Anti-apoptotic actions of the platelet-activating factor acetylhydrolase I alpha2 catalytic subunit.

Platelet-activating factor (PAF) is an important mediator of cell loss following diverse pathophysiological challenges, but the manner in which PAF transduces death is not clear. Both PAF receptor-dependent and -independent pathways are implicated. In this study, we show that extracellular PAF can be internalized through PAF receptor-independent mechanisms and can initiate caspase-3-dependent apoptosis when cytosolic concentrations are elevated by approximately 15 pM/cell for 60 min. Reducing cytosolic PAF to less than 10 pM/cell terminates apoptotic signaling. By pharmacological inhibition of PAF acetylhydrolase I and II (PAF-AH) activity and down-regulation of PAF-AH I catalytic subunits by RNA interference, we show that the PAF receptor-independent death pathway is regulated by PAF-AH I and, to a lesser extent, by PAF-AH II. Moreover, the anti-apoptotic actions of PAF-AH I are subunit-specific. PAF-AH I alpha1 regulates intracellular PAF concentrations under normal physiological conditions, but expression is not sufficient to reduce an acute rise in intracellular PAF levels. PAF-AH I alpha2 expression is induced when cells are deprived of serum or exposed to apoptogenic PAF concentrations limiting the duration of pathological cytosolic PAF accumulation. To block PAF receptor-independent death pathway, we screened a panel of PAF antagonists (CV-3988, CV-6209, BN 52021, and FR 49175). BN 52021 and FR 49175 accelerated PAF hydrolysis and inhibited PAF-mediated caspase 3 activation. Both antagonists act indirectly to promote PAF-AH I alpha2 homodimer activity by reducing PAF-AH I alpha1 expression. These findings identify PAF-AH I alpha2 as a potent anti-apoptotic protein and describe a new means of pharmacologically targeting PAF-AH I to inhibit PAF-mediated cell death.

Oligodendrocyte progenitor enrichment in the connexin32 null-mutant mouse.

Before the establishment of chemical synapses, neural progenitors are often coupled by connexin-mediated gap junctions providing a robust mechanism for cell-cell communication in developing brain. The present study was undertaken to determine whether alterations in junctional coupling also affect neural progenitor proliferation, survival, and differentiation in adult brain. We localized the connexin32 gap junction protein to a subset of NG2+ and platelet-derived growth factor alpha receptor+ early oligodendrocyte progenitors in the dentate gyrus of adult mice. In connexin32-deficient mice, we found an increase in the total number of proliferating nestin+ and NG2+ progenitors in the subgranular zone, hilus, and polymorphonuclear layer of the dentate gyrus in vivo and in the total number of nestin+ progenitors capable of clonogenic expansion in vitro. By bromodeoxyuridine labeling, lineage analysis, and terminal deoxynucleotidyl nick end labeling, we demonstrate that turnover of these cells is constitutively enhanced in the connexin32 knock-out dentate gyrus reflecting a dynamic defect in oligodendrogenesis in this population. Analyses of surviving bromodeoxyuridine-labeled cells at 1, 3, 7, and 28 d after injection demonstrate that this transient amplifying population fails to terminally differentiate and is deleted by an apoptotic-like mechanism within 3 d of labeling. These data provide empirical evidence to support the hypothesis that connexin expression influences adult progenitor number and specifically implicate connexin32-mediated signaling in the activation, survival, and differentiation of a subset of early oligodendrocyte progenitors in postnatal brain.