For the safe clinical application of embryonic stem cells (ESCs) for neurological diseases, it is critical to evaluate the tumorigenicity and function of human ESC (hESC)-derived neural cells in primates. We have herein, for the first time, compared the growth and function of hESCderived cells with different stages of neural differentiation implanted in the brains of primate models of Parkinson's disease. We herein show that residual undifferentiated cells expressing ESC markers present in the cell preparation can induce tumor formation in the monkey brain. In contrast, a cell preparation matured by 42-day culture with brain-derived neurotrophic factor/glial cell linederived neurotrophic factor (BDNF/GDNF) treatment did not form tumors and survived as primarily dopaminergic (DA) neurons. In addition, the monkeys with such grafts showed behavioral improvement for at least 12 months. These results support the idea that hESCs, if appropriately matured, can serve as a source for DA neurons without forming any tumors in a primate brain. STEM CELLS 2012;30:935-945 Disclosure of potential conflicts of interest is found at the end of this article.

Enhanced matrix metalloproteinases (MMPs) can cause vasogenic edema and hemorrhagic transformation after cerebral ischemia, and affect the extent of ischemic injury. We hypothesized that the endogenous MMP inhibitors, tissue inhibitor of MMPs (TIMPs), were essential to protect against blood-brain barrier (BBB) disruption after ischemia by regulating the activities of MMPs. We confirmed the transition of MMP-2 and MMP-9, and the TIMPs family after 30 mins of middle cerebral artery occlusion, and elucidated the function of TIMP-1 and TIMP-2 in focal ischemia, using TIMP-1 À/À and TIMP-2 À/À mice. TIMP-1 mRNA expression was gradually increased until 24 h after reperfusion. In TIMP-1 À/À mice, MMP-9 protein expression and gelatinolytic activity were significantly more augmented after cerebral ischemia than those in WT mice, and were accompanied by exacerbated BBB disruption, neuronal apoptosis, and ischemic injury. In contrast, TIMP-2 gene deletion mice exhibited no significant difference in MMP expressions and the degree of ischemic injury despite an increased Evans blue leakage. These results suggest that TIMP-1 inhibits MMP-9 activity and can play a neuroprotective role in cerebral ischemia.

Embryonic stem (ES) cells are a promising donor source for transplantation therapy, but several problems must be solved before they can be clinically useful. One of these is the host immune reaction to allogeneic grafts. In this article, we examine the effect of the host immune reaction on survival and differentiation of grafted ES cell-derived neural precursor cells (NPCs). We induced NPCs from mouse ES cells by stromal cell-derived inducing activity and then transplanted them into mouse brains with or without administering the immunosuppressant cyclosporine A (CsA). Two and 8 weeks following transplantation, the accumulation of host-derived microglia/macrophages and lymphocytes was observed around the graft. This effect was reduced by CsA treatment, although no significant difference in graft volume was observed. These data suggest that an immune response occurs in allografts of ES cell-derived NPCs. Intriguingly, however, the ratio of neurons to astrocytes in the graft was higher in immunosuppressed mice. Because inflammatory or immune cells produce various cytokines, we examined the effect of IL-1beta, IL-6, IFN-gamma, and TNF-alpha on the differentiation of NPCs in vitro. Only IL-6 promoted glial cell fate, and this effect could be reversed by the addition of an IL-6 neutralizing antibody. These results suggest that allogeneic ES cell-derived NPCs can cause an immune response by the host brain, but it is not strong enough to reject the graft. More important, activated microglia and lymphocytes can suppress neuronal differentiation of grafted NPCs in vivo by producing cytokines such as IL-6.

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