Xinjie Xia

Development of a real‐time PCR method for Firmicutes and Bacteroidetes in faeces and its application to quantify intestinal population of obese and lean pigs

Aims:u2002 To investigate whether the relative abundance of the Bacteroidetes and Firmicutes divisions in pigs is different between obese and lean animals.

A novel rice calmodulin-like gene, OsMSR2, enhances drought and salt tolerance and increases ABA sensitivity in Arabidopsis

Many abiotic stimuli, such as drought and salt stresses, elicit changes in intracellular calcium levels that serve to convey information and activate adaptive responses. Ca2+ signals are perceived by different Ca2+ sensors, and calmodulin (CaM) is one of the best-characterized Ca2+ sensors in eukaryotes. Calmodulin-like (CML) proteins also exist in plants, but their functions at the physiological and molecular levels are largely unknown. In this report, we present data on OsMSR2 (Oryza sativa L. Multi-Stress-Responsive gene 2), a novel calmodulin-like protein gene isolated from rice Pei’ai 64S (Oryza sativa L.). Expression of OsMSR2 was strongly up-regulated by a wide spectrum of stresses, including cold, drought, and heat in different tissues at different developmental stages of rice, as revealed by both microarray and quantitative real-time RT-PCR analyses. Analysis of the recombinant OsMSR2 protein demonstrated its potential ability to bind Ca2+ in vitro. Expression of OsMSR2 conferred enhanced tolerance to high salt and drought in Arabidopsis (Arabidopsis thaliana) accompanied by altered expression of stress/ABA-responsive genes. Transgenic plants also exhibited hypersensitivity to ABA during the seed germination and post-germination stages. The results suggest that expression of OsMSR2 modulated salt and drought tolerance in Arabidopsis through ABA-mediated pathways.

Selenoprotein Gene Expression in Thyroid and Pituitary of Young Pigs Is Not Affected by Dietary Selenium Deficiency or Excess

Expression and function of selenoproteins in endocrine tissues remain unclear, largely due to limited sample availability. Pigs have a greater metabolic similarity and tissue size than rodents as a model of humans for that purpose. We conducted 2 experiments: 1) we cloned 5 novel porcine selenoprotein genes; and 2) we compared the effects of dietary selenium (Se) on mRNA levels of 12 selenoproteins, activities of 4 antioxidant enzymes, and Se concentrations in testis, thyroid, and pituitary with those in liver of pigs. In Experiment 1, porcine Gpx2, Sephs2, Sep15, Sepn1, and Sepp1 were cloned and demonstrated 84-94% of coding sequence homology to human genes. In Experiment 2, weanling male pigs (n = 30) were fed a Se-deficient (0.02 mg Se/kg) diet added with 0, 0.3, or 3.0 mg Se/kg as Se-enriched yeast for 8 wk. Although dietary Se resulted in dose-dependent increases (P < 0.05) in Se concentrations and GPX activities in all 4 tissues, it did not affect the mRNA levels of any selenoprotein gene in thyroid or pituitary. Testis mRNA levels of Txnrd1 and Sep15 were decreased (P < 0.05) by increasing dietary Se from 0.3 to 3.0 mg/kg. Comparatively, expressions of Gpx2, Gpx4, Dio3, and Sep15 were high in pituitary and Dio1, Sepp1, Sephs2, and Gpx1 were high in liver. In conclusion, the mRNA abundances of the 12 selenoprotein genes in thyroid and pituitary of young pigs were resistant to dietary Se deficiency or excess.

A high-selenium diet induces insulin resistance in gestating rats and their offspring.

Although supranutrition of selenium (Se) is considered a promising anti-cancer strategy, recent human studies have shown an intriguing association between high body Se status and diabetic risk. This study was done to determine if a prolonged high intake of dietary Se actually induced gestational diabetes in rat dams and insulin resistance in their offspring. Forty-five 67-day-old female Wistar rats (n=15/diet) were fed a Se-deficient (0.01 mg/kg) corn-soy basal diet (BD) or BD+Se (as Se-yeast) at 0.3 or 3.0mg/kg from 5 weeks before breeding to day 14 postpartum. Offspring (n=8/diet) of the 0.3 and 3.0mg Se/kg dams were fed with the same respective diet until age 112 days. Compared with the 0.3mg Se/kg diet, the 3.0mg/kg diet induced hyperinsulinemia (P<0.01), insulin resistance (P<0.01), and glucose intolerance (P<0.01) in the dams at late gestation and/or day 14 postpartum and in the offspring at age 112 days. These impairments concurred with decreased (P<0.05) mRNA and/or protein levels of six insulin signal proteins in liver and muscle of dams and/or pups. Dietary Se produced dose-dependent increases in Gpx1 mRNA or GPX1 activity in pancreas, liver, and erythrocytes of dams. The 3.0mg Se/kg diet decreased Selh (P<0.01), Sepp1 (P=0.06), and Sepw1 (P<0.01), but increased Sels (P<0.05) mRNA levels in the liver of the offspring, compared with the 0.3mg Se/kg diet. In conclusion, supranutrition of Se as a Se-enriched yeast in rats induced gestational diabetes and insulin resistance. Expression of six selenoprotein genes, in particular Gpx1, was linked to this metabolic disorder.

The Selenium Deficiency Disease Exudative Diathesis in Chicks Is Associated with Downregulation of Seven Common Selenoprotein Genes in Liver and Muscle

Fast-growing broiler chicks are susceptible to Se deficiency diseases including exudative diathesis (ED). Our objective was to determine if ED could be induced by feeding a current, practical diet and if the incidence was related to selenogenome expression in liver and muscle of chicks. Four groups of day-old broiler chicks (n = 60/group) were fed a corn-soy basal diet (BD; 14 μg Se/kg; produced in the Se-deficient area of Sichuan, China and not supplemented with Se or vitamin E), the BD and all-rac-α-tocopheryl acetate at 50 mg/kg and Se (as sodium selenite) at 0.3 mg/kg, or both of these nutrients for 6 wk. A high incidence of ED and mortality of chicks were induced by the BD. The incidences and mortality were completely prevented by supplemental dietary Se but were only partially decreased by supplemental α-tocopherol acetate. Dietary Se deficiency decreased (P < 0.05) mRNA levels of 7 common selenoprotein genes (Gpx1, Gpx4, Sepw1, Sepn1, Sepp1, Selo, and Selk) in muscle and liver. Whereas supplementing α-tocopherol acetate enhanced (P < 0.05) only the muscle Sepx1 mRNA level, it actually decreased (P < 0.05) hepatic Gpx1, Seli, Txnrd1, and Txnrd2 mRNA levels. In conclusion, dietary Se protected chicks from the Se deficiency disease ED, probably by upregulating selenoprotein genes coding for oxidation- and/or lesion-protective proteins. The protection by vitamin E might be mediated via selenoproteins not assayed in this study and/or Se-independent mechanisms. The inverse relationship between hepatic expression of 4 redox-related selenoprotein genes and vitamin E status revealed a novel interaction between Se and vitamin E in vivo.

Prolonged Dietary Selenium Deficiency or Excess Does Not Globally Affect Selenoprotein Gene Expression and/or Protein Production in Various Tissues of Pigs

We previously determined the effects of dietary selenium (Se) deficiency or excess on mRNA abundance of 12 selenoprotein genes in pig tissues. In this study, we determined the effect of dietary Se on mRNA levels of the remaining porcine selenoprotein genes along with protein production of 4 selenoproteins (Gpx1, Sepp1, Selh, and Sels) and body glucose homeostasis. Weanling male pigs (n = 24) were fed a Se-deficient (<0.02 mg Se/kg), basal diet supplemented with 0, 0.3, or 3.0 mg Se/kg as Se-enriched yeast (Angel Yeast) for 16 wk. Although mRNA abundance of the 13 selenoproteins in 10 tissues responded to dietary Se in 3 patterns, there was no common regulation for any given gene across all tissues or for any given tissue across all genes. Dietary Se affected (P < 0.05) 2, 3, 3, 5, 6, 7, 7, and 8 selenoprotein genes in muscle, hypothalamus, liver, kidney, heart, spleen, thyroid, and pituitary, respectively. Protein abundance of Gpx1, Sepp1, Selh, and Sels in 6 tissues was regulated (P < 0.05) by dietary Se concentrations in 3 ways. Compared with those fed 0.3 mg Se/kg, pigs fed 3.0 mg Se/kg became hyperinsulinemic (P < 0.05) and had lower (P < 0.05) tissue levels of serine/threonine protein kinase. In conclusion, dietary Se exerted no global regulation of gene transcripts or protein levels of individual selenoproteins across porcine tissues. Pigs may be a good model for studying mechanisms related to the potential prodiabetic risk of high-Se intake in humans.

Real-time PCR quantification of the predominant bacterial divisions in the distal gut of Meishan and Landrace pigs

Firmicutes and Bacteroidetes are closely related to body fat in humans and mice, which are the two dominant bacterial divisions of gut microbiota in mammals. Here real-time PCR analysis indicated that Meishan pigs had a 34% reduction in percentage Bacteroidetes (P = 0.008) and a significantly lower proportion of Bacteroides (P = 0.013) than Landrace pigs. The percentage of Bacteroidetes or Bacteroides had a negative correlation with body fat (R2 was 0.63 for Bacteroidetes and 0.57 for Bacteroides, P < 0.05). There was a trend that the percentage of Firmicutes in Meishan pigs was higher in numerical value than in Landrace pigs, although this difference was not statistically significant (P = 0.290) between the two breeds. These suggested that body fat correlated with the percentage of Bacteroidetes division of the gut microbiota in the common pig breeds, and the differences of gut microbial ecology in obese versus lean animals may be analogous.

Structure and evolution of a highly repetitive DNA sequence from Brassica napus.

A Hind III family of highly repetitive DNA sequences, canrep (canola repeat), was cloned from the nuclear DNA of canola (Brassica napus cv. Westar). The basic units of this family of repeats consists of 176 bp and are arranged in clusters of tandem direct repeats. Each canrep repeat is composed of three related subrepeats of ca. 60 bp. Each subrepeat contains two inverted repeats of about 23 bp and another unrelated sequence of about 12 bp. Based on the internal structure, a possible scheme for the evolution of canrep is proposed. At least two subfamilies of the canrep sequences are present in the genome, as revealed by sequence analyses. In situ hybridization showed that canrep sequences are mainly clustered at centromeric regions of chromosomes. Northern hybridizations indicate that there are no transcripts related to canrep in the total RNAs extracted from plant seedlings.

Enhanced water-holding capacity of meat was associated with increased Sepw1 gene expression in pigs fed selenium-enriched yeast

To study the effect of selenium-enriched yeast (SeY) level on selenoprotein genes expression and the relation between gene expression and antioxidant status and meat quality, 30 selenium (Se)-depleted pigs (7-week old, 10.30±0.68 kg) were randomly divided into 3 groups and fed a basal diet plus 0, 0.3 and 3.0 mg Se/kg as SeY for 8 weeks. Results showed that dietary SeY supplementation improved the antioxidant status in muscle. The increased levels of SeY decreased (P<0.05) the drip loss and the concentration of thiobarbituric acid reactive substances in the muscle and meat. However, increased dietary SeY intake quadratically increased (P<0.01) the mRNA level of Sepw1 gene among the 12 selenoprotein genes examined in muscle. Statistical analysis showed drip loss was negatively correlated with the mRNA level of Sepw1 gene. These suggested that the enhanced water-holding capacity of meat was associated with the increased expression of Sepw1 gene.

Porcine Serum Can Be Biofortified with Selenium to Inhibit Proliferation of Three Types of Human Cancer Cells

Our objectives were to determine if porcine serum could be enriched with selenium (Se) by feeding pigs with high concentrations of dietary Se and if the Se-biofortified serum inhibited proliferation of 3 types of human cancer cells. In Expt. 1, growing pigs (8 wk old, n = 3) were fed 0.02 or 3.0 mg Se/kg (as sodium selenite) for 16 wk and produced serum with 0.5 and 5.4 μmol/L Se, respectively. In Expt. 2, growing pigs (5 wk old, n = 6) were fed 0.3 or 1.0 mg Se/kg (as Se-enriched yeast) for 6 wk and produced serum with 2.6 and 6.2 μmol/L Se, respectively. After the Se-biofortified porcine sera were added at 16% in RPMI 1640 to treat NCI-H446, DU145, and HTC116 cells for 144 h, they decreased (P < 0.05) the viability of the 3 types of human cancer cells by promoting apoptosis, compared with their controls. This effect was replicated only by adding the appropriate amount of methylseleninic acid to the control serum and was mediated by a downregulation of 8 cell cycle arrest genes and an upregulation of 7 apoptotic genes. Along with 6 previously reported selenoprotein genes, selenoprotein T (Selt), selenoprotein M (Selm), selenoprotein H (Selh), selenoprotein K (Selk), and selenoprotein N (Sepn1) were revealed to be strongly associated with the cell death-related signaling induced by the Se-enriched porcine serum. In conclusion, porcine serum could be biofortified with Se to effectively inhibit the proliferation of 3 types of human cancer cells and the action synchronized with a matrix of coordinated functional expression of multiple selenoprotein genes.