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Expression and localization of mitochondrial ferritin mRNA in Alzheimer's disease cerebral cortex.

Wang L, Yang H, Zhao S, Sato H, Konishi Y, Beach TG, Abdelalim EM, Bisem NJ, Tooyama I - PLoS ONE (2011)

Bottom Line: The neuroprotective effect of MtF on oxidative stress induced by H(2)O(2) was measured by MTT assay.Real-time PCR and western-blot confirmed that MtF expression levels in the cerebral cortex were significantly higher in AD cases than that in control cases at both the mRNA and the protein level.Finally, MtF expression showed a significant neuroprotective effect against H2O2-induced oxidative stress (p<0.05).

View Article: PubMed Central - PubMed

Affiliation: Molecular Neuroscience Research Center, Shiga University of Medical Science, Otsu, Japan.

ABSTRACT
Mitochondrial ferritin (MtF) has been identified as a novel ferritin encoded by an intron-lacking gene with specific mitochondrial localization located on chromosome 5q23.1. MtF has been associated with neurodegenerative disorders such as Friedreich ataxia and restless leg syndrome. However, little information is available about MtF in Alzheimer's disease (AD). In this study, therefore, we investigated the expression and localization of MtF messenger RNA (mRNA) in the cerebral cortex of AD and control cases using real-time polymerase chain reaction (PCR) as well as in situ hybridization histochemistry. We also examined protein expression using western-blot assay. In addition, we used in vitro methods to further explore the effect of oxidative stress and β-amyloid peptide (Aβ) on MtF expression. To do this we examined MtF mRNA and protein expression changes in the human neuroblastoma cell line, IMR-32, after treatment with Aβ, H2O2, or both. The neuroprotective effect of MtF on oxidative stress induced by H(2)O(2) was measured by MTT assay. The in situ hybridization studies revealed that MtF mRNA was detected mainly in neurons to a lesser degree in glial cells in the cerebral cortex. The staining intensity and the number of positive cells were increased in the cerebral cortex of AD patients. Real-time PCR and western-blot confirmed that MtF expression levels in the cerebral cortex were significantly higher in AD cases than that in control cases at both the mRNA and the protein level. Cell culture experiments demonstrated that the expression of both MtF mRNA and protein were increased by treatment with H2O2 or a combination of Aβ and H2O2, but not with Aβ alone. Finally, MtF expression showed a significant neuroprotective effect against H2O2-induced oxidative stress (p<0.05). The present study suggests that MtF is involved in the pathology of AD and may play a neuroprotective role against oxidative stress.

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Effect of MtF expression on cell viability after treatment with H2O2.An MtF protein band exists with an apparent molecular mass of 22 kDa on SDS-PAGE; weak bands were detected in IMR-32 and vector-IMR-32 cells (B). Wild-type IMR-32 cells, empty vector transfectants (Vector-IMR-32), and MtF transfectants (MtF-IMR-32) were treated with 300 µM H2O2 for 30 min (A). Cell viability was measured by MTT assay. A remarkable decrease in the viability of IMR-32 and Vector- IMR-32 cells (about 50%; p<0.01, compared with control groups) was observed after treatment with 300 µM H2O2 for 30 min (A). The viability of MtF- IMR-32 cells under treatment decreased about 30%, however, the cell viability was much higher than in the control group (p<0.05). ** p<0.01 vs. non-treated cells; * p<0.05 vs. the H2O2-treated control cells.
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pone-0022325-g007: Effect of MtF expression on cell viability after treatment with H2O2.An MtF protein band exists with an apparent molecular mass of 22 kDa on SDS-PAGE; weak bands were detected in IMR-32 and vector-IMR-32 cells (B). Wild-type IMR-32 cells, empty vector transfectants (Vector-IMR-32), and MtF transfectants (MtF-IMR-32) were treated with 300 µM H2O2 for 30 min (A). Cell viability was measured by MTT assay. A remarkable decrease in the viability of IMR-32 and Vector- IMR-32 cells (about 50%; p<0.01, compared with control groups) was observed after treatment with 300 µM H2O2 for 30 min (A). The viability of MtF- IMR-32 cells under treatment decreased about 30%, however, the cell viability was much higher than in the control group (p<0.05). ** p<0.01 vs. non-treated cells; * p<0.05 vs. the H2O2-treated control cells.

Mentions: In order to investigate the possible roles of MtF in neuroprotection, wild type IMR-32 cells (IMR-32), empty vector transfectants (vector-IMR-32) and MtF transfectants (MtF-IMR-32) were incubated with or without 300 µM H2O2 for 30 min. Figures 7A and B show the results of cell viability and western blots of MtF, respectively. Similar to previous reports [15], [17], [18], an MtF protein band with an apparent molecular mass of 22 kDa was detected by SDS-PAGE. Weak bands were detected in IMR-32 and vector-IMR-32 cells (Fig. 7B). A remarkable decrease in the viability of IMR-32 and Vector-IMR-32 cells (about 50%; p<0.01, compared with the control groups) was observed after treatment with 300 µM H2O2 for 30 min (Fig. 7A). The viability of MtF-IMR-32 cells decreased about 30% after H2O2 treatment, therefore cell viability was much higher than in control groups (p<0.05). These results show that overexpression of MtF reduces the rate of cell death after H2O2 treatment.


Expression and localization of mitochondrial ferritin mRNA in Alzheimer's disease cerebral cortex.

Wang L, Yang H, Zhao S, Sato H, Konishi Y, Beach TG, Abdelalim EM, Bisem NJ, Tooyama I - PLoS ONE (2011)

Effect of MtF expression on cell viability after treatment with H2O2.An MtF protein band exists with an apparent molecular mass of 22 kDa on SDS-PAGE; weak bands were detected in IMR-32 and vector-IMR-32 cells (B). Wild-type IMR-32 cells, empty vector transfectants (Vector-IMR-32), and MtF transfectants (MtF-IMR-32) were treated with 300 µM H2O2 for 30 min (A). Cell viability was measured by MTT assay. A remarkable decrease in the viability of IMR-32 and Vector- IMR-32 cells (about 50%; p<0.01, compared with control groups) was observed after treatment with 300 µM H2O2 for 30 min (A). The viability of MtF- IMR-32 cells under treatment decreased about 30%, however, the cell viability was much higher than in the control group (p<0.05). ** p<0.01 vs. non-treated cells; * p<0.05 vs. the H2O2-treated control cells.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3140525&req=5

pone-0022325-g007: Effect of MtF expression on cell viability after treatment with H2O2.An MtF protein band exists with an apparent molecular mass of 22 kDa on SDS-PAGE; weak bands were detected in IMR-32 and vector-IMR-32 cells (B). Wild-type IMR-32 cells, empty vector transfectants (Vector-IMR-32), and MtF transfectants (MtF-IMR-32) were treated with 300 µM H2O2 for 30 min (A). Cell viability was measured by MTT assay. A remarkable decrease in the viability of IMR-32 and Vector- IMR-32 cells (about 50%; p<0.01, compared with control groups) was observed after treatment with 300 µM H2O2 for 30 min (A). The viability of MtF- IMR-32 cells under treatment decreased about 30%, however, the cell viability was much higher than in the control group (p<0.05). ** p<0.01 vs. non-treated cells; * p<0.05 vs. the H2O2-treated control cells.
Mentions: In order to investigate the possible roles of MtF in neuroprotection, wild type IMR-32 cells (IMR-32), empty vector transfectants (vector-IMR-32) and MtF transfectants (MtF-IMR-32) were incubated with or without 300 µM H2O2 for 30 min. Figures 7A and B show the results of cell viability and western blots of MtF, respectively. Similar to previous reports [15], [17], [18], an MtF protein band with an apparent molecular mass of 22 kDa was detected by SDS-PAGE. Weak bands were detected in IMR-32 and vector-IMR-32 cells (Fig. 7B). A remarkable decrease in the viability of IMR-32 and Vector-IMR-32 cells (about 50%; p<0.01, compared with the control groups) was observed after treatment with 300 µM H2O2 for 30 min (Fig. 7A). The viability of MtF-IMR-32 cells decreased about 30% after H2O2 treatment, therefore cell viability was much higher than in control groups (p<0.05). These results show that overexpression of MtF reduces the rate of cell death after H2O2 treatment.

Bottom Line: The neuroprotective effect of MtF on oxidative stress induced by H(2)O(2) was measured by MTT assay.Real-time PCR and western-blot confirmed that MtF expression levels in the cerebral cortex were significantly higher in AD cases than that in control cases at both the mRNA and the protein level.Finally, MtF expression showed a significant neuroprotective effect against H2O2-induced oxidative stress (p<0.05).

View Article: PubMed Central - PubMed

Affiliation: Molecular Neuroscience Research Center, Shiga University of Medical Science, Otsu, Japan.

ABSTRACT
Mitochondrial ferritin (MtF) has been identified as a novel ferritin encoded by an intron-lacking gene with specific mitochondrial localization located on chromosome 5q23.1. MtF has been associated with neurodegenerative disorders such as Friedreich ataxia and restless leg syndrome. However, little information is available about MtF in Alzheimer's disease (AD). In this study, therefore, we investigated the expression and localization of MtF messenger RNA (mRNA) in the cerebral cortex of AD and control cases using real-time polymerase chain reaction (PCR) as well as in situ hybridization histochemistry. We also examined protein expression using western-blot assay. In addition, we used in vitro methods to further explore the effect of oxidative stress and β-amyloid peptide (Aβ) on MtF expression. To do this we examined MtF mRNA and protein expression changes in the human neuroblastoma cell line, IMR-32, after treatment with Aβ, H2O2, or both. The neuroprotective effect of MtF on oxidative stress induced by H(2)O(2) was measured by MTT assay. The in situ hybridization studies revealed that MtF mRNA was detected mainly in neurons to a lesser degree in glial cells in the cerebral cortex. The staining intensity and the number of positive cells were increased in the cerebral cortex of AD patients. Real-time PCR and western-blot confirmed that MtF expression levels in the cerebral cortex were significantly higher in AD cases than that in control cases at both the mRNA and the protein level. Cell culture experiments demonstrated that the expression of both MtF mRNA and protein were increased by treatment with H2O2 or a combination of Aβ and H2O2, but not with Aβ alone. Finally, MtF expression showed a significant neuroprotective effect against H2O2-induced oxidative stress (p<0.05). The present study suggests that MtF is involved in the pathology of AD and may play a neuroprotective role against oxidative stress.

Show MeSH
Related in: MedlinePlus