Neuroprotective Effect Of Astragaloside IV To Parkinson’s Disease

Parkinson’s disease (PD) is a central nervous system degenerative diseasewith the characteristic pathological changes of mesencephalic substantia nigradopaminergic neuron degeneration and necrosis and its main clinicalmanifestations are myotonia, resting tremor, motor retardation and attitudedisturbance and so on. The reasons of nigral neuronal cell death in Parkinsondisease are still not entirely clear, generally thought to be related toenvironmental toxins, gene mutation, genetic factors, oxidative stress, immunesystem abnormalities, iron ion aggregation and neuronal excitotoxicity. With theintensive aging of the population, represented by the PD central nervousdegenerative disease has become one of the major lethal diseases only aftercardiovascular diseases, malignant tumor and stroke, causing serious burden tothe patients and the whole society. Therefore, to explore the pathogenesis of the nervous system degenerative disease and to find the best treatment strategieshave become the current hot topic in the field of neuroscience research.As the commonly used traditional Chinese medicine Astragaluscharacteristic constituents, Astragaloside IV has proven to have a variety ofpositive pharmacological effect. Several researches showed that AstragalosideIV played an important role in anti-inflammatory, antioxidant and anti-apoptosis.We explored the function of Astragaloside IV in2006and found thatAstragaloside IV had a protective effect on blood-brain barrier injury caused bybrain ischemia reperfusion, and even had anti-inflammatory effects. Recentstudies have shown, Astragaloside IV contributes to CNS cell differentiation andaxon growth, and protects against the neurotoxin induced oxidative injury.Whether Astragaloside IV can protect mesencephalic substantia nigradopaminergic neuron against apoptosis in Parkinson’s disease? What is thespecific molecular mechanism?We used C57BL mouse to establish PD model by intraperitoneal injection ofMPTP, and used SH-SY5Y cells treated with MPP~+to be the Parkinson model invitro. Both of the two PD models were treated with Astragaloside IVrespectively; and we detected PD mouse behavior changes through the tractiontest and open field test; then we used immunohistochemistry, Western blot assayto detect the neuroprotective effect and its mechanism of Astragaloside IV onPD model.Firstly, we succeed prepared the PD model induced by MPTP in C57BLmice, and we observed that Astragaloside IV could improve the behavioralimpairment symptoms caused by MPTP on PD mice. In the traction experiment,the control group of mice could grasp the wire with four limbs, but the PDmodel group could only use the forepaw to grip the wire; the group treated with Astragaloside IV could grasp the wire with one or two after claws, withsignificantly higher score than those in the model group; and the score is higherin high dose group than that in low dose group. In the open field test, theexercise frequency and the number of standing significantly reduced in MPTPtreated group, however the exercise frequency and standing times increasedsignificantly in the group treated with Astragaloside IV compared with themodel group.We performed immunohistochemical staining in the substantianigra compacta, and found that the dopamine neurons increased in both lowdose and high dose group treated with Astragaloside IV compared with MPTPmodel group. We used Western blot for further analysis, and found that theexpression of TH was higher in Astragaloside IV treated group than in theMPTP model group.In the second part, we used SH-SY5Y cells induced by MPP~+to establish theParkinson model in vitro to be the research object, and further explored theneuronal protection effects of Astragaloside IV. The results showed that thecellular viability in SH-SY5Y cells treated with0-6mM MPP~+was decreasedwith dose decrease. The cell viability was50%treated with3mM MPP~+for24h.There was no obvious toxicity effect on SH-SY5Y treated with1-50μMAstragaloside IV. The pretreatment of25μM or50μM Astragaloside IV onSH-SY5Y cells could significantly reduce cell damage induced by MPP~+. TheHoechst33258staining results showed that SH-SY5Y cell nucleus pyknosisafter MPP~+treatment, and the nuclear morphous recovered after treated withAstragaloside IV.We further explored neuroprotective mechanism of Astragaloside IV onanimal tissue at the third part. Through Western blot analysis, we found thatAstragaloside IV treatment group could obviously inhibit MPTP induced the expression of apoptosis precursor protein Bax increased, and reduced Bcl-2expression level, and also could inhibit the expression level of downstreamapoptosis protein caspase-3. We did the same experiments in the cell model, andAstragaloside IV treatment could also obviously inhibit the expression of Bax inMPP~+induced SH-SY5Y cells, reduce the expression of Bcl-2, and inhibit theactivation of caspase-3. In order to confirm the exact mechanism of theneuroprotection effects of Astragaloside IV further, we designed BAX siRNAaccording to Lee’s report, and downregulated the expression of the endogenousBAX. Our results showed that Astragaloside IV prevented dopaminergicneuronal death via the inhibition of Bax-mediated pathways. In order to studythe neuroprotective mechanisms of Astragaloside IV, we also measured the ROSactivity in SH-SY5Y cells. The results showed that, Astragaloside IV couldobvious inhibit the excessive production of ROS in SH-SY5Y cells induced byMPP~+.In a word, in this work we used morphology, immunology, and molecularbiology experiment methods to confirm the neuroprotection effects ofAstragaloside IV on Parkinson’s disease at the first time, and explored the exactmolecular mechanism. It will lay the foundation for further research onneuroprotection effects and application of Astragaloside IV in Parkinson’sdisease or other neuron degeneration disease.