Protecting And Mechanism Studies Of Isatin Derivative HKL-A027-182-2 On Aβ-Induced Nerve Cell Injury

Alzheimer’s disease(AD)is a hidden onset neurodegenerative disorder.Its complicated pathogenesis remains unknown.There’s still absent of effective strategies.Currently,β-amyloid(Aβ)caused neurotoxicity thought to be critical for inducing and promoting AD,and has been recognized as one of the potential therapeutic targets for AD prevention and treatment.Therefore,we established AD cell models by Aβ-induced nerve cell injury.As a result,we screened out the isatin derivative HKL-A027-182-2,which has a better protective effect on nerve cell damage,and further studied the protecting mechanism of the compound.Firstly,we selected PC-12 cells and SH-SY5Y cells as the research objects to establish AD injury model induced by Aβ25-35/Aβ1-42.Cell viability was measured by MTT.Based on these AD cell models,the best inhibitor HKL-A027-182-2 was selected,which is a kind of isatin derivative.Secondly,when studying the protecting mechanism of HKL-A027-182-2 on nerve cells,we found that:HKL-A027-182-2 could inhibit the activity of AChE at the cellular level,which resulted in the normal release of ACh and the normal development of nerve impulses.Thirdly,antioxidant capacity test results show that HKL-A027-182-2 not only has certain total reducing power and DPPH+ scavenging ability,but also can improve the activity of SOD,CAT and GSH in cells,and reduce the level of LDH and ROS,which indicates that HKL-A027-182-2 can increase the antioxidant capacity of cells.Fourthly,using flow cytometry and western blot,we found that HKL-A027-182-2 restores the decrease of mitochondrial membrane potential by reducing ROS level,thereby inhibiting the activation of Bax by Aβ,and up regulating the expression of Bcl-2 and Bcl-xL,thus preventing cell apoptosis.Fifthly,we used western blot to study its effect on Tau protein phosphorylation.The results showed that HKL-A027-182-2 inhibited the expression of p-p38 MAPK,down-regulated the phosphorylation expression of Tau at Ser396.And finally it inhibited Tau hyperphosphorylation induced by Aβ.Sixthly,HKL-A027-182-2 can down-regulate the expression of BACE and PS1,which leads to decreased expression ofβ-secretase and y-secretase,thus inhibiting the amyloid degradation pathway of APP and reducing the production of Aβ.Furthermore,HKL-A027-182-2 reduced the formation ofβ-pleated sheet and Aβ oligomer.HKL-A027-182-2 ultimately protected neurons.Finally,we studied the blood-brain barrier of HKL-A027-182-2 at the animal level.The results showed that the efficiency of HKL-A027-182-2 through the blood-brain barrier was 0.129%,so the drug concentration did not reach the therapeutic concentration.Therefore,the plan for the next research in this subject is to improve the efficiency of HKL-A027-182-2 through the blood-brain barrier,which is also a difficult problem for many drugs for treating brain diseases.In conclusion,the isatin derivative HKL-A027-182-2 can inhibit nerve cell damage caused by Aβ through multiple targets,which provides a potential treatment option for Alzheimer’s disease.This study also provides important data support for further research on Alzheimer’s disease drugs in the future.