Effects Of ADAM10Deletion On Notch-1Signaling Pathway And Neuronal Differentiation In Adult Mouse Brain

Objective As we know, Notch-1signaling pathway is essential for the embryonicdevelopment of mice. The recent study showed that ADAM10is responsible for theactivation of Notch-1signaling pathway in embryo. However, how ADAM10regulates Notch-1signaling pathway and plays a role in the neuronal cells of adultbrain are unclear. So, we use the established adult neuronal-specific ADAM10conditional knockout mice (ADAM10cKO) to investigate the activation of Notch-1signaling and the effects of ADAM10deletion on neuronal differentiation andproliferation in adult mice brain.Methods At First, we used western blotting to detect the amount of S2-site cleavagefragments of Notch-1receptors and the activated NICD in the cortex andhippocampus of4-month-old ADAM10cKO mice (n=5) and ADAM10loxp/loxpmice(n=5), respectively. Then, quantitative RT-PCR was used to detect the transcription ofNotch-1downstream target genes Hes1, Hes5, Hey1and Hey2in cortex andhippocampus of ADAM10cKO (n=4) mice and ADAM10loxp/loxpmice (n=4) at the ageof four months. Finally, immunohistochemistry tests were performed to detect theneuronal differentiation and proliferation in5-month-old ADAM10cKO (n=6) andADAM10loxp/loxpcontrol mice (n=6).Results Western blotting results showed that the S2-site cleavage fragments andNICD of Notch-1decreased by88.88%and71.36%in the hippocampus of ADAM10cKO mice compared with controls, respectively. Moreover, the S2-site cleavagefragments in the cortex of ADAM10cKO mice were reduced by73.86%, and therewas no significant difference in NICD between ADAM10cKO and control mice. TheS1-site cleavage fragments of Notch-1have no significant difference in cortex andhippocampus of ADAM10cKO mice and the controls. Subsequently, quantitativeRT-PCR showed that Notch-1downstream target genes hes1, hes5, hey1and hey2 were downregulated and decreased by56.6%,55.5%,39.3%and43.5%in thehippocampus of cKO mice, respectively. Immunohistochemical analysis showed anincrease of mature neurons in hippocampus dentate gyrus and a reduction of immatureneurons in the hippocampal dentate gyrus subgranular zone of ADAM10cKO micecompared with controls.Conclusion These results demonstrated that Notch-1signaling and neuronaldifferentiation and proliferation were significantly affected by ADAM10deletion inthe neurons of adult mice. The activation of Notch-1signaling was prohibited and thelateral inhibition of Notch-1signaling was removed, resulting in the precociousneuronal differentiation and the depletion of neuronal stem cells in the hippocampusof ADAM10cKO mice. In summary, ADAM10proteinase plays an essential role inthe activation of Notch-1signaling and in the maintenance of adult neuronalhomeostasis.