Analysis Of Pd-related Phenotypes And Mechanisms Of A53T ?-Syn And G2019S LRRK2 Trangenic Mice

Parkinson's disease(PD)is the second most common neurodegenerative disease and is characterized by loss of dopaminergic neurons in the substantia nigra and misfolding of?-Synuclein,which may be caused by mitochondrial dysfunction,oxidative stress,Excitatory poisoning,apoptosis,inflammatory response and proteasome disorders.Mutations in LRRK2 and ?-Syn can cause familial autosomal dominant PD.The underlying pathophysiological interaction between LRRK2 and ?-Syn is still unclear.LRRK2 can impair the microtubule dynamics,Golgi complex structure,ubiquitin-proteasome pathway,and indirectly promote ?-Syn accumulation and aggregation.Resveratrol has anti-tumor,cardiovascular,anti-inflammatory,anti-apoptotic,anti-oxidation,anti-free radical effects,and can cross the blood-brain barrier into the brain,and is expected to become an effective dopaminergic neuron protective agent.However,the specific mechanism remains to be studied.Objective:(1)To investigate the effect of G2019 S LRRK2 on the pathological changes of ?-Syn in A53 T ?-Syn transgenic mice,and the effect of mutations in the G2019 S LRRK2 and A53 T ?-Syn mutations on PD mice.(2)The protective mechanism of resveratrol on A53T/G2019 S double transgenic mice.Methods:(1)The behaviors of 4-18 months of WT,A53 T HET,G2019 S HOMO,G2019 S HET,and A53T/G2019 S transgenic mice were detected by using open field test,rotarod test,and pole test,respectively.(2)The contents of DA,DOPAC and HVA in the striatum of 5 genotype mice were detected by HPLC-ECD.(3)The expression of P-LRRK2,TH,DAT and VMAT2 protein in the striatum and substantia nigra of 5genotype mice was detected by Western blot.(4)Stereotactic injection of HSV G2019 S to construct a virus model.(5)The behaviors of the control groups and the GS2 groups were tested by using open field test,rotarod test,and pole test,respectively.(6)The IHC method was used for TH staining of the striatum and substantia nigra regions.(7)Western blot wasused to detect the expression of TH,?-Synuclein,DAT,LC3,Nrf2 and other proteins in the striatum and substantia nigra of A53T/G2019 S mice before and after drug delivery.Results:(1)Behavioral experiments showed that the A53 T HET,G2019 S HOMO,G2019 S HET,and A53T/G2019 S transgenic mice had lower movement distances and fewer central entries than the WT groups,and took a long time to fall down.The contents of DA,DOPAC,HVA,and TH were lower than those of WT groups,but the levels of DA synthesis and metabolism(DOPAC/DA),metabolism and release(HVA/DA)were higher than those of WT groups.(2)The behavioral performance and DA,DOPAC,TH,DAT and VMAT2 of G2019 S HOMO groups were lower than those of G2019 S HET groups,and the behavioral performance was the worst among the five groups.(3)The A53T/G2019 S groups had lower behavioral performance,DA,DOPAC,HVA,TH,DAT and VMAT2 levels than the A53 T HET groups and were the lowest among the five groups and showed a tendency to decrease with age,but did not have the corresponding lowest behavioral performance.The early mortality rate of A53T/G2019 S groups was the highest among the five groups.(4)The GS2 virus models had lower behavioral performance than the control groups,and the loss of TH neurons in the substantia nigra was evident,and the DA fiber density in the striatum was reduced.(5)Western blot showed increase of Beclin1,decrease of P62,activation of LC3-II,degradation of ?-Synuclein,decrease of P-LRRK2 and increase of TH,Nrf2 and GCLC after administration in A53T/G2019 S groups.Conclusions:(1)The activities of autonomous activity,exploration ability and excitability,DA and TH contents in A53 T HET,G2019 S HOMO,G2019 S HET and A53T/G2019 S transgenic mice were lower than those in WT groups.(2)The PD symptoms in the G2019 S HOMO groups were significantly higher than that in G2019 S HET groups,indicating that homozygotes were more pathogenic.(3)The behavior,DA and TH contents of A53T/G2019 S groups were lower than that of A53 T HET groups,and the early mortality was higher,which may be mediated by G2019 S LRRK2.(4)The virus injection models has good PD symptoms and can be used for future experimental studies.(5)Resveratrol activates the autophagic pathway that has been inhibited to a certain extent,enhances the antioxidant capacity,reduces the deposition of ?-Syn protein and phosphorylation of LRRK2,and thus alleviates the PD symptoms of A53T/G2019 Stransgenic mice.The protective effect may be better with optimized administration time and earlier mice age.