Effects And Mechanism Of Lipid On Hippocampal Neurons And PC12Cells Apoptosis Mediated By PCSK9

Alzheimer’s disease(AD) is one of the most common type of dementia, caused by a varietyof factors and highly correlated with age. Progressive cognitive and memory damage incentral nervous system is the major characteristic of AD. Neuronal apoptosis is one of itsmain pathological features. Hyperlipidemia is a risk factor of AD and other neurodegenerativediseases, but the mechanism of hyperlipidemia and neurodegenerative diseases have not beenfully clarified. Our research focus on the effect and mechanism of neuronal apoptosis inhyperlipidemia, to explore the mechanism between hyperlipidemia and AD.AIM: To explore the role and mechanism of PCSK9in hippocampal neuron apoptosis ofapoE (-/-) mice induced by hyperlipidemia and observe expression of PCSK9in PC12cellsapoptosis induced by different concentration of oxLDL. To study molecular mechanism ofPCSK9mediated neuron apoptosis induced by lipid via inhibiting PCSK9expression of PC12cells with oxLDL treatment.Methods: The experiment was divided into two parts: in vivo experiment and in vitroexperiment. Eighteen apoE (-/-) mice of six weeks old were randomly divided into thenormal diet group and high fat diet group. After twelve weeks of feeding, the hippocamptissue was taken out for frozen section. HE staining, oil red O staining, Hoechst33258,immunohistochemistry staining and Bielschowsky staining were used to observe the changesof hippocamp. Different concentrations of oxLDL were used to treat PC12cells24h. Theapoptosis of PC12cells was observed by Hoechst33258staining. RT-PCR and Western blotwere used to detect mRNA and protein level of PCSK9and BACE1, respectively. The contentof Aβ40and Aβ42were detected by ELISA. After48hours transfected by PCSK9siRNAs,PC12cell was incubated for24hours with oxLDL, Hoechst33258staining and flowcytometry were used to detect apoptosis change. Western blot was used to examine PCSK9, BACE1, Caspase3, Caspasee9, Bcl-2and Bax expression. The content of Aβ40and Aβ42were detected also by ELISA.Results: Compared with the normal diet group, pyramidal cells in hippocamp of apoE (-/-)mice of high fat diet group was disordered. In addition the intercellular space was increased,cell swelled and nuclear size became smaller and hyperchromatic, showed karyopyknosis,especially in CA3region. CA1region lesions was not significant change. Lipid and senileplaque of apoE (-/-) mice hippocamp of high fat diet group increased slightly. Furthermoreapoptosis, PCSK9, BACE1, Caspase3and Bax expression increased obviously, only a smallincrease of Bcl-2expression.75mg/L oxLDL incubated PC12cells24h caused large cellapoptosis. Meanwhile, mRNA and protein level of PCSK9was increased in a dose dependentmanner, and the most obvious concentration of oxLDL increased PCSK9expression was75mg/L. The mRNA and protein level of BACE1and Aβ content were decreased in a dosedependent manner, and the most obvious concentration of oxLDL decreased BACE1expression was75mg/L. After transfection for48h, PC12cells were treated with oxLDL for24h, Hoechst33258staining and flow cytometer showed PCSK9-siRNA reduced apoptosismarkedly. Moreover, Caspase3, Caspasee9, Bax protein expression were dropped, BACE1,Bcl-2protein expression and Aβ40, Aβ42content were raised.Conclusion: Hyperlipidemia lead to hippocampal neuron apoptosis of apoE (-/-) mice byincreasing PCSK9expression that gave rise to activating of Bcl-2/Bax-Caspase3signalpathway, raising BACE1and Aβ levels. OxLDL has a dual regulation on neuron apoptosis,the mechanism is that PCSK9is up-regulated by a high level of oxLDL then to induce neuronapoptosis through Bcl-2/Bax-Caspase (9,3) signaling pathways, moreover, the degradation ofBACE1can inhibit the APP/Aβ metabolic pathway to reduce Aβ generation, therebyinhibiting Aβ-induced neuron apoptosis induced by Aβ.