Up-regulation of divalent metal transporter 1 is involved in

Apoptosis has been identified as one of the important mechanisms involved in the degeneration of dopaminergic neurons in Parkinson’s disease (PD). Our previous study showed increased iron levels in the substantia nigra as well as loss of dopaminergic neur

Up-regulation of divalent metal transporter 1 is involved in

1-methyl-4-phenylpyridinium (MPP+)-induced apoptosis

in MES23.5 cells

Zhang Shuzhen，Wang Jun，Song Ning，Xie Junxia，Jiang Hong

Department of Physiology, Medical College of Qingdao University, Qingdao (266071)

E-mail:

Abstract

Apoptosis has been identified as one of the important mechanisms involved in the degeneration of dopaminergic neurons in Parkinson’s disease (PD). Our previous study showed increased iron levels in the substantia nigra as well as loss of dopaminergic neurons in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine-induced PD mouse models. 1-Methyl-4-phenylpyridinium (MPP+) is commonly used to establish a cellular model of PD. Although intracellular iron plays a crucial role in MPP+-induced apoptosis, the molecular mechanism linking increased iron and MPP+-induced neurodegeneration is largely unknown. In the present study, we investigate the involvement of divalent metal transporter 1 (DMT1) that accounts for the ferrous iron transport in MPP+-treated MES23.5 cells. In the treated cells, a significant influx of ferrous iron was observed. This resulted in a decreased mitochondrial membrane potential. Additionally, an elevated level of ROS production and activation of caspase-3 were also detected, as well as the subsequent cell apoptosis. These effects could be fully abolished by using iron chelator desferal (DFO). Increased DMT1 (―IRE) expression but not DMT1 (+IRE) accounted for the increased iron influx. However, there were no changes for iron regulatory protein 1 (IRP1), despite decreased expression of IRP2. Iron itself had no effect on IRP1 and IRP2 expression. Our data suggest that although DMT1 mRNA contains an iron responsive element, its expression is not totally controlled by this. MPP+ could up-regulate the expression of DMT1 (―IRE) in an IRE/IRP-independent manner. Our findings also show that MPP+-induced apoptosis in MES23.5 cells involves DMT1-dependent iron influx and mitochondria dysfunction.

Keywords: Divalent metal transporter 1 (DMT1); Apoptosis; Parkinson's disease; Iron; Iron regulatory protein; Iron chelator

1. Introduction

Elevated iron levels are found in the substantia nigra (SN), the brain region composed of

the dopaminergic neurons that undergo selective neurodegeneration in Parkinson’s disease (PD) (Dexter et al., 1989, Riederer et al., 1989, Dexter et al., 1993, Rouault, 2001, Xie et al., 2003, Gotz et al., 2004, Wang et al., 2004, Gal et al., 2005, Hardy et al., 2005, Berg and Hochstrasser, 2006, Jiang et al., 2006, Jiang et al., 2007, Wang et al., 2007). Accessible intracellular ferrous iron can react with hydrogen peroxide to produce the hydroxyl radicals that in turn can damage proteins, nucleic acids and lipids, leading to cell death (Kamp et al., 2002, Andersen, 2004). Since apoptosis has been implicated as one of the important mechanisms leading to the death of dopaminergic neurons in PD, 1-methyl-4-phenylpyridinium (MPP+) is employed to produce a PD cell model in vitro in this study. Our aim is to elucidate whether iron is involved in MPP+-induced apoptosis. MPP+, a metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), is selectively transported into dopaminergic neurons through the dopamine transporter and concentrated into mitochondria. MPP+ induced cell apoptosis involves multiple mechanisms, including complex I inhibition, inhibition of calcium homeostasis, opening of the mitochondrial transition pore, etc (Kalivendi et al., 2003, Deguil et al., 2007). In our previous study, we found elevated iron levels in the SN of the MPTP-induced PD mouse models, as well as the loss of dopaminergic neurons. Up-regulation of divalent metal transporter 1 (DMT1) was also observed (Jiang et al., 2003). Although transferrin receptor (TfR)-induced iron uptake has been reported to play a role in MPP+ toxicity (Kalivendi et al., 2003), whether other iron transporters are involved is still unknown.

DMT1, also known as natural resistance associated macrophage protein 2 (Nramp2), is a

Apoptosis has been identified as one of the important mechanisms involved in the degeneration of dopaminergic neurons in Parkinson’s disease (PD). Our previous study showed increased iron levels in the substantia nigra as well as loss of dopaminergic neur

widely expressed transmembrane protein (Gunshin et al., 1997). Four DMT1 isoforms are distinguished arising from their variant mRNA transcripts that vary both at their 5'- untranslational region (UTR) (starting from exon 1A or exon 1B) and their 3'-UTR (depending on the presence or absence of the iron responsive element (IRE) in the 3'-ends) (Lee et al., 1998, Hubert and Hentze, 2002, Mackenzie et al., 2007). High levels of DMT1 expression are found in some nuclei of the basal ganglia, particularly the caudate nucleus, putamen, and SN, indicating that DMT1 may account for the high iron levels in these regions (Gunshin et al., 1997, Burdo et al., 2001, Huang et al., 2004, Knutson et al., 2004, Ke et al., 2005). DMT1 is also highly expressed in the SN in PD (Andrews et al., 1999, Jiang et al., 2003, Moos and Morgan, 2004), indicating that disrupted expression of DMT1 might be involved in the iron accumulation in this area.

Due to the suggested involvement of DMT1 in PD, we investigate the expression of

DMT1 (+IRE) and DMT1 (―IRE) in MPP+-treated MES23.5 cells, a dopaminergic cell line hybridized from murine neuroblastoma-glioma N18TG2 cells with rat mesencephalic neurons exhibits several properties similar to the primary neurons originated in the SN (Crawford et al., 1992 …… 此处隐藏：44976字，全部文档内容请下载后查看。喜欢就下载吧 ……