Experimental Studies On The Synergistic Toxic Effect Of Paraquat And Lactacystin On The Nigrostriatal System And The Protective Effect Of MSCs

The exact pathogenesis of Parkinson' disease (PD) is still unknown. Thepathological hallmarks of PD are reduction or loss of dopaminergic neurons in themesencephalon and the presence of Lewy body in the degenerated neurons. It wasreported recently that the inability of ubiquitin-proteasome system to degradealpha-synuclein, ubiquitin and other unwanted proteins in substantia nigra inidiopathic PD may contribute to the formation of Lewy body. Epidemiological studiesindicate that the exposure to environmental herbicides or pesticides may increase therisk for the development of PD. Whether the chrinic exposure to paraquat (0.5mg/kg.d)can increase the risk for PD of rats with lightly impaired proteasome by promotingalpha-synuclein aggregation is not reported until now. MSCs can secret brain-derivedneurotrophic factor (BDNF) which has specific protective role for dopaminegicneurons in vitro. Whether the effect of transplanted MSCs is derived from its abilityof secreting BDNF is not reported at present. The present study was focused on: 1) thesetting up lactacystin-PD model with Lewy body and the role of proteasomedysfunction in the pathogenesis of PD, 2) the synergistic effect of paraquat andlactacystin on the development of PD, 3) the therapeutic effect and mechanism ofMSCs for the treatment of lactacystin-PD. Part Ⅰ To set up lactacystin-PD rat model with Lewy body 70 SD rats was randomly divided into five groups(n=14): NS group, 6-OHDAgroup and lactacystin group( including 0.2μg, 2μg and 8μg 3 groups), which wererespectively injected with NS, 16μg 6-OHDA and 0.2μg,2μg,8μg lactacystin bystereotaxic unilateral injection into the substantia nigral pars compacta. Thespontaneous and apomorphine-induced contralateral behaviors of rats were observed;nigral degeneration and Lewy body were viewed by HE staining and electronmicroscope; expressions of alpha-synuclein and tyrosine hydroxylase in nigral cellswere investigated by immunohistochemistry; RT-PCR was used to determine themRNA of TH; contents of dopamine(DA) and homovanillic acid(HVA) were 8博士研究生毕业论文determined by HPLC. The results showed that animals treated with NS or 0.2μglactacystin was normal. 2μg group showed a little abnormal behavior. The animals of6-OHDA group and 8 group developed progressively bradykinetic and displayedcontralateral head tilting and tremor and showed notebly apomorphine-inducedcontralateral behavior. With the increasing dosage of lactacystin, the number of nigralcells and contents of DA and HVA in striatum were gradually decreased but theexpression of alpha-synuclein increased compared to control group. The levels of THprotein and mRNA in 2μg and 8μg groups were decreased to 63.34%(P<0.001),60.55%(P<0.05)and 81.95%(P<0.001),89.53%(P<0.01) respectively comparedto control. Some of nigral cells in 8μg group contained acidophilic Lewy body withintense immune response to alpha-synuclein , however, the expression ofalpha-synuclein in 6-OHDA group reduced and had no Lewy body. Part Ⅱ The synergistic toxic effect on nigrostriatal system of paraquat and lactacystin 56 SD rats was randomly divided into four groups (n=14): NS group, lactacystingroup(calling lact group for short), paraquat group, double factors group. Animals ofparaquat group and double factors group were fed with paraquat (0.5mg/kg) for 8weeks. Animals of NS group and lact group were fed with sodium saline as control.Then, animals of lact group and double factors group were injected with 2μglactacystin by stereotaxic unilateral injection into the substantia nigral pars compacta;animals of paraquat group and NS group were injected with sodium saline as control;The behaviors, TH positive cells, expression of alpha-synuclein, contents of DA andHVA of ra...