SUMMARYBrassinosteroids (BRs) are essential for plant growth and development; however, whether and how they promote stomatal closure is not fully clear. In this study, we report that 24-epibrassinolide (EBR), a bioactive BR, induces stomatal closure in Arabidopsis (Arabidopsis thaliana) by triggering a signal transduction pathway including ethylene synthesis, the activation of Ga protein, and hydrogen peroxide (H 2 O 2 ) and nitric oxide (NO) production. EBR initiated a marked rise in ethylene, H 2 O 2 and NO levels, necessary for stomatal closure in the wild type. These effects were abolished in mutant bri1-301, and EBR failed to close the stomata of gpa1 mutants. Next, we found that both ethylene and Ga mediate the inductive effects of EBR on H 2 O 2 and NO production. EBR-triggered H 2 O 2 and NO accumulation were canceled in the etr1 and gpa1 mutants, but were strengthened in the eto1-1 mutant and the cGa line (constitutively overexpressing the G protein a-subunit AtGPA1). Exogenously applied H 2 O 2 or sodium nitroprusside (SNP) rescued the defects of etr1-3 and gpa1 or etr1 and gpa1 mutants in EBR-induced stomatal closure, whereas the stomata of eto1-1/AtrbohF and cGa/AtrbohF or eto1-1/nia1-2 and cGa/nia1-2 constructs had an analogous response to H 2 O 2 or SNP as those of AtrbohF or Nia1-2 mutants. Moreover, we provided evidence that Ga plays an important role in the responses of guard cells to ethylene. Ga activator CTX largely restored the lesion of the etr1-3 mutant, but ethylene precursor ACC failed to rescue the defects of gpa1 mutants in EBR-induced stomatal closure. Lastly, we demonstrated that Ga-activated H 2 O 2 production is required for NO synthesis. EBR failed to induce NO synthesis in mutant AtrbohF, but it led to H 2 O 2 production in mutant Nia1-2. Exogenously applied SNP rescued the defect of AtrbohF in EBR-induced stomatal closure, but H 2 O 2 did not reverse the lesion of EBR-induced stomatal closure in Nia1-2. Together, our results strongly suggest a signaling pathway in which EBR induces ethylene synthesis, thereby activating Ga, and then promotes AtrbohF-dependent H 2 O 2 production and subsequent Nia1-catalyzed NO accumulation, and finally closes stomata.

Two experiments were conducted to evaluate the efficacy of feeding an Escherichia coli (E. coli) derived phytase to pigs fed P deficient, corn-soybean meal diets. In Exp. 1, one hundred and twenty crossbred piglets (9.53±0.84 kg) were allocated to one of five treatments which consisted of four low P diets (0.61% Ca, 0.46% total P and 0.24% non-phytate P) supplemented with 0, 500, 1,000, or 20,000 FTU/kg E. coli phytase as well as a positive control formulated to be adequate in all nutrients (0.77% Ca, 0.62% total P and 0.42% non-phytate P). The treatments were applied to six pens with four pigs per pen for 28 days. In Exp. 2, ten crossbred pigs (19.66±1.16 kg) fitted with ileal T-cannula were used in a nutrient balance study. The pigs were assigned to treatments similar to those used in Exp. 1 in a doubly replicated 5×4 incomplete Latin square design (5 diets with 4 periods). Each period consisted of a 5-d adjustment period followed by a 3-d total collection of feces and urine and then a 2-d collection of ileal digesta. Supplementation with phytase linearly increased (p<0.05) weight gain, feed intake, feed efficiency, bone breaking strength and fat-free dry and ash bone weight. There were linear increases (p<0.01) in the apparent ileal digestibility (AID) of DM, GE, CP, Ca, total P, inositol hexaphosphate (IP6) and some AA with increasing dose of E. coli phytase. Pigs fed 20,000 FTU/kg had a greater (p<0.05) AID of IP6 (80% vs 59% or 64%, respectively) than pigs fed diets with 500 or 1,000 FTU/kg phytase. There were linear increases (p<0.05) in the total tract digestibility of Ca, total P, Na, K, Mg, and Zn as well as in the retention of Mg and Zn with increased phytase dose. The retention and utilization of Cu, and the total tract digestibility of CP and Cu quadratic increased (p<0.05) with increased phytase dose. In conclusion, supplementation of 500 FTU of phytase/kg and above effectively hydrolyzed phytate in low-P corn-soybean diets for pigs. In addition, a super dose of phytase (20,000 FTU/kg) hydrolyzed most of the IP6 and consequently further improved mineral use, protein utilization and performance.

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Two experiments were conducted to determine the digestible energy (DE) and metabolizable energy (ME) content of 19 rice bran samples and to develop prediction equations for DE and ME based on their chemical composition. The 19 rice bran samples came from different rice varieties, processing methods and regions. The basal diet was formulated using corn and soybean meal (74.43% corn and 22.91% soybean meal and 2.66% vitamins and minerals). The 19 experimental diets based on a mixture of corn, soybean meal and 29.2% of each source of rice bran, respectively. In Exp. 1, 108 growing barrows (32.1±4.2 kg) were allotted to 1 of 18 treatments according to a completely randomized design with 6 pigs per treatment. The treatment 1 was the control group which was fed with basal diet. The treatments 2 to 18 were fed with experimental diets. In Exp. 2, two additional rice bran samples were measured to verify the prediction equations developed in Exp. 1. A control diet and two rice bran diets were fed to 18 growing barrows (34.6±3.5 kg). The control and experimental diets formulations were the same as diets in Exp. 1. The results showed that the DE ranged from 14.48 to 16.85 (mean 15.84) MJ/kg of dry matter while the ME ranged from 12.49 to 15.84 (mean 14.31) MJ/kg of dry matter. The predicted values of DE and ME of the two additional samples in Exp. 2 were very close to the measured values.

Although the detrimental effects of diabetes mellitus/hyperglycemia have been observed in many liver disease models, the function and mechanism of hyperglycemia regulating liver-resident macrophages, Kupffer cells (KCs), in thioacetamide (TAA)-induced liver injury remain largely unknown. In this study, we evaluated the role of hyperglycemia in regulating NOD-like receptor family pyrin domain-containing 3 protein (NLRP3) inflammasome activation by inhibiting autophagy induction in KCs in the TAA-induced liver injury model. Type I diabetes/hyperglycemia was induced by streptozotocin treatment. Compared with the control group, hyperglycemic mice exhibited a significant increase in intrahepatic inflammation and liver injury. Enhanced NLRP3 inflammasome activation was detected in KCs from hyperglycemic mice, as shown by increased gene induction and protein levels of NLRP3, cleaved caspase-1, apoptosis-associated speck-like protein containing a caspase recruitment domain and interleukin-1b, compared with control mice. NLRP3 inhibition by its antagonist CY-09 effectively suppressed inflammasome activation in KCs and attenuated liver injury in hyperglycemic mice. Furthermore, inhibited autophagy activation was revealed by transmission electron microscope detection, decreased LC3B protein expression and p-62 protein degradation in KCs isolated from TAA-stressed hyperglycemic mice. Interestingly, inhibited 5 0 AMP-activated protein kinase (AMPK) but enhanced mammalian target of rapamycin (mTOR) activation was found in KCs from TAA-stressed hyperglycemic mice. AMPK activation by its agonist 5aminoimidazole-4-carboxamide ribonucleotide (AICAR) or mTOR signaling knockdown by small interfering RNA restored autophagy activation, and subsequently, inhibited NLRP3 inflammasome activation in KCs, leading to ultimately reduced TAA-induced liver injury in the hyperglycemic mice. Our findings demonstrated that hyperglycemia aggravated TAA-induced acute liver injury by promoting liver-resident macrophage NLRP3 inflammasome activation via inhibiting AMPK/mTOR-mediated autophagy. This study provided a novel target for prevention of toxin-induced acute liver injury under hyperglycemia.

BackgroundThis experiment was conducted to determine the nutritive value of corn from the north of China for growing pigs. The experiment examined corn variety (LS1, LS2, LS3 and LS4) grown in one location, drying method (sun dried and artificially dried) and different drying temperatures. Corn harvested at 20-25% moisture was dried to about 12% moisture by sun drying and artificially drying at 80, 100, or 120°C in a fluidized bed dryer. Ninety-six barrows (average BW of 33.4 ± 2.7 kg) were housed in individual metabolism crates to facilitate separate collection of feces and urine. A five-day collection period followed a seven-day diet acclimation period.ResultsThe results indicated that variety significantly influenced (P < 0.01) the 1,000 kernel weight of corn but not the bulk weight. Variety also influenced the available energy content (digestible energy of dry matter, P < 0.01; metabolisable energy of dry matter, P < 0.01) and digestibility of organic matter (P < 0.01), as well as dry matter (P < 0.01) and gross energy (GE) content (P < 0.02). The drying method of corn significantly influenced the 1,000 kernel weight (P < 0.01), bulk weight (P < 0.01) and digestibility of ether extract (EE) (P < 0.01). No effect of drying temperature on the digestibility of organic matter, dry matter (DM), crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF) and gross energy was observed, but gelatinization (P < 0.05) and test weight (P < 0.01) decreased with an increase in temperature.ConclusionsVariety has a significant impact on the nutritive value of corn for growing pigs, and greater attention needs to be paid to these influences in the assignment of the nutritive value of corn given to growing pigs.

This experiment was conducted to determine the optimal standardized ileal digestible lysine (SID Lys) level in diets fed to primiparous sows during lactation. A total of 150 (Landrace × Large White) crossbred gilts (weighing 211.1 ± 3.5 kg with a litter size of 11.1 ± 0.2) were fed lactation diets (3325 kcal metabolizable energy (ME)/kg) containing SID Lys levels of 0.76, 0.84, 0.94, 1.04 or 1.14%, through 28 days lactation. Gilts were allocated to treatments based on their body weight and backfat thickness 48 h after farrowing. Gilt body weight loss was significantly (P < 0.05) decreased by increasing dietary SID Lys levels. Fitted broken-line (P < 0.05) and quadratic plot (P < 0.05) analysis of body weight loss indicated that the optimal SID Lys for primiparous sows was 0.85 and 1.01%, respectively. Average daily feed intake (ADFI), weaning-to-estrus interval and subsequent conception rate were not affected by dietary SID Lys levels. Increasing dietary lysine had no effect on litter performances. Protein content in milk was increased by dietary SID Lys (P < 0.05). Dietary SID Lys tended to increase concentrations of serum insulin-like growth factor I (P = 0.066). These results of this experiment indicate that the optimal dietary SID Lys for lactating gilts was at least 0.85%, which approaches the recommendation of 0.84% that is estimated by the National Research Council (2012).

Although diabetes mellitus/hyperglycemia is a risk factor for acute liver injury, the underlying mechanism remains largely unknown. Liver-resident macrophages (Kupffer cells, KCs) and oxidative stress play critical roles in the pathogenesis of toxin-induced liver injury. Here, we evaluated the role of oxidative stress in regulating KC polarization against acetaminophen (APAP)-mediated acute liver injury in a streptozotocin-induced hyperglycemic murine model. Compared to the controls, hyperglycemic mice exhibited a significant increase in liver injury and intrahepatic inflammation. KCs obtained from hyperglycemic mice secreted higher levels of the proinflammatory factors, such as TNF-α and IL-6, lower levels of the anti-inflammatory factor IL-10. Furthermore, enhanced oxidative stress was revealed by increased levels of reactive oxygen species (ROS) in KCs from hyperglycemic mice post APAP treatment. In addition, ROS inhibitor NAC resulted in a significant decrease of ROS production in hyperglycemic KCs from mice posttreated with APAP. We also analyzed the role of hyperglycemia in macrophage M1/M2 polarization. Interestingly, we found that hyperglycemia promoted M1 polarization, but inhibited M2 polarization of KCs obtained from APAP-exposed livers, as evidenced by increased MCP-1 and inducible NO synthase (iNOS) gene induction but decreased Arg-1 and CD206 gene induction accompanied by increased STAT1 activation and decreased STAT6 activation. NAC restored Arg-1, CD206 gene induction, and STAT6 activation. To explore the mechanism how hyperglycemia regulates KCs polarization against APAP-induced acute liver injury, we examined the AMPK/PI3K/AKT signaling pathway and found decreased AMPK activation and increased AKT activation in liver and KCs from hyperglycemic mice post APAP treatment. AMPK activation by its agonist AICAR or PI3K inhibition by its antagonist LY294002 inhibited ROS production in KCs from hyperglycemic mice post APAP treatment and significantly attenuated APAP-induced liver injury in the hyperglycemic mice, compared to the control mice. Our results demonstrated that hyperglycemia exacerbated APAP-induced acute liver injury by promoting liver-resident macrophage proinflammatory response via AMPK/PI3K/AKT-mediated oxidative stress.