Selenomethionine alleviated Ochratoxin A induced pyroptosis and renal fibrotic factors expressions in MDCK cells.

Ochratoxin A (OTA) is universally known to induce nephrotoxicity via inducing oxidative stress and apoptosis, inhibiting protein synthesis and activating autophagy. Our previous studies have proved that OTA induces nephrotoxicity in vitro and in vivo by adjusting the NOD-like receptor protein 3 (NLRP3) inflammasome activation and caspase-1-dependent pyroptosis. Based on these findings, we further investigated the protective role of selenomethionine (SeMet) on OTA-caused nephrotoxicity using the Madin-Darby canine kidney (MDCK) epithelial cells as an in vitro model, proposing to offer a new way for remedying OTA-induced nephrotoxicity by nutritional manipulation. We measured the cell vitality, lactate dehydrogenase (LDH) activity and the expression of renal fibrotic genes, NLRP3 inflammasome and pyroptosis related genes. MTT and LDH results indicated that SeMet supplementation significantly mitigated 2.0 µg/ml OTA-induced cytotoxicity in MDCK cells (p < 0.05). Meanwhile, SeMet alleviated OTA induced increase of reactive oxygen species in MDCK cells. Then, the expressions of α-SMA, Vimentin, and TGF-ß were detected both in mRNA and protein levels. The results indicated 8 µM SeMet supplementation could significantly downregulate the expression of OTA-induced renal fibrosis-related genes (p < 0.05). In addition, the upregulation of OTA-induced NLRP3 inflammasome and pyroptosis downstream genes was also significantly inhibited by 8 µM of SeMet (p < 0.05). In summary, SeMet could alleviate OTA-induced renal fibrotic genes expression and reduce NLRP3-caspase-1-dependent pyroptosis. Therefore, SeMet supplementation may become an effective approach for preserving animals from renal injury exposed to OTA.

Ochratoxin A induces nephrotoxicity in vitro and in vivo via pyroptosis.

Ochratoxin A (OTA), a prevalent nephrotoxic mycotoxin contaminant in food and feedstuff, has been reported to induce renal injury. To disclose the nephrotoxicity of continuous administration of OTA and to investigate potential mechanisms related to pyroptosis, male C57BL/6 mice were intraperitoneally injected with 1.0 and 2.0 mg/kg B.W. OTA every other day for 14 days. At 2.0 mg/kg B.W. OTA administration significantly increased histological injury and renal fibrosis molecules (α-SMA, Vimentin, TGF-ß) and activated the NOD-like receptor protein 3 (NLRP3) inflammasome and induced pyroptosis compared with control. In the in vitro tests, Madin-Darby canine kidney (MDCK) epithelial cells were exposed to 0-4.0 µg/ml OTA for 24 h in serum-free medium. Data showed that OTA dose-dependently affected cell viability and significantly up-regulated renal fibrosis genes (α-SMA, Vimentin, TGF-ß). 2.0 µg/ml OTA significantly induced NLRP3 inflammasome activation and caspase-1-dependent pyroptosis, increasing the expression and secretion of pro-inflammatory cytokines (IL-6, TNF-α) and pyroptosis-related genes (GSDMD, IL-1ß, IL-18) in MDCK cells. These outcomes were significantly abrogated after inhibiting NLRP3 activation with inhibitor MCC950 and silencing NLRP3 with small interfering RNA (siRNA). Furthermore, knockdown of caspase-1 also ameliorated OTA-induced renal fibrosis via the inhibition of pyroptosis. Collectively, the chosen doses of OTA-triggered nephrotoxicity through NLRP3 inflammasome activation and caspase-1-dependent pyroptosis both in vitro and in vivo.

PCV2 replication promoted by oxidative stress is dependent on the regulation of autophagy on apoptosis.

Porcine circovirus type 2 (PCV2) is an economically important swine pathogen but some extra trigger factors are required for the development of PCV2-associated diseases. By evaluating cap protein expression, viral DNA copies and the number of infected cells, the present study further confirmed that oxidative stress can promote PCV2 replication. The results showed that oxidative stress induced autophagy in PCV2-infected PK15 cells. Blocking autophagy with inhibitor 3-methyladenine or ATG5-specific siRNA significantly inhibited oxidative stress-promoted PCV2 replication. Importantly, autophagy inhibition significantly increased apoptosis in oxidative stress-treated PK15 cells. Suppression of apoptosis by benzyloxycarbonyl-Val-Ala-Asp fluoromethylketone in conditions of autophagy inhibition restored PCV2 replication. Taken together, autophagy protected host cells against potential apoptosis and then contributed to PCV2 replication promotion caused by oxidative stress. Our findings can partly explain the pathogenic mechanism of PCV2 related to the oxidative stress-induced autophagy.

Nephrotoxicity instead of immunotoxicity of OTA is induced through DNMT1-dependent activation of JAK2/STAT3 signaling pathway by targeting SOCS3.

Ochratoxin A (OTA) is reported to induce nephrotoxicity and immunotoxicity in animals and humans. However, the underlying mechanism and the effects of OTA on DNA damage have not been reported until now. The present study aims to investigate OTA-induced cytotoxicity and DNA damage and the underlying mechanism in PK15 cells and PAMs. The results showed that OTA at 2.0-8.0 µg/mL for 24 h induced cytotoxicity and DNA damage in PK15 cells and PAMs as demonstrated by decreasing cell viabilities and mRNA levels of DNA repair genes (OGG1, NEIL1 and NEIL3), increasing LDH release, Annexin V staining cells, apoptotic nuclei and the accumulation of γ-H2AX foci. OTA at 2.0-8.0 µg/mL increased DNMT1 and SOCS3 mRNA expressions about 2-4 fold in PK15 cells or 1.3-2 fold in PAMs. OTA at 2.0-8.0 µg/mL increased DNMT1, SOCS3, JAK2 and STAT3 protein expressions in PK15 cells or PAMs. DNMT inhibitor (5-Aza-2-dc), promoted SOCS3 expression, inhibited JAK2 and STAT3 expression, alleviated cytotoxicity, apoptosis and DNA damage induced by OTA at 4.0 µg/mL in PK15 cells. While, in PAMs, 5-Aza-2-dc had no effects on SOCS3 expression induced by OTA at 4.0 µg/mL, but inhibited JAK2 and STAT3 expression, and alleviated cytotoxicity, apoptosis and DNA damage induced by OTA. JAK inhibitor (AG490) or STAT3-siRNA alleviated OTA-induced cytotoxicity and DNA damage in PK15 cells or PAMs. Taken together, nephrotoxicity instead of immunotoxicity of OTA is induced by targeting SOCS3 through DNMT1-mediated JAK2/STAT3 signaling pathway. These results provide a scientific and new explanation of the underlying mechanism of OTA-induced nephrotoxicity and immunotoxicity.

Regulation of expression and activity of selenoenzymes by different forms and concentrations of selenium in primary cultured chicken hepatocytes.

The expression and activity of selenoenzymes are regulated by Se. In the present study, the effects of different forms and concentrations of Se on the regulation of glutathione peroxidase (GPx) activity and phospholipid hydroperoxide GPx (GPx4) and type I deiodinase (D1) mRNA levels in chicken hepatocytes were evaluated. Primary cultured chicken hepatocyte monolayers derived from male White Leghorn chickens (aged 30-40 d) were incubated for 24 h with 0 (control), 0.5, 1, 1.5, 2, 3, 4 or 5 µmol/l of Se supplied as dl-selenomethionine (Se-Met), κ-selenocarrageenan (Se-Car) or sodium selenite (Na2SeO3). Compared with the control, Se significantly increased GPx activity in all the hepatocytes, but the activity was not increased in the hepatocytes treated with 5 µmol/l of Na2SeO3, with maximal effects being observed at 2 µmol/l of Se-Met or Se-Car and at 1.5 µmol/l of Na2SeO3, respectively. Significant decreases in GPx4 mRNA levels were observed in all the hepatocytes treated with Se (v. control). The D1 mRNA levels were significantly increased in all the groups treated with Se (v. control), with maximal effects being observed at 1.5 µmol/l of Se-Met and at 0.5 µmol/l of Se-Car or Na2SeO3, respectively. Se-Met at doses of 1.5-5 µmol/l had a greater effect on D1 mRNA than Se-Car and Na2SeO3 at equivalent doses. After resulting in a maximal effect, higher Se supplementation led to a dose-dependent reduction in GPx activity and D1 mRNA levels in all the hepatocytes treated with Se. These results suggest that in chicken hepatocytes, the regulations of GPx and D1 by different forms and concentrations of Se vary.

Regulation of taurine in OTA-induced apoptosis and autophagy.

Ochratoxin A (OTA), one of the most deleterious mycotoxins, could cause a variety of toxicological effects especially nephrotoxicity in animals and humans. Taurine, a wide-distributed cytoprotective amino acid, plays an important role as a basic factor for maintaining cellular integrity homeostasis. However, the potential effect of taurine in OTA-induced nephrotoxicity remains unknown. In the present study, we demonstrated that OTA treatment at 4.0-8.0 µM increased apoptosis in PK-15 cells as shown by increased the ratio of apoptosis and protein expression of Bax and cleaved-caspase-3, decreased protein expression of Bcl-2. Meantime, OTA treatment triggered autophagy, as indicated by markedly increased the protein expression of LC3-II and fluorescence intensity of GFP-LC3 dots. Taurine supplementation decreased OTA-induced cytotoxicity and attenuated apoptosis as shown by the decreased Annexin V/PI staining and the decreased expression of apoptosis-related proteins including Bax and caspase-3. Meanwhile, taurine attenuated OTA-induced autophagy by decreased the protein expression of LC3-II and fluorescence intensity of GFP-LC3 dots to maintain cellular homeostasis. In conclusion, taurine treatment could alleviate OTA-induced apoptosis and inhibit the triggered autophagy in PK-15 cells. Our study provides supportive data for the potential roles of taurine in reducing OTA-induced renal toxicity.

The aggravating effect of selenium deficiency on T-2 toxin-induced damage on primary cardiomyocyte results from a reduction of protective autophagy.

Selenium deficiency and T-2 toxin exposure may contribute to the development of Keshan disease characterized by congestive cardiomyopathy. The aim of this study was to explore the role of autophagy in the aggravation of selenium deficiency on T-2 toxin-induced damages on primary cardiomyocyte. Our present study demonstrated that 0.25-1 µM T-2 toxin damaged primary cardiomyocytes and selenium deficiency exacerbated T-2 toxin-induced damages by measuring the levels of MTT, lactate dehydrogenase and cleaved-caspase 3. T-2 toxin triggered autophagy in primary cardiomyocytes, as indicated by markedly increased expressions of LC3-Ⅱ and Beclin-1 mRNA levels. Rapamycin (autophagy agonist) treatment increased autophagy levels and decreased the cytotoxicity caused by T-2 toxin while 3-methyladenine (autophagy inhibitor) treatment reduced autophagy levels and enhanced the cytotoxicity of T-2 toxin, suggesting that autophagy protect primary cardiomyocytes from the cytotoxicity of T-2 toxin. Selenium deficiency lowered cytoprotective autophagy in the primary cardiomyocytes treated by T-2 toxin. It can be concluded that autophagy induced by T-2 toxin plays a role in protecting primary cardiomyocyte, but selenium deficiency decreases the protective autophagy and then exacerbate T-2 toxin-induced damages. Our finding may partly interpret the combination effects of selenium deficiency and T-2 toxin on the development of Keshan disease.

Zinc supplementation alleviates OTA-induced oxidative stress and apoptosis in MDCK cells by up-regulating metallothioneins.

AIMS: The present study was to investigate the protective effects of Zn supplementation in OTA-induced apoptosis of Madin-Darby canine kidney (MDCK) epithelial cells and explore the potential mechanisms. Aiming to provides a new insight into the treatment strategy of OTA-induced nephrotoxicity by nutritional regulation. MAIN METHODS: Initially, through MTT and LDH assay revealed that Zn supplementation significantly suppressed OTA-induced cytotoxicity in MDCK cells. Then, the production of reactive oxygen species (ROS) was detected by using a DCFH-DA assay. Annexin V-FITC/PI, Hoechst 33258 staining and Flow cytometry were used to detect the apoptosis. The expressions of apoptosis-related molecules were determined by RT-PCR, Western blotting. Interestingly, OTA treatment slightly increased the levels of Metallothionein-1 (MT-1) and Metallothionein-2 (MT-2) by using RT-PCR, Western blotting assay; while Zn supplementation further improved the increase of MT-1 and MT-2 induced by OTA. However, the inhibitive effects of Zn supplementation were significantly blocked after double knockdown of MT-1 and MT-2 by using Small Interfering RNA (siRNA) Transfection method. KEY FINDINGS: Our study provides supportive data for the potential roles of Zn in reducing OTA-induced oxidative stress and apoptosis in MDCK cells. SIGNIFICANCE: Zn is one of the key structural components of many proteins, which plays an important role in several physiological processes such as cell survival and apoptosis. This metal is expected to contribute to the conservative and adjuvant treatment of kidney disease and should therefore be investigated further.

Bush sophora root polysaccharide could help prevent aflatoxin B1-induced hepatotoxicity in the primary chicken hepatocytes.

The aim of this study was to evaluate the effects of bush sophora root polysaccharide (BSRPS) on the aflatoxin B1 (AFB1)-induced hepatotoxicity and to explore the underlying mechanisms. The primary chicken hepatocytes were used as the model in the present experiment. The results showed that AFB1 induced hepatotoxicity of chicken hepatocytes in a dose dependent manner as demonstrated by decreasing cell viability and increasing LDH activity, ALT and AST levels. AFB1 at 0.16 µM significantly increased the levels of hepatic cytochrome P450 1A5 (CYP450 1A5) mRNA and malondialdehyde (MDA) and decreased the activity and mRNA level of manganese superoxide dismutase(SOD2) and the glutathione peroxidases (GSH-Px) activity in the hepatocytes compared with the blank control. BSRPS at 8.93 µM, 17.86 µM, and 35.72 µM supplementation could significantly reverse the above-mentioned changes induced by AFB1, and 17.86 µM of BSRPS has the largest effects on protecting the AFB1-induced hepatocytes damage. Knock-down of SOD2 by SOD2-specific siRNA significantly eliminated the protective effects of BSRPS on AFB1-induced the increase of CYP450 1A5 mRNA levels and hepatotoxicity. These results suggested that the BSRPS has protective effects on AFB1-induced hepatotoxicity by down-regulating CYP450 1A5 mRNA level via up-regulating SOD2 expression in the primary chicken hepatocytes.

Lycium barbarum polysaccharides improve CCl4-induced liver fibrosis, inflammatory response and TLRs/NF-kB signaling pathway expression in wistar rats.

Lycium barbarum polysaccharides (LBPs) have multiple biological and pharmacological functions, including antioxidant, anti-inflammatory and anticancer activities. This research was conducted to evaluate whether LBPs could alleviate carbon tetrachloride (CCl4)-induced liver fibrosis and the underlying signaling pathway mechanism. Fifty male wistar rats were randomly allocated to five groups (n=10): control, CCl4 and CCl4 with 400, 800 or 1600mg/kg LBPs, respectively. Each wistar rat from each group was used for blood and tissue collections at the end of experiment. The results showed that CCl4 induced liver fibrosis as demonstrated by increasing histopathological damage, α-smooth muscle actin expression, aspartate transaminase activities, alkaline phosphatase activities and alanine aminotransferase activities. LBPs supplementation alleviated CCl4-induced liver fibrosis as demonstrated by reversing the above parameters. In addition, CCl4 treatment induced the oxidative injury, increased the mRNA levels of tumor necrosis factor-α, monocyte chemoattractant protein-1 and interleukin-1ß, and up-regulated the protein expressions of toll-like receptor 4 (TLR4), TLR2, myeloid differentiation factor 88, nuclear factor-kappa B (NF-kB) and p-p65. LBPs supplementation alleviated CCl4-induced oxidative injury, inflammatory response and TLRs/NF-kB signaling pathway expression by reversing the above some parameters. These results suggest that the alleviating effects of LBPs on CCl4-induced liver fibrosis in wistar rats may be through inhibiting the TLRs/NF-kB signaling pathway expression.

Dietary Supplementation of Selenium-Enriched Probiotics Enhances Meat Quality of Broiler Chickens (Gallus gallus domesticus) Raised Under High Ambient Temperature.

We investigated the effects of selenium-enriched probiotics (SP) on broiler meat quality under high ambient temperature and explore their underlying mechanisms. A total of 200 1-day-old male broiler chicks (Ross 308) were randomly allotted to four treatment groups, each with five replicates, in groups of ten birds. These birds were fed a corn-soybean basal diet (C), a basal diet plus probiotics supplementation (P), a basal diet plus Se supplementation in the form of sodium selenite (SS, 0.30 mg Se/kg), and a basal diet with the addition of selenium-enriched probiotics (SP, 0.30 mg Se/kg). The experiment lasted for 42 days. The birds were sacrificed by cervical dislocation, and the breast muscles were removed for further process. Our results showed that SP diet significantly increased (p < 0.05) the physical (pH, colors, water holding capacity, drip loss, shear force) and sensory characteristics of breast meat. All P, SS, and SP supplementation enhanced the antioxidant system by increasing (p < 0.05) the Se concentrations, glutathione (GSH) levels, activities of glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD) whereas decreasing (p < 0.05) malondialdehyde (MDA) levels, with SP being higher than P and SS. Moreover, SP diet significantly upregulated (p < 0.05) the mRNA levels of glutathione peroxidase genes (GPx1, GPx4) while it downregulated heat stress biomarkers such as heat shock protein (HSP) 70 as compared to C, P, and SS diets. In conclusion, our findings suggest that SP may function as beneficial nutritive supplement that is capable of improving meat quality during the summer season.

Effect of selenium source and level in hen's diet on tissue selenium deposition and egg selenium concentrations.

The present study was conducted in a 2 x 4 factorial arrangement in a randomized complete block (RCB) design to compare the effects of a commercial inorganic Se source (sodium selenite, SS) with a commercial organic Se source (Se-enriched yeast, SY) on tissue Se distribution and blood and whole-egg Se concentrations in laying hens. Both Se sources were added into the basal diet at 0, 0.2, 0.5, and 1.0 mg/kg of Se. Seven hundred 68 week old Rohman laying hens were fed with a basal diet containing 0.15 mg/kg DM (dry matter) of Se for 2 weeks, and then, they were allocated randomly into seven groups and were investigated for 28 days. Each group was replicated five times with five cages of four hens per cage in each replicate. During the experiment, two eggs per replicate from each treatment were collected every 7 days and blood was sampled on days 0, 14, and 28 for whole-egg and whole-blood Se analyses. At the end of the experiment, two hens per replicate from each treatment were slaughtered, and muscle (cardiac and breast muscles), liver, spleen, and kidney were sampled for the determination of Se concentrations. The results showed that the addition of Se from either source caused a significant increase in whole-egg and whole-blood Se concentrations (p < 0.01) and Se concentrations in liver, kidney, spleen, and cardiac and breast muscles (p < 0.05) of hens in comparison to the control. Both Se sources and Se levels significantly influenced (p < 0.01) Se concentrations in egg, blood, and the above-mentioned tissues. There was a more significant increase in the Se concentrations in egg (p < 0.01), spleen (p < 0.05), and breast muscle (p < 0.01) and a decrease (p < 0.01) in whole-blood and kidney from hens fed SY than those from hens fed SS. The order of Se distribution was liver > kidney > spleen > cardiac muscle > egg > blood > breast muscle, irrespective of the addition level or source. It was concluded that meat and eggs from hens fed commercial SY are a potential source of Se for humans.

Comparison of viability and infectivity of Cryptosporidium parvum oocysts stored in potassium dichromate solution and chlorinated tap water.

The present study was undertaken to compare the viability and infectivity of Cryptosporidium parvum oocysts that had been stored for 1, 4, 7, 10, 13, 16, 20, 25 and 30 months at 4 degrees C in 2.5% potassium dichromate (Cr) or chlorinated tap water, respectively. An excystation protocol was performed in vitro to evaluate viability. One hundred and eighty female BABL/c mice were used to evaluate the infectivity of oocysts by investigating the prepatent period of C. parvum infection, the quantity of oocysts excreted, and the number of parasites that colonized the villi of the ileum. The results showed that C. parvum oocysts preserved in Cr for 1-16 months or in water for 1-13 months were capable of excystation in vitro and infection of mice. The excystation rates of oocysts and the prepatent periods in mice infected by oocysts stored in Cr and water were not significantly different (p>0.05), and there was a strong correlation between prepatent period and duration of oocyst storage (Cr: R2=0.92; water: R2=0.98). There were no significant differences in oocyst shedding from feces or parasitism of the terminal ilea of mice by Cryptosporidia between the two storage media (p>0.05). In conclusion, C. parvum oocysts may be stored at 4 degrees C in water instead of Cr for the purposes of laboratory research. However, the presence of viable C. parvum oocysts in water is a severe challenge to the drinking water treatment industry.

Selenomethionine Alleviates AFB1-Induced Damage in Primary Chicken Hepatocytes by Inhibiting CYP450 1A5 Expression via Upregulated SelW Expression.

This study aims to evaluate the protective effects of selenomethionine (SeMet) on aflatoxin B1 (AFB1)-induced hepatotoxicity in primary chicken hepatocytes. Cell viability and lactic dehydrogenase activity assays revealed the dose dependence of AFB1 toxicity to chicken hepatocytes. AFB1 concentrations of >0.05 µg/mL significantly reduced glutathione and total superoxide dismutase levels and increased the malondialdehyde concentration and cytochrome P450 enzyme 1A5 (CYP450 1A5) mRNA levels (P < 0.05). AFB1, however, did not affect CYP450 3A37 mRNA levels. Supplementation with 2 µM SeMet protected against AFB1-induced changes and significantly increased selenoprotein W (SelW) mRNA levels (P < 0.05). Additionally, SelW knockdown attenuated the protective effect of SeMet on AFB1-induced damage and significantly increased the level of CYP450 1A5 expression (P < 0.05). Therefore, SeMet alleviates AFB1-induced damage in primary chicken hepatocytes by improving SelW expression, thus inhibiting CYP450 1A5 expression.

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.

Effects of selenium on proliferation, interleukin-2 production and selenoprotein mRNA expression of normal and dexamethasone-treated porcine splenocytes.

Porcine splenocytes were isolated in vitro, treated with different levels of dexamethasone (DEX), and stimulated by concanavalin A. Further, the normal (non-DEX-supplemented) or DEX-treated (0.01 µmol/L) splenocytes were incubated with 0, 0.5, 2, and 5 µmol/L Na2SeO3. The splenocyte proliferation, IL-2 production, intracellular glutathione peroxidase 1 (GPx1) mRNA level and activity and thioredoxin reductase 1 mRNA level were measured. The results showed that addition of 0.5 or 2 µmol/L Na2SeO3 significantly promoted normal and DEX-treated splenocyte proliferation, IL-2 production and GPx1 mRNA expression and activity (P < 0.05), respectively. The maximum effect was observed in DEX-treated splenocytes with 0.5 µmol/L Na2SeO3. Thus, our results show that the immune state modulation of Se is stronger in DEX-treated splenocytes than normal splenocytes. The mechanism underlying this effect may be increased in GPx1 expression induced by Se. Our results explain the controversy of varying reports on the immune state modulation induced by Se.

The intravenous glucose tolerance test in water buffalo.

OBJECTIVE: The response to intravenous glucose loading in the buffalo using the intravenous glucose tolerance test (IGTT) was investigated to provide a reference for intravenous glucose injection in buffaloes. METHOD: Twelve healthy, fasted, male swamp buffaloes were divided into three groups. Group I: six buffaloes were given 50% glucose at a dosage of 1 g/kg body weight via the jugular vein. Group II: three buffaloes received normal saline. Group III: three buffaloes were not injected. Blood samples were taken from the opposite vein at 60 and 10 min pre-injection (pre60 and pre10), and at 1, 5, 10, 30, 60, 120, 180, 240, 300, 360 and 420 min post-glucose injection (PGI). Plasma glucose was analyzed by the oxidase method. Insulin and glucagon were soon determined with a human radioimmunoassay kit. The insulin (pmol/l)/glucose (mmol/l) ratios (IGR) were also calculated for each sampling time. RESULTS: Mean plasma glucose, insulin and glucagon concentrations of buffaloes in groups II and III were similar at all the sampling times (p > 0.05) and the curves of the IGR for group II and group III were flat throughout. Group I Buffaloes showed an immediate 20 times increase in the mean plasma glucose concentration PGI, over the pre60 and pre10. The peak plasma insulin concentration occurred at 30 min PGI. The mean plasma glucose and insulin concentrations remained above pre-administration levels until 420 min PGI (p < 0.05). However, the mean plasma glucagon concentrations were different only at 1 and 5 min PGI sampling times. The curve of the IGR for group I showed an initial decrease at 1 min PGI, and fluctuated from 10.18 to 25.55 for the remainder of the sampling period. The correlation analysis showed that the mean plasma glucose concentration was positively correlated with insulin level (r = 0.73, p < 0.005), and significantly negatively correlated with mean plasma glucagon (r = -0.58, p < 0.05). The mean plasma insulin level did not show significant correlation with the glucagon (r = 0.06, p > 0.05). CONCLUSION: The hyperglycemia, high insulin, and protracted glucose and insulin curves, the initial decrease in the insulin/glucose ratio indicates that there was an unexpected glucose tolerance to acute intravenous glucose loading in water buffalo compared with other ruminants. The possibly suggested intravenous glucose load in buffaloes is about 5.09-8.28 mmol/l.

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.

Sam68 is a novel marker for aggressive neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is the most common solid extracranial tumor in children. However, the molecular mechanism and progression of NB is largely unknown, and unfortunately, the prognosis is poor. Src-associated in mitosis with a molecular weight of 68 kDa (Sam68) is associated with carcinogenesis and neurogenesis. The present study aimed to investigate the clinical and prognostic significance of Sam68 in NB. METHODS: The expression of Sam68 in immortalized normal epithelial cells, NB cell lines, and in four cases of paired NB tissue and adjacent normal tissue from the same patient was examined using Western blotting, reverse transcription-polymerase chain reaction (PCR) and real-time reverse transcription-PCR. The proliferation of NB cells was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Furthermore, Sam68 protein expression was analyzed in 90 NB cases characterized as clinicopathological using immunohistochemistry. Statistical analyses were applied to evaluate the diagnostic value and associations of Sam68 with clinical parameters. RESULTS: Western blotting and reverse transcription-PCR showed that the expression level of Sam68 was markedly higher in NB cell lines than in the immortalized normal epithelial cells at both messenger RNA and protein levels. The MTT assay revealed that Sam68 expression supported proliferation of NB cells. Sam68 expression levels were significantly up-regulated in tumor tissues in comparison to the matched adjacent normal tissues from the same patient. Sam68 protein level was positively correlated with clinical stage (P<0.001), tumor histology (P<0.001), and distant metastasis (P=0.029). Patients with higher Sam68 expression had shorter overall survival time, whereas those with lower tumor Sam68 expression had longer survival time. CONCLUSION: Our results suggest that Sam68 expression is associated with neuroblastoma progression and may represent a novel and valuable predictor for prognostic evaluation of neuroblastoma patients.

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.