selenite) at 0.15 mg/kg for 55 d. Dietary Se deficiency resulted in accelerated (P < 0.05) cell apoptosis that was associated with decreased glutathione peroxidase activity and elevated lipid peroxidation in these muscles. All these responses were stronger in the pectoral muscle than in the thigh and wing muscles (P < 0.05). Relative distribution of the 4 ER resident selenoprotein gene mRNA amounts and their responses to dietary Se deficiency were consistent with the resultant oxidative stress and cell apoptosis in the 3 muscles. Expression of Sepn1, Sels, and Selt in these muscles was correlated with (r > 0.72; P < 0.05) that of Sepsecs encoding a key enzyme for biosynthesis of selenocysteine (selenocysteinyl-tRNA synthase). In conclusion, the pectoral muscle demonstrated unique expression patterns of the ER resident selenoprotein genes and GPx activity, along with elevated susceptibility to oxidative cell death, compared with the other skeletal muscles. These features might help explain why it is a primary target of Se deficiency diseases in chicks.

Selenoprotein W serves as an antioxidant in chicken myoblasts

BACKGROUND Selenoprotein W (SelW) was thought to play an antioxidant role in mammals. Because chicken SelW has no cysteine (Cys) at the residue 37 (Cys37) that is required for the presumed antioxidant function in mammals, this study was conducted to determine whether chicken SelW possessed the same function. METHODS Small interfering RNAs (siRNAs) technology was applied to suppress the SelW expression in chicken embryonic myoblasts. Thereafter, these myoblasts were treated with different concentrations of H2O2 and assayed for cell viability, apoptosis rate, reactive oxygen species (ROS) status, and expression levels of apoptosis-related genes and proteins (Bax, Bcl-2, and caspase-3). RESULTS Silencing of the myoblast SelW gene decreased their cell viability, and increased their apoptosis rate and susceptibility to H2O2. While the knockout down of SelW up-regulated Bax and caspase-3 and down-regulated Bcl-2, the induced oxidative injuries were alleviated by treatment with a ROS scavenger, N-acetyl-l-cysteine (NAC). CONCLUSION Chicken SelW protected embryonic myoblasts against cell apoptosis mediated by endogenous and exogenous H2O2. GENERAL SIGNIFICANCE Chicken SelW possesses antioxidant function similar to the mammalian homologues despite the lack of Cys37 in the peptide.

Different responses of selenoproteins to the altered expression of selenoprotein W in chicken myoblasts

The aim of the present study was to examine the role of Selenoprotein W (Sepw1) in modulating the expression of other selenoproteins. In the present study, we silenced and overexpressed the expression of Sepw1 in chicken myoblasts and subsequently treated the myoblasts with a reactive oxygen species (ROS) scavenger, N-acetyl-L-cysteine (NAC), and H2O2. Thereafter, the levels of expression of 25 selenoproteins and the activities of certain antioxidative enzymes, glutathione peroxidase (Gpx), superoxide dismutase (SOD), and catalase (CAT) were analyzed. In addition, principal component analysis (PCA) was used to define the most important parameters that could be used as key factors. The results indicated that as a highly expressed selenoprotein (only lower than Gpx1, Selk, Sels and Sep15), Sepw1 could interact with H2O2 (P < 0.05) and influence the expression of some selenoproteins (Gpx3, Gpx4, Txnrd1, Selt, Selh, Sepp1, Sels and Sep15, P < 0.05) and the sensitivity of the cells to H2O2. Both the overexpression and silencing of Sepw1 influenced the mRNA levels of selenoproteins. However, the responses of selenoproteins to altered Sepw1 expression were different. The results indicated that Sepw1 played a special role in H2O2 metabolism and may modulate the expression of certain selenoproteins through the redox pathway. Therefore, these results indicate that Sepw1 is an essential antioxidative selenoprotein in chicken myoblasts.

Effects of atrazine and chlorpyrifos on cytochrome P450 in common carp liver.

Atrazine (ATR) and chlorpyrifos (CPF), widely used in agriculture, have resulted in a series of toxicological and environmental problems. We investigated the activities of the biotransformation enzymes ethoxyresorufin-O-deethylase (EROD) and pentoxyresorufin-O-deethylase (PROD), total cytochrome P450 (CYP), CYP1A mRNA level and level of tissue ATR, CPF, and their metabolites in the liver of common carp (Cyprinus carpio L.) after a 40-d exposure to CPF and ATR, alone or in combination, and a 20-d recovery. In the present study, juvenile common carp was exposed to ATR (at concentrations of 4.28, 42.8 and 428 μg L(-1)), CPF (1.16, 11.6 and 116 μg L(-1)), and ATR/CPF mixture (at concentrations of 1.13, 11.3 and 113 μg L(-1)). A general increasing trend for the activity of the biotransformation enzymes (EROD and PROD), CYP and CYP1A mRNA level was observed in the liver of common carp exposed to ATR, CPF and the ATR/CPF mixture. In addition, ATR, CPF, and their metabolites demonstrated a high accumulation in the liver. These results demonstrated that the CYP system in fish could be used as a biomarkers in evaluating the impact of ATR and CPF exposure on the common carp.

Effects of cold stress on mRNA expression of immunoglobulin and cytokine in the small intestine of broilers

To investigate the effects of cold stress on mRNA expression of immunoglobulin and cytokine in small intestine of broilers, eighty-four 15-day-old male chickens were randomly divided into 12 groups. There were 1 control (25°C) and 5 acute stress groups (under the temperature of 12 ± 1°C) for 1, 3, 6, 12 and 24h, 3 control (25°C) and 3 chronic cold stress groups (under the temperature of 12 ± 1°C) for 5, 10, and 20d. The mRNA expression levels of IL-2, IL-4, IL-7, IL-10, IL-17, IFN-γ, IgA, IgM, IgG, plgR, and TGF-β4 in duodenum, jejunum and ileum were detected by real-time PCR. The results showed that expression levels of IgM, IgA, IgG, plgR and IL-7 had an increased tendency in acute and chronic cold stress, especially plgR that was markedly increased in the duodenum than jejunum and ileum in the acute cold stress. In addition, the mRNA expression levels of IL-2, IFN-γ, IL-4, IL-17 and TGF-β4 had a first increased then decreased tendency in acute and chronic cold stress groups, however, expression levels of IL-4 were higher in the stress groups than control groups. The histopathological detect showed that issues in cold stress group was seriously injured. These results demonstrated that cold stress could cause the change of immune function in chicken intestinal.

Selenium Deficiency Inhibits the Conversion of Thyroidal Thyroxine (T4) to Triiodothyronine (T3) in Chicken Thyroids

Selenium (Se) influences the metabolism of thyroid hormones in mammals. However, the role of Se deficiency in the regulation of thyroid hormones in chickens is not well known. In the present study, we examined the levels of thyroidal triiodothyronine (T3), thyroidal thyroxine (T4), free triiodothyronine, free thyroxine (FT4), and thyroid-stimulating hormone in the serum and the mRNA expression levels of 25 selenoproteins in chicken thyroids. Then, principal component analysis (PCA) was performed to analyze the relationships between the selenoproteins. The results indicated that Se deficiency influenced the conversion of T4 to T3 and induced the accumulation of T4 and FT4. In addition, the mRNA expression levels of the selenoproteins were generally decreased by Se deficiency. The PCA showed that eight selenoproteins (deiodinase 1 (Dio1), Dio2, Dio3, thioredoxin reductase 2 (Txnrd2), selenoprotein i (Seli), selenoprotein u (Selu), glutathione peroxidase 1 (Gpx1), and Gpx2) have similar trends, which indicated that they may play similar roles in the metabolism of thyroid hormones. The results showed that Se deficiency inhibited the conversion of T4 to T3 and decreased the levels of the crucial metabolic enzymes of the thyroid hormones, Dio1, Dio2, and Dio3, in chickens. In addition, the decreased selenoproteins (Dio1, Dio2, Dio3, Txnrd2, Seli, Selu, Gpx1, and Gpx2) induced by Se deficiency may indirectly limit the conversion of T4 to T3 in chicken thyroids. The information presented in this study is helpful to understand the role of Se in the thyroid function of chickens.

Selenoprotein W redox-regulated Ca 2+ channels correlate with selenium deficiency-induced muscles Ca 2+ leak

Selenium (Se) deficiency induces Ca2+ leak and calcification in mammal skeletal muscles; however, the exact mechanism is still unclear. In the present study, both Se-deficient chicken muscle models and selenoprotein W (SelW) gene knockdown myoblast and embryo models were used to study the mechanism. The results showed that Se deficiency-induced typical muscular injuries accompanied with Ca2+ leak and oxidative stress (P < 0.05) injured the ultrastructure of the sarcoplasmic reticulum (SR) and mitochondria; decreased the levels of the Ca2+ channels, SERCA, SLC8A, CACNA1S, ORAI1, STIM1, TRPC1, and TRPC3 (P < 0.05); and increased the levels of Ca2+ channel PMCA (P < 0.05). Similarly, SelW knockdown also induced Ca2+ leak from the SR and cytoplasm; increased mitochondrial Ca2+ levels and oxidative stress; injured SR and mitochondrial ultrastructure; decreased levels of SLC8A, CACNA1S, ORA1, TRPC1, and TRPC3; and caused abnormal activities of Ca2+ channels in response to inhibitors in myoblasts and chicken embryos. Thus, both Se deficiency and SelW knockdown induced Ca2+ leak, oxidative stress, and Ca2+ channel reduction. In addition, Ca2+ levels and the expression of the Ca2+ channels, RyR1, SERCA, CACNA1S, TRPC1, and TRPC3 were recovered to normal levels by N-acetyl-L-cysteine (NAC) treatment compared with SelW knockdown cells. Thus, with regard to the decreased Ca2+ channels, SelW knockdown closely correlated Se deficiency with Ca2+ leak in muscles. The redox regulation role of SelW is crucial in Se deficiency-induced Ca2+ leak in muscles.

SelW regulates inflammation-related cytokines in response to H2O2 in Se-deficient chicken liver

Selenium (Se) deficiency-induced liver damage is related to oxidative stress, and the alternative transcription of cytokines has been linked to liver disease. Selenoprotein W (SelW) is known to play an antioxidant role in mammals, but chicken SelW does not possess a cysteine at residue 37 that is required for the presumed antioxidant function in mammals. The relationship between SelW and cytokines in H2O2-mediated chicken liver damage remains unclear. To investigate the protective role of SelW against H2O2 by influencing the expression of cytokine mRNAs, SelW was overexpressed or/and knocked down in cultured chicken liver cells. After the cells were exposed to H2O2, the levels of interleukin (IL)-1, IL-6, IL-8, IL-17, IL-4, IL-10 and IFN-γ were lower in the SelW over-expressing cells but higher in the SelW knockdown cells compared to the H2O2-only treatment group. Meanwhile, chicken were fed with a Se-deficient granulated diet containing 0.032 mg kg−1 Se for 15, 25, 35, 45 and 55 days, and the results showed that IL-1, IL-6, IL-8, IL-17, IL-4, IL-10 and IFN-γ in Se-deficient chicken liver and serum were dramatically higher than the normal control group. In conclusion, our findings suggested that SelW regulated inflammation-related cytokines during H2O2-dependent chicken liver damage.

The disruption of mitochondrial metabolism and ion homeostasis in chicken hearts exposed to manganese

Exposure to high levels of manganese (Mn) can result in cardiotoxicity in animals. However, little is known about the effect of excess Mn on poultry hearts. The aim of this study was to investigate the effect of dietary Mn on chicken cardiac injuries and the possible mechanisms of this process. In the present study, 400 fifty-day-old Hy-line brown cocks were randomly divided into four groups, and were fed either a commercial diet (containing 100mg/kg Mn) or a Mn-supplemented diet containing 600mg/kg, 900mg/kg, or 1800mg/kg Mn for 30, 60 or 90 days, respectively. Next, we examined several biomarkers of cardiac injury, including biochemical blood serum analyses, electrocardiogram assays, histological analyses, ultra-structural assays and apoptosis assays. To investigate the possible mechanisms of Mn-induced cardiotoxicity, we examined the effect of MnCl(2) on mitochondrial function and metal ion homeostasis. We found that subchronic MnCl(2) exposure induced damage in chicken hearts. Further investigations indicated that possible mechanisms for Mn-induced chicken cardiac injury included the disruption of mitochondrial metabolism and the alteration of ion homeostasis.

Pro- and anti-inflammatory cytokine expression in immune organs of the common carp exposed to atrazine and chlorpyrifos

Atrazine (ATR) and chlorpyrifos (CPF) are toxic and subject to long-term in vivo accumulation in different aquatic species throughout the world. The purpose of the present study was to examine the effect of ATR, CPF and combined ATR/CPF exposure on cytokines in the head kidney and spleen of common carp (Cyprinus carpio L.). The carp were sampled after a 40-d exposure to CPF and ATR, individually or in combination, followed by a 40-d recovery to measure the mRNA expression of IL-6fam (IL-6), IL-8, TNF-α, IL-10 and TGF-β1 (TGF-β) in the head kidney and spleen tissues. These results showed that the expression of cytokines IL-6, IL-8 and TNF-α in the head kidney and spleen was upregulated following ATR, CPF and mixed ATR/CPF exposure compared with the control group. The expression of IL-10 and TGF-β mRNA was significantly inhibited in both head kidney and spleen of carp exposed to ATR, CPF and the ATR/CPF mixture. The results suggested that long-term exposure of ATR, CPF and the ATR/CPF mixture in aquatic environments can induce the dysregulation of pro-/anti-inflammatory cytokine expression. The information regarding the effects of ATR and CPF on cytokine mRNA expression generated in this study will be important information for pesticides toxicology evaluation.