| Purpose: Parkinson's disease(PD)is a common degenerative disease of the central nervous system(CNS).Its main pathological feature is the loss of dopaminergic neurons and the appearance of Lewy bodies in the substantia nigra pars compacta(SNpc),thus leading to dopamine(DA)depletion in the striatum.More and more evidences have suggested that iron play a key role in the pathogenesis of PD.The oxidative stress caused by abnormal accumulation of iron in the substantia nigra(SN)eventually leads to the death of neurons,which is one of the major pathogenic mechanisms of PD.There are two pathways for iron uptake in the brain.The first one is transferrin(Tf)-Tf receptor(TfR)pathway,the other one is Non-transferrin pathway which is mainly mediated by divalent metal transporter 1(DMT1).Iron release is through ferroportin1(FPN1),which is the only iron export protein up to now.For oligodendrocytes,no matter the undifferentiated or differentiated type,TfR1 is responsible for iron uptake.In the central nervous system,oligodendrocytes are the major iron-staining positive cells.During the maturation of oligodendrocytes,a large amount of iron is needed.The mature oligodendrocytes in PD SN stained iron more than any other cells.Mature oligodendrocytes differentiated from oligodendrocyte precursor cells(OPCs).Current studies have found that OPCs account for more than 70% of the dividing cells of the central nervous system,and OPCs require a large amount of iron during maturation.Therefore,it is of great significance to study the iron metabolism of oligodendrocytes to understand the iron accumulation in PD.Previous studies in our laboratory showed that 6-hydroxydopamine(6-OHDA)up-regulated DMT1 and down-regulated FPN1,which was caused by IRPs activation in neurons;6-OHDA promoted iron traffic in astrocytes by up-regulating both iron import protein DMT1 and iron export proteinFPN1 through hypoxia-inducible factors-2?(HIF-2?);When 6-OHDA treated in microglia,the expression of DMT1 increased,FPN1 expression remained unchanged,DMT1 was regulated by IRP1 activation,while FPN1 was regulated by IRP1 and hepcidin.Then,how will 6-OHDA regulate iron metabolism in oligodendrocytes? What is the role of oligodendrocytes differentiation in the development of PD?Method: In this study,we used MTT,immunofluorescence staining,real-time fluorescence quantitative PCR,laser confocal microscopy,and western blots in human MO3.13 oligodendrocytes,as well as primary cultured rat OPCs to investigate the effec t of 6-OHDA on the expressions of iron-related proteins in oligodendrocytes and its possible mechanisms.Results:1.After treatment of undifferentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,The protein expression of IRP1 was significantly increased,compared with the control group(10 ?M,P<0.05,n=6;100 ?M,P<0.01,n=3).The protein expression of FPN1 was significantly decreased,compared with the control group(10 ?M,P<0.001,n=7;100 ?M,P<0.001,n=3).The protein expression of TfR1 was significantly increased,compared with the control group(10 ?M,P<0.05,n=4;100 ?M,P<0.001,n=4).2.After treatment of undifferentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,the mRN A expression of FPN1 was significantly decreased,compared with the control group(10 ?M,P<0.001,n=4;100 ?M,P<0.001,n=4)and 10?M group(P<0.01,n=4).The mRN A expression o f TfR1 was significantly increased,compared with the control group(10 ?M,P<0.05,n=4;100 ?M,P<0.05,n=4)and 10 ?M group(P<0.05,n=4).3.After treatment of undifferentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,iron uptake in undifferentiated MO3.13 cells was increased compared with the control group(10 ?M,P<0.001,n=4;100 ?M,P<0.001,n=4).4.After treatment of undifferentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,the mRNA expression of IL-1? in the experimental group at 10 ?M did not change;the mRNA expression of IL-1? in the 100 ?M 6-OHDA group increased significantly,compared with the control group(P<0.01,n=5)and 10 ?M group(P<0.05,n=5).The mRNA expression of TNF-? was significantly increased,compared with the control group(10 ?M,P<0.05,n=3;100 ?M,P<0.01,n=4).5.After treatment of differentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,the protein expression of IRP1 was significantly decreased,compared with the control group(10 ?M,P<0.05,n=3;100 ?M,P<0.01,n=4).The protein expression of FPN1 was significantly increased,compared with the control group(10 ?M,P<0.05,n=4;100 ?M,P<0.01,n=4).The protein expression of TfR1 was significantly decreased,compared with the control group(10 ?M,P<0.01,n=4;100 ?M,P<0.05,n=3).6.After treatment of differentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,the mRNA expression of FPN1 was significantly increased,compared with the control group(10 ?M,P<0.01,n=4;100 ?M,P<0.05,n=3).The mRN A expression of TfR1 was significantly decreased,compared with the control group(10 ?M,P<0.05,n=3;100 ?M,P<0.05,n=5).7.After treatment of differentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,iron uptake in differentia ted MO3.13 cells was decreased compared with the control group(10 ?M,P<0.01,n=5;100 ?M,P<0.001,n=5).8.After treatment of differentiated MO3.13 cells with 10 ?M and 100 ?M 6-OHDA for 24 h,the mRN A expression of IL-1? in the experimental group at 10 ?M did not change;the mRNA expression of IL-1? in 100 ?M 6-OHDA group increased significantly,compared with the control group(P<0.01,n=5)and 10 ?M group(P<0.01,n=3).The mRNA expression of TNF-? was significantly decreased,compared with the control group(10 ?M,P<0.01,n=5;100 ?M,P<0.001,n=4).9.The numbers of OPCs of SN in the lesioned side of 6-OHDA unilaterally lesioned rats were significantly increased compared with the control group(P<0.05,n=5).10.After treatment of primary cultured rat OPCs with 10 ?M 6-OHDA for 24 h,the protein expression of IRP1 was significantly increased,compared with the control group(P<0.05,n=3).The protein expression of FPN1 was significantly decreased,compared with the control group,(P<0.05,n=5).The protein expression of TfR1 was significantly increased,compared with the control group(P<0.01,n=4).In conclusion: In summary,in undifferentiated MO3.13 oligodendrocytes,6-OHDA up-regulated TfR1 and down-regulated FPN1,which were induced by IRP1 activation.In differentiated MO3.13 oligodendrocytes,6-OHDA down-regulated TfR1 and up-regulated FPN1,which were induced by IRP1 inactivation.6-OHDA increased the expression of IL-1? and TNF-? mRNA in undifferentiated MO3.13 oligodendrocytes,however,6-OHDA increased the expression of IL-1? mRNA but decreased the expression of TNF-? mRNA in differentiated MO3.13 oligodendrocytes.6-OHDA promoted proliferation of OPCs.The results suggest that,6-OHDA could increase iron levels and pro-inflammatory factors,promote proliferation of OPCs.When oligodendrocytes have differentiated,the need for iron decreases and they will not uptake lots of iron anymore.Therefore,iron deposition in PD SN is mainly in oligodendrocytes,and its mechanism may be related to oxidative stress-induced proliferation and differentiation of OPCs,leading to increased cellular iron demand. |
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