The Preliminary Experimental Study Of Ad-hSYN1-DHCR24 Recombinant Adenovirus On Pharmacodynamics Of Alzheimer’s Disease

Many of studies have proposed Alzheimer’s disease(AD)is “type 3 diabetes”recent years.These studies pointed out that insulin-related cell survival signaling pathway was impaired or abnormal in the brain of AD patients.Clinical studies also showed that the intranasal insulin therapy may play a therapeutic effect for AD patients and the patients with mild cognitive impairment.3β-Hydroxysteroid-Δ24reductase(DHCR24)is a multifunctional enzyme that possesses anti-apoptotic and cholesterol-synthesizing activities.DHCR24 could play an important role in insulin-Akt cell survival signaling through maintaining the cholesterol biosynthesis and normal structure and function of caveolae in mouse embryonic fibroblasts.To explore the possibility of DHCR24 as a target of gene therap of AD,we up-regulated DHCR24 expression by Adenoviruses especially in neuron at cellular and animal level,and then analyzed the changes of structures of caveolae in neuronal cell membrane and insulin/IGF-1 cell signalings which are related to Alzheimer Disease.First of all,we cultured N2 A cells,rat embryonic cortical primary culture cells and infected them with adenovirus Ad-h SYN1-DHCR24,or intracerebroventricular injected the adenobiruses to the brain of Rat.The adenovirus Ad-lac Z were used as the control.The Western blotting results demonstrated that the Ad-h SYN1-DHCR24 could up-regulate the expression of DHCR24 at cellular and animal level.Next,we studied the effect of Ad-h SYN1-DHCR24 on rescuing neuron from apoptosis.We found that Ad-h SYN1-DHCR24 infected cells were resistant to Amyloid β(Aβ)-induced apoptosis,if compared to the control.The similar results were observed when using the TM as the Endoplastic reticulum stress-inducer and H2O2 as the oxidative stress-inducer.Our previous studies have shown that lower cholesterol levels may destroy the structure of caveolae on plasma membrane and cause cell apoptosis in mouse embryonic fibroblasts.Therefore,we focused on the mechanism of how Ad-h SYN1-DHCR24 plays a cytoprotective role on caveolae and insulin signaling pathways.We observed that Ad-h SYN1-DHCR24 could significantlyimprove the neuron-protective effect of insulin like growth factor IGF-1 in N2 A cells.Western Blotting results show that the protective effect of Ad-h SYN1-DHCR24 is through enhancing the activation of IGF-IRS-Akt cell survival signaling pathway.Additionally,intracellular cholesterol level was also elevated in the Ad-h SYN1-DHCR24-infected cells,accompanied by a well-organized formation of caveolae on the plasma membrane,and improved colocalization of caveolin-1 and insulin-like growth factor 1 receptor in N2 A cells.Finaly,we proved for the first time that IGF-1 receptor can well locate in caveolae in the hippocampus of rat brain.Taken together,the present study has proved that the Ad-h SYN1-DHCR24 could up-regulate the expression of DHCR24 at cellular and animal level.The overexpression of DHCR24 can protect neuronal cells from apoptosis induced by ER stress,oxidative stress and A treatment.DHCR24 could also improve the neuron-protective effects of IGF-1 through elevating cellular cholesterol levels thus maintaining caveolae structure and enhancing IGF-IRS-Akt signaling pathway.The present study not only provides the evidences of DHCR24 as a new gene therapy of AD but provides a new insight for developing Ad-h SYN1-DHCR24 to a new drug for gene therapy of AD.