Abstract 199: Pre-treatment Of Human Embryonic-derived Neural Progenitor Cells With Bdnf Increases Migration Of Cells To The Brain And Elicits Functional Recovery In A Mouse Model Of Hypoxic-ischemia

Abstract

Intro: Increasing cell migration through upregulation of chemokine and adhesion molecule receptors could improve intravascular cell treatment for stroke and BDNF has been shown to induce these pathways. Therefore we tested whether BDNF cell-pretreatment would improve cell migration and functional recovery in an experimental stroke model. Methods: hES-derived NPCs (5x10 5 in 5µl saline) pre-treated with BDNF for 5 hours and harboring a reporter gene construct containing renilla luciferase and eGFP in serum free media, non-treated hES-derived NPCs (5×10 5 in 5[l saline) in media, and media control with BDNF were delivered to the brain via the ipsilateral carotid artery at 3 days after hypoxic-ischemic stroke in NODSCID mice (n=11/group). Cell engraftment was monitored by in-vivo bioluminescence imaging (BLI). The ladder test was used to assess behavioral recovery throughout a 4 week time course. Brain homogenates from animals at 28 days were analyzed using RT-qPCR for common chemokines, adhesion molecules, and neurotrophins. Mechanisms of cell migration were evaluated by assessing cell receptor expression of chemokines and adhesion molecules on hES-derived NPC and by analyzing the change in expression profile in the mouse brain at 3 days after stroke. Boyden-chamber migration assays were used to evaluate cell migratory potential in vitro . RESULTS: One day after cell transplantation the subset of animals transplanted with BDNF-pretreated cells showed significantly higher BLI signal at 1 (p=0.021) and 7 days after transplantation (p=0.002). Histological analysis also revealed engraftment of hES-derived NPC at 1 week after transplantation. Behavioral assessment revealed significant functional recovery in the BDNF pre-treated group throughout the 28 day time course (ANOVA, p<0.05). BDNF-pretreatment of hES-derived NPCs upregulated CXCR4 expression 12.5 times and in vitro led to significantly greater migration in response to CXCL12 (CXCR4 ligand) compared to untreated cells. At 28 days after transplantation, neurotrophic factors IL6, IL10, Ntrk1 were upregulated 3.3, 3.4, and 3.3 times. Common T-cell and neutrophil cytokine receptors IL8rb, IL8ra and IL1a were all downregulated, while several chemokines that increase migration of inflammatory cells were downregulated including CCL2, CCL5, CCL8, and CCL12. Anti-Angiogenic factor Adamts8 was also downrgulated in the brains of animals transplanted with BDNF pre-treated cells. Lastly, MMP3, MMP8, and MMP9 were downregulated at 28 days after stroke indicating increased blood brain barrier integrity. Conclusion: Intravascular transplantation of BDNF pre-treated hES-derived NPCs elicits functional gains via increased migration of cells, immunomodulation, increased BBB integrity, and by influencing the upregulation of neur0protective factors.