GSE131065 bulk RNA-seq of human nucleusus pulposus from scoliosis patients

This study aimed to define strategies to derive NPCs from human ESC/iPSC. Human ESC3, ESC9 and IMR-90-iPSC were used for a three-step protocol to direct differentiation toward mesodermal, subsequently notochordal and finally NPCs. Our results showed that notochordal-like cells (NCCs) were successfully derived from the first two-step of protocol confirmed by immunofluorescence staining of Noto, Brachyury (T) and Foxa2. RT-PCR results showed that the expression of Noto, T and Foxa2 was the highest at day 5 after differentiation. Furthermore, these NCCs can be differentiated into NPCs via the final step of the protocol. These NPCs expressed the tyrosine kinase receptor Tie2 (Tie2), disialoganglioside 2 (GD2), collagen II and aggrecan. Genome-wide transcriptomic analyses by sequencing (RNA-seq) further reveals the expression of a wide array of extra-cellular matrix (ECM) genes, up-stream regulators and pathways. Cross-comparison of our RNA-seq profiles with public NPC data confirms the differentiated products are much closer to NPCs than are ESC/iPSC. Co-culturing NPCs with Light2 cell line, a luciferase-based reporter responsive to sonic hedgehog (Shh) secreted protein, showed NPCs synthesized Shh. Transplantation of NPCs effectively attenuated the spinal injury in a puncture-induced disc degeneration (PDD) model in rat. Furthermore, we utilized CRISPR/Cas9 technology to seamlessly knock in an enhanced fluorescent protein (GFP) to the loci of the noto gene in ESC9. Our study dem...
Source: GEO: Gene Expression Omnibus - Category: Genetics & Stem Cells Tags: Expression profiling by high throughput sequencing Homo sapiens Source Type: research