Abstract
Despite increased neurogenic differentiation markers in the hippocampal CA1 in Alzheimer disease, neurons are not replaced in CA1 and the neocortex in the disease. β-Amyloid (Aβ) might cause deterioration of the brain microenvironment supporting neurogenesis and the survival of immature neurons. To test this possibility, we examined whether Aβ alters the expression of cell fate determinants in cerebral cortical cultures and in an Alzheimer disease mouse model (PrP-APPSW). Up-regulation of Mash1 and down-regulation of Olig2 were found in cerebral cortical cultures treated with Aβ-(1-42). Mash1 was expressed in nestin-positive immature cells. The majority of Mash1-positive cells in untreated cortical culture co-expressed Olig2. Aβ increased the proportion of Olig2-negative/Mash1-positive cells. A decrease in Olig2+ cells was also observed in the cerebral cortex of adult PrP-APPSW mice. Cotransfection experiments with Mash1 cDNA and Olig2 siRNA revealed that overexpression of Mash1 in neurosphere cells retaining Olig2 expression enhanced neural differentiation but accelerated death of Olig2-depleted cells. Growth factor deprivation, which down-regulated Olig2, accelerated death of Mash1-overexpressing neurosphere cells. We conclude that cooperation between Mash1 and Olig2 is necessary for neural stem/progenitor cells to develop into fully mature neurons and that down-regulation of Olig2 by Aβ in Mash1-overexpressing cells switches the cell fate to death. Maintaining Olig2 expression in differentiating cells could have therapeutic potential.
Highlights
Adult neural stem/progenitor cells continue to generate neurons in the brain throughout life, and there are high expectations that it may be possible to repair the nervous system in neurodegenerative diseases such as Alzheimer disease by stimulating endogenous progenitor cells or by stem cell transplantation
We examined whether A alters the expression of cell fate determinants in cerebral cortical cultures and in an Alzheimer disease mouse model (PrP-APPSW)
We examined the expression of basic helix-loop-helix factors, which are involved in neurogenesis during development, in A-treated cultures, and in the cerebral cortex of an Alzheimer disease (AD) mouse model (PrP-APPSW, Tg2576)
Summary
Cell Culture—For neuronal cell culture, cerebral cortices dissected from day E17 embryonic rats were dissociated by incubation with 0.08% trypsin, 0.008% DNase I at 37 °C for 10 min and passed through a 62-m nylon mesh. Brains were post-fixed in phosphate-buffered 4% paraformaldehyde for 24 h at 4 °C. Northern Blot Analysis—Poly(A)ϩRNA from cultured cortical neurons or neurosphere cells was isolated using a MicroFastTrack 2.0 kit (Invitrogen). RT-PCR Cloning of cDNA and Quantitative Real-time PCR (QRT-PCR)—First-strand cDNA was synthesized from poly(A)ϩRNA of cultured cortical neurons using SuperScript II and oligo(dT) primers (Invitrogen). Quantitative RT-PCR analysis was performed using an iCycler iQ detection system (Bio-Rad) with ELONGase enzyme mix (Invitrogen), SYBR Green I (1/50000 dilution, Takara) and 400 nM gene-specific primers (Table 1). The results were evaluated with the ICYCLER IQ Real Time Detection System software (Bio-Rad)
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