The Journal of Neuroscience, November 15, 2006, ():
Leukemia Inhibitory Factor Promotes Neural Stem Cell Self-Renewal in the Adult Brain
J. Neurosci. Bauer and Patterson
26: 12089
Supplemental Data
Files in this Data Supplement:
- supplemental material
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Supplemental Figure S1. LIF is biologically active in vivo after injection of Ad:LIF into the lateral ventricle. (A) LIF activity is visualized by phosphorylated Stat3 (pStat3) immunohistochemistry. Nuclear staining is detected in regions adjacent to the lateral venctricle (LV), prominently in the striatum (St) and to a lesser extent in the corpus callosum (CC). Staining intensity appears stronger in areas immediately adjacent to the ventricle, while it decreases further away from the ventricle in a gradient-like pattern. No staining is detected in animals injected with Ad:LacZ as a control. (B) Higher magnification pictures in a region of the striatum (St) adjacent to the lateral ventricle (LV) show that the majority of pStat3+ nuclei also express the mature neuron marker NeuN. Scale bars: A, 150 μm; B, 50 μm.
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Supplemental Figure S2. LIF receptor (LIFR) is expressed in the SVZ by neuroblasts (type A cells) and transit amplifying cells (type C cells). (A) Immunohistochemical staining for LIFR co-localizes with doublecortin (DCX) and ß-III-tubulin (TuJ1) (arrows), two neuroblast markers. Pictures are 3 consecutive confocal 1 µm-thick optical sections. LV= lateral ventricle. (B) LIFR staining co-localizes with doublecortin (DCX; arrows), a neuroblast marker. Pictures are 3 consecutive confocal 1 µm-thick optical sections. (C) LIFR staining co-localizes with EGF receptor (EGFR; arrows), a marker of transit amplifying cells. Pictures correspond to a projection of z-stack (8 confocal 1 µm-thick optical sections). Scale bars: A, 30 μm; B, 15 μm; C, 30 μm.
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Supplemental Figure S3. LIF impairs neurosphere formation in vitro and SVZ regeneration in vivo. (A) Adult SVZ-derived neurospheres do not form in vitro when single cells are continuously exposed to LIF. Pictures depict neurospheres obtained after 8 days in vitro (DIV) in EGF only or in EGF supplemented with LIF (20 ng/ml). LIF impairs sphere formation even at very low dose. Single cells were incubated with various concentration of LIF and neurospheres were quantified after 8 DIV. ***P < 0.001 compared to control without LIF (Student’s t-Test). Scale bar: 300 μm. (B) A transient exposition of neurospheres to LIF (20 ng/ml) for the last 2 DIV (starting at 6 DIV) does not affect the number of neurospheres at 8 DIV. (C) Comparison of the effects of LIF between sphere formation in vitro and SVZ regeneration in vivo. Dissociated neurosphere cells were plated at 20,000 cells/well in EGF+bFGF, with or without LIF (20 ng/ml), and were quantified after 2 and 4 DIV. In vitro cell numbers were divided 1000-fold before plotting (right axis). For in vivo SVZ regeneration, data are the number of BrdU+ cells (left axis) quantified in the SVZ at 1 and 3 days after AraC removal (data also shown in Fig. 5).
