title: Enhanced expression of glucose transporter-1 in vascular smooth muscle cells via the Akt/tuberous sclerosis complex subunit 2 (TSC2)/mammalian target of rapamycin (mTOR)/ribosomal S6 protein kinase (S6K) pathway in experimental renal failure.
abstract: BACKGROUND
  Chronic renal failure (CRF) is associated with increased cardiovascular mortality, and medial vascular smooth muscle cell (VSMC) hypertrophy, proliferation, and calcification play a pivotal role in uremic vasculopathy. Glucose transporter-1 (GLUT1) facilitates the transport of glucose into VSMCs, and GLUT1 overexpression associated with high glucose influx leads to a stimulation of VSMC proliferation. However, the role of GLUT1 in uremic vasculopathy remains unclear. This study aimed to identify changes in the expression of GLUT1 in VSMCs in the setting of experimental uremia and investigate whether Akt/tuberous sclerosis complex subunit 2 (TSC2)/mammalian target of rapamycin (mTOR)/ribosomal S6 protein kinase (S6K) signaling, which plays a crucial role in VSMC proliferation and glucose metabolism, is involved in the regulation of GLUT1 expression.
  
  
  METHODS
  In vivo experimental CRF was induced in Wistar rats by 5/6 nephrectomy, and the GLUT1 expression in aortic tissue was determined by the reverse transcriptase-polymerase chain reaction, immunoblotting, and immunohistochemical staining. Indoxyl sulfate (IS) is a uremic retention solute proven with pro-proliferative effect on rat VSMCs, and we further studied the expression of GLUT1 in rat A7r5 rat embryonic aortic cells stimulated by IS in the presence or absence of phloretin, a GLUT1 inhibitor, to explore the pathogenic role of GLUT1 in uremic vasculopathy. The contribution of Akt/TSC2/mTOR/S6K signaling in modifying the GLUT1 expression was also assessed.
  
  
  RESULTS
  Eight weeks after 5/6 nephrectomy, aortic tissue obtained from CRF rats exhibited increased wall thickness and VSMC hypertrophy, hyperplasia, and degeneration. Compared with the sham-operated control group, the messenger (m)RNA and protein abundance of GLUT1 were both markedly increased in CRF rats. In vitro, IS induced a significant increase in expression of GLUT1 protein as well as pro-proliferative cyclin D1 and p21 mRNA and a modest increase in expression of antiapoptotic p53 mRNA in A7r5 cells, whereas inhibition of GLUT1 mediated glucose influx reduced the pro-proliferative and antiapoptotic effects of IS. In addition to increased GLUT1 expression, IS significantly suppressed Akt and TSC2 phosphorylation after 6-hour and 12-hour treatment, but increased S6K phosphorylation after 3-hour treatment. Inactivation of mTOR downstream signaling by rapamycin treatment inhibited S6K phosphorylation and abolished the stimulatory effect of IS on GLUT1 expression.
  
  
  CONCLUSIONS
  In vivo and in vitro experimental CRF displayed prominent GLUT1 upregulation in VSMCs. The uremic toxin IS stimulated proliferation of VSMCs possibly through induction of GLUT1 expression. The Akt/TSC/mTOR/S6K signaling pathway may be one of the mechanisms underlying the upregulation of GLUT1 expression in uremic VSMCs.
authors.affiliation: Division of Cardiac Surgery, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan .
authors.forename: Chih-Yuan
authors.initials: CY
authors.lastname: Lin
authors.forename: Shih-Che
authors.initials: SC
authors.lastname: Hsu
authors.forename: Herng-Sheng
authors.initials: HS
authors.lastname: Lee
authors.forename: Shih-Hua
authors.initials: SH
authors.lastname: Lin
authors.forename: Chien-Sung
authors.initials: CS
authors.lastname: Tsai
authors.forename: Shih-Ming
authors.initials: SM
authors.lastname: Huang
authors.forename: Chun-Che
authors.initials: CC
authors.lastname: Shih
authors.forename: Yu-Juei
authors.initials: YJ
authors.lastname: Hsu
mesh_terms: D000818:Animals; D001011:Aorta / Q000201:enzymology; D017209:Apoptosis; D015153:Blotting, Western; D002460:Cell Line; D049109:Cell Proliferation; D019938:Cyclin D1 / Q000235:genetics / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D005947:Glucose / Q000378:metabolism; D051272:Glucose Transporter Type 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006965:Hyperplasia; D006984:Hypertrophy; D007150:Immunohistochemistry; D007200:Indican / Q000378:metabolism; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D009392:Nephrectomy; D010693:Phloretin / Q000494:pharmacology; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D051437:Renal Insufficiency / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism*; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013997:Time Factors; D000077005:Tuberous Sclerosis Complex 2 Protein; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D015854:Up-Regulation; D014511:Uremia / Q000201:enzymology
chemical_list: C530106:Ccnd1 protein, rat; C493958:Cdkn1a protein, rat; D050759:Cyclin-Dependent Kinase Inhibitor p21; D051272:Glucose Transporter Type 1; D012333:RNA, Messenger; C495679:Slc2a1 protein, rat; C000624655:Tsc2 protein, rat; D000077005:Tuberous Sclerosis Complex 2 Protein; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D019938:Cyclin D1; C546845:mTOR protein, rat; D051057:Proto-Oncogene Proteins c-akt; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D005947:Glucose; D007200:Indican; D010693:Phloretin; D020123:Sirolimus