title: Diabetic cardiomyopathy is associated with defective myocellular copper regulation and both defects are rectified by divalent copper chelation.
abstract: BACKGROUND
  Heart disease is the leading cause of death in diabetic patients, and defective copper metabolism may play important roles in the pathogenesis of diabetic cardiomyopathy (DCM). The present study sought to determine how myocardial copper status and key copper-proteins might become impaired by diabetes, and how they respond to treatment with the Cu (II)-selective chelator triethylenetetramine (TETA) in DCM.
  
  
  METHODS
  Experiments were performed in Wistar rats with streptozotocin (STZ)-induced diabetes with or without TETA treatment. Cardiac function was analyzed in isolated-perfused working hearts, and myocardial total copper content measured by particle-induced x-ray emission spectroscopy (PIXE) coupled with Rutherford backscattering spectrometry (RBS). Quantitative expression (mRNA and protein) and/or activity of key proteins that mediate LV-tissue-copper binding and transport, were analyzed by combined RT-qPCR, western blotting, immunofluorescence microscopy, and enzyme activity assays. Statistical analysis was performed using Student's t-tests or ANOVA and p-values of < 0.05 have been considered significant.
  
  
  RESULTS
  Left-ventricular (LV) copper levels and function were severely depressed in rats following 16-weeks' diabetes, but both were unexpectedly normalized 8-weeks after treatment with TETA was instituted. Localized myocardial copper deficiency was accompanied by decreased expression and increased polymerization of the copper-responsive transition-metal-binding metallothionein proteins (MT1/MT2), consistent with impaired anti-oxidant defences and elevated susceptibility to pro-oxidant stress. Levels of the high-affinity copper transporter-1 (CTR1) were depressed in diabetes, consistent with impaired membrane copper uptake, and were not modified by TETA which, contrastingly, renormalized myocardial copper and increased levels and cell-membrane localization of the low-affinity copper transporter-2 (CTR2). Diabetes also lowered indexes of intracellular (IC) copper delivery via the copper chaperone for superoxide dismutase (CCS) to its target cuproenzyme, superoxide dismutase-1 (SOD1): this pathway was rectified by TETA treatment, which normalized SOD1 activity with consequent bolstering of anti-oxidant defenses. Furthermore, diabetes depressed levels of additional intracellular copper-transporting proteins, including antioxidant-protein-1 (ATOX1) and copper-transporting-ATPase-2 (ATP7B), whereas TETA elevated copper-transporting-ATPase-1 (ATP7A).
  
  
  CONCLUSIONS
  Myocardial copper deficiency and defective cellular copper transport/trafficking are revealed as key molecular defects underlying LV impairment in diabetes, and TETA-mediated restoration of copper regulation provides a potential new class of therapeutic molecules for DCM.
authors.forename: Shaoping
authors.initials: S
authors.lastname: Zhang
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.forename: Greeshma V
authors.initials: GV
authors.lastname: Amarsingh
authors.forename: Carlos C H
authors.initials: CC
authors.lastname: Cheung
authors.forename: Sebastian
authors.initials: S
authors.lastname: Hogl
authors.forename: Umayal
authors.initials: U
authors.lastname: Narayanan
authors.forename: Lin
authors.initials: L
authors.lastname: Zhang
authors.forename: Selina
authors.initials: S
authors.lastname: McHarg
authors.forename: Jingshu
authors.initials: J
authors.lastname: Xu
authors.forename: Deming
authors.initials: D
authors.lastname: Gong
authors.forename: John
authors.initials: J
authors.lastname: Kennedy
authors.forename: Bernard
authors.initials: B
authors.lastname: Barry
authors.forename: Yee Soon
authors.initials: YS
authors.lastname: Choong
authors.forename: Anthony R J
authors.initials: AR
authors.lastname: Phillips
authors.affiliation: The School of Biological Sciences, Faculty of Science, University of Auckland, Auckland, New Zealand. g.cooper@auckland.ac.nz.
authors.forename: Garth J S
authors.initials: GJ
authors.lastname: Cooper
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology / Q000627:therapeutic use*; D003300:Copper / Q000172:deficiency / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000188:drug therapy / Q000378:metabolism*; D058065:Diabetic Cardiomyopathies / Q000188:drug therapy / Q000378:metabolism*; D008297:Male; D051381:Rats; D017208:Rats, Wistar
chemical_list: D002614:Chelating Agents; D003300:Copper