The Role of Neural Precursor Cell Expressed Developmentally Down-Regulated Protein 4.1- Mediated Insulin-Like Growth Factor-1R? Ubiquitination in the Pathogenesis of Alzheimer?s disease

Abstract

To study the role of Neural precursor cell expressed developmentally down-regulated protein 4.1 on ubiquitination of central Insulin-like growth factor 1β pathway in Alzheimer’s Disease, and to clarify the molecular mechanism of Neural precursor cell expressed developmentally down-regulated protein 4.1 and Insulin-like growth factor 1β for central insulin signalling pathway damage. Mouse hippocampal neurons cells were isolated from Intracerebral haemorrhage mice. 30 μM or 300 μM S-Nitroso-l-cysteine were incubated with hippocampal neurons cells for 4 h. pCMV-Neural precursor cell expressed developmentally down-regulated protein 4.1 or siR- Neural precursor cell expressed developmentally down-regulated protein 4.1 were synthesized and transfected into S-Nitroso-l-cysteine treated hippocampal neurons cells. Cycloheximide and MG132 were incubated with hippocampal neuron cells after S-Nitroso-l-cysteine stimulation. Insulin resistant brain state model were constructed by streptozotocin injection. Western blot were used to test the expression of neural precursor cell expressed developmentally down-regulated protein 4.1, Insulin-like growth factor-1Rβ, Protein kinase B and p- Protein kinase B, Glycogen synthase kinase 3β and p- Glycogen synthase kinase 3β, Tau protein and p-Tau protein in vitro and in vivo. Coimmunoprecipitation were used to analyse Neural precursor cell expressed developmentally downregulated protein 4.1 and Insulin-like growth factor-1Rβ. Neural precursor cell expressed developmentally down-regulated protein 4.1 in the hippocampus of Tg2576 mice increased significantly compared to the control mice (p<0.05), while the expression of Insulin-like growth factor-1Rβ decreased significantly compared to the control mice (p<0.05). S-Nitroso-l-cysteine stimulated hippocampal neuron cells to increase the ubiquitination of Insulin-like growth factor-1Rβ (p<0.05) and decrease the expression of Insulin-like growth factor-1Rβ (p<0.05). Down-regulation of Neural precursor cell expressed developmentally downregulated protein 4.1 can decreased the ubiquitination of Insulin-like growth factor-1Rβ significantly (p<0.05), and rescued the S-Nitroso-l-cysteine inhibition effect on expression of Insulin-like growth factor-1Rβ. Over-expression Neural precursor cell expressed developmentally down-regulated protein 4.1 could increase the ubiquitination level of Insulin-like growth factor-1Rβ (p<0.05). MG132 and cyclohexide both can reverse the degradation of Insulin-like growth factor-1Rβ induced by S-Nitroso-l-cysteine. The phosphorylation degree of Protein kinase B and Glycogen synthase kinase 3β in neurons cells treated with S-Nitroso-l-cysteine decreased significantly (p<0.05), while Tau phosphorylation level in hippocampal neuron cells treated with Insulin-like growth factor-1 increased significantly (p<0.05). S-Nitroso-lcysteine can improve Tau protein phosphorylation (p<0.05) in neurons cells. C1060 site is necessary for its ubiquitination and degradation. Insulin-like growth factor-1Rβ were significantly lower (p<0.05) in the Streptozotocin model while Neural precursor cell expressed developmentally down-regulated protein 4.1 and Aβ were significantly higher (p<0.05) in the Streptozotocin model. p- Protein kinase B/Protein kinase B and p- Glycogen synthase kinase 3β/Glycogen synthase kinase 3β in the Streptozotocin dementia rats were significantly lower (p<0.05), and p-Tau/Tau were significantly increased (p<0.05). Abnormal increasing neural precursor cell expressed developmentally down-regulated protein 4.1 in brain injury tissue induced the rising ubiquitination of Insulin-like growth factor-1Rβ and rising of Aβ amount and Tau protein phosphorylation. C1060 site in Insulin-like growth factor-1Rβ was necessary for the Neural precursor cell expressed developmentally down-regulated protein 4.1 of ubiquitination and degradation of Insulin-like growth factor-1Rβ. In addition, Neural precursor cell expressed developmentally down- regulated protein 4.1 affects Insulin-like growth factor-1 signal pathway in brain, such as inhibiting Protein kinase B and Glycogen synthase kinase 3β phosphorylation, promoting Tau protein phosphorylation, and promoting β-amyloid secretions. The Role of Neural Precursor Cell Expressed Developmentally Down-Regulated Protein 4.1- Mediated Insulin-Like Growth Factor-1Rβ Ubiquitination in the Pathogenesis of Alzheimer’s disease