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Wang J, Ren W, Li Z, Li L, Wang R, Ma S, Zeng Y, Meng J, Yao X. Regulatory Mechanisms of Yili Horses During an 80 km Race Based on Transcriptomics and Metabolomics Analyses. Int J Mol Sci 2025; 26:2426. [PMID: 40141070 PMCID: PMC11942362 DOI: 10.3390/ijms26062426] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2025] [Revised: 02/27/2025] [Accepted: 03/06/2025] [Indexed: 03/28/2025] Open
Abstract
Equine endurance exercise induces physiological changes that alter metabolism and molecular pathways to maintain balance after intense physical activity. However, the specific regulatory mechanisms remain under debate. Identifying differentially expressed genes (DEGs) and differential metabolites (DMs) associated with equine endurance is essential for elucidating these regulatory mechanisms. This study collected blood samples from six Yili horses before and after an 80 km race and conducted transcriptomics and metabolomics analyses, yielding 722 DEGs and 256 DMs. These DEGs were primarily enriched in pathways related to amino acid biosynthesis, cellular senescence, and lipid metabolism/atherosclerosis. The DMs were predominantly enriched in fatty acid biosynthesis and the biosynthesis of unsaturated fatty acids. The integrative transcriptomics and metabolomics analyses of DEGs and DMs highlight functional changes during the endurance race. The findings offer a holistic understanding of the regulatory mechanisms underlying equine endurance and a solid foundation for formulating training programs to optimize horse performance in endurance racing.
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Affiliation(s)
- Jianwen Wang
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
- Xinjiang Key Laboratory of Equine Breeding and Exercise Physiology, Urumqi 830052, China
| | - Wanlu Ren
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
- Xinjiang Key Laboratory of Equine Breeding and Exercise Physiology, Urumqi 830052, China
| | - Zexu Li
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
| | - Luling Li
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
| | - Ran Wang
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
| | - Shikun Ma
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
| | - Yaqi Zeng
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
- Xinjiang Key Laboratory of Equine Breeding and Exercise Physiology, Urumqi 830052, China
| | - Jun Meng
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
- Xinjiang Key Laboratory of Equine Breeding and Exercise Physiology, Urumqi 830052, China
| | - Xinkui Yao
- College of Animal Science, Xinjiang Agricultural University, Urumqi 830052, China; (J.W.); (W.R.); (Z.L.); (L.L.); (R.W.); (S.M.); (Y.Z.)
- Xinjiang Key Laboratory of Equine Breeding and Exercise Physiology, Urumqi 830052, China
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Thomsen KL, Eriksen PL, Kerbert AJC, De Chiara F, Jalan R, Vilstrup H. Role of ammonia in NAFLD: An unusual suspect. JHEP Rep 2023; 5:100780. [PMID: 37425212 PMCID: PMC10326708 DOI: 10.1016/j.jhepr.2023.100780] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/08/2022] [Revised: 03/29/2023] [Accepted: 03/30/2023] [Indexed: 07/11/2023] Open
Abstract
Mechanistically, the symptomatology and disease progression of non-alcoholic fatty liver disease (NAFLD) remain poorly understood, which makes therapeutic progress difficult. In this review, we focus on the potential importance of decreased urea cycle activity as a pathogenic mechanism. Urea synthesis is an exclusive hepatic function and is the body's only on-demand and definitive pathway to remove toxic ammonia. The compromised urea cycle activity in NAFLD is likely caused by epigenetic damage to urea cycle enzyme genes and increased hepatocyte senescence. When the urea cycle is dysfunctional, ammonia accumulates in liver tissue and blood, as has been demonstrated in both animal models and patients with NAFLD. The problem may be augmented by parallel changes in the glutamine/glutamate system. In the liver, the accumulation of ammonia leads to inflammation, stellate cell activation and fibrogenesis, which is partially reversible. This may be an important mechanism for the transition of bland steatosis to steatohepatitis and further to cirrhosis and hepatocellular carcinoma. Systemic hyperammonaemia has widespread negative effects on other organs. Best known are the cerebral consequences that manifest as cognitive disturbances, which are prevalent in patients with NAFLD. Furthermore, high ammonia levels induce a negative muscle protein balance leading to sarcopenia, compromised immune function and increased risk of liver cancer. There is currently no rational way to reverse reduced urea cycle activity but there are promising animal and human reports of ammonia-lowering strategies correcting several of the mentioned untoward aspects of NAFLD. In conclusion, the ability of ammonia-lowering strategies to control the symptoms and prevent the progression of NAFLD should be explored in clinical trials.
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Affiliation(s)
- Karen Louise Thomsen
- Department of Hepatology and Gastroenterology, Aarhus University Hospital, Denmark
- UCL Institute of Liver and Digestive Health, University College London, United Kingdom
| | - Peter Lykke Eriksen
- Department of Hepatology and Gastroenterology, Aarhus University Hospital, Denmark
| | - Annarein JC. Kerbert
- UCL Institute of Liver and Digestive Health, University College London, United Kingdom
| | - Francesco De Chiara
- UCL Institute of Liver and Digestive Health, University College London, United Kingdom
| | - Rajiv Jalan
- UCL Institute of Liver and Digestive Health, University College London, United Kingdom
- European Foundation for the Study of Chronic Liver Failure, Barcelona, Spain
| | - Hendrik Vilstrup
- Department of Hepatology and Gastroenterology, Aarhus University Hospital, Denmark
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Jung EH, Nguyen J, Nelson C, Brauner CJ, Wood CM. Ammonia transport is independent of PNH 3 gradients across the gastrointestinal epithelia of the rainbow trout: A role for the stomach. JOURNAL OF EXPERIMENTAL ZOOLOGY. PART A, ECOLOGICAL AND INTEGRATIVE PHYSIOLOGY 2023; 339:180-192. [PMID: 36369634 DOI: 10.1002/jez.2670] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/25/2022] [Revised: 10/01/2022] [Accepted: 10/05/2022] [Indexed: 11/13/2022]
Abstract
Although the gastrointestinal tract (GIT) is an important site for nitrogen metabolism in teleosts, the mechanisms of ammonia absorption and transport remain to be elucidated. Both protein catabolism in the lumen and the metabolism of the GIT tissues produce ammonia which, in part, enters the portal blood through the anterior region of the GIT. The present study examined the possible roles of different GIT sections of rainbow trout (Oncorhynchus mykiss) in transporting ammonia in its unionized gas form-NH3 -by changing the PNH3 gradient across GIT epithelia using in vitro gut sac preparations. We also surveyed messenger RNA expression patterns of three of the identified Rh proteins (Rhbg, Rhcg1, and Rhcg2) as potential NH3 transporters and NKCC as a potential ammonium ion (NH4 + ) transporter along the GIT of rainbow trout. We found that ammonia absorption is not dependent on the PNH3 gradient despite expression of Rhbg and Rhcg2 in the intestinal tissues, and Rhcg2 in the stomach. We detected no expression of Rhbg in the stomach and no expression of Rhcg1 in any GIT tissues. There was also a lack of correlation between ammonia transport and [NH4 + ] gradient despite NKCC expression in all GIT tissues. Regardless of PNH3 gradients, the stomach showed the greatest absorption and net tissue consumption of ammonia. Overall, our findings suggest nitrogen metabolism zonation of GIT, with stomach serving as an important site for the absorption, handling and transport of ammonia that is independent of the PNH3 gradient.
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Affiliation(s)
- Ellen H Jung
- Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Jessica Nguyen
- Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Charlotte Nelson
- Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Colin J Brauner
- Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada
| | - Chris M Wood
- Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada
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Paulusma CC, Lamers W, Broer S, van de Graaf SFJ. Amino acid metabolism, transport and signalling in the liver revisited. Biochem Pharmacol 2022; 201:115074. [PMID: 35568239 DOI: 10.1016/j.bcp.2022.115074] [Citation(s) in RCA: 63] [Impact Index Per Article: 21.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2022] [Revised: 04/28/2022] [Accepted: 04/29/2022] [Indexed: 11/02/2022]
Abstract
The liver controls the systemic exposure of amino acids entering via the gastro-intestinal tract. For most amino acids except branched chain amino acids, hepatic uptake is very efficient. This implies that the liver orchestrates amino acid metabolism and also controls systemic amino acid exposure. Although many amino acid transporters have been identified, cloned and investigated with respect to substrate specificity, transport mechanism, and zonal distribution, which of these players are involved in hepatocellular amino acid transport remains unclear. Here, we aim to provide a review of current insight into the molecular machinery of hepatic amino acid transport. Furthermore, we place this information in a comprehensive overview of amino acid transport, signalling and metabolism.
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Affiliation(s)
- Coen C Paulusma
- Tytgat Institute for Liver and Intestinal Research, Amsterdam University Medical Centers, University of Amsterdam, Amsterdam, Netherlands; Department of Gastroenterology and Hepatology, Amsterdam University Medical Centers, Amsterdam, Netherlands; Amsterdam Gastroenterology Endocrinology Metabolism, Amsterdam University Medical Centers, Amsterdam, The Netherlands
| | - Wouter Lamers
- Tytgat Institute for Liver and Intestinal Research, Amsterdam University Medical Centers, University of Amsterdam, Amsterdam, Netherlands; Department of Gastroenterology and Hepatology, Amsterdam University Medical Centers, Amsterdam, Netherlands; Department of Anatomy & Embryology, Maastricht University, Maastricht, The Netherlands
| | - Stefan Broer
- Department of Gastroenterology and Hepatology, Amsterdam University Medical Centers, Amsterdam, Netherlands; Research School of Biology, Australian National University, Canberra, Australia
| | - Stan F J van de Graaf
- Tytgat Institute for Liver and Intestinal Research, Amsterdam University Medical Centers, University of Amsterdam, Amsterdam, Netherlands; Department of Gastroenterology and Hepatology, Amsterdam University Medical Centers, Amsterdam, Netherlands; Amsterdam Gastroenterology Endocrinology Metabolism, Amsterdam University Medical Centers, Amsterdam, The Netherlands; Department of Anatomy & Embryology, Maastricht University, Maastricht, The Netherlands.
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Levitt MD, Levitt DG. Use Of Quantitative Modelling To Elucidate The Roles Of The Liver, Gut, Kidney, And Muscle In Ammonia Homeostasis And How Lactulose And Rifaximin Alter This Homeostasis. Int J Gen Med 2019; 12:367-380. [PMID: 31686894 PMCID: PMC6798813 DOI: 10.2147/ijgm.s218405] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2019] [Accepted: 09/24/2019] [Indexed: 12/18/2022] Open
Abstract
Humans must eliminate approximately 1M of ammonia per day while maintaining the blood concentration of this potent neurotoxin at a concentration of only about 30 µM. The mechanisms producing such effective ammonia homeostasis are poorly understood by clinicians due to the multiple organs (liver, gut, kidney and muscle) involved in ammonia homeostasis. Based on literature values we present a novel, simplified description of normal and disordered ammonia and the potential mechanisms whereby the drugs used to treat hepatic encephalopathy, lactulose and rifaximin, lower the blood ammonia concentration. Concepts discussed include the following: 1) only about 44 mmol of ammonia/day (4.4% of total production) reaches the peripheral circulation due to the efficient linkage of amino deamination and the urea cycle in hepatic mitochondria; 2) the gut and kidney contribute roughly equally to delivery of this 44 mmol/day to systemic blood; 3) the bulk of gut ammonia production seemingly originates in the small bowel from bacterial deamination of urea by bacteria and mucosal deamination of circulating and ingested glutamine; 4) the apparent production of ammonia in the small bowel markedly exceeds that quantity that enters the portal blood, indicating that ammonia disposal mechanisms in the small bowel play a major role in ammonia homeostasis. With regard to the hyperammonemia of chronic liver disease: 1) shunting of portal blood around the liver, by itself, can account for commonly observed ammonia elevations; 2) severe portal hypertension causes an increased release of ammonia by the kidney; 3) high blood ammonia is associated with an unexplained massive increase in the muscle uptake of ammonia that could play an important role in limiting hyperammonemia; and 4) a major action of lactulose administration may be the enhancement of ammonia uptake by small bowel bacteria, while the mechanism of action of rifaximin is unclear.
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Affiliation(s)
- Michael D Levitt
- Research Service, Veterans Affairs Medical Center, Minneapolis, MN 55417, USA
| | - David G Levitt
- Department of Integrative Biology and Physiology, University of Minnesota, Minneapolis, MN 55455, USA
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Huang J, Jia Y, Li Q, Burris WR, Bridges PJ, Matthews JC. Hepatic glutamate transport and glutamine synthesis capacities are decreased in finished vs. growing beef steers, concomitant with increased GTRAP3-18 content. Amino Acids 2018; 50:513-525. [PMID: 29392419 PMCID: PMC5917004 DOI: 10.1007/s00726-018-2540-8] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2017] [Accepted: 01/22/2018] [Indexed: 01/17/2023]
Abstract
Hepatic glutamate uptake and conversion to glutamine is critical for whole-body N metabolism, but how this process is regulated during growth is poorly described. The hepatic glutamate uptake activities, protein content of system [Formula: see text] transporters (EAAC1, GLT-1) and regulatory proteins (GTRAP3-18, ARL6IP1), glutamine synthetase (GS) activity and content, and glutathione (GSH) content, were compared in liver tissue of weaned Angus steers randomly assigned (n = 8) to predominantly lean (growing) or predominantly lipid (finished) growth regimens. Steers were fed a cotton seed hull-based diet to achieve final body weights of 301 or 576 kg, respectively, at a constant rate of growth. Liver tissue was collected at slaughter and hepatic membranes fractionated. Total (75%), Na+-dependent (90%), system [Formula: see text]-dependent (abolished) glutamate uptake activity, and EAAC1 content (36%) in canalicular membrane-enriched vesicles decreased as steers developed from growing (n = 6) to finished (n = 4) stages, whereas Na+-independent uptake did not change. In basolateral membrane-enriched vesicles, total (60%), Na+-dependent (60%), and Na+-independent (56%) activities decreased, whereas neither system [Formula: see text]-dependent uptake nor protein content changed. EAAC1 protein content in liver homogenates (n = 8) decreased in finished vs. growing steers, whereas GTRAP3-18 and ARL6IP1 content increased and GLT-1 content did not change. Concomitantly, hepatic GS activity decreased (32%) as steers fattened, whereas GS and GSH contents did not differ. We conclude that hepatic glutamate uptake and GS synthesis capacities are reduced in livers of finished versus growing beef steers, and that hepatic system [Formula: see text] transporter activity/EAAC1 content is inversely proportional to GTRAP3-18 content.
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Affiliation(s)
- J Huang
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA
| | - Y Jia
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA
| | - Q Li
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA
| | - W R Burris
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA
| | - P J Bridges
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA
| | - J C Matthews
- Department of Animal and Food Sciences, University of Kentucky, Lexington, 40546, USA.
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7
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Jia Y, Li Q, Burris WR, Aiken GE, Bridges PJ, Matthews JC. Forms of selenium in vitamin-mineral mixes differentially affect serum prolactin concentration and hepatic glutamine synthetase activity of steers grazing endophyte-infected tall fescue. J Anim Sci 2018; 96:715-727. [PMID: 29385471 DOI: 10.1093/jas/skx068] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2017] [Accepted: 12/19/2017] [Indexed: 01/11/2023] Open
Abstract
The goal of this study was to test the hypothesis that sodium selenite (ISe), SEL-PLEX (OSe), vs. an 1:1 blend (MIX) of ISe and OSe in a basal vitamin-mineral (VM) mix would differentially affect metabolic parameters and performance of growing steers grazing toxic endophyte-infected tall fescue mixed forage (E+) pasture. Predominately-Angus steers (BW = 183 ± 34 kg) were randomly selected from herds of fall-calving cows grazing E+ pasture and consuming VM mixes that contained 35 ppm Se as ISe, OSe, and MIX forms. Steers were weaned, depleted of Se for 98 d, and subjected to summer-long common grazing of an E+ pasture (0.51 ppm total ergovaline per ergovalinine; 10.1 ha). Steers were assigned (n = 8 per treatment) to the same Se-form treatments upon which they were raised. Selenium treatments were administered by daily top-dressing 85 g of VM mix onto 0.23 kg soyhulls, using in-pasture Calan gates. The PROC MIXED procedure of SAS was used to assess effect of Se-form treatments on whole blood Se (ng/mL) and serum prolactin (ng/mL) at day 0, 22, 43, 64, and 86, and caudal arterial area (mm2) at day -7, 43, and 86. The effect of Se treatment on ADG (day 86), and liver glutamine synthetase (GS) mRNA, protein, and activity (nmol/mg wet tissue/min) were assessed using the PROC GLM procedure of SAS. Fisher's protected LSD procedure was used to separate treatment means. Whole blood Se increased (P < 0.01) for all treatments from day 0 to 22 and then did not change (P ≥ 0.17), and was greater (P ≤ 0.04) for MIX and OSe steers. Serum prolactin decreased (P < 0.01) over time and was greater (P < 0.05) for MIX and OSe steers. Liver GS mRNA content was 66% and 59% greater (P < 0.05) in MIX and OSe steers, respectively, than ISe steers. Liver GS protein content in MIX steers was 94% more (P < 0.01) than ISe steers. Moreover, MIX and OSe steers had 99% and 55% more (P ≤ 0.01) liver GS activity, respectively, than ISe steers. ADG was not affected (P = 0.36) by Se treatments. We conclude that consumption of 3 mg Se/d as OSe or MIX forms of Se in VM mixes increased 1) whole blood Se content, an indicator of greater whole-body Se assimilation; 2) serum prolactin, the reduction of which is a hallmark of fescue toxicosis; and 3) hepatic GS activity, indicating greater hepatic assimilation of acinar ammonia. However, 4) these positive effects on metabolic parameters were not accompanied by increased growth performance.
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Affiliation(s)
- Yang Jia
- Department of Animal and Food Sciences, University of Kentucky, Lexington, KY
| | - Qing Li
- Department of Animal and Food Sciences, University of Kentucky, Lexington, KY
| | - W R Burris
- Department of Animal and Food Sciences, University of Kentucky, Lexington, KY
| | - Glenn E Aiken
- ARS-USDA, Forage-Animal Production Research Unit, Lexington, KY
| | - Phillip J Bridges
- Department of Animal and Food Sciences, University of Kentucky, Lexington, KY
| | - James C Matthews
- Department of Animal and Food Sciences, University of Kentucky, Lexington, KY
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Moran ET. Gastric digestion of protein through pancreozyme action optimizes intestinal forms for absorption, mucin formation and villus integrity. Anim Feed Sci Technol 2016. [DOI: 10.1016/j.anifeedsci.2016.05.015] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
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Dietary glutamine supplementation effects on amino acid metabolism, intestinal nutrient absorption capacity and antioxidant response of gilthead sea bream (Sparus aurata) juveniles. Comp Biochem Physiol A Mol Integr Physiol 2015; 191:9-17. [PMID: 26424608 DOI: 10.1016/j.cbpa.2015.09.012] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2015] [Revised: 09/17/2015] [Accepted: 09/18/2015] [Indexed: 01/04/2023]
Abstract
A study was undertaken to evaluate dietary glutamine supplementation effects on gilthead sea bream performance, intestinal nutrient absorption capacity, hepatic and intestinal glutamine metabolism and oxidative status. For that purpose gilthead sea bream juveniles (mean weight 13.0g) were fed four isolipidic (18% lipid) and isonitrogenous (43% protein) diets supplemented with 0, 0.5, 1 and 2% glutamine for 6weeks. Fish performance, body composition and intestinal nutrient absorption capacity were not affected by dietary glutamine levels. Hepatic and intestinal glutaminase (GlNase), glutamine synthetase (GSase), alanine aminotransferase, aspartate aminotransferase and glutamate dehydrogenase activities were also unaffected by dietary glutamine supplementation. In the intestine GlNase activity was higher and GSase/GlNase ratio was two-fold lower than in the liver, suggesting a higher use of glutamine for energy production by the intestine than by the liver. The liver showed higher catalase and glucose-6-phosphate dehydrogenase activities, while the intestine presented higher glutathione peroxidase and glutathione reductase activities and oxidised glutathione content, which seems to reveal a higher glutathione dependency of the intestinal antioxidant response. Total and reduced glutathione contents in liver and intestine and superoxide dismutase activity in the intestine were enhanced by dietary glutamine, though lipid peroxidation values were not affected. Overall, differences between liver and intestine glutamine metabolism and antioxidant response were identified and the potential of dietary glutamine supplementation to gilthead sea bream's antioxidant response was elucidated.
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Díaz-Herrero MM, del Campo JA, Carbonero-Aguilar P, Vega-Pérez JM, Iglesias-Guerra F, Periñán I, Miñano FJ, Bautista J, Romero-Gómez M. THDP17 decreases ammonia production through glutaminase inhibition. A new drug for hepatic encephalopathy therapy. PLoS One 2014; 9:e109787. [PMID: 25329718 PMCID: PMC4201470 DOI: 10.1371/journal.pone.0109787] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/08/2014] [Accepted: 09/03/2014] [Indexed: 11/29/2022] Open
Abstract
Ammonia production is implicated in the pathogenesis of hepatic encephalopathy (HE), being intestinal glutaminase activity the main source for ammonia. Management of ammonia formation can be effective in HE treatment by lowering intestinal ammonia production. The use of glutaminase inhibitors represents one way to achieve this goal. In this work, we have performed a search for specific inhibitors that could decrease glutaminase activity by screening two different groups of compounds: i) a group integrated by a diverse, highly pure small molecule compounds derived from thiourea ranging from 200 to 800 Daltons; and ii) a group integrated by commonly use compounds in the treatment of HE. Results shown that THDP-17 (10 µM), a thiourea derivate product, could inhibit the intestinal glutaminase activity (57.4±6.7%). Inhibitory effect was tissue dependent, ranging from 40±5.5% to 80±7.8% in an uncompetitive manner, showing Vmax and Km values of 384.62 µmol min−1, 13.62 mM with THDP-17 10 µM, respectively. This compound also decreased the glutaminase activity in Caco-2 cell cultures, showing a reduction of ammonia and glutamate production, compared to control cultures. Therefore, the THDP-17 compound could be a good candidate for HE management, by lowering ammonia production.
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Affiliation(s)
- M. Mar Díaz-Herrero
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
| | - José A. del Campo
- Unidad de Gestión Clínica de Enfermedades Digestivas & Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Hospital Universitario de Valme, Sevilla, Spain
| | - Pilar Carbonero-Aguilar
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
| | - José M. Vega-Pérez
- Departamento de Química Orgánica y Química Farmacéutica, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
| | - Fernando Iglesias-Guerra
- Departamento de Química Orgánica y Química Farmacéutica, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
| | - Ignacio Periñán
- Departamento de Química Orgánica y Química Farmacéutica, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
| | - Francisco J. Miñano
- Unidad de Farmacología Experimental y Clínica (UFEC), Hospital Universitario de Valme, Universidad de Sevilla, Sevilla, Spain
| | - Juan Bautista
- Departamento de Bioquímica y Biología Molecular, Facultad de Farmacia, Universidad de Sevilla, Sevilla, Spain
- * E-mail: (JDB); (MRG)
| | - Manuel Romero-Gómez
- Unidad de Gestión Clínica de Enfermedades Digestivas & Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Hospital Universitario de Valme, Sevilla, Spain
- * E-mail: (JDB); (MRG)
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Cabrera RA, Usry JL, Arrellano C, Nogueira ET, Kutschenko M, Moeser AJ, Odle J. Effects of creep feeding and supplemental glutamine or glutamine plus glutamate (Aminogut) on pre- and post-weaning growth performance and intestinal health of piglets. J Anim Sci Biotechnol 2013; 4:29. [PMID: 23916292 PMCID: PMC3765720 DOI: 10.1186/2049-1891-4-29] [Citation(s) in RCA: 45] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2013] [Accepted: 08/01/2013] [Indexed: 01/16/2023] Open
Abstract
BACKGROUND Creep feeding is used to stimulate piglet post-weaning feed consumption. L-Glutamine (GLN) is an important source of fuel for intestinal epithelial cells. The objective of this study was to determine the impact of creep feeding and adding GLN or AminoGut (AG; containing glutamine + glutamate) to pre- and post-weaning diets on pig performance and intestinal health. Litters (N = 120) were allotted to four treatments during 14-21 d of lactation: 1) No creep feed (NC, n = 45); 2) creep fed control diet (CFCD, n = 45); 3) creep fed 1% GLN (CFGLN, n = 15); 4) creep fed .88% AG (CFAG, n = 15). After weaning, the NC and CFCD groups were sub-divided into three groups (n = 15 each), receiving either a control nursery diet (NC-CD, CFCD-CD) or a diet supplemented with either GLN (NC-GLN, CFCD-GLN) or with AG (NC-AG, CFCD-AG). Litters that were creep fed with diets containing GLN or AG also were supplemented with those amino acids in the nursery diets (CFGLN-GLN, CFAG-AG). Glutamine was added at 1% in all three post-weaning diet phases and AG was added at .88% in phase 1 and 2 and at .66% in phase 3. RESULTS Feed conversion (feed/gain) showed means among treatment means close to significance (P = 0.056) and Tukey's test for pairwise mean comparisons showed that Pigs in the CFGLN-GLN group had the best feed conversion (feed/gain) in the first three-week period post-weaning, exceeding (P = 0.044) controls (CFCD-CD) by 34%. The NC-AG group had (P = 0.02) the greatest feed intake in the last three week of the study, exceeding controls (CFCD-CD) by 12%. CFGLN-GLN, CFCD-GLN and sow reared (SR) pigs had the greatest (P = 0.049) villi height exceeding the CFCD-AG group by 18%, 20% and 19% respectively. The CFAG-AG group had the deepest (P = 0.001) crypts among all treatments. CFGLN-GLN, CFCD-GLN and SR groups had the greatest (P = 0.001) number of cells proliferating (PCNA) exceeding those in the NC-CD group by 43%, 54% and 63% respectively. Sow reared pigs showed the greatest (P = 0.001) intestinal absorption capacity for xylose and mannitol. CONCLUSION Supplementation of creep feed and nursery diets with GLN and/or AminoGut in the first three week improved feed conversion possibly due to improved intestinal health.
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Affiliation(s)
- Rafael A Cabrera
- Laboratory of Developmental Nutrition, Department of Animal Science, North Carolina State University, 101 Polk Hall, North Carolina State University, Raleigh, NC 27695, USA
- Author current employment: Huvepharma USA, 525 West Park Drive Suite 230, Peachtree City, GA 30269, USA
| | - James L Usry
- Ajinomoto Heartland Lysine, Chicago, IL 60631, USA
- Author current employment: Micronutrients, 1550 Research Way, Indianapolis, IN 46231, USA
| | - Consuelo Arrellano
- Department of Statistics, North Carolina State University, Raleigh, NC 27695-8203, USA
| | - Eduardo T Nogueira
- Ajinomoto do Brasil. Ajinomoto Animal Nutrition, São Paulo, SP 04015-001, Brazil
| | - Marianne Kutschenko
- Ajinomoto do Brasil. Ajinomoto Animal Nutrition, São Paulo, SP 04015-001, Brazil
| | - Adam J Moeser
- Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695, USA
| | - Jack Odle
- Laboratory of Developmental Nutrition, Department of Animal Science, North Carolina State University, 101 Polk Hall, North Carolina State University, Raleigh, NC 27695, USA
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12
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Ghosh SP, Singh R, Chakraborty K, Kulkarni S, Uppal A, Luo Y, Kaur P, Pathak R, Kumar KS, Hauer-Jensen M, Cheema AK. Metabolomic changes in gastrointestinal tissues after whole body radiation in a murine model. MOLECULAR BIOSYSTEMS 2013; 9:723-31. [PMID: 23403731 PMCID: PMC3601576 DOI: 10.1039/c3mb25454b] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
Exposure to ionizing radiation (IR) elicits a set of complex biological responses involving gene expression and protein turnover that ultimately manifest as dysregulation of metabolic processes representing the cellular phenotype. Although radiation biomarkers have been reported in urine and serum, they are not informative about IR mediated tissue or organ specific injury. In the present study we report IR induced metabolic changes in gastrointestinal (GI) tissue of CD2F1 mice using ultra-performance liquid chromatography (UPLC) coupled with electrospray time-of-flight mass spectrometry. Post-radiation GI injury is a critical determinant of survival after exposure to IR. Our results show a distinct dose and time dependent response to GI tissue injury.
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Affiliation(s)
- Sanchita P Ghosh
- Armed Forces Radiobiology Research Institute, USUHS, Bethesda, MD 20889-5603, USA
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13
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Zhong X, Li W, Huang X, Wang Y, Zhang L, Zhou Y, Hussain A, Wang T. Effects of glutamine supplementation on the immune status in weaning piglets with intrauterine growth retardation. Arch Anim Nutr 2012; 66:347-56. [PMID: 22962945 DOI: 10.1080/1745039x.2012.683325] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Neonates with intrauterine growth retardation (IUGR) often suffer from impaired cellular immunity, and weaning may further aggravate adverse effects of IUGR on development and function of the immune system. In this study, we investigated effects of glutamine supplementation on immune status in the intestines of weaning pigs with IUGR, focusing on molecular mechanisms underlying altered immune response. Piglets with IUGR were weaned at 21 days of age and received orally 1.22 g alanine or 1 g glutamine per kg body weight every 12 h. Weight gain and intestinal weight of weaning piglets were increased by glutamine supplementation. Levels of serum IgG in piglets supplemented with glutamine were increased compared with Control piglets. The production of IL-1 and IL-8 in the serum and jejunum was decreased by glutamine supplementation, whereas the levels of IL-4 in the serum and the concentrations of IL-4 and IL-10 in the jejunum were increased. The expression of heat shock protein 70 (Hsp70) in the jejunum was increased by glutamine supplementation, but the degradation of inhibitor κB and the activity of nuclear factor-κB (NF-κB) were decreased. In conclusion, glutamine supplementation enhanced immune response in weaning piglets with IUGR. The effects of glutamine in IUGR are associated with increased Hsp70 expression and suppression of NF-κB activation.
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Affiliation(s)
- Xiang Zhong
- College of Animal Science and Technology , Nanjing Agricultural University, Nanjing, Jiangsu, China
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14
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Zhong X, Zhang XH, Li XM, Zhou YM, Li W, Huang XX, Zhang LL, Wang T. Intestinal growth and morphology is associated with the increase in heat shock protein 70 expression in weaning piglets through supplementation with glutamine. J Anim Sci 2011; 89:3634-42. [PMID: 21705630 DOI: 10.2527/jas.2010-3751] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
The objectives of this study were to determine the effects of oral Gln supplementation on growth performance, intestinal morphology, and expression of heat shock protein (Hsp) 70 in weaning piglets. A total of 65 piglets after weaning at 21 d of age (d 0) were used in this experiment. Five piglets were randomly selected and euthanized initially at d 0 to determine baseline values for the expression of Hsp70 in the small intestine. The remaining piglets were randomly assigned to 1 of 2 treatments and received 0 or 1 g of oral Gln/kg of BW every 12 h. After piglets were humanely killed at d 3, 7, and 14 postweaning, the duodenum, jejunum, and ileum of piglets were sampled to evaluate intestinal morphology and the expression and localization of Hsp70. The results indicated that oral Gln supplementation increased plasma concentrations of Gln compared with those in control piglets (P < 0.05). Average daily gain and ADFI were greater in piglets orally supplemented with Gln than in control piglets during the whole period (P < 0.05). The incidence of diarrhea in piglets orally supplemented with Gln was 24% less than (P = 0.064) that in control piglets at 8 to 14 d after weaning. The weights of the jejunum and ileum were greater in piglets orally supplemented with Gln compared with those of control piglets relative to BW on d 14 postweaning (P < 0.05). The villus height and the villus height:crypt depth ratio in the jejunum and the ileum were greater in piglets receiving oral Gln on d 14 postweaning (P < 0.05) than in control piglets. These results indicate that Gln supplementation can influence the intestinal morphology of weaned piglets. The expression of hsp70 mRNA and Hsp70 proteins in the duodenum and jejunum was greater in piglets supplemented with Gln than in control piglets (P < 0.05). However, Gln supplementation had no effect on the expression of hsp70 mRNA and Hsp70 proteins in the ileum. Moreover, the localization of Hsp70 in the cytoplasm indicated that Hsp70 has a cytoprotective role in epithelial cell function and structure. These results indicate that Gln supplementation may be beneficial for intestinal health and development and may thus mitigate diarrhea and improve growth performance. The protective mechanisms of Gln in the intestine may be associated with the increase in Hsp70 expression.
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Affiliation(s)
- X Zhong
- College of Animal Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
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15
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Transcriptome atlas of glutamine family amino acid metabolism-related genes in eight regenerating liver cell types. Cell Biol Int 2011; 34:1189-98. [PMID: 20716061 DOI: 10.1042/cbi20090352] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
To explore glutamine family amino acid metabolism of eight liver cell types in rat liver regeneration, eight kinds of rat regenerating liver cells were isolated by using the combination of Percoll density gradient centrifugation and immunomagnetic bead methods, then Rat Genome 230 2.0 Array was used to detect the expression profiles of the genes associated with metabolism of glutamine family amino acid in rat liver regeneration and finally how these genes involved in activities of eight regenerating liver cell types were analysed by the methods of bioinformatics and systems biology. The results showed that in the priming stage of liver regeneration, hepatic stellate cells and sinusoidal endothelial cells transformed proline and glutamine into glutamate; hepatocytes, hepatic stellate cells, sinusoidal endothelial cells and dendritic cells catabolized glutamate to 2-oxoglutarate or succinate; hepatic stellate cells and sinusoidal endothelial cells catalysed glutamate into glutamyl-tRNA for protein synthesis; urea cycle, which degraded from arginine, was enhanced in biliary epithelia cells, sinusoidal endothelial cells and dendritic cells; synthesis of polyamines from arginine was enhanced in biliary epithelia cells, sinusoidal endothelial cells, Kupffer cells and dendritic cells; the content of NO was increased in sinusoidal endothelial cells and dendritic cells; degradation of proline was enhanced in hepatocytes and biliary epithelia cells. In the progress stage, biliary epithelia cells converted glutamine into GMP and glucosamine 6-phosphate; oval cells converted glutamine into glucosamine 6-phosphate; hepatic stellate cells converted glutamine into NAD; the content of NO, which degraded from arginine, was increased in biliary epithelia cells, oval cells, pit cells and dendritic cells. In the termination stage, oval cells converted proline into glutamate; glutamate degradation, which degraded from arginine, was enhanced in hepatocytes and dendritic cells; the content of NO was increased in oval cells, sinusoidal endothelial cells, pit cells and dendritic cells. The synthesis of creatine phosphate was enhanced in hepatocytes, biliary epithelia cells, pit cells and dendritic cells in both progress and termination stages. In summary, glutamine family amino acid metabolism has some differences in liver regeneration in different liver cells.
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Wright G, Noiret L, Olde Damink SWM, Jalan R. Interorgan ammonia metabolism in liver failure: the basis of current and future therapies. Liver Int 2011; 31:163-75. [PMID: 20673233 DOI: 10.1111/j.1478-3231.2010.02302.x] [Citation(s) in RCA: 105] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/13/2023]
Abstract
Hepatic encephalopathy complicates the course of both acute and chronic liver disease and its treatment remains an unmet clinical need. Ammonia is thought to be central in its pathogenesis and remains an important target of current and future therapeutic approaches. In liver failure, the main detoxification pathway of ammonia metabolism is compromised leading to hyperammonaemia. In this situation, the other ammonia-regulating pathways in multiple organs assume important significance. The present review focuses upon interorgan ammonia metabolism in health and disease describing the role of the key enzymes, glutamine synthase and glutaminase. Better understanding of these alternative pathways are leading to the development of new therapeutic approaches.
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Affiliation(s)
- Gavin Wright
- UCL Institute of Hepatology, Division of Medicine, University College London, London, UK
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17
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Masola B, Ngubane NP. The activity of phosphate-dependent glutaminase from the rat small intestine is modulated by ADP and is dependent on integrity of mitochondria. Arch Biochem Biophys 2010; 504:197-203. [PMID: 20831857 DOI: 10.1016/j.abb.2010.09.002] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2010] [Revised: 08/29/2010] [Accepted: 09/01/2010] [Indexed: 11/20/2022]
Abstract
The effect of adenine nucleotides and phosphate on rat small intestine phosphate-dependent glutaminase (PDG) activity was investigated in intact mitochondria. Disruption of the integrity of mitochondria by sonication or freeze-thawing resulted in loss of enzyme activity. ADP was the strongest adenine nucleotide activator of the enzyme giving a V(max) that was over 5-fold of that for AMP or ATP. The sigmoid activation curve of PDG by ADP became hyperbolic in presence ATP. ADP also lowered the K(m) for glutamine and increased V(max) and these effects were further enhanced by the presence of ATP. Activation of PDG by phosphate and ADP was not completely additive suggesting some antagonism between the activators. There was no clear relationship between changing ATP/ADP ratios and PDG activity in presence of a constant concentration of phosphate. However, ratios of approximately 1:4 and 4:1 gave the highest and lowest activities, respectively. The pH dependence of PDG activity was affected by phosphate concentration and results suggest that the divalent ion is the activating species.
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Affiliation(s)
- B Masola
- Department of Biochemistry, University of KwaZulu-Natal, Durban, South Africa.
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18
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Varma S, Eskin MNA, Bird R, Dolenko B, Raju J, Ijare OB, Bezabeh T. Potential of magnetic resonance spectroscopy in assessing the effect of fatty acids on inflammatory bowel disease in an animal model. Lipids 2010; 45:843-54. [PMID: 20721632 DOI: 10.1007/s11745-010-3455-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2010] [Accepted: 07/29/2010] [Indexed: 12/12/2022]
Abstract
People with inflammatory bowel disease (IBD) are at risk for developing colorectal cancer, and this risk increases at a rate of 1% per year after 8-10 years of having the disease. Saturated and omega-6 polyunsaturated fatty acids (PUFAs) have been implicated in its causation. Conversely, omega-3 PUFAs may have the potential to confer therapeutic benefit. Since proton magnetic resonance spectroscopy ((1)H MRS) combined with pattern recognition methods could be a valuable adjunct to histology, the objective of this study was to analyze the potential of (1)H MRS in assessing the effect of dietary fatty acids on colonic inflammation. Forty male Sprague-Dawley rats were administered one of the following dietary regimens for 2 weeks: low-fat corn oil (omega-6), high-fat corn oil (omega-6), high-fat flaxseed oil (omega-3) or high-fat beef tallow (saturated fatty acids). Half of the animals were fed 2% carrageenan to induce colonic inflammation similar to IBD. (1)H MRS and histology were performed on ex vivo colonic samples, and the (1)H MR spectra were analyzed using a statistical classification strategy (SCS). The histological and/or MRS studies revealed that different dietary fatty acids modulate colonic inflammation differently, with high-fat corn oil being the most inflammatory and high-fat flaxseed oil the least inflammatory. (1)H MRS is capable of identifying the biochemical changes in the colonic tissue as a result of inflammation, and when combined with SCS, this technique accurately differentiated the inflamed colonic mucosa based on the severity of the inflammation. This indicates that MRS could serve as a valuable adjunct to histology in accurately assessing colonic inflammation. Our data also suggest that both the type and the amount of fatty acids in the diet are critical in modulating IBD.
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Affiliation(s)
- Sonal Varma
- National Research Council Institute for Biodiagnostics, 435 Ellice Ave., Winnipeg, MB, R3B 1Y6, Canada
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19
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Wilkinson DJ, Smeeton NJ, Watt PW. Ammonia metabolism, the brain and fatigue; revisiting the link. Prog Neurobiol 2010; 91:200-19. [PMID: 20138956 DOI: 10.1016/j.pneurobio.2010.01.012] [Citation(s) in RCA: 115] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2009] [Revised: 01/27/2010] [Accepted: 01/29/2010] [Indexed: 12/15/2022]
Abstract
This review addresses the ammonia fatigue theory in light of new evidence from exercise and disease studies and aims to provide a view of the role of ammonia during exercise. Hyperammonemia is a condition common to pathological liver disorders and intense or exhausting exercise. In pathology, hyperammonemia is linked to impairment of normal brain function and the onset of the neurological condition, hepatic encephalopathy. Elevated blood ammonia concentrations arise due to a diminished capacity for removal via the liver and lead to increased exposure of organs, such as the brain, to the toxic effects of ammonia. High levels of brain ammonia can lead to deleterious alterations in astrocyte morphology, cerebral energy metabolism and neurotransmission, which may in turn impact on the functioning of important signalling pathways within the neuron. Such changes are believed to contribute to the disturbances in neuropsychological function, in particular the learning, memory, and motor control deficits observed in animal models of liver disease and also patients with cirrhosis. Hyperammonemia in exercise occurs as a result of an increased production by contracting muscle, through adenosine monophosphate (AMP) deamination (the purine nucleotide cycle) and branched chain amino acid (BCAA) deamination prior to oxidation. Plasma concentrations of ammonia during exercise often achieve or exceed those measured in liver disease patients, resulting in increased cerebral uptake. In this article we propose that exercise-induced hyperammonemia may lead to concomitant disturbances in brain function, potentially through similar mechanisms underpinning pathology, which may impact on performance as fatigue or reduced function, especially during extreme exercise.
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Affiliation(s)
- Daniel J Wilkinson
- Department of Sport and Exercise Science, Chelsea School, University of Brighton, 30 Carlisle Road, Eastbourne, UK.
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20
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Taylor J, Cooper C, Mommsen T. Implications of GI function for gas exchange, acid–base balance and nitrogen metabolism. FISH PHYSIOLOGY 2010. [DOI: 10.1016/s1546-5098(10)03006-2] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
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21
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Blachier F, Boutry C, Bos C, Tomé D. Metabolism and functions of L-glutamate in the epithelial cells of the small and large intestines. Am J Clin Nutr 2009; 90:814S-821S. [PMID: 19571215 DOI: 10.3945/ajcn.2009.27462s] [Citation(s) in RCA: 219] [Impact Index Per Article: 13.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/11/2023] Open
Abstract
l-Glutamate is one of the most abundant amino acids in alimentary proteins, but its concentration in blood is among the lowest. This is largely because l-glutamate is extensively oxidized in small intestine epithelial cells during its transcellular journey from the lumen to the bloodstream and after its uptake from the bloodstream. This oxidative capacity coincides with a high energy demand of the epithelium, which is in rapid renewal and responsible for the nutrient absorption process. l-Glutamate is a precursor for glutathione and N-acetylglutamate in enterocytes. Glutathione is involved in the enterocyte redox state and in the detoxication process. N-acetylglutamate is an activator of carbamoylphosphate synthetase 1, which is implicated in l-citrulline production by enterocytes. Furthermore, l-glutamate is a precursor in enterocytes for several other amino acids, including l-alanine, l-aspartate, l-ornithine, and l-proline. Thus, l-glutamate can serve both locally inside enterocytes and through the production of other amino acids in an interorgan metabolic perspective. Intestinal epithelial cell capacity to oxidize l-glutamine and l-glutamate is already high in piglets at birth and during the suckling period. In colonocytes, l-glutamate also serves as a fuel but is provided from the bloodstream. Alimentary and endogenous proteins that escape digestion enter the large intestine and are broken down by colonic bacterial flora, which then release l-glutamate into the lumen. l-Glutamate can then serve in the colon lumen as a precursor for butyrate and acetate in bacteria. l-Glutamate, in addition to fiber and digestion-resistant starch, can thus serve as a luminally derived fuel precursor for colonocytes.
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Affiliation(s)
- François Blachier
- INRA, CRNH-IdF, UMR Nutrition Physiology and Ingestive Behavior, Paris, France.
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22
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Odongo NE, Greenwood SL, Or-Rashid MM, Radford D, AlZahal O, Shoveller AK, Lindinger MI, Matthews JC, McBride BW. Effects of nutritionally induced metabolic acidosis with or without glutamine infusion on acid-base balance, plasma amino acids, and plasma nonesterified fatty acids in sheep1. J Anim Sci 2009; 87:1077-84. [DOI: 10.2527/jas.2008-1165] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
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Romero-Gómez M, Jover M, Galán JJ, Ruiz A. Gut ammonia production and its modulation. Metab Brain Dis 2009; 24:147-57. [PMID: 19067141 DOI: 10.1007/s11011-008-9124-3] [Citation(s) in RCA: 59] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/02/2008] [Accepted: 10/28/2008] [Indexed: 12/12/2022]
Abstract
Systemic hyperammonemia has been largely found in patients with cirrhosis and hepatic encephalopathy, and ammonia plays a major role in the pathogenesis of hepatic encephalopathy. However, controversial points remain: a) the correlation between plasma ammonia levels and neurophysiological impairment. The lack of correlation between ammonia levels and grade of hepatic encephalopathy in some cases has been considered a weakness of the ammonia hypothesis, but new methods for ammonia measurements and the implication of systemic inflammation in the modulation of ammonia neurotoxicity could explain this gap; b) the source of ammonia production. Hyperammonemia has been considered as derived from urea breakdown by intestinal bacteria and the majority of treatments were targeted against bacteria-derived ammonia from the colon. However, some data suggest an important role for small intestine ammonia production: 1) the hyperammonemia after porto-caval shunted rats has been found similar in germ-free than in non-germ-free animals. 2) In cirrhotic patients the greatest hyperammonemia was found in portal drained viscera and derived mainly from glutamine deamination. 3) The amount of time required to increase of ammonia (less than one hour) after oral glutamine challenge supports a small intestine origin of the hyperammonemia. As the main source of ammonia in cirrhotics derives from portal drained viscera owing to glutamine deamidation, increased glutaminase activity in the intestine seems to be responsible for systemic hyperammonemia. Lastly, some genetic alterations in the glutaminase gene such as the haplotype TACC could modulate intestinal ammonia production and the risk of overt hepatic encephalopathy in cirrhotics.
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Affiliation(s)
- Manuel Romero-Gómez
- Unit for the Clinical Management of Digestive Diseases & ciberehd, Hospital Universitario de Valme, Universidad de Sevilla, 41014 Sevilla, Spain.
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24
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Xue H, Sawyer MB, Field CJ, Dieleman LA, Baracos VE. Nutritional modulation of antitumor efficacy and diarrhea toxicity related to irinotecan chemotherapy in rats bearing the ward colon tumor. Clin Cancer Res 2008; 13:7146-54. [PMID: 18056195 DOI: 10.1158/1078-0432.ccr-07-0823] [Citation(s) in RCA: 58] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
PURPOSE To evaluate and compare the influence of dietary elements on cancer progression, chemotherapy efficacy, and toxicity, particularly severe, late-onset diarrhea related to irinotecan (CPT-11) treatment. EXPERIMENTAL DESIGN We used laboratory rats fed a standardized basal diet, Ward colon tumor, and CPT-11 therapy for the study of CPT-11-induced diarrhea. Dietary interventions were selected from nutrients already established to modify other forms of colitis and which have been hypothesized to mitigate chemotherapy-induced gastrointestinal injury (glutamine, n-3 fatty acids, prebiotic oligosaccharides). Animals adapted to test diets were treated with CPT-11 at the maximum tolerated dose (125 mg/kg x 3 days) and diarrhea was followed continuously for 1 week. RESULTS The inclusion of n-3 fatty acids in the diet (5%, w/w of total fat) suppressed tumor growth and enhanced CPT-11's efficacy; this treatment did not affect the incidence or severity of diarrhea. By contrast, oral glutamine bolus (0.75 g/kg) administered prior to each CPT-11 treatment reduced the incidence of severe diarrhea (34.1 +/- 4.7% versus 53.8 +/- 4.2%, P < 0.005) and decreased the area under the curve of diarrhea score (16.5 +/- 1.0 versus 18.8 +/- 0.5, P < 0.05). Identical results were obtained with i.v. bolus glutamine administration. Glutamine treatment did not alter CPT-11's antitumor efficacy. The addition of prebiotic oligosaccharides to the diet (8%, w/w of diet) did not mitigate the severity of diarrhea, and it raised the activity of beta-glucuronidase in cecal contents, a key bacterial enzyme mediating CPT-11-related intestinal toxicity. CONCLUSION Our experiments suggest that glutamine and n-3 fatty acids might be potentially useful adjuncts to CPT-11 treatment.
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Affiliation(s)
- Hongyu Xue
- Department of Oncology, University of Alberta, Edmonton, Alberta, Canada
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25
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Niva CC, Lee JM, Myohara M. Glutamine synthetase gene expression during the regeneration of the annelid Enchytraeus japonensis. Dev Genes Evol 2008; 218:39-46. [PMID: 18183418 PMCID: PMC2265772 DOI: 10.1007/s00427-007-0198-4] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2007] [Accepted: 11/27/2007] [Indexed: 11/29/2022]
Abstract
Enchytraeus japonensis is a highly regenerative oligochaete annelid that can regenerate a complete individual from a small body fragment in 4–5 days. In our previous study, we performed complementary deoxyribonucleic acid subtraction cloning to isolate genes that are upregulated during E. japonensis regeneration and identified glutamine synthetase (gs) as one of the most abundantly expressed genes during this process. In the present study, we show that the full-length sequence of E. japonensis glutamine synthetase (EjGS), which is the first reported annelid glutamine synthetase, is highly similar to other known class II glutamine synthetases. EjGS shows a 61–71% overall amino acid sequence identity with its counterparts in various other animal species, including Drosophila and mouse. We performed detailed expression analysis by in situ hybridization and reveal that strong gs expression occurs in the blastemal regions of regenerating E. japonensis soon after amputation. gs expression was detectable at the cell layer covering the wound and was found to persist in the epidermal cells during the formation and elongation of the blastema. Furthermore, in the elongated blastema, gs expression was detectable also in the presumptive regions of the brain, ventral nerve cord, and stomodeum. In the fully formed intact head, gs expression was also evident in the prostomium, brain, the anterior end of the ventral nerve cord, the epithelium of buccal and pharyngeal cavities, the pharyngeal pad, and in the esophageal appendages. In intact E. japonensis tails, gs expression was found in the growth zone in actively growing worms but not in full-grown individuals. In the nonblastemal regions of regenerating fragments and in intact worms, gs expression was also detected in the nephridia, chloragocytes, gut epithelium, epidermis, spermatids, and oocytes. These results suggest that EjGS may play roles in regeneration, nerve function, cell proliferation, nitrogenous waste excretion, macromolecule synthesis, and gametogenesis.
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Affiliation(s)
- Cintia Carla Niva
- Invertebrate Gene Function Research Unit, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan.
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Nieto R, Obitsu T, Fernández-Quintela A, Bremner D, Milne E, Calder AG, Lobley GE. Glutamine metabolism in ovine splanchnic tissues: effects of infusion of ammonium bicarbonate or amino acids into the abomasum. Br J Nutr 2007. [DOI: 10.1079/bjn2002525] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
This study investigates the effects of increased NH3or amino acid supply on glutamine utilisation and production by the splanchnic tissues of fed sheep. Six sheep, prepared with vascular catheters in the aorta, mesenteric, portal and hepatic veins, were fed grass pellets to 1·1×energy maintenance requirements. Each treatment involved a 4 d abomasal infusion, of either ammonium bicarbonate (AMM; 23·4 μmol/kg0·75per min), water (CONT), or a mixture of amino acids that excluded glutamine and glutamate (AA; 46·8 μmol amino acid-N/kg0·75per min). The treatments simulated nutritional extremes in terms of the balance of absorbed N. Kinetics across the whole gut and the liver were monitored during an intra-jugular infusion of [5-15N]glutamine. Blood flow across the whole gut or liver were unaffected by treatment. Both AMM and AA infusions doubled the hepatic release of urea-N compared with CONT (P<0·02). AA infusion decreased arterial glutamine concentration by 26 % (P<0·01) and 23 % (P<0·05) compared with AMM and CONT respectively. Despite this, whole-body glutamine flux was not affected by treatment. In contrast, AMM infusion increased hepatic glutamine production by 40 % compared with CONT (P<0·02). This provided a mechanism to ensure NH3supply to the periphery was maintained within the normal low physiological levels. Hepatic glutamine utilisation tended to increase during AA infusion, probably to ensure equal inflows of N to the ornithine cycle. Between 6 and 10 % of NH3absorbed across the digestive tract was derived from the amido-N of glutamine. Overall, splanchnic glutamine utilisation accounted for 45–70 % of whole-body glutamine flux.
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Martin G, Ferrier B, Conjard A, Martin M, Nazaret R, Boghossian M, Saadé F, Mancuso C, Durozard D, Baverel G. Glutamine gluconeogenesis in the small intestine of 72 h-fasted adult rats is undetectable. Biochem J 2007; 401:465-73. [PMID: 17002601 PMCID: PMC1820798 DOI: 10.1042/bj20061148] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/02/2023]
Abstract
Recent reports have indicated that 48-72 h of fasting, Type 1 diabetes and high-protein feeding induce gluconeogenesis in the small intestine of adult rats in vivo. Since this would (i) represent a dramatic revision of the prevailing view that only the liver and the kidneys are gluconeogenic and (ii) have major consequences in the metabolism, nutrition and diabetes fields, we have thoroughly re-examined this question in the situation reported to induce the highest rate of gluconeogenesis. For this, metabolically viable small intestinal segments from 72 h-fasted adult rats were incubated with [3-13C]glutamine as substrate. After incubation, substrate utilization and product accumulation were measured by enzymatic and NMR spectroscopic methods. Although the segments utilized [13C]glutamine at high rates and accumulated 13C-labelled products linearly for 30 min in vitro, no substantial glucose synthesis could be detected. This was not due to the re-utilization of [13C]glucose initially synthesized from [13C]glutamine. Arteriovenous metabolite concentration difference measurements across the portal vein-drained viscera of 72 h-fasted Wistar and Sprague-Dawley rats clearly indicated that glutamine, the main if not the only gluconeogenic precursor taken up, could not give rise to detectable glucose production in vivo. Therefore we challenge the view that the small intestine of the adult rat is a gluconeogenic organ.
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Affiliation(s)
- Guy Martin
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Bernard Ferrier
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Agnès Conjard
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Mireille Martin
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Rémi Nazaret
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Michelle Boghossian
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Fadi Saadé
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Claire Mancuso
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Daniel Durozard
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
| | - Gabriel Baverel
- Institut National de la Santé et de la Recherche Médicale, UMR 499, Animet, Faculté de Médecine RTH Laennec, Université Lyon 1, Rue G. Paradin, 69372 Lyon Cedex 08, France
- To whom correspondence should be addressed (email )
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van Straaten HWM, He Y, van Duist MM, Labruyère WT, Vermeulen JLM, van Dijk PJ, Ruijter JM, Lamers WH, Hakvoort TBM. Cellular concentrations of glutamine synthetase in murine organs. Biochem Cell Biol 2006; 84:215-31. [PMID: 16609703 DOI: 10.1139/o05-170] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
Glutamine synthetase (GS) is the only enzyme that can synthesize glutamine, but it also functions to detoxify glutamate and ammonia. Organs with high cellular concentrations of GS appear to function primarily to remove glutamate or ammonia, whereas those with a low cellular concentration appear to primarily produce glutamine. To validate this apparent dichotomy and to clarify its regulation, we determined the GS concentrations in 18 organs of the mouse. There was a >100-fold difference in GS mRNA, protein, and enzyme-activity levels among organs, whereas there was only a 20-fold difference in the GS protein:mRNA ratio, suggesting extensive transcriptional and posttranscriptional regulation. In contrast, only small differences in the GS enzyme activity : protein ratio were found, indicating that posttranslational regulation is of minor importance. The cellular concentration of GS was determined by relating the relative differences in cellular GS concentration, detected using image analysis of immunohistochemically stained tissue sections, to the biochemical data. There was a >1000-fold difference in cellular concentrations of GS between GS-positive cells in different organs, and cellular concentrations were up to 20x higher in subpopulations of cells within organs than in whole organs. GS activity was highest in pericentral hepatocytes (approximately 485 micromol.g(-1).min-(1), followed in descending order by epithelial cells in the epididymal head, Leydig cells in the testicular interstitium, epithelial cells of the uterine tube, acid-producing parietal cells in the stomach, epithelial cells of the S3 segment of the proximal convoluted tubule of the kidney, astrocytes of the central nervous tissue, and adipose tissue. GS activity in muscle amounted to only 0.4 micromol.g(-1).min(-1). Our findings confirmed the postulated dichotomy between cellular concentration and GS function.
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Romero-Gomez M, Jover M, Diaz-Gomez D, de Teran LC, Rodrigo R, Camacho I, Echevarria M, Felipo V, Bautista JD. Phosphate-activated glutaminase activity is enhanced in brain, intestine and kidneys of rats following portacaval anastomosis. World J Gastroenterol 2006; 12:2406-11. [PMID: 16688834 PMCID: PMC4088079 DOI: 10.3748/wjg.v12.i15.2406] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To assess whether portacaval anastomosis (PCA) in rats affects the protein expression and/or activity of glutaminase in kidneys, intestines and in three brain areas of cortex, basal ganglia and cerebellum and to explain the neurological alterations found in hepatic encephalopathy (HE).
METHODS: Sixteen male Wistar rats weighing 250-350 g were grouped into sham-operation control (n = 8) or portacaval shunt (n = 8). Twenty-eight days after the procedure, the animals were sacrificed. The duodenum, kidney and brain were removed, homogenised and mitochondria were isolated. Ammonia was measured in brain and blood. Phosphate-activated glutaminase (PAG) activity was determined by measuring ammonia production following incubation for one hour at 37 °C with O-phthalaldehyde (OPA) and specific activity expressed in units per gram of protein (µkat/g of protein). Protein expression was measured by immunoblotting.
RESULTS: Duodenal and kidney PAG activities together with protein content were significantly higher in PCA group than in control or sham-operated rats (duodenum PAG activity was 976.95±268.87 µkat/g of protein in PCA rats vs 429.19±126.92 µkat/g of protein in sham-operated rats; kidneys PAG activity was 1259.18 ± 228.79 µkat/g protein in PCA rats vs 669.67± 400.8 µkat/g of protein in controls, P < 0.05; duodenal protein content: 173% in PCA vs sham-operated rats; in kidneys the content of protein was 152% in PCA vs sham-operated rats). PAG activity and protein expression in PCA rats were higher in cortex and basal ganglia than those in sham-operated rats (cortex: 6646.6 ± 1870.4 µkat/g of protein vs 3573.8 ± 2037.4 µkat/g of protein in control rats, P < 0.01; basal ganglia, PAG activity was 3657.3 ± 1469.6 μkat/g of protein in PCA rats vs 2271.2 ± 384 μkat/g of protein in sham operated rats, P < 0.05; In the cerebellum, the PAG activity was 2471.6 ± 701.4 μkat/g of protein vs 1452.9 ± 567.8 μkat/g of protein in the PCA and sham rats, respectively, P < 0.05; content of protein: cerebral cortex: 162% ± 40% vs 100% ± 26%, P < 0.009; and basal ganglia: 140% ± 39% vs 100% ± 14%, P < 0.05; but not in cerebellum: 100% ± 25% vs 100% ± 16%, P = ns).
CONCLUSION: Increased PAG activity in kidney and duodenum could contribute significantly to the hyperammonaemia in PCA rats, animal model of encephalopathy. PAG is increased in non-synaptic mitochondria from the cortex and basal ganglia and could be implicated in the pathogenesis of hepatic encephalopathy. Therefore, PAG could be a possible target for the treatment of HE or liver dysfunction.
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Affiliation(s)
- Manuel Romero-Gomez
- Unit for Clinical Management of Digestive Diseases, Hospital Universitario de Valme, ctra Cadiz s/n, 41014 Seville, Spain.
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Abstract
Disturbed body nitrogen homeostasis due to impaired hepatic urea synthesis leads to an alteration in inter-organ ammonia trafficking, resulting in hyperammonemia. Glutamine (Gln) synthase is the alternative pathway for ammonia detoxification. Gln taken up by several organs is split by the intramitochondrial phosphate-activated enzyme glutaminase (PAG) into glutamate (Glu) and ammonia. In cirrhotic patients with portosystemic intrahepatic shunt, the main source of systemic hyperammonemia is the small intestine, and ammonia derives mainly from Gln deamidation. Recently, PAG has been found increased in cirrhotics showing minimal hepatic encephalopathy and, therefore, could be implicated in the production of systemic hyperammonemia in these patients. Intestinal PAG activity correlates with psychometric test and magnetic resonance spectroscopy findings. Moreover, nitric oxide and tumor necrosis factor seem to be the major factors regulating intestinal ammonia production in cirrhotics. In the brain, PAG localized into the astrocytes is responsible for ammonia and free-radical production. The blockade of PAG, using 6-oxo-5-norleucine, avoids the toxic effects of Gln accumulation in the brain. These data support an important role for intestinal and brain glutaminase in the pathogenesis of hepatic encephalopathy and could be a new target for future therapies.
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Boelens PG, van Leeuwen PAM, Dejong CHC, Deutz NEP. Intestinal renal metabolism of L-citrulline and L-arginine following enteral or parenteral infusion of L-alanyl-L-[2,15N]glutamine or L-[2,15N]glutamine in mice. Am J Physiol Gastrointest Liver Physiol 2005; 289:G679-85. [PMID: 15933220 DOI: 10.1152/ajpgi.00026.2005] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Previously, we observed increased plasma arginine (ARG) concentrations after glutamine (GLN)-enriched diets, in combination with clinical benefits. GLN delivers nitrogen for ARG synthesis, and the present study was designed to quantify the interorgan relationship of exogenous L-GLN or GLN dipeptide, by enteral or parenteral route, contributing to intestinal citrulline (CIT) and renal de novo ARG synthesis in mice. To study this, we used a multicatheterized mouse model with Swiss mice (n = 43) in the postabsorptive state. Stable isotopes were infused into the jugular vein or into the duodenum {per group either free L-[2,(15)N]GLN or dipeptide L-ALA-L-[2,(15)N]GLN, all with L-[ureido-(13)C-(2)H(2)]CIT and L-[guanidino-(15)N(2)-(2)H(2)]ARG} to establish renal and intestinal ARG and CIT metabolism. Blood flow was measured using (14)C-para-aminohippuric acid. Net intestinal CIT release, renal uptake of CIT, and net renal ARG efflux was found, as assessed by arteriovenous flux measurements. Quantitatively, more de novo L-[2,(15)N]CIT was produced when free L-[2,(15)N]GLN was given than when L-ALA-L-[2,(15)N]GLN was given, whereas renal de novo L-[2,(15)N]ARG was similar in all groups. In conclusion, the intestinal-renal axis is hereby proven in mice in that L-[2,(15)N]GLN or dipeptide were both converted into de novo renal L-[2,(15)N]ARG; however, not all was derived from intestinal L-[2,(15)N]CIT production. In this model, the feeding route and form of GLN did not influence de novo renal ARG production derived from GLN.
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Affiliation(s)
- Petra G Boelens
- Dept. of Surgery, Vrije Universiteit University Medical Center, 1007 MB Amsterdam, The Netherlands
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Lenaerts K, Mariman E, Bouwman F, Renes J. Differentiation stage-dependent preferred uptake of basolateral (systemic) glutamine into Caco-2 cells results in its accumulation in proteins with a role in cell-cell interaction. FEBS J 2005; 272:3350-64. [PMID: 15978041 DOI: 10.1111/j.1742-4658.2005.04750.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Glutamine is an essential amino acid for enterocytes, especially in states of critical illness and injury. In several studies it has been speculated that the beneficial effects of glutamine are dependent on the route of supply (luminal or systemic). The aim of this study was to investigate the relevance of both routes of glutamine delivery to in vitro intestinal cells and to explore the molecular basis for proposed beneficial glutamine effects: (a) by determining the relative uptake of radiolabelled glutamine in Caco-2 cells; (b) by assessing the effect of glutamine on the proteome of Caco-2 cells using a 2D gel electrophoresis approach; and (c) by examining glutamine incorporation into cellular proteins using a new mass spectrometry-based method with stable isotope labelled glutamine. Results of this study show that exogenous glutamine is taken up by Caco-2 cells from both the apical and the basolateral side. Basolateral uptake consistently exceeds apical uptake and this phenomenon is more pronounced in 5-day-differentiated cells than in 15-day-differentiated cells. No effect of exogenous glutamine supply on the proteome was detected. However, we demonstrated that exogenous glutamine is incorporated into newly synthesized proteins and this occurred at a faster rate from basolateral glutamine, which is in line with the uptake rates. Interestingly, a large number of rapidly labelled proteins is involved in establishing cell-cell interactions. In this respect, our data may point to a molecular basis for observed beneficial effects of glutamine on intestinal cells and support results from studies with critically ill patients where parenteral glutamine supplementation is preferred over luminal supplementation.
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Affiliation(s)
- Kaatje Lenaerts
- Maastricht Proteomics Center, Nutrition and Toxicology Research Institute Maastricht (NUTRIM), Department of Human Biology, Maastricht University, the Netherlands.
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Handlogten ME, Hong SP, Zhang L, Vander AW, Steinbaum ML, Campbell-Thompson M, Weiner ID. Expression of the ammonia transporter proteins Rh B glycoprotein and Rh C glycoprotein in the intestinal tract. Am J Physiol Gastrointest Liver Physiol 2005; 288:G1036-47. [PMID: 15576624 DOI: 10.1152/ajpgi.00418.2004] [Citation(s) in RCA: 82] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Ammonia metabolism is important in multiple aspects of gastrointestinal physiology, but the mechanisms of ammonia transport in the gastrointestinal tract remain incompletely defined. The present study examines expression of the ammonia transporter family members Rh B glycoprotein (RhBG) and Rh C glycoprotein (RhCG) in the mouse gastrointestinal tract. Real-time RT-PCR amplification and immunoblot analysis identified mRNA and protein for both RhBG and RhCG were expressed in stomach, duodenum, jejunum, ileum, and colon. Immunohistochemistry showed organ and cell-specific expression of both RhBG and RhCG. In the stomach, both RhBG and RhCG were expressed in the fundus and forestomach, but not in the antrum. In the forestomach, RhBG was expressed by all nucleated squamous epithelial cells, whereas RhCG was expressed only in the stratum germinativum. In the fundus, RhBG and RhCG immunoreactivity was present in zymogenic cells but not in parietal or mucous cells. Furthermore, zymogenic cell RhBG and RhCG expression was polarized, with apical RhCG and basolateral RhBG immunoreactivity. In the duodenum, jejunum, ileum, and colon, RhBG and RhCG immunoreactivity was present in villous, but not in mucous or crypt cells. Similar to the fundic zymogenic cell, RhBG and RhCG expression in villous epithelial cells was polarized when apical RhCG and basolateral RhBG immunoreactivity was present. Thus the ammonia transporting proteins RhBG and RhCG exhibit cell-specific, axially heterogeneous, and polarized expression in the intestinal tract suggesting they function cooperatively to mediate gastrointestinal tract ammonia transport.
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Affiliation(s)
- Mary E Handlogten
- Medical Service, North Florida/South Georgia Veterans Health System, USA
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Arnaud A, Ramírez M, Baxter JHJH, Angulo AJAJ. Absorption of enterally administered N-acetyl-l-glutamine versus glutamine in pigs. Clin Nutr 2004; 23:1303-12. [PMID: 15556252 DOI: 10.1016/j.clnu.2004.04.004] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2003] [Accepted: 04/05/2004] [Indexed: 11/27/2022]
Abstract
BACKGROUND AND AIMS Glutamine instability in liquid media suggests that evaluation of reasonable enteral nutrition sources of glutamine is needed. N-acetyl-l-glutamine offers no instability and no intolerance problems. This research was conducted to study the absorption and apparent digestibility of glutamine versus N-acetyl-l-glutamine. METHODS Two pig models were used. (1) In a clamped jejunal loop experiment, we measured the concentrations of glutamine and N-acetyl-l-glutamine in the intestinal infused solutions, intestinal mucosa, and portal and peripheral blood. (2) In a feeding experiment, we determined their apparent digestibility. RESULTS N-acetyl-l-glutamine ( approximately 76%) was slightly less absorbed than glutamine ( approximately 85%) from the intestinal lumen into mucosa, where it was not detected as intact molecule, suggesting almost complete hydrolysis during absorption. Virtually no intact N-acetyl-l-glutamine was observed in the blood compartments; glutamine from lumenal N-acetyl-l-glutamine had the same behavior as that from lumenal-free glutamine in portal and peripheral blood. The apparent ileal digestibility of N-acetyl-l-glutamine was lower than that of free glutamine, as N-acetyl-l-glutamine was probably retained in the intestinal lumen to a greater extent than glutamine. CONCLUSION N-acetyl-l-glutamine appeared to be a good candidate for glutamine fortification of enteral nutrition formulas.
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Affiliation(s)
- Alexandra Arnaud
- Abbott Laboratories, Ross Products Division, International R and D Department, Camino de Purchil 68, Granada 18004, Spain
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Tuhacek LM, Mackey AD, Li N, DeMarco VG, Stevens G, Neu J. Substitutes for glutamine in proliferation of rat intestinal epithelial cells. Nutrition 2004; 20:292-7. [PMID: 14990271 DOI: 10.1016/j.nut.2003.11.003] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
OBJECTIVES Glutamine (Gln) is important for intestinal epithelial proliferation. The purpose of this study was to determine whether glutamate (Glu), a mixture of nucleotide monophosphates, arginine, or glucosamine could support proliferation of rat intestinal crypt cells (IEC-6) in the absence of Gln. METHODS Glu with added ammonia acetate, glucosamine, arginine, and nucleotide monophosphates were tested at concentrations that were isonitrogenous with respect to Gln. To determine whether de novo synthesis of Gln was affected by these nutrients, a duplicate set of treatment groups was also tested with 1.0 mM/L of methionine sulfoximine, an inhibitor of Gln synthetase. RESULTS Gln + methionine sulfoximine-treated cells showed suboptimal proliferation below 0.6 mM/L but normal proliferation between 0.6 and 4.0 mM/L of Gln. In the absence of exogenous Gln, isonitrogenous concentrations of Glu, glucosamine, arginine, or nucleotide monophosphates yielded similar proliferation as Gln. Cells treated with Glu, glucosamine, arginine, or nucleotide monophosphate mixture showed a decrease in proliferation compared with cells treated with Gln across all treatment doses (P < 0.03). CONCLUSIONS The importance of these results is that, in the presence of active Gln synthetase, these nutrients can maintain intestinal epithelial proliferation similar to that observed with Gln.
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Affiliation(s)
- Lauren M Tuhacek
- Department of Pediatrics, University of Florida College of Medicine, Gainesville, Florida, USA
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Romero-Gómez M, Ramos-Guerrero R, Grande L, de Terán LC, Corpas R, Camacho I, Bautista JD. Intestinal glutaminase activity is increased in liver cirrhosis and correlates with minimal hepatic encephalopathy. J Hepatol 2004; 41:49-54. [PMID: 15246207 DOI: 10.1016/j.jhep.2004.03.021] [Citation(s) in RCA: 80] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/01/2003] [Revised: 02/08/2004] [Accepted: 03/26/2004] [Indexed: 12/13/2022]
Abstract
BACKGROUND/AIMS We performed the current study to assess the intestinal activity of enterocyte phosphate-activated glutaminase (PAG) in cirrhosis. METHODS Forty-nine cirrhotic patients and 36 control subjects underwent endoscopic duodenal biopsies. Minimal hepatic encephalopathy (MHE) was evaluated using three psychometric tests. Oral glutamine challenge (OGC) was performed and MELD, Child-Pugh and the presence of esophageal varices were recorded. PAG was measured by enzymatic methods. Cerebral magnetic resonance spectroscopy was performed in 10 cirrhotics. RESULTS PAG was found to be higher in cirrhotics than control subjects 2.4+/-1.51 vs. 0.68+/-0.57IU/mg protein (P<0.001). PAG was also increased in patients with MHE and correlated with MELD, INR, esophageal varices and serum bile acids. A negative correlation was observed between PAG activity and intra-cerebral choline/creatine ratio (r=-0.67; P=0.035) and a positive correlation with glutamine plus glutamate/creatine ratio (r=0.78; P=0.007). In multivariate analysis using backward logistic regression, presence of MHE was the only variable independently related to altered enterocyte PAG. CONCLUSIONS Enterocyte PAG is increased in cirrhotic patients and correlates with MHE. These data support a possible role for intestinal glutaminase in the pathogenesis of hepatic encephalopathy (HE) and could be a new target for future therapies.
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Affiliation(s)
- Manuel Romero-Gómez
- Hepatology Unit, Hospital Universitario de Valme, Ctra Cádiz s/n, 41014 Sevilla, Spain.
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Mommsen TP, Busby ER, von Schalburg KR, Evans JC, Osachoff HL, Elliott ME. Glutamine synthetase in tilapia gastrointestinal tract: zonation, cDNA and induction by cortisol. J Comp Physiol B 2003; 173:419-27. [PMID: 12783264 DOI: 10.1007/s00360-003-0350-z] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 04/01/2003] [Indexed: 10/26/2022]
Abstract
Glutamine synthetase, an enzyme generally associated with ammonia detoxication in the vertebrate brain and with hepatic nitrogen turnover in mammals, shows substantial activities in the gastrointestinal tract of teleostean fishes. Enzyme activity is highest in the central area of the stomach and reveals a distinct distribution pattern in stomach and along the intestine of tilapia (Oreochromis niloticus), rainbow trout (Oncorhynchus mykiss) and copper rockfish (Sebastes caurinus). In all three species, intestinal activity peaks in the distal region of the intestine. The brain contains the highest titre of the enzyme (46 U g(-1) in tilapia brain versus 15 U g(-1) in tilapia stomach), but because of the relative mass of the stomach, the largest glutamine synthetase pool in tilapia body appears to be localized in the stomach. Activities in white and red muscle are very modest at 0.1% of the brain. Independent of distribution, peak activities of glutamine synthetase in selected areas of tilapia stomach and intestine are significantly (two- to fourfold) increased after a 5-day treatment with an intraperitoneal cortisol deposit. Cortisol also increases glutamine synthetase activity in tilapia liver, white and red muscle, while activities in brain remain unaffected. We cloned and sequenced the predominant transcript of tilapia stomach glutamine synthetase (about 1.9 kb), encoding a 371-amino acid peptide. The open reading frame shows considerable identity with glutamine synthetase in toadfish (92% at peptide level, 87% at nucleotide level), but possesses a longer 3'-untranslated region than the toadfish. The tilapia glutamine synthetase mRNA contains a remnant of a putative mitochondrial leader sequence, but without a conserved second site for initiation of translation. We also find evidence for additional transcripts of glutamine synthetase in tilapia, suggesting multiple genes. Finally, we present evidence for similar abundance of glutamine synthetase transcripts in all regions of rockfish intestine. The physiological significance of the presence of glutamine synthetase in teleostean intestine is discussed.
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Affiliation(s)
- T P Mommsen
- Department of Biochemistry and Microbiology, University of Victoria, P.O. Box 3055, Victoria, B.C., V8W 3P6, Canada.
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Abstract
Since the pioneering work of Windmueller and Spaeth, the importance of glutamine to the support of intestinal mucosal metabolic function has become generally accepted. Nevertheless, the mechanisms underlying this role still remain obscure. This paper explores a number of questions: 1) Is glutamine essential for intestinal function? 2) To what extent does this relate to its intermediary metabolism? 3) What is the importance of glutamine as a biosynthetic precursor? 4) Is glutamine supplementation of the nutrient mixture presented to patients of any metabolic or clinical benefit? As a result of this exploratory exercise, the following general conclusions were reached: 1) Much suggestive biochemical and physiologic evidence exists that implies that glutamine, especially systemic glutamine, supports the function of the intestinal mucosal system. 2) Despite the extensive metabolism of this amino acid by the intestinal tissues, most evidence suggests that if glutamine does play a physiologic role in the bowel, it is not compellingly related to its intermediary metabolism. 3) There is, on the other hand, evidence that the mucosal cells not only utilize extracellular glutamine but synthesize the amino acid. Given that inhibition of glutamine synthesis inhibits both proliferation and differentiation of mucosal cell cultures, this suggests some more subtle regulatory role. This notion is supported by the demonstration that glutamine will activate a number of genes associated with cell cycle progression in the mucosa. 4) Despite the accumulated evidence, the mechanisms underlying glutamine's function and the question whether glutamine supplementation uniformly benefits mucosal health remain equivocal at best.
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Affiliation(s)
- P J Reeds
- Department of Animal Sciences, University of Illinois, Urbana, IL 61801, USA.
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Abstract
The amino acid glutamine has become one of the most intensively studied nutrients in the field of nutrition and metabolic support. A variety of studies in cell culture systems, animal models of gut mucosal atrophy, injury/repair and adaptation and a limited number of clinical trials demonstrate trophic and cytoprotective effects of glutamine in small bowel and colonic mucosal cells. Although the routine clinical use of glutamine-enriched parenteral and enteral nutrient solutions remains controversial, available data demonstrate both the safety and metabolic and clinical efficacy of glutamine treatment in selected patient groups. Basic investigations are elucidating underlying mechanisms of glutamine action in intestinal cells. These will inform preclinical and clinical investigations designed to determine glutamine efficacy in selected gastrointestinal disorders. Emerging clinical trials will further define the utility of adjunctive glutamine supplementation as a component of specialized nutrition support in gastrointestinal disease.
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Affiliation(s)
- T R Ziegler
- Department of Medicine, Emory University School of Medicine, Atlanta, Georgia, USA.
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Walker SJ, Byrne JP, Birbeck N. What's new in the pathology, pathophysiology and management of benign esophageal disorders? Dis Esophagus 2000; 12:219-37. [PMID: 10631918 DOI: 10.1046/j.1442-2050.1999.00056.x] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Affiliation(s)
- S J Walker
- Department of Surgery, Blackpool Victoria Hospital, Lancs, UK
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