An activated protein C analog stimulates neuronal production by human neural progenitor cells via a PAR1-PAR3-S1PR1-Akt pathway

J Neurosci. 2013 Apr 3;33(14):6181-90. doi: 10.1523/JNEUROSCI.4491-12.2013.

Abstract

Activated protein C (APC) is a protease with anticoagulant and cell-signaling activities. In the CNS, APC and its analogs with reduced anticoagulant activity but preserved cell signaling activities, such as 3K3A-APC, exert neuroprotective, vasculoprotective, and anti-inflammatory effects. Murine APC promotes subependymal neurogenesis in rodents in vivo after ischemic and traumatic brain injury. Whether human APC can influence neuronal production from resident progenitor cells in humans is unknown. Here we show that 3K3A-APC, but not S360A-APC (an enzymatically inactive analog of APC), stimulates neuronal mitogenesis and differentiation from fetal human neural stem and progenitor cells (NPCs). The effects of 3K3A-APC on proliferation and differentiation were comparable to those obtained with fibroblast growth factor and brain-derived growth factor, respectively. Its promoting effect on neuronal differentiation was accompanied by inhibition of astroglial differentiation. In addition, 3K3A-APC exerted modest anti-apoptotic effects during neuronal production. These effects appeared to be mediated through specific protease activated receptors (PARs) and sphingosine-1-phosphate receptors (S1PRs), in that siRNA-mediated inhibition of PARs 1-4 and S1PRs 1-5 revealed that PAR1, PAR3, and S1PR1 are required for the neurogenic effects of 3K3A-APC. 3K3A-APC activated Akt, a downstream target of S1PR1, which was inhibited by S1PR1, PAR1, and PAR3 silencing. Adenoviral transduction of NPCs with a kinase-defective Akt mutant abolished the effects of 3K3A-APC on NPCs, confirming a key role of Akt activation in 3K3A-APC-mediated neurogenesis. Therefore, APC and its pharmacological analogs, by influencing PAR and S1PR signals in resident neural progenitor cells, may be potent modulators of both development and repair in the human CNS.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Analysis of Variance
  • Animals
  • Apoptosis / drug effects
  • Blood Coagulation Factors / agonists*
  • Brain-Derived Neurotrophic Factor / pharmacology
  • Bromodeoxyuridine
  • Cell Differentiation / drug effects
  • Cell Movement / drug effects
  • Cells, Cultured
  • Cerebral Ventricles / cytology
  • Dactinomycin / analogs & derivatives
  • Dactinomycin / metabolism
  • Enzyme-Linked Immunosorbent Assay
  • Female
  • Fetus
  • Flow Cytometry
  • Humans
  • Intermediate Filament Proteins / metabolism
  • Ki-67 Antigen / metabolism
  • Lysophospholipids / metabolism
  • Male
  • Mutation / physiology
  • Nerve Tissue Proteins / metabolism
  • Nestin
  • Neural Stem Cells / drug effects
  • Neural Stem Cells / physiology*
  • Neurogenesis / drug effects*
  • Oligopeptides / pharmacology*
  • Oncogene Protein v-akt / metabolism
  • RNA Interference / physiology
  • RNA, Small Interfering / pharmacology
  • Receptors, Cell Surface / agonists*
  • Receptors, Lysosphingolipid / metabolism
  • Receptors, Proteinase-Activated / metabolism
  • Serine / metabolism
  • Signal Transduction / drug effects*
  • Sphingosine / analogs & derivatives
  • Sphingosine / metabolism
  • Sphingosine-1-Phosphate Receptors
  • Time Factors

Substances

  • Blood Coagulation Factors
  • Brain-Derived Neurotrophic Factor
  • Intermediate Filament Proteins
  • Ki-67 Antigen
  • Lysophospholipids
  • NES protein, human
  • Nerve Tissue Proteins
  • Nestin
  • Oligopeptides
  • RNA, Small Interfering
  • Receptors, Cell Surface
  • Receptors, Lysosphingolipid
  • Receptors, Proteinase-Activated
  • S1PR1 protein, human
  • Sphingosine-1-Phosphate Receptors
  • activated protein C receptor
  • brain-derived growth factor
  • protein C activator peptide
  • Dactinomycin
  • sphingosine 1-phosphate
  • Serine
  • 7-aminoactinomycin D
  • Oncogene Protein v-akt
  • Bromodeoxyuridine
  • Sphingosine