Leukemia inhibitory factor is essential for subventricular zone neural stem cell and progenitor homeostasis as revealed by a novel flow cytometric analysis.

Abstract

Stem cells rely on extracellular signals produced by the niche, which dictate their ability to self-renew, expand and differentiate. It is essential to have sensitive and reproducible methods of either quantifying or isolating these stem cells and progenitors to understand their intrinsic properties and how extrinsic signals regulate their development. However, stem cells are difficult to distinguish from multipotential progenitors, which may look and act like them. Here we define a 4-color flow cytometry panel using CD133, LeX, CD140a, NG2 to define a neural stem cell (NSC) as well as 4 classes of multipotential progenitors and 3 classes of bipotential progenitors, several of which have not been described previously. We performed gain and loss of function studies for leukemia inhibitory factor (LIF) and showed a depletion of NSCs, a subset of multipotential neural precursors and immature oligodendrocytes in LIF null mice. Gain of function studies showed that LIF increased the abundance of these precursors. Our studies also show that these NPs have differential requirements for LIF and ciliary neurotrophic factor (CNTF) and for epidermal growth factor (EGF), fibroblast growth factor (FGF-2) and platelet-derived growth factor (PDGF) for their propagation in vitro. Surprisingly, the related cytokine, CNTF, was less potent than LIF in increasing the NSCs and more potent than LIF in increasing the PDGF responsive multipotential precursors. Finally, we show that LIF increases the expression of the core transcription factors: Klf4, Fbx15, Nanog, Sox2 and c-Myc. Altogether our FACS (fluorescence-activated cell sorter) analyses reveal that the neonatal subventricular zone is far more heterogeneous than previously suspected and our studies provide new insights into the signals and mechanisms that regulate their self-renewal and proliferation.