Gene Expression Profiling of an Immortalized Human Neural Stem Cell Line HB1.F3

Neural stem cells (NSC) with self-renewal and multipotent properties serve as an ideal cell source for transplantation to treat spinal cord injury, stroke, and neurodegenerative diseases. To efficiently induce neuronal lineage cells from NSC for neuron replacement therapy, we should clarify the intrinsic genetic programs involved in a time and place-specific regulation of human NSC differentiation. Recently, we established an immortalized human NSC clone HB1.F3 to provide an unlimited NSC source applicable to genetic manipulation for cell-based therapy. To investigate a role of neurogenin 1 (Ngn1), a proneural basic helix-loop-helix (bHLH) transcription factor, in human NSC differentiation, we established a clone derived from F3 stably overexpressing Ngn1. Genome-wide gene expression profiling identified 250 upregulated genes and 338 downregulated genes in Ngn1-overexpressing F3 cells (F3-Ngn1) versus wild-type F3 cells (F3-WT). Notably, leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a novel stem cell marker, showed a robust increase in F3-Ngn1. Primary cultures of fetal human telencephalon cells were transformed with a retroviral vector pLSNmyc carrying the v-myc oncogene and the neomycin resistance gene. Following selection with G418, a single continuously dividing clone with a capacity to self-renew and differentiate into neurons and glial cells both in vitro and in vivo was isolated and designated HB1.F3. It carried normal human karyotype of 46 XX. After transducing a retroviral vector pBabePNgn1 carrying the open-reading frame (ORF) of the human Ngn1 gene and the puromycin resistance gene into HB1.F3 cells, a single puromycin-resistant clone was selected, expanded, and designated HB1.F3-Ngn1. In the present study, the wild-type HB1.F3 cells and the HB1.F3-Ngn1 cells are abbreviated as F3-WT and F3-Ngn1. Total RNA of F3-WT and F3-Ngn1 isolated from subconfluent cells was processed for two times different hybridization experiments, named as 1st and 2nd experiments.