Abstract 3575: Oligodendrocyte Proliferation and Improved Myelination Following Human Embryonic-Derived Neural Stem Cell Treatment in an Experimental Model of Cerebral Palsy

Abstract

Introduction: Cerebral palsy (CP) is the most common cause of motor disability in children, and chronic deficits are associated with white matter injury. Neonatal hypoxic-ischemic insults, an important cause of CP, induce oligodendrocyte apoptosis and impair normal myelin development. No CP treatments target myelination making regenerative medicine a promising research frontier. We investigated the effect of human embryonic-derived neural stem cell (NSC) treatment on oligodendrocytes and myelination following hypoxia-ischemia (HI). Methods: Neonatal Wistar rat pups underwent left CCA ligation followed by placement in 8% O2 at 37°C on post-natal day 7 (P7). Following T2w MRI on P9, immunosuppressed pups received intra-arterial transplant of 500k fLuc/eGFP transduced NSCs or saline on P10. BrdU was administered intraperitoneally from P11 - P18. In vivo bioluminescence images (BLI) were obtained 1 - 10 days (d) after injection. Myelination was evaluated using luxol fast blue (LFB) and myelin basic protein (MBP) staining 10 and 30 d after treatment. Oligodendrogenesis was quantified using BrdU staining in conjunction with Olig2, NG2, and CC1. RT-qPCR was performed on neonatal brain isolates and NSC mRNA. Luminex immunological assay was used to quantify NSC protein secretion. Functional recovery was assessed using the novel object recognition (NOR) task at P30. Results: Stroke size between groups was not significantly different 3, 10, and 30 d after treatment. BLI demonstrated significant NSC homing to the ischemic hemisphere days 1 - 7 (p=0.001) after transplant. Histology confirmed initial NSC localization to corpus callosum and cortex with migration into external capsule and corona radiata 30 d after transplant. NSC-treated pups had significantly more BrdU+ cells near the lateral ventricle (p=0.036) and in the corpus callosum (p=0.020) than controls. In addition to more Olig2+ and NG2+ cells in the striatum, NSC-treated pups had significantly more BrdU+/Olig2+ cells in the corpus callosum (p<0.05) than controls 30 d after treatment. LFB and MBP staining demonstrated greater myelination 10 and 30 d after treatment in corpus callosum (p=0.022, p<0.05) and striatum (p=0.017, p=0.001) of NSC-treated pups. Stat3 (2.82), IL-6 (1.48), and IL-6Rβ (1.73) mRNA was upregulated in the brains of NSC-treated pups. Proteomic and mRNA data confirm NSC expression of VEGF (17.9pg/mL, 4.35) and CXCL1 (3.6pg/mL, 10.27). NSC-treated pups performed better on NOR (p=0.016). Conclusions: Intra-arterial NSC transplant after hypoxia-ischemia results in NSC engraftment into white matter tracts, increased oligodendrocyte proliferation, and improved myelination. NSC-derived proteins may drive the distinct changes in gene expression occurring in the brain after NSC treatment and may mediate functional recovery via activation of endogenous self-repair mechanisms, including oligodendrogenesis.