| Objective:To explore the protective effect of carnosic acid on MPTP-induced Parkinson’s model in vivo(C57BL/6 mouse model)and in vitro(neuron model).Methods:1.Culture SH-SY5Y cells in vitro,add different drugs(drugs A,B,C,D,E,carnosic acid and solvent),use MPTP to damage the cells,and use cell viability detection kit(CCK-8)to detect and treat After the cell viability,an active drug(carnosic acid)was screened out.SH-SY5Y cells were cultured in vitro and divided into 9 groups,namely control group,model group,drug group(CA low,medium,and high dose group),blank control group and positive control group.The CCK-8 kit was used for detection and treatment After the cell viability.2.Take C57BL/6 pregnant mice from 18 days of gestation and enter the cell culture room through the delivery window.After the pregnant rats were killed by necking them,they were sprayed with 75%alcohol for disinfection,and then the fetal rats were taken out,and the hippocampus and cortex of the fetal rats’brains were taken out under aseptic operation to isolate and purify neurons.Culture neurons in an incubator,change the medium half after 24 hours(replace a fresh medium with half the volume of the medium),add cytarabine(2.5 mg/L)after 72 hours to continue the culture,and change to fresh after 120 hours B-27 plus neurobasal medium medium,continue to cultivate.Used for experiments after 7 days.Divide neurons into 9 groups.Cell viability detection kit(CCK-8)was used to detect neuronal viability.Observe the cell morphology under a confocal microscope to detect the release of lactate dehydrogenase(LDH),the content of superoxide dismutase(SOD)and the content of malondialdehyde(MDA).The expression of neuronal apoptosis-related genes caspase-3 mRNA,total caspase-3 protein and spliced caspase-3 protein were detected.3.Divide 90 C57BL/6 mice into six groups randomly,namely the control group,the drug group(CA low,medium,and high dose groups),the positive control group and the negative control group(MPTP group).The administration was continued for 28 day s,and the mice were weighed daily to observe the changes in the weight of the mice;the open field test was used to detect the autonomous activity level of the mice on the 1,7,and 28 days after the administration of the model.Climbing rod experiment was used to detect the coordination level of the mouse limbs on days 1,7,and 28 after administration of the model;Western Blot was used to analyze the brains of mice in each group on the first 1,7,and 28 days after administration of the model.The expression of tyrosine hydroxylase(TH),α-Synuclein and caspase-3 protein on 28 days.Results:1.Carnosic acid can protect MPTP-induced SH-SY5Y cell damage;carnosic acid can protect MPTP-induced neuronal damage,reduce LDH leakage in the culture medium,reduce intracellular MD A production,enhance SOD activity,and prevent caspase-3 Abnormal activation of mRNA and spliced caspase-3.2.The weight loss of Parkinson’s disease mice after MPTP modeling,carnosic acid can partially improve the weight loss of MPTP-induced Parkinson’s disease mice;carnosic acid can increase the autonomous activity level of Parkinson’s disease mice and improve Parkinson’s disease The level of limb movement coordination of diseased mice;it can improve the abnormal expression of α-Synuclein,caspase-3 mRNA and TH protein in the brain of Parkinson’s disease miceConclusions:1.Carnosic acid can protect SH-SY5Y cells and dopamine neurons from damage induced by MPTP.The mechanism may be related to inhibiting neuronal oxidative damage and preventing the abnormal increase of caspase-3.2.Carnosic acid can improve the pathological changes of MPTP-induced Parkinson’s mouse model and improve behavioral abnormalities.The mechanism may be related to inhibiting neuronal oxidative damage and preventing the abnormal increase of apoptosis-related protein caspase-3. |
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