Experimental Study On Effect And Mechanism Of Erythropoietin On Dopaminergic Neurons In Substantia Nigra Of Rat With Parkinson's Disease Induced By 6-hydroxydopamine In Vitro

PartⅠExperimental study on effect of preconditioning with recombinant human erythropoietin on prognosis of dopaminergic neurons in substantia nigra of rat with Parkinson's disease induced by 6-hydroxydopamine in vitro.Object:Study the effect of preconditioning with recombinant human erythropoietin (rhEPO) on the survival of dopaminergic(DA) neurons in substantia nigra(SN) of rat with Parkinson's disease (PD) induced by 6-hydroxydopamine(6-OHDA) in vitro. Methods: The healthy Sprague-Dawley(SD) rats were divided into 5 groups:control group, 6-OHDA group, 6-OHDA+1u/ml rhEPO group,6-OHDA+6u/ml rhEPO group, 6-OHDA+10u/ml rhEPO group. PD models in vitro were established by incubating rat midbrain slices containing SN in artificial cerebrospinal fluid II(ACSFII) containing 6-OHDA (0.05mM).The midbrain slices were preconditioned with the different concentration of rhEPO for different duration. Then observed the change of the number of tyrosine hydroxylase–immunoreactive(TH–IR) positive neurons between the different groups Results:The preconditioning with rhEPO can ameliorate the damage which was produced by 6-OHDA to DA neurons in SN of PD models in vitro. The protection of the preconditioning group with rhEPO at concentration of 6u/ml ,where the midbrain slices were incubated for 3 hours or 6 hours, is superior to the protection of the preconditioning group with rhEPO at concentration of 1u/ml where the midbrain slices were incubated for 3 hours or 6 hours ( P<0.01, P<0.05,respectively ) ;and so is the protection of the preconditioning group with rhEPO at concentration of 10u/ml ( P<0.01, P<0.05, respectively). But there is no different protection between 6u/ml rhEPO and 10u/ml rhEPO(P>0.05). Conclusion: Preconditioning with rhEPO can induce neuroprotection of DA neurons in SN of rat with PD induced by 6- OHDA in vitro. PartⅡExperimental study on effect of preconditioning with recombinant human erythropoietin on dopaminergic neuronal apoptosis in substantia nigra of rat with Parkinson's disease induced by 6-hydroxydopamine in vitro.Object: The last experimental result have showed that the preconditioning with recombinant human erythropoietin (rhEPO)induced neuroprotection of dopaminergic neurons in substantia nigra (SN) of rat with Parkinson's disease (PD) induced by 6-Hydroxydopamine (6-OHDA) in vitro.Here, study the mechanism of neuroprotection of preconditioning with rhEPO on dopaminergic neurons in SN of rat with PD induced by 6-OHDA in vitro. Methods:Eighteen healthy SD rats were divided into 3 groups:control group,6-OHDA group, 6-OHDA+6u/ml rhEPO group, and then the midbrain slices containing SN were incubated in artificial cerebrospinal fluid II (ACSFII) for 6 hours, respectively. Immunohistochemical technique was used to detect the expression of Caspase-3 in SN,and at the same time,TUNEL method was used to detect dopaminergic neuronal apoptosis.Results: The Caspase-3-IR positive cells and TUNEL positive cells in 6-OHDA+rhEPO group were less than those in 6-OHDA group respectively (P<0.01, P<0.01,respectively).Conclusion:Preconditioning with rhEPO can ameliorate the damage, which was produced by 6-OHDA,to DA neurons in SN of rat with PD induced by 6-OHDA in vitro,it seems likely that protection of rhEPO is associated with inhibition of DA neuronal apoptosis in SN. PartⅢExperimental study on effet of preconditioning with recombinant human erythropoietin on oxidative stress of dopaminergic neurons in substantia nigra of rat with Parkinson's disease induced by 6-hydroxydopamine in vitroObject: The first experimental result have showed that the preconditioning with recombinant human erythropoietin (rhEPO)induced neuroprotection of dopaminergic neurons in substantia nigra (SN) of rat with Parkinson's disease (PD) induced by 6-Hydroxydopamine (6-OHDA) in vitro.Here, study further the mechanism of neuroprotection of preconditioning with rhEPO on dopaminergic neurons in SN of rat with PD induced by 6-OHDA in vitro. Methods: PD models in vitro were established by incubating rat midbrain slices containing SN in artificial cerebrospinal fluid II(ACSFII) containing 6-OHDA (0.05mM).The midbrain slices were preconditioned with rhEPO at the concentration of 6u/ml.Then observed the change of Maleic Dialdehyde(MDA), Glutathione(GSH),Superoxide Dismutase(SOD) and Glutathione Peroxidase(GSH-PX) between the different groups. Results: The MDA decreased, and the GSH,SOD and GSH-PX increased in the preconditioning groups with rhEPO at concentration of 6u/ml when compared with those in the 6-OHDA group. Conclusion: The neuroprotection induced by preconditioning with rhEPO is possibly due to ameliorating oxidative stress.