Acid Transporter mRNA Research Articles

Impact of rosiglitazone on uric acid metabolism and its mechanism

Objective To observe whether high-level insulin increases serum uric acid level and rosiglitazone improves hyperuricemia, and to explore the mechanism. Methods OLETF rats with spontaneous type 2 diabetes complicated with metabolic syndrome and normal control LETO rats were randomly divided into three groups (n=20 each). The animals were fed with standard chow diet in control group, high-purine diet and adenine administered intragastrically in experimental group, and rosiglitazone in interventional group. Body weight, serum levels of uric acid, insulin, triglyceride ( TG ) , and total cholesterol ( TC ) were measured after 3 weeks. Urate transporter 1 ( URAT1 ) and uric acid transporter (UAT) mRNA expressions in renal cortex were examined. HK-2 cells were incubated with various concentrations of insulin for 24 hours. UAT mRNA expression in HK-2 cells was examined. Results ( 1 ) In control group, the insulin level of OLETF rats was significantly higher than that of LETO rats ( P<0. 05 ), and there was no significant difference in serum uric acid level between OLETF and LETO rats. (2)In experimental groups, the insulin level in OLETF rats was significantly higher than that in LETO rats [(61.83±12.13 vs 36.73±12.73 )μIU/ml ,P<0. 05], and the incidence of hyperuricemia (76.92% vs 36.13%,P<0.01 ) and serum uric acid level[( 327.75 ±45.73 vs 264.40±36.32 ) μmol/L, P<0. 01]in OLETF rats were significantly higher than those in LETO rats. (3) Insulin[(41.3± 10.2 vs 61.8±12. 1 )μIU/ml,P<0. 05]and uric acid[( 198.0±45.4 vs 236.9±29.30 ) μmol/L, P<0. 05]levels in OLETF rats in interventional group were significantly lower than those of OLETF rats in experimental group, meanwhile the amount of urinary uric acid excretion was significantly increased[(5 644±371 vs 4 692±278 ) μ mol/L, P<0. 05]. (4) There was no significant difference in insulin level and the expressions of URAT1 and UAT mRNA in renal cortex between OLETF rats in control group and experimental group. URAT1 mRNA expression of OLETF rats in interventional group was significantly decreased, while UAT mRNA expression was significantly increased. (5)With the increase of insulinconcentration in culture medium, the expression of UAT mRNA expression in HK-2 cells was gradually decreased. Conclusions Rosiglitazone may alleviate hyperinsulinemia-induced hyperuricemia via regulating UAT and URAT1 mRNA expression. Key words: Rosiglitazone; Insulin; Hyperuricemia; Urate transporter 1; Uric acid transporter

Skeletal Muscle Amino Acid Transporter mRNA Expression Is Increased In Young And Older Humans Following Resistance Exercise

INTRODUCTION: Amino acid availability is important for increasing muscle protein synthesis. However, for amino acids to be taken up and utilized by the cell, adequate transport of amino acids must occur. Interestingly, amino acid transporters have been linked to mTOR cellular signaling- a major regulator of muscle protein synthesis. PURPOSE: Therefore, we evaluated the expression levels of select amino acid transporters in young and older humans following a single bout of heavy resistance exercise. METHODS: Young (n=5) and Older (n=5) male subjects participated in a single bout of resistance exercise. Muscle biopsies were sampled at Basal, 6 and 24h post exercise. Muscle biopsies were analyzed for LAT1/SLC7A5, CD98/SLC3A2, SNAT2/SLC38A2 and PAT1/SLC36A1 mRNA expression utilizing real-time PCR. RESULTS: LAT1, CD98 and PAT1 mRNA expression were significantly elevated at 6h post in young and older subjects. However, LAT1 response was greater and was prolonged (24h) for the older subjects. Additionally, SNAT2 mRNA expression was elevated at 6 and 24h post in the older while unchanged for the younger subjects. CONCLUSIONS: These data support a rapid upregulation of amino acid transporters gene expression which may support enhanced protein synthesis following a bout of exercise. However, the robust and sustained transporter expression in the old may reflect a greater export of amino acids due to accelerated protein breakdown and muscle remodeling. Supported by ACSM Research Endowment Grant (MJD), P30 AG-024832 (MJD) and NIH/NIAMS grant R01 AR049877 (BBR)

Preparing for Migration? The Effects of Photoperiod and Exercise on Muscle Oxidative Enzymes, Lipid Transporters, and Phospholipids in White‐Crowned Sparrows

The extreme energetic demands of avian migration result in various physiological changes that can be observed during the migratory period. However, the degree to which birds alter muscle physiology in advance of migration has been poorly studied. We studied the effects of "migratory" photoperiod and exercise on metabolic enzymes, fatty acid transporter mRNA expression, and muscle phospholipid fatty acid composition in captive white-crowned sparrows (Zonotrichia leucophrys). Ten sparrows were held on short photoperiod (8L:16D) for 58 d then switched to long days (16L:8D) for 3 wk before sampling. Increased nightly activity indicated that the birds were indeed in migratory condition. Another 13 birds were held on short days during the entire experiment, and a subset (5) were exercised for 1 h every other day for the last 2 wk. "Migratory" photoperiod did not change the activities of citrate synthase, carnitine palmitoyl transferase, and 3-hydroxyacyl-CoA dehydrogenase or the expression of FAT/CD36, FABPpm, and H-FABP mRNA in pectoralis muscle, suggesting that these cannot be increased in advance of migratory flight. Docosahexaenoic acid increased in pectoralis muscle phospholipids with exercise but was negatively correlated with catabolic enzyme activity, indicating that the presence of this fatty acid may not aid migratory performance as suggested by other studies.

An increase in essential amino acid availability upregulates amino acid transporter expression in human skeletal muscle

Essential amino acids (EAA) stimulate skeletal muscle mammalian target of rapamycin complex 1 (mTORC1) signaling and protein synthesis. It has recently been reported that an increase in amino acid (AA) transporter expression during anabolic conditions is rapamycin-sensitive. The purpose of this study was to determine whether an increase in EAA availability increases AA transporter expression in human skeletal muscle. Muscle biopsies were obtained from the vastus lateralis of seven young adult subjects (3 male, 4 female) before and 1-3 h after EAA ingestion (10 g). Blood and muscle samples were analyzed for leucine kinetics using stable isotopic techniques. Quantitative RT-PCR, and immunoblotting were used to determine the mRNA and protein expression, respectively, of AA transporters and members of the general AA control pathway [general control nonrepressed (GCN2), activating transcription factor (ATF4), and eukaryotic initiation factor (eIF2) alpha-subunit (Ser(52))]. EAA ingestion increased blood leucine concentration, delivery of leucine to muscle, transport of leucine from blood into muscle, intracellular muscle leucine concentration, ribosomal protein S6 (Ser(240/244)) phosphorylation, and muscle protein synthesis. This was followed with increased L-type AA transporter (LAT1), CD98, sodium-coupled neutral AA transporter (SNAT2), and proton-coupled amino acid transporter (PAT1) mRNA expression at 1 h (P < 0.05) and modest increases in LAT1 protein expression (3 h post-EAA) and SNAT2 protein expression (2 and 3 h post-EAA, P < 0.05). Although there were no changes in GCN2 expression and eIF2 alpha phosphorylation, ATF4 protein expression reached significance by 2 h post-EAA (P < 0.05). We conclude that an increase in EAA availability upregulates human skeletal muscle AA transporter expression, perhaps in an mTORC1-dependent manner, which may be an adaptive response necessary for improved AA intracellular delivery.

Dietary Protein Intake and Stage of Lactation Differentially Modulate Amino Acid Transporter mRNA Abundance in Porcine Mammary Tissue1–3

To test the hypothesis that under restricted and surfeit protein intake the mammary gland undergoes adaptive regulation, changes in mammary tissue mRNA abundance of cationic amino acid (AA) transporter (CAT)-1, CAT-2B, alanine/serine/cysteine/threonine transporter 1 (ASCT1), and broad specificity transporter for neutral and cationic AA (ATB0,+), and CAT-1 protein abundance were investigated at 2 stages of lactation. Eighteen sows were allocated to a 2 × 3 randomized incomplete block design with 2 stages of lactation (early and peak) and 3 protein levels: deficient (D), adequate (A), or in excess (E) of lactation requirement. In early lactation, compared with A, sows fed E had lower (P = 0.05) piglet growth rate and sows fed D or E had lower (P = 0.05) casein yield. In early lactation, piglet growth rate and milk protein and casein yield increased from D to A and decreased from A to E (quadratic, P = 0.095, P < 0.05, and P < 0.01, respectively). Protein intake did not affect CAT-1, ASCT1, ATB0,+ mRNA abundance, or CAT-1 protein level. Overall, CAT-2B mRNA abundance decreased linearly with increasing protein intake (P < 0.05). Compared with A, E decreased CAT-2B mRNA abundance (P < 0.05). Compared with early lactation, peak lactation did not increase CAT-1 mRNA abundance or relative CAT-1 protein content, but increased abundance of ASCT1 and ATB0,+ mRNA (P < 0.01). Mammary CAT-2B appears to be adaptively regulated in vivo at the transcription level, whereas ASCT1 and ATB0,+ mRNA abundances are associated only with stage of lactation. Neither protein intake nor stage of lactation affects porcine mammary CAT-1 gene expression in vivo.

Ruminal and abomasal starch hydrolysate infusions selectively decrease the expression of cationic amino acid transporter mRNA by small intestinal epithelia of forage-fed beef steers

Although cationic amino acids (CAA) are considered essential to maximize optimal growth of cattle, transporters responsible for CAA absorption by bovine small intestinal epithelia have not been described. This study was conducted to test 2 hypotheses: 1) the duodenal, jejunal, and ileal epithelia of beef cattle differentially express 7 mRNA associated with 4 mammalian amino acid (AA) transport activities: y+ (CAT1), B0,+ (ATB0,+), b0,+ (b0,+AT and rBAT), and y+L (y+LAT1, y+LAT2, and 4F2hc), and 2) the expression of these mRNA is responsive to small intestinal luminal supply of AA substrates (derived from ruminal microbes) or glucose-derived energy (from starch hydrolysate, SH), or both. Eighteen ruminally and abomasally catheterized Angus steers (body weight = 260±17kg) fed an alfalfa cube-based diet at 1.33×net energy for maintenance requirement were assigned to 3 treatments (n=6): ruminal and abomasal water infusion (control); ruminal SH and abomasal water infusion; and ruminal water and abomasal SH infusion. The dosage of SH infusion amounted to 20% of metabolizable energy intake. After 14 or 16 d of infusion, steers were slaughtered, duodenal, jejunal, and ileal epithelia were harvested, and total RNA was extracted. The relative amounts of mRNA expressed by epithelia were quantified using real-time reverse transcription-PCR. All 7 mRNA species were expressed by the epithelium from each region, but their abundance differed among the regions. Specifically, duodenal expression of CAT1 and ATB0,+ mRNA was greater than jejunal or ileal expression; ileal expression of b0,+AT, rBAT, and y+LAT1 mRNA was greater than jejunal or duodenal expression, whereas the expression of y+LAT2 and 4F2hc mRNA did not differ among the 3 epithelia. With regard to SH infusion effect, ruminal infusion down-regulated or tended to down-regulate the jejunal expression of CAT1, rBAT, y+LAT2, and 4F2hc mRNA. Abomasal infusion down-regulated the jejunal expression of y+LAT2 mRNA and tended to down-regulate the jejunal expression of 4F2hc mRNA. This study characterized the pattern of CAA transporter mRNA expressed by growing beef cattle fed an alfalfa-based diet. Moreover, this study demonstrated that increasing the luminal supply of microbe-derived AA (by ruminal supplementation of SH) results in a reduced capacity of apical and basolateral membrane to transport of CAA, whereas increasing luminal glucose supply (by abomasal supplementation of SH) reduces only the basolateral transport capacity, assuming that CAA transporter mRNA content represents functional capacity.

Segmental distribution and expression of two heterodimeric amino acid transporter mRNAs in the intestine of pigs during different ages

AbstractBACKGROUND: Segmental distribution and expression of two heterodimeric amino acid transporter mRNAs during different ages were evaluated in pigs along the horizontal axis of the intestine.RESULTS: The intestinal distribution of b0, + AT and y+LAT1 mRNA at day 90 showed that for b0, + AT the mRNA levels were increased from the proximal to distal part of the small intestine (P &lt; 0.05) and dropped dramatically in colon (P &lt; 0.05). The ileum had the highest b0, + AT mRNA abundance and the ileum had the lowest. Compared to b0, + AT, the y+ LAT1 mRNA levels were decreased from the proximal to distal part of the small intestine (P &lt; 0.05). The duodenum has the highest y+ LAT1 mRNA abundance. The mRNA abundance of b0, + AT mRNA increased linearly (P &lt; 0.0001) from day 1 to day 150 in ileum.CONCLUSION: The mRNA expression of heterodimeric amino acid transporters (b0, + AT and y+LAT1) was not only differentially regulated by age but also differentially distributed along the intestine of piglets at early stages and growing stages of life, which may be related to luminal substrate concentration as well as amino acid requirement and hormones. Copyright © 2008 Society of Chemical Industry

Expression profiles of lipid metabolism-related genes in liver of apoE(-/-)/LDLR(-/-) mice

To explore the relationship between the expression characteristics of lipid metabolism-related genes in the liver and early atherosclerotic lesions in apolipoprotein E and low density lipoprotein receptor gene double knockout (apoE(-/-)/LDLR(-/-)) mice. RT-PCR was used to detect the differential expression of lipid metabolism-related genes in the liver of apoE(-/-)/LDLR(-/-) and wild type (WT) mice. Serum total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) level as well as aortic morphology were also analyzed. Among the 11 lipid metabolism-related genes, apolipoprotein B100 (apoB100) mRNA levels were significantly higher in apoE(-/-)/LDLR(-/-)mice compared with WT mice. At 14 days, 1, 2 and 3 months of age, the level of mRNA expression were 1.55, 1.47, 1.50 and 2.42 folds of those of the age matched WT mice respectively. The fatty acid transporter (FAT/CD36) mRNA expression levels were higher in 14-day and 3-month old mice at 1.30 and 1.35 folds of those of the age matched WT mice, respectively. Apolipoprotein A IV (apoA IV) and Apolipoprotein AV (apoAV) mRNA levels were significantly down-regulated (0.89 fold decrease in 14-day, and 0.90 folds decrease in 3-month, respectively). The mRNA expression levels of apolipoprotein AI (apo AI), apolipoprotein F (apo F), peroxidase proliferator-activated receptor alpha (PPAR-alpha), liver X receptor alpha (LXRalpha), angiopoietin-like protein 3 (ANGPTL3), acyl-coenzymeA oxidase 1 (ACOX1) and carnitine palmitoyl transferase 1 (CPT1) had no significant changes. Serum TC, TG and LDL-C were higher than those of age matched WT mice at 7, 2 and 30 folds, respectively. Furthermore, apoE(-/-)/LDLR(-/-) mice demonstrated typical early atherosclerotic lesions at sinus and root regions of aorta in an age dependent manner. Alterations of the expression of lipid metabolism-related genes in liver play important roles in the development of AS in the apoE(-/-)/LDLR(-/-) mice at early ages.

Regulation of an amino acid transporter mRNA in Trypanosoma brucei

Trypanosoma brucei regulates gene expression by post-transcriptional mechanisms, such as mRNA turnover and translation control. This regulation frequently requires specific sequences located in the 3′-untranslated region. Microarray analysis and Northern blot hybridization showed that the amino acid transporter 11 mRNA is up-regulated in insect stages of the parasite. By RT-PCR and sequencing, the AATP11 polyadenylation site was mapped. We show that this 3′-UTR causes higher expression of the chloramphenicol acetyltransferase (CAT) reporter gene in procyclic trypanosomes than in bloodstream forms. Results of deletion analysis suggested that multiple elements located between nucleotides 141 and 618 of the 3′-untranslated region are required for this control.

Activation of a system A amino acid transporter, ATA1/SLC38A1, in human hepatocellular carcinoma and preneoplastic liver tissues

The expression of amino acid transporter (AT) mRNAs including A system (ATA1/SNAT1/SLC38A1, ATA2/SNAT2/SLC38A2 and ATA3/SNAT3/SLC38A4), L system (LAT1/SLC7A5 and LAT2/SLC7A8), and y+ (CAT2/SLC7A2) genes, were compared among hepatocellular carcinoma (HCC) and non-cancerous liver cells. Among them the ATA1 mRNA expression was significantly elevated in all HCC cell lines (HepG2, HLF, HuH7 and JHH4) examined compared with normal liver tissue. We further discovered that the expression of ATA1 mRNA was significantly activated in HCC tissues and also elevated in pre-malignant cirrhotic livers from HCC patients, compared with normal livers from non-HCC patients. The ATA1 protein was extensively accumulated in the cytoplasm of pre-malignant liver and most HCCs, while being weak or undetectably low in normal liver tissues. SiRNA-mediated suppression of endogenous ATA1 lowered the viability of HepG2 cells. Thus, the activation of ATA1 confers growth and survival advantages in pre-malignant and malignant liver lesions.

Distribution and abundance of nutrient transporter mRNA in the intestinal tract of the black bear, Ursus americanus

End products of digestion are absorbed by the body through the action of transporter proteins expressed on the apical membrane of intestinal epithelial cells. We investigated the mRNA abundance and distribution of a peptide transporter (PepT1), a glucose transporter (SGLT1), two amino acid transporters (NBAT and b o,+AT), and a digestive enzyme, aminopeptidase N (APN), in the intestinal tract of black bears ( Ursus americanus). Intestinal total RNA was isolated from 10 bears and abundance of PepT1, SGLT1, NBAT, b o,+AT, and APN mRNA were determined by Northern blots. Abundance of PepT1 ( P < 0.05), APN ( P < 0.05), and SGLT1 ( P < 0.0001) changed quadratically from the proximal to distal intestine with abundance being greatest in the midregion. Abundance of b o,+AT mRNA increased linearly ( P < 0.05) from the proximal to distal intestine. The number of molecules of mRNA/ng of total RNA for each gene was determined using Real-Time PCR. PepT1 mRNA was present at 10-fold or greater levels than amino acid transporter mRNA in all segments of the intestine, suggesting that di- and tripeptides constitute a major form in which amino acids are absorbed in the black bear. The abundance of NBAT and b o,+AT mRNA was greater towards the distal intestine, suggesting a role in salvaging unabsorbed amino acids.

Tissue-Specific and Fatty Acid Transporter-Specific Changes in Heart and Soleus Muscle Over a 1-yr Period

Rates of fatty acid oxidation increase rapidly in both rat heart and skeletal muscle in the early postnatal period. Therefore, we examined in heart and soleus muscle, (a) whether there were rapid changes in fatty acid transporter (FAT/CD36, FABPpm) mRNA and protein expression early in life (days 10 -36) and thereafter (days 84, 160, 365), and (b) whether the rates of fatty acid transport and the plasmalemmal content of FAT/CD36 and FABPpm were altered. Protein expression was altered rapidly from day 10-36 in both heart (FAT/CD36 only, +21%, P < 0.05)) and soleus muscle (FAT/CD36 + 100%, P < 0.05; FABPpm -20%, P < 0.05), with no further changes thereafter (P < 0.05). Rates of fatty acid transport (day 10 vs day 160) were increased in heart (+33%, P < 0.05) and muscle (+85%, P < 0.05), and were associated with concomitant increases in plasmalemmal FABPpm (+44%, P < 0.05) and FAT/CD36 (+16%, P < 0.05) in the heart, and only plasmalemmal FAT/CD36 in muscle (+90%, P < 0.05). Therefore, known changes in the rates of fatty acid oxidation in heart and muscle early in life appear to be accompanied by a concurrent upregulation in the rates of fatty acid transport and the expression of FAT/CD36 in heart and muscle, as well as an increase in plasmalemmal FAT/CD36 and FABPpm in the heart, and only plasmalemmal FAT/CD36 in soleus muscle. We speculate that the rapid upregulation of fatty acid transport rates in heart and muscle are needed to support the increased rates of fatty oxidation that have been previously observed in these tissues.

Eicosapentaenoic acid (20:5 n-3) increases fatty acid and glucose uptake in cultured human skeletal muscle cells

This study was conducted to evaluate the chronic effects of eicosapentaenoic acid (EPA) on fatty acid and glucose metabolism in human skeletal muscle cells. Uptake of [14C]oleate was increased >2-fold after preincubation of myotubes with 0.6 mM EPA for 24 h, and incorporation into various lipid classes showed that cellular triacylgycerol (TAG) and phospholipids were increased 2- to 3-fold compared with control cells. After exposure to oleic acid (OA), TAG was increased 2-fold. Insulin (100 nM) further increased the incorporation of [14C]oleate into all lipid classes for EPA-treated myotubes. Fatty acid beta-oxidation was unchanged, and complete oxidation (CO2) decreased in EPA-treated cells. Basal glucose transport and oxidation (CO2) were increased 2-fold after EPA, and insulin (100 nM) stimulated glucose transport and oxidation similarly in control and EPA-treated myotubes, whereas these responses to insulin were abolished after OA treatment. Lower concentrations of EPA (0.1 mM) also increased fatty acid and glucose uptake. CD36/FAT (fatty acid transporter) mRNA expression was increased after EPA and OA treatment compared with control cells. Moreover, GLUT1 expression was increased 2.5-fold by EPA, whereas GLUT4 expression was unchanged, and activities of the mitogen-activated protein kinase p38 and extracellular signal-regulated kinase were decreased after treatment with OA compared with EPA. Together, our data show that chronic exposure of myotubes to EPA promotes increased uptake and oxidation of glucose despite a markedly increased fatty acid uptake and synthesis of complex lipids.

Lysine deficiency and feed restriction independently alter cationic amino acid transporter expression in chickens ( Gallus gallus domesticus)

The effect of a lysine-deficient diet on cationic amino acid transporter (CAT1-3) mRNA expression was determined in broiler chickens. Chicks consumed a lysine-adequate (LA; 1.3% lysine) or lysine-deficient (LD; 0.7% lysine) diet. Pair-fed chicks consumed the LA diet in an amount equal to that consumed by LD chicks during the previous day (PLA). CAT 1–3 mRNA expression in the liver, pectoralis and bursa of LD chicks were lower than that of LA and PLA chicks ( P < 0.05), and levels were not detectable in LD chick thymus. High affinity CAT mRNA expression in isolated bursacytes was 16-fold higher in LD chicks than that of LA chicks ( P < 0.001). Thymocyte high affinity CAT mRNA expression was 5-fold lower than that of LA chicks ( P < 0.05). The summed amount of high affinity CAT-1 and CAT-3 mRNA expression in chicks fed a lysine adequate diet was highly correlated ( r 2 = 0.51; P < 0.001) to a tissue's growth during a lysine deficiency or feed restriction. In the thymus and bursa of LD chicks, CAT mRNA levels differed between resident lymphocytes and their surrounding tissues. By expressing high affinity CAT isoforms, developing lymphocytes may have a greater ability to obtain lysine than their surrounding tissue during a lysine deficiency.

Apical sodium bile acid transporter and ileal lipid binding protein in gallstone carriers

Although a cholesterol supersaturation of gallbladder bile has been identified as the underlying pathophysiologic defect, the molecular pathomechanism of gallstone formation in humans remains poorly understood. A deficiency of the apical sodium bile acid transporter (ASBT) and ileal lipid binding protein (ILBP) in the small intestine may result in bile acid loss into the colon and might promote gallstone formation by reducing the bile acid pool and increasing the amount of hydrophobic bile salts. To test this hypothesis, protein levels and mRNA expression of ASBT and ILBP were assessed in ileal mucosa biopsies of female gallstone carriers and controls. Neither ASBT nor ILBP levels differed significantly between gallstone carriers and controls. However, when study participants were subgrouped by body weight, ASBT and ILBP protein were 48% and 67% lower in normal weight gallstone carriers than in controls (P < 0.05); similar differences were found for mRNA expression levels. The loss of bile transporters in female normal weight gallstone carriers was coupled with a reduction of protein levels of hepatic nuclear factor 1alpha and farnesoid X receptor. In conclusion, in normal weight female gallstone carriers, the decreased expression of ileal bile acid transporters may form a molecular basis for gallstone formation.

The acute phase response alters cationic amino acid transporter expression in growing chickens ( Gallus gallus domesticus)

The effect of an acute phase response (APR) on cationic amino acid transporter (CAT1–3) mRNA expression in liver, muscle, bursa and thymus was determined in broiler strain chickens. The APR was initiated by injecting Salmonella typhimurium lipopolysaccharide subcutaneously (LPS; 1 mg/kg bw). In Experiment 1, CAT1–3 mRNA expression was determined at multiple time points following LPS administration. LPS increased bursa and liver total and high affinity CAT mRNA expression ( P < 0.05) and transiently increased pectoralis total CAT mRNA expression ( P < 0.05). Total CAT mRNA expression in the thymus decreased 7.7-fold from 0 to 8 h after LPS injection ( P < 0.05). In Experiment 2, fasted chicks were uninjected or LPS-injected. LPS increased total and high affinity CAT mRNA 2-fold in both the bursa and liver ( P < 0.05) and did not change thymus total and high affinity CAT mRNA expression ( P > 0.05). LPS increased liver weight only ( P < 0.05) and did not alter the plasma lysine and arginine concentration ( P > 0.05). In Experiments 3 and 4, thymocyte proliferation and total protein content were dependent upon the media lysine concentration ( P < 0.001). The inability of the thymus to compete for lysine and arginine during the APR may limit the ability of thymocytes to develop during infections.

Expressional reprogramming of survival pathways in rat cardiocytes by neuregulin-1β

Neuregulin/ErbB2-induced kinase signaling provides essential survival and protection clues for functional integrity of the adult heart and skeletal muscle. To define the regulatory pathways involved in neuregulin-dependent muscle cell survival, we set out to map the largely unknown transcript targets of this growth/differentiation factor in cardiocytes. Freshly isolated adult primary rat cardiocytes were treated for 24 h with recombinant human neuregulin-1beta (NRG-1beta, 30 ng/ml). Transcript level alterations in NRG-1beta-treated and control cardiocytes (n = 6) were identified with Atlas Rat Toxicology 1.2 cDNA arrays (BD Clontech) and established permutation L1 regression analysis. Selected transcriptional adjustments were confirmed by RT-PCR and Western blotting. Involvement of MAPK pathways was verified with the inhibitor PD-98059. Application of the single dose of NRG-1beta to quiescent cardiocytes induced expressional reprogramming of distinct cellular processes. This response included a prominent 50-100% increase in transcripts of multiple redox systems. It also involved a comparable mRNA augmentation of protein synthetic and folding factors together with augmented message for the trigger of cardiac hypertrophy, cyclin D1 (CCND1). First evidence for a role of neuregulin in promotion of mitochondrial turnover, voltage-gated ion channel expression, and the suppression of fatty acid transporter mRNAs was revealed. Subsequent analysis confirmed a corresponding upregulation of redox factor proteins thioredoxin and the thioredoxin reductase 1, GSTP-1, and CCND1 and demonstrated downregulation of the related transcripts by PD-98059 in neuregulin-stimulated cultures. These MAPK-dependent expressional adjustments point to novel oxidative defense and hypertrophy pathways being involved in the longer lasting protective function of neuregulin in the heart.

Altered Epithelial Cell Lineage Allocation and Global Expansion of the Crypt Epithelial Stem Cell Population Are Associated with Ileitis in SAMP1/YitFc Mice

Crohn's disease is characterized by cycles of mucosal injury and ulceration followed by epithelial regeneration and restoration of normal epithelial function. In this study, we examined whether ileitis in SAMP1/YitFc mice, a recombinant-inbred line that spontaneously develops ileitis resembling human Crohn's disease, was associated with alterations in normal patterns of epithelial differentiation or changes in epithelial regeneration after experimental injury. Increased numbers of Paneth, goblet, and intermediate cells were present focally in the ileum of SAMP1/YitFc mice by 4 weeks of age, before any histological evidence of acute or chronic inflammation. This increase in secretory cells became more pronounced at sites of ileitis with increasing age and inflammation. Additionally, there was mispositioning of Paneth and intermediate cells along the crypt-to-villus unit. A concomitant reduction in the number of absorptive enterocytes was observed. In contrast to the ileal-specific changes in lineage allocation, crypt stem cell numbers began to increase in both the ileum and proximal jejunum at the onset of inflammation in SAMP1/YitFc mice. These data suggest that the alterations in epithelial cell differentiation and increases in the size of the crypt stem cell population observed in SAMP1/YitFc mice are regulated by distinct mechanisms. We speculate that these epithelial alterations may play a role in the pathogenesis of ileitis in this murine model of Crohn's disease.

Increased Expression of Specific Intestinal Amino Acid and Peptide Transporter mRNA in Rats Fed by TPN Is Reversed by GLP-2

Intestinal function depends on the presence of luminal nutrients and is altered during starvation and refeeding. Amino acids are essential for enterocytes, but the luminal supply is compromised with changes in dietary intake. To test the hypothesis that during periods of restricted luminal nutrient availability mucosal cells undergo adaptations aimed toward preserving amino acid supply, the expression of amino acid and peptide transporter mRNAs was quantified in rats with no oral intake, whose nutritional status was maintained with total parenteral nutrition (TPN). The role of the intestinotrophic hormone glucagon-like peptide-2 (GLP-2) was investigated in the adaptive responses. Rats were administered TPN with or without exogenous GLP-2. Amino acid and peptide transporter mRNAs in small intestine mucosa were measured by semiquantitative RT-PCR. Compared with orally fed rats, removal of luminal nutrition increased the expression of ASCT1, SAT2, and GLYT1 mRNAs in the duodenum and of ASCT2, EAAC1, NBAT, and PepT1 mRNAs in the ileum. CAT1, PAT1, and SN2 mRNA abundances were unaffected. GLP-2 reversed these effects. Three subgroups of transporters were identified by regional differences in response to TPN. This may reflect differing roles for substrates of transporters located apically and basally and along the proximal-distal axis of the intestine. The importance of maintaining amino acid supply for intestinal mucosal cells is illustrated.

Glucose and cationic amino acid transporter expression in growing chickens ( Gallus gallus domesticus)

Tissue glucose transporter (GLUT1–3) and cationic amino acid transporter (CAT1–3) mRNA expression was determined in growing broiler chicks posthatch. In two experiments, tissues were either collected on days 1, 3 and 7 or days 1 and 14 posthatch. Heart and liver were the only tissues expressing a GLUT isoform on day 1. All tissues expressed a GLUT isoform on day 7 except for the thymus. Most tissues expressing a CAT isoform on day 1 decreased mRNA levels through day 7 ( P<0.05), except for bursa CAT-1 which tended to increase ( P=0.05). The thymus and spleen did not express any CAT isoform mRNA until day 7. The liver was the only tissue expressing GLUT-2 mRNA through day 14. On day 14, GLUT-1, CAT-1 and CAT-2 mRNA were differentially expressed across tissues ( P<0.05). High-affinity GLUT and CAT mRNA expression was highest in the heart and bursa, respectively ( P<0.05). Total CAT mRNA expression was greatest in the bursa ( P<0.05). The thymus had the lowest high affinity GLUT and total CAT mRNA expression on day 14 posthatch. Therefore, T lymphocytes within the thymus may be most susceptible to glucose and cationic amino acid supply.