Wence Wang

Molecular cloning, tissue distribution and ontogenetic expression of the amino acid transporter b0,+ cDNA in the small intestine of Tibetan suckling piglets

The small intestine is the main absorption place of peptides and free amino acids in mammals. The amino acid transporter system b(0,+) mediates apical uptake of basic amino acids, especially lysine, arginine and cysteine. The aim of the current study was to clone Tibetan porcine amino acid transporter b(0,+)AT (SLC7A9) for comparing the sequences of Tibetan and common (Sus scrofa) pigs, and investigating the tissue distribution and ontogenetic expression in the small intestine of Tibetan suckling piglets. The Tibetan porcine SLC7A9 gene was first cloned from the porcine small intestine and found to encode the amino acid transporter b(0,+)AT. The entire open reading frame (ORF) of the SLC7A9 is 1464 bp and codes for 487 amino acid residues, with a higher degree of sequence similarity with common pig (99.59%) and horse counterparts (91.2%) than with monkey (89.5%) or human (88.7%). The deduced protein has 12 putative transmembrane domains. In this study, SLC7A9 mRNA was detected in brain, kidney, duodenum, jejunum, ileum, heart, liver, lung and muscle from Tibetan pigs at 7 and 21 days by PCR. We also investigated the age-dependent expression of b(0,+)AT in Tibetan suckling piglets in duodenum, anterior jejunum, posterior jejunum, ileum and kidney from day 1 to 35. The abundance of SLC7A9 mRNA in duodenum and jejunum was highest and lowest, respectively. Expression patterns were similar in duodenum and anterior jejunum, where the mRNA level was decreased before the suckling period and increased until day 35. Posterior jejunum expression was increasing steadily with age, except on day 7. The ileum has the highest expression at day 14 and became steady after day 28. The mRNA abundance in the kidney is opposite to duodenum, increasing until day 14 and reducing thereafter. Our results showed the pattern of b(0,+)AT expressed in small intestine of Tibetan pig and lay the foundation for in depth investigations of the regulation of b(0,+)AT in vivo.

Effects of Dietary Arginine and Glutamine on Alleviating the Impairment Induced by Deoxynivalenol Stress and Immune Relevant Cytokines in Growing Pigs

Deoxynivalenol (DON) is a mycotoxin that reduces feed intake and animal performance, especially in swine. Arginine and glutamine play important roles in swine nutrition. The objective of this study was to determine the effects of dietary supplementation with arginine and glutamine on both the impairment induced by DON stress and immune relevant cytokines in growing pigs. A total of forty 60-d-old healthy growing pigs with a mean body weight of 16.28±1.54 kg were randomly divided into 5 groups, and assigned to 3 amino acid treatments fed 1.0% arginine (Arg), 1.0% glutamine (Gln) and 0.5% Arg+0.5% Gln, respectively, plus a toxin control and a non-toxin control. Pigs in the 3 amino acid treatments were fed the corresponding amino acids, and those in non-toxin control and toxin control were fed commercial diet with 1.64% Alanine as isonitrogenous control for 7 days. The toxin control and amino acid treatments were then challenged by feeding DON-contaminated diet with a final DON concentration of 6 mg/kg of diet for 21 days. No significant differences were observed between toxin control and the amino acid groups with regard to the average daily gain (ADG), although the values for average daily feed intake (ADFI) in the amino acid groups were significantly higher than that in toxin control (P<0.01). The relative liver weight in toxin control was significantly greater than those in non-toxin control, arginine and Arg+Glu groups (P<0.01), but there were no significant differences in other organs. With regard to serum biochemistry, the values of BUN, ALP, ALT and AST in the amino acid groups were lower than those in toxin control. IGF1, GH and SOD in the amino acid groups were significantly higher than those in toxin control (P<0.01). The IL-2 and TNFα values in the amino acid groups were similar to those in non-toxin control, and significantly lower than those in toxin control (P<0.01). These results showed the effects of dietary supplementation with arginine and glutamine on alleviating the impairment induced by DON stress and immune relevant cytokines in growing pigs.

Chemerin regulates proliferation and differentiation of myoblast cells via ERK1/2 and mTOR signaling pathways.

Obesity in human is an alarming major public health crisis worldwide and insulin resistance is a hallmark of it. The negative cross-talk between skeletal muscle and adipose tissue through adipokines is now accepted as one of the leading cause of insulin resistance. Chemerin is a novel adipokine previously reported to induce insulin resistance in primary human skeletal muscle cells. To investigate the role of chemerin in myogenesis, C2C12 cells were used and treated with chemerin in proliferation and differentiation stages. Our results showed that chemerin promoted proliferation and suppressed differentiation of C2C12 cells through extracellular-signal regulated kinase-1/2 (ERK1/2) and mammalian target of rapamycin (mTOR) signaling pathways, and these two pathways were interacted with each other in C2C12 cells treated with chemerin. It is concluded from this in vitro study that chemerin which expression is increased during myoblast differentiation appears to be able, likely in an autocrine/paracrine manner, to increase myoblast proliferation and decrease myoblast differentiation.

Impacts of Birth Weight on Plasma, Liver and Skeletal Muscle Neutral Amino Acid Profiles and Intestinal Amino Acid Transporters in Suckling Huanjiang Mini-Piglets

Genetic selection strategies towards increased prolificacy have resulted in more and more increased littler size and incidences of impaired fetal development. Low birth weight (LBW) piglets, with long-term alterations in structure, physiology and metabolism, have lower survival rates and poor growth performance. The aim of the study was to compare the plasma, liver and skeletal muscle contents of neutral amino acids (NAA) and the intestinal expression of NAA transporters between LBW and high birth weight (HBW) suckling Huanjiang mini-piglets. Forty piglets with either LBW or HBW (20 piglets per group) were sampled on day 0, 7, 14 and 21 of age to give 5 observations per day per group. The contents of NAA in plasma, liver and skeletal muscle were measured, and jejunal expression of transporters for NAA, including Slc6a19 (B0AT1) and Slc1a5 (ASCT2), were determined by real-time RT-PCR and Western Blot, respectively. Results showed that the suckling piglets with LBW had higher contents of Thr, Ser, Gly, Ala, Val, Met, Ile, Leu, Tyr, Phe and Pro in liver, and Gly in skeletal muscle, whereas lower contents of Met, Ser and Ala in plasma when compared with the HBW littermates. Consistent with the content differences in plasma NAA, the jejunal expression profiles of both Slc6a19 (B0AT1) and Slc1a5 (ASCT2) in the LBW piglets were lower in compared with the HBW littermates during the early suckling period. These findings suggested that intestinal dysfunction in the LBW piglets may be one of the reasons in altered physiology and metabolism states of other organs, which result in lower survival and growth rate.

EAAT3 promotes amino acid transport and proliferation of porcine intestinal epithelial cells

Excitatory amino acid transporter 3 (EAAT3, encoded by SLC1A1) is an epithelial type high-affinity anionic amino acid transporter, and glutamate is the major oxidative fuel for intestinal epithelial cells. This study investigated the effects of EAAT3 on amino acid transport and cell proliferation through activation of the mammalian target of the rapamycin (mTOR) pathway in porcine jejunal epithelial cells (IPEC-J2). Anionic amino acid and cystine (Cys) transport were increased (P<0.05) by EAAT3 overexpression and decreased (P<0.05) by EAAT3 knockdown rather than other amino acids. MTT and cell counting assays suggested that IPEC-J2 cell proliferation increased (P<0.05) with EAAT3 overexpression. Phosphorylation of mTOR (Ser2448), ribosomal protein S6 kinase-1 (S6K1, Thr389) and eukaryotic initiation factor 4E-binding protein-1 (4EBP1, Thr70) was increased by EAAT3 overexpression and decreased by EAAT3 knockdown (P<0.05), as were levels of activating transcription factor 4 (ATF4) and cystine/glutamate antiporter (xCT) (P<0.05). Our results demonstrate for the first time that EAAT3 facilitates anionic amino acid transport and activates the mTOR pathway, promoting Cys transport and IPEC-J2 cell proliferation.

Systematical investigation and physical mechanism of HfO 2 gate stacks band alignment, VFB shift and Fermi level pinning

Band alignment of TiN/HfO₂/SiO₂/Si stack is systematically investigated by X-ray photoelectron spectroscopy. The differences of Si 2p binding energies between SiO₂ and Si substrate are experimentally found to decrease with the sequence of SiO₂/Si, 4 nm HfO₂/SiO₂/Si and 2 nm HfO₂/SiO₂/Si stacks. The p-type Schottky barrier heights at TiN/HfO₂ interface of TiN/HfO₂/SiO₂/Si stack are experimentally estimated to increase with thicker HfO₂ thickness. A physical model based on band alignment of TiN/HfO₂/SiO₂/Si stack is employed to successfully explain these experimental results. The positive VFB shift of TiN/HfO₂/SiO₂/Si stack and Fermi level pinning are also physically demonstrated by this model and attributed to interface induced gap states at TiN/HfO₂ and HfO₂/SiO₂ interfaces.