Fang Gan

Selenium-Enriched Probiotics Improve Antioxidant Status, Immune Function, and Selenoprotein Gene Expression of Piglets Raised under High Ambient Temperature

This research was conducted to evaluate the effects of selenium-enriched probiotics (SP) on growth performance, antioxidant status, immune function, and selenoprotein gene expression of piglets under natural high ambient temperature in summer. Forty-eight crossbred weanling piglets randomly allocated to four groups were fed for 42 days ad libitum a basal diet without (Con, 0.16 mg Se/kg) and with supplementation of probiotics (P, 0.16 mg Se/kg), sodium selenite (SS, 0.46 mg Se/kg), and SP (0.46 mg Se/kg). From each group, three piglets were randomly selected for blood collection on days 0, 14, 28, and 42 and tissue collection on day 42. The SP improved growth performance of piglets. Both SS and SP increased blood glutathione peroxidase activity and tissue thioredoxin reductase 1 mRNA expression, with SP being higher than SS. All P, SS, and SP supplementation increased the superoxide dismutase activity (40.1, 53.0, and 64.5%), glutathione content (84.6, 104, and 165%), TCR-induced T lymphocyte proliferation (20.8, 26.4, and 50.0%), and IL-2 concentration (24.9, 27.2, and 46.2%) and decreased malondialdehyde content (25.1, 26.3, and 49.3%), respectively. The greatest effects of SP supplementation suggest that SP may serve as a better feed additive than P or SS for piglets under high-temperature environments.

Reactive oxygen species regulate the replication of porcine circovirus type 2 via NF-κB pathway

Intracellular redox state has been suggested to have various effects on the replication of different viruses within host cells. The aim of the present study was to investigate the influence of reactive oxygen species (ROS) on replication of porcine circovirus type 2 (PCV2), in PK15 cells. Following PCV2 infection there was a time-dependent increase in ROS. Antioxidant N-acetyl-l-cysteine treatment of cells resulted in lower ROS levels and lower PCV2 replication. In contrast, treatment by buthionine sulfoximine (BSO), a GSH synthesis inhibitor, resulted in elevation of ROS levels and increased PCV2 replication. Furthermore, inhibiting the activity of NF-κB, a redox-responsive transcription factor, suppressed BSO-mediated increase of PCV2 replication, indicating that increased PCV2 replication likely occurs via ROS activation of NF-κB. Taken together, our results indicate that the generation of ROS during PCV2 infection is involved in its replication and this progression is associated with the alteration in NF-κB activity induced by ROS.

Effects of selenium-enriched probiotics on heat shock protein mRNA levels in piglet under heat stress conditions.

The effects of selenium-enriched probiotics (SP) on tissue selenium (Se) deposition, glutathione peroxidase-1 (GPx1) activity and mRNA level, and heat shock protein (Hsp) mRNA levels of piglets under heat stress conditions were investigated. A total of 48 crossbred ([Landrace × Yorkshire] × Duroc) piglets were randomly divided into 4 groups and fed a basal diet (Con, 0.16 mg Se/kg) or basal diets with added probiotics (P, 0.16 mg Se/kg), sodium selenite (SS, 0.46 mg Se/kg), or SP (0.46 mg Se/kg), respectively, for 42 days. Three piglets were randomly selected from each group for blood sample collection at days 0, 14, 28, and 42 and for liver, kidney, and spleen sample collection at day 42. The results showed that P, SS, and SP could significantly down-regulate the average mRNA levels of Hsp70 (17.3, 23.7, and 40.1%) and Hsp27 (22.4, 24.4, and 44.7%) of the tissues, respectively (P < 0.05), whereas SS and SP could significantly elevate Se concentration, GPx1 activity and mRNA level (P < 0.05). The maximal effects of these parameters were observed in SP. It was concluded that SP is a feasible dietary supplementation of piglets under heat stress conditions during the summer season.

Selenium Promotes T-Cell Response to TCR-Stimulation and ConA, but Not PHA in Primary Porcine Splenocytes

There is controversy in the literature over whether the selenium (Se) influences cellular immune responses, and the mechanisms possibly underlying these effects are unclear. In this study, the effects of Se on T-cell proliferation and IL-2 production were studied in primary porcine splenocytes. Splenocytes were treated with different mitogens in the presence of 0.5–4 µmol/L sodium selenite. Se significantly promoted T-cell receptor (TCR) or concanavalin A (ConA)-induced T-cell proliferation and IL-2 production but failed to regulate T-cell response to phytohemagglutinin (PHA). In addition, Se significantly increased the levels of cytosolic glutathione peroxidase (GPx1) and thioredoxin reductase 1 (TR1) mRNA, the activity of GPx1 and the concentration of reduced glutathione (GSH) in the unstimulated, or activated splenocytes. These results indicated that Se improved the redox status in all splenocytes, including unstimulated, TCR, ConA and PHA -stimulated, but only TCR and ConA-induced T-cell activation was affected by the redox status. N-acetylcysteine (NAC), a pharmacological antioxidant, increased T-cell proliferation and IL-2 production by TCR and ConA stimulated splenocytes but had no effect on the response to PHA in primary porcine splenocytes confirming that PHA-induced T-cell activation is insensitive to the redox status. We conclude that Se promotes GPx1 and TR1 expression and increases antioxidative capacity in porcine splenocytes, which enhances TCR or ConA -induced T-cell activation but not PHA-induced T-cell activation. The different susceptibilities to Se between the TCR, ConA and PHA -induced T-cell activation may help to explain the controversy in the literature over whether or not Se boosts immune responses.

Astragalus polysaccharides inhibits PCV2 replication by inhibiting oxidative stress and blocking NF-κB pathway

Porcine circovirus type 2 (PCV2) is the primary causative agent of porcine circovirus-associated disease (PCVAD). Astragalus polysaccharide (APS), as one kind of biological macromolecule extracted from Astragalus, has antiviral activities. This study was undertaken to explore the effect of APS on PCV2 replication in vitro and the underlying mechanisms. Our results showed that adding APS before PCV2 infection decreased significantly PCV2 DNA copies, the number of infected cells, MDA level, ROS level and NF-κB activation in PK15 cells and increased significantly GSH contents and SOD activity compared to control without APS. Oxidative stress induced by BSO could eliminate the effect of PCV2 replication inhibition by APS. LPS, as a NF-κB activator, could attenuate the effect of PCV2 replication inhibition by APS. BAY 11-7082, as a NF-κB inhibitor, could increase the effect of PCV2 replication inhibition by APS. In conclusion, APS inhibits PCV2 replication by decreasing oxidative stress and the activation of NF-κB signaling pathway, which suggests that APS might be employed for the prevention of PCV2 infection.

Ochratoxin A promotes porcine circovirus type 2 replication in vitro and in vivo

Abstract Ochratoxin A (OTA), a worldwide mycotoxin found in food and feeds, is a potent nephrotoxin in animals and humans. Porcine circovirus-associated disease (PCVAD), including porcine dermatitis and nephropathy syndrome, is a worldwide swine disease. To date, little is known concerning the relationship between OTA and porcine circovirus type 2 (PCV2), the primary causative agent of PCVAD. The effects of OTA on PCV2 replication and their mechanisms were investigated in vitro and in vivo. The results in vitro showed that low doses of OTA significantly increased PCV2 DNA copies and the number of infected cells. Maximum effects were observed at 0.05μg/ml OTA. The results in vivo showed that PCV2 replication was significantly increased in serum and tissues of pigs fed 75μg/kg OTA compared with the control group and pigs fed 150μg/kg OTA. In addition, low doses of OTA significantly depleted reduced glutathione and mRNA expression of NF-E2-related factor 2 and γ-glutamylcysteine synthetase; increased reactive oxygen species, oxidants, and malondialdehyde; and induced p38 and ERK1/2 phosphorylation in PK15 cells. Adding N-acetyl-l-cysteine reversed the changes induced by OTA. Knockdown of p38 and ERK1/2 by their respective specific siRNAs or inhibition of p38 and ERK1/2 phosphorylation by their respective inhibitors (SB203580 and U0126) eliminated the increase in PCV2 replication induced by OTA. These data indicate that low doses of OTA promoted PCV2 replication in vitro and in vivo via the oxidative stress-mediated p38/ERK1/2 MAPK signaling pathway. This suggests that low doses of OTA are potentially harmful to animals, as they enhance virus replication, and partly explains why the morbidity and severity of PCVAD vary significantly in different pig farms.

Selenium Alleviates Aflatoxin B1-Induced Immune Toxicity through Improving Glutathione Peroxidase 1 and Selenoprotein S Expression in Primary Porcine Splenocytes

Selenium (Se) is generally known as an essential micronutrient and antioxidant for humans and animals. Aflatoxin B1 (AFB1) is a frequent contaminant of food and feed, causing immune toxicity and hepatotoxicity. Little has been done about the mechanisms of how Se protects against AFB1-induced immune toxicity. The aim of this present study is to investigate the protective effects of Se against AFB1 and the underlying mechanisms. The primary splenocytes isolated from healthy pigs were stimulated by anti-pig-CD3 monoclonal antibodies and treated by various concentrations of different Se forms and AFB1. The results showed that Se supplementation alleviated the immune toxicity of AFB1 in a dose-dependent manner, as demonstrated by increasing T-cell proliferation and interleukin-2 production. Addition of buthionine sulfoximine abrogated the protective effects of SeMet against AFB1. SeMet enhanced mRNA and protein expression of glutathione peroxidase 1 (GPx1), selenoprotein S (SelS), and thioredoxin reductase 1 without and with AFB1 treatments. Furthermore, knockdown of GPx1 and SelS by GPx1-specific siRNA and SelS-specific siRNA diminished the protective effects of SeMet against AFB1-induced immune toxicity. It is concluded that SeMet diminishes AFB1-induced immune toxicity through increasing antioxidant ability and improving GPx1 and SelS expression in splenocytes. This study suggests that organic selenium may become a promising supplementation to protect humans and animals against the decline in immunity caused by AFB1.

Selenium alleviates porcine nephrotoxicity of ochratoxin A by improving selenoenzyme expression in vitro.

Ochratoxin A (OTA), a mycotoxin, is a potent nephrotoxin in humans and animals. Selenium (Se) is an essential micronutrient for humans and animals, and plays a key role in antioxidant defense. To date, little is known about the effect of Se on OTA-induced nephrotoxicity. In this study, the protective effects of selenomethionine against OTA-induced nephrotoxicity were investigated using the porcine kidney 15 (PK15) cells as a model. The results showed that OTA induced nephrotoxicity in a dose-dependent manner. Se at 0.5, 1, 2 and 4 μM had significant protective effects against OTA-induced nephrotoxicity. Furthermore, selenomethionine enhanced the activity and mRNA and protein expression of glutathione peroxidase 1 (GPx1), mRNA expression of GPx4, and mRNA expression of thioredoxin reductase 1 in the presence and absence of OTA. Among them, promoting effect of selenomethionine on GPx1 was maximal. Knock-down of GPx1 by using a GPx1-specific siRNA eliminated the protective effects of selenomethionine against OTA-induced nephrotoxicity. The results suggest that selenomethionine alleviates OTA-induced nephrotoxicity by improving selenoenzyme expression in PK15 cells. Therefore, selenomethionine supplementation may be an attractive strategy for protecting humans and animals from the risk of kidney damage induced by OTA.

Interaction of porcine circovirus type 2 replication with intracellular redox status in vitro

Abstract Objectives Redox status influences replication of some viruses but its effect on porcine circovirus type 2 (PCV2), the primary causative agent of the emerging swine disease post-weaning multisystemic wasting syndrome is not known. The interaction of PCV2 replication with intracellular redox status in PK15 cells was examined in this study. Methods Intracellular glutathione (GSH) was measured spectrophotometrically by reaction with 5, 5′-dithiobis (2-nitrobenzoic acid). Total superoxide dismutase activity (SOD) was assayed by inhibition of oxyamine oxidation by the xanthine oxidase system. Malondialdehyde (MDA) was assayed spectrophotometrically using the thiobarbituric acid reaction. Both quantification of PCV2 DNA by real-time polymerase chain reaction and indirect immunofluorescence of PCV2-infected cells were used to evaluate the replication of PCV2. Results Both GSH and SOD decreased significantly at 48 hours after PCV2 infection, whereas MDA concentration increased significantly after 48 hour post-infection. Furthermore, PCV2 replication in PK15 cells was significantly impaired after the elevation of intracellular GSH through treatment with the antioxidant N-acetyl-l-cysteine (NAC), a precursor in GSH synthesis. In contrast, PCV2 replication in PK15 cells was enhanced after reduction of GSH levels through H2O2-mediated oxidation. In addition, NAC treatment blocked the increase of virus replication induced by H2O2. Conclusions This study suggests that PCV2 infection induces oxidative stress and that intracellular redox status influences PCV2 replication in PK15 cells.

Nephropathy and hepatopathy in weaned piglets provoked by natural ochratoxin A and involved mechanisms

Ochratoxin A (OTA) contamination is a worldwide problem in pig industry. The objectives of the present study were to investigate the toxicity of natural OTA in weaned piglets and to further explore the underlying mechanisms. Totally, 36 crossbred ([Landrace × Yorkshire] × Duroc) piglets were randomly divided into 3 groups (three replicates per group, 4 piglets per replicate), and fed a basal diet (Con group) and basal diets added with 0.4 mg (OTA-L group) or 0.8 mg OTA/kg (OTA-H group), respectively for 42 days. The results showed that growth performance was significantly decreased (P<0.05) in OTA added groups compared with Con group. OTA concentration was relatively high in serum and OTA concentration in kidney was higher than in liver, respectively. AST, creatinine and urea in serum of OTA added groups were significantly increased (P<0.05), while glucose, total protein, albumin and globulin in serum of OTA added groups were significantly decreased (P<0.05) compared with Con group. Degenerative changes were observed in the epithelial cells of proximal tubules and in hepatocytes of OTA added groups. Antioxidant capacities in blood of OTA added groups and in kidney of OTA-H group were significantly decreased (P<0.05) compared with Con group. The mRNA expressions of bcl-2 were up-regulated, mRNA expressions of bax were down-regulated and the ratio of bcl-2 and bax was increased in kidney and liver of OTA added groups compared with Con group. In conclusion, OTA could reduce antioxidant capacity and suppress apoptosis in tissues and cause degenerative changes in the epithelial cells in proximal tubules and hepatic cells, which may have a negative effect on the growth performance of piglets.