Cytological And In Vivo Studies On The Relationship Between Selenoprotein MSelK And Alzheimer's Disease

Alzheimer's Disease?AD?,commonly known as Alzheimer's disease,is a multiple disease in the elderly population that can cause deterioration of cognitive function.The prevention and treatment of Alzheimer's disease is a problem that needs to be solved urgently.The pathological features of Alzheimer's disease are senile plaques formed by the accumulation of A??Amyloid beta?protein as a core,dystrophic neuronal fibers entangled around them,and a series of manifestations associated with neuronal loss,resulting in cognitive dysfunction.Many studies have found that aggregated chemotaxis of microglial cells surrounding A?and reacting with A?to produce inflammatory cytokines.Therefore,microglial cells are thought to play an important role in the occurrence and development of Alzheimer's disease.Selenoprotein mSelK is a kind of high expression of selenoprotein in the human brain.The level of mSelK expression has a great influence on the phagocytosis and migration of microglial cells.Whether mSelK is associated with AD,and what is the signaling pathway of mSelK affecting the phagocytosis and migration of microglial cells are key scientific problems to be solved in this experiment.This subject has studied the above problems through in vitro and in vivo experiments in mice.Signalling pathway studies have shown that the knockdown/overexpression of mSelK?mouse mSelK?significantly reduces/increases the expression of the endoplasmic reticulum calcium ion release channel protein IP₃R?Inositol 1,4,5-trisphosphate receptor?,and cytosolic free calcium levels are significantly reduced/increased.After adding the calcium channel antagonist 2-APB to the medium or infecting the microglial cells with the IP₃R knockdown adenoviral expression vector,the elevation of the migration,phagocytosis and the free calcium level in the cytoplasm of microglial cells caused by mSelK overexpression could be reduced,these evidences further proved that mSelK plays a role through regulating the expression of IP₃R,a calcium release channel in the endoplasmic reticulum.The CRISPR-Cas9 technique was used to construct mSelK knockout mice.First,the mSelK gene was knocked out by CRISPR-Cas9 technology to obtain heterozygotes of mSelK knockout mice.The knockout homozygotes of mSelK knockout mice were then obtained by hybridization between a 1 bp deletion and a 1bp deletion heterozygote of mSelK knockout mice,a 2 bp deletion and a 2 bp deletion heterozygote of mSelK knockout mice.Homozygous mice were identified by PCR amplification and sequencing.A batch of mSelK knockout mice were obtained.Brain injections of A?1-42 and AlCl₃ were performed on old mice.Water maze experiments showed that the injection of AlCl₃ and A?1-42 could impair the cognitive ability and behavior of mice,and the incidence of AD increased significantly.A?1-42 activation of microglial cells was observed by microglial immunohistochemical experiments,suggesting that an inflammatory reaction was produced in the brain.Western Blot experiments showed that AlCl₃ and A?1-42reduced the expression of selenoprotein mSelK in the brain of injected mice,while peroxidase GPx1?Glutathione peroxidase 1?did not change significantly.These studies laid the foundation for the further researches on the role of selenoprotein mSelK in the prevention of Alzheimer's disease.Through the above experimental study,the following conclusions are drawn:Selenium affects the microglial cells mainly through mSelK.The increase/decrease of mSelK expression increases/decreases the level of free calcium ion in the cells by regulating the increase/decrease of IP₃R,further increases/decreases the phagocytic and migrating ability of microglial cells,and then affects the scavenging capacity of microglial cells to A?polypeptide.The injection of AlCl₃ and A?1-42 reduced the expression of mSelK in brain.Reduced expression of mSelK in mice will make the mice more susceptible to AD,and selenium supplementation may help prevent AD development.The above studies revealed the important role of microglial cells and selenoprotein mSelK in the development of AD and the significance of selenium supplementation in humans,which contributes to the prevention and treatment of AD.