title: C-peptide enhances insulin-mediated cell growth and protection against high glucose-induced apoptosis in SH-SY5Y cells.
abstract: BACKGROUND
  We have previously reported that C-peptide exerts preventive and therapeutic effects on diabetic neuropathy in type 1 diabetic BB/Wor-rats and that it prevents duration-dependent hippocampal apoptosis in the same animal model. In the present study, we examined human neuroblastoma SH-SY5Y cells to examine whether C-peptide stimulates cell proliferation/neurite outgrowth and whether it has antiapoptotic effects.
  
  
  METHODS
  For neurite outgrowth, serum-starved cultures were treated with C-peptide and/or insulin or IGF-1. Neurites were visualized with NF-L antibody and measured morphometrically. Cell numbers were determined using an electronic cell counter. Scrambled C-peptide was used as a negative control. For assessment of apoptosis, SH-SY5Y cells were incubated with 100 mM glucose for 24 h, and the effects of C-peptide and/or insulin or IGF-1 were examined. Apoptosis was demonstrated by transferase-mediated dUTP nick-end labeling (TUNEL)/4,6-diamidino-2-phenylindole (DAPI) stainings, flow cytometry and changes in the expression of Bcl2. Activation of insulin signaling intermediaries was determined by Western blots. Translocation of NF-kappaB was demonstrated immunocytochemically.
  
  
  RESULTS
  C-peptide but not scrambled C-peptide stimulated cell proliferation and neurite outgrowth. In the presence of 4 nM insulin, 3 nM C-peptide significantly increased autophosphorylation of the insulin receptor (IR) but not that of the insulin-like growth factor 1 receptor (IGF-1R). It stimulated phosphoinositide 3-kinase (PI-3 kinase) and p38 mitogen-activated protein (MAP) kinase activation, enhanced the expression and translocation of nuclear factor-kappaB (NF-kappaB), promoted the expression of Bcl2 and reduced c-jun N-terminal kinase (JNK) phosphorylation in excess of that of insulin alone.
  
  
  CONCLUSIONS
  C-peptide in the presence of insulin exerts synergistic effects on cell proliferation, neurite outgrowth and has in the presence of insulin an antiapoptotic effect on high glucose-induced apoptosis but less so on hyperosmolar-induced apoptosis. These effects are likely to be mediated via interactions with the insulin signaling pathway.
authors.affiliation: Department of Pathology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
authors.forename: Zhen-Guo
authors.initials: ZG
authors.lastname: Li
authors.forename: Weixian
authors.initials: W
authors.lastname: Zhang
authors.forename: Anders A F
authors.initials: AA
authors.lastname: Sima
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D002096:C-Peptide / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects*; D045744:Cell Line, Tumor; D004357:Drug Synergism; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008353:Mannitol / Q000494:pharmacology; D016501:Neurites / Q000187:drug effects / Q000502:physiology; D009447:Neuroblastoma
chemical_list: D002096:C-Peptide; D007328:Insulin; D008353:Mannitol; D005947:Glucose