FIGURE 5.

Lrp1 knock-out NSPCs proliferate and differentiate in vitro. A, schematic diagram outlining the generation of Lrp1 knock-out NSPCs. NSPCs obtained from Lrp1^flox/flox mice or Lrp1^wt/wt littermates were treated with cell-permeant Cre recombinase, expanded as neurospheres and differentiated. B, scheme of the Lrp1 gene locus illustrating the location of the loxP sites and the primers used to assess recombination. C, exemplary PCR verifying successful recombination in Cre-treated cells. D, phase contrast image of neurospheres derived from Cre-treated Lrp1^flox/flox or Lrp1^wt/wt cells. E, Western blot demonstrating the reduction of LRP1 protein in NSPCs after recombination. F, LRP1 expression permanently eliminated in Cre-treated Lrp1^flox/flox NSPCs even when passaged over a time period of 6 weeks. G–J, immunostainings against cell type-specific biomarkers after differentiation of Lrp1 knock-out. Nestin (neural stem precursor cells (NSPCs)); GFAP, glial fibrillary acidic protein (astrocytes); O4 (oligodendrocytes); βIII-tubulin, (young neurons). Scale bar, 50 μm.