Purpose: Volatile anesthetics are a potential risk for cognitive impairment in the developing brain. Isoflurane causes cell death, reduces neurogenesis, and changes neuronal differentiation. In this study, the effects of a single isoflurane exposure on the developing human brain were evaluated using human embryonic stem cell (hESC)-derived neural progenitor cells (NPCs).
Methods: Multipotent NPCs were derived from hESCs and randomly exposed to either 5 vol% isoflurane (4 h) or no isoflurane (control group). The cells were fixed after 1, 3, 5, and 7 days to evaluate differentiation using the ratio of β-III tubulin to nestin. Neuronal cell survival and proliferation were assessed using the WST-1 and bromodeoxyuridine (BrdU) cell proliferation assays, respectively. To evaluate the mechanism of isoflurane neurotoxicity, we added TAT-Pep5, a p75 neurotrophic receptor (p75NTR) inhibitor, to each of the groups.
Results: Isoflurane had minimal or no effect on the cell survival of NPCs in vitro. Proliferation, assessed on the basis of BrdU incorporation, was inhibited in the isoflurane group on days 3 (p = 0.033) and 5 (p = 0.001). The ratios of β-III tubulin to nestin in the isoflurane group on days 1 and 3 were significantly lower (p = 0.004 and p = 0.029, respectively) than those in the control group. Anti-proliferative and differentiation-reducing effect did not persist. TAT-Pep5 pretreatment significantly blocked the isoflurane-mediated decrease in the β-III tubulin to nestin ratio (p = 0.012) on day 1.
Conclusion: Exposing NPCs to isoflurane hampered proliferation and differentiation but not neuronal survival. Isoflurane may be a powerful neuronal modulator during the early developmental period, partly mediated by activation of p75NTR.
Keywords: Embryonic stem cells; Isoflurane; Neurodevelopment; Neurotoxicity.