title: Sublytic C5b-9-stimulated Schwann cell survival through PI 3-kinase-mediated phosphorylation of BAD.
abstract: Sublytic C5b-9 induces cell cycle activation, proliferation, and rescue from apoptosis in Schwann cells. The signaling pathways for C5b-9-mediated rescue were investigated. Following serum withdrawal, DNA fragmentation, detected by TUNEL and FACS analysis, was 56.7% +/- 7.3 and 91.9% +/- 2.4 in cultured sciatic nerve Schwann cells from 6-day-old rats after 18 h and 24 h, respectively. Apoptosis was confirmed by inhibition of DNA fragmentation in a dose-dependent manner by DMQD-CHO, a caspase-3 inhibitor. Treatment with sublytic C5b-9 generated with purified components (C5*9) or Ab+C7-depleted serum (C7dHS)+C7 rescued 89% and 86% of Schwann cells, respectively, as compared with cells treated with C5*6, C8, C9, or Ab+C7dHS. Sublytic C5b-9 increased Schwann cell PI-3 kinase and Akt activity maximally at 5 min 3.14 +/- 0.5-fold and 3.56 +/- 0.4-fold, respectively, over controls. ERK-1 activity was maximally stimulated 2.98-fold at 15 min. Inhibition of PI-3 kinase by LY294002 abrogated the C5b-9-mediated Schwann cell rescue from apoptosis, while inhibition of ERK-1 with PD098,059 did not. PI-3 kinase-Akt pathway activation by C5b-9 induced, within 15 min, a 6.34 +/- 1.2-fold increase in BAD phosphorylation at Ser 136, but not at Ser 112. Downstream Bcl-x(L) protein was increased 2.61-fold +/- 0.34-fold by 18 h and 3.9-fold +/- 0.84-fold by 24 h over controls. LY294002 prevented both BAD phosphorylation at Ser 136 and Bcl-x(L) protein induction, while PD098,059 did not. Our data indicated that sublytic C5b-9 rescued Schwann cell from apoptosis via activation of PI-3 kinase-Akt, BAD phosphorylation on Ser 136 and increased expression of Bcl-x(L). Sublytic C5b-9 detected on Schwann cell in vivo during inflammatory neuropathy may facilitate survival of Schwann cell capable of remyelination.
authors.affiliation: Department of Neurology, University of Maryland School of Medicine, Baltimore, Maryland, USA.
authors.forename: S
authors.initials: S
authors.lastname: Hila
authors.forename: L
authors.initials: L
authors.lastname: Soane
authors.forename: C L
authors.initials: CL
authors.lastname: Koski
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002352:Carrier Proteins / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured / Q000166:cytology / Q000378:metabolism; D015938:Complement Membrane Attack Complex / Q000378:metabolism* / Q000494:pharmacology; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020287:In Situ Nick-End Labeling; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011129:Polyradiculoneuropathy / Q000378:metabolism / Q000503:physiopathology; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012583:Schwann Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D051025:bcl-Associated Death Protein; D051020:bcl-X Protein
chemical_list: D002352:Carrier Proteins; D015938:Complement Membrane Attack Complex; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D019253:Proto-Oncogene Proteins c-bcl-2; D051025:bcl-Associated Death Protein; D051020:bcl-X Protein; C494821:Bad protein, rat; C494813:Bcl2l1 protein, rat; C494920:Akt1 protein, rat