Study On The Mechanism Of PGC-1α Improving Alzheimer’s Disease By Regulating The Expression Of VDR And Inhibiting Oxidative Stress

Alzheimer’s disease（AD）is a common chronic age-related neurodegenerative disease with many inducements and complex pathogenesis,and it is characterized by progressive impairment of cognition and behavior.The accumulation and deposition of Amyloidβ（Aβ）is a typical histomathological feature of AD patients.Notably,Aβdeposition is also considered to be a key factor of the disease occurrence.Recent studies have shown that Vitamin D receptor（VDR）gene polymorphism is associated with AD susceptibility,and neurons are more susceptible to be attacked by reactive oxygen species after VDR inhibition.Being one of the transcriptional coactivators of VDR,Peroxisome proliferator-activated receptorγcoactivator-1（PGC-1α）could protect cells against oxidative stress mediated neurodegeneration by regulating the generation of mitochondria and the gene transcription of antioxidant enzymes.However,the role of PGC-1αin AD,as well as its precise involvement of VDR in the neuroprotective strategy are largely unknown.Objectives（1）To investigate VDR and oxidative stress level in the cerebral cortex and the hippocampus of AD mice;（2）To explore the relationship between PGC-1αexpression and pathology effect of AD;（3）To study the target intervention effect of PGC-1αto VDR,as well as how PGC-1regulating the synaptic plasticity and oxidative stress in AD;（4）To investigate whether neuroprotective function of PGC-1αis mediated,partly by activating VDR expression and improving oxidative damage in AD.Methods（1）Using the APP/PS-1 double transgenic（2x Tg-AD）mice at 6 months of age,we studied the levels of Aβdeposition in the hippocampus and the cortex by immunohischemistry and VDR expression by immunofluorescence and Western Blotting,as well as examined antioxidative enzymes,including Superoxide dismutase2（SOD₂）and Glutathione peroxidase 1（GPX₁）by Western blotting to investigate the effect of VDR in AD,and to explore effectiveness of AD transgenic model.（2）Further,expression levels of PGC-1αin the cortex and the hippocampus of APP/PS-1 mice were examined by immunofluorescence and Western Blotting.（3）A specific mouse line,Dlx5/6-Cre:PGC-1α^fl/fl in which the PGC-1αdeficiency was limited to the hippocampus and the cortex were generated by Cre/Loxp mediated gene recombination technology,the mitochondrial ultrastructure and synaptic plasticity in the cortex and the hippocampus were detected by transmission electron microscopy.And then we used immunohistochemistry to measured the expression of 8-oxo-7,8-dihydro-20-deoxyguanine（8-oxo-d G）,a product of DNA oxidative damage in the cortex and the hippocampus.VDR levels in the cortex and hippocampus were also detected by Western Blotting.（4）Microinjection technique in targeted brain region was used to force PGC-1αoverexpression in hippocampal CA1 region of APP/PS-1 mice（HE staining was used to check the accuracy of the injection site）.After three weeks of virus expression,we firstly detect the HA tag and PGC-1αprotein expression level by immunofluorescence and Western Blotting to verify whether PGC-1αwas successfully overexpressed.Secondly,we used immunofluorescence and immunohistochemistry to detect the amount and area fraction of Aβdeposition in APP/PS-1 mice.Next,we also detected the expression of VDR by Western Blotting,and the expression of 8-oxo-d G was measured by immunohistochemistry.Therefore,clarifing whether PGC-1αcan improve the oxidative stress and pathological abnormality of AD by regulating the expression of VDR.Results（1）A remarkable rise in levels of Aβdeposition and a significant reduction in VDR and antioxidatiative enzymes,including SOD₂ and GPX₁ were found in APP/PS-1 mice at 6 months of age,the varies were specifically observed in cerebral cortex and hippocampus,especially in its CA1 and DG regions;（2）The expression of PGC-1αwas reduced in cerebral cortex and hippocampus,especially in its CA1 and DG regions of AD mice;（3）Abnormal mitochondrial morphology was observed in the cortex and the hippocampus of PGC-1αconditioned knockout mice,reduced synapse,broadened synaptic gap and decreased PSD were available in the cortex of the genotype mice.Further,the increased oxidative damage,as revealed by increased expression of DNA damage marker 8-oxo-d G,and reduced expression of VDR protein were found in PGC-1αconditioned knockout mice;（4）PGC-1αcould protect cells against oxidative stress.Specifically,overexpressed PGC-1αincrease the expression of VDR protein,decreased oxidative stress damage and Aβdeposition in the cortex and the hippocampus,which is possibly mediated by activating PGC-1α/VDR pathway.ConclusionsThe occurrence of AD is accompanied by downregulation of PGC-1αand VDR and oxidative stress.Further,the distribution and expression of PGC-1αand VDR in the central nervous system are consistent,and PGC-1αcan regulate the expression of VDR,forced overexpression of PGC-1αcan improve oxidative stress and Aβdeposition in AD by targeting VDR.PGC-1α/VDR signaling pathway provides a new research idea for the early diagnosis,intervention and treatment of AD.