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Prion Protein Does Not Confer Resistance to Hippocampus-Derived Zpl Cells against the Toxic Effects of Cu2+, Mn2+, Zn2+ and Co2+ Not Supporting a General Protective Role for PrP in Transition Metal Induced Toxicity.

Cingaram PK, Nyeste A, Dondapati DT, Fodor E, Welker E - PLoS ONE (2015)

Bottom Line: By employing a cell viability assay, we examined the effects of various concentrations of Cu2+, Zn2+, Mn2+, and Co2+ on Zpl (Prnp-/-) and ZW (Prnp+/+) hippocampus-derived mouse neuronal cells.However, when we introduced PrP or only the empty vector into Zpl cells, we could not discern any protective effect associated with the presence of PrP.Thus, our results on this mouse cell culture model do not seem to support a strong protective role for PrP against transition metal toxicity and also emphasize the necessity of extreme care when comparing cells derived from PrP knock-out and wild type mice.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.

ABSTRACT
The interactions of transition metals with the prion protein (PrP) are well-documented and characterized, however, there is no consensus on their role in either the physiology of PrP or PrP-related neurodegenerative disorders. PrP has been reported to protect cells from the toxic stimuli of metals. By employing a cell viability assay, we examined the effects of various concentrations of Cu2+, Zn2+, Mn2+, and Co2+ on Zpl (Prnp-/-) and ZW (Prnp+/+) hippocampus-derived mouse neuronal cells. Prnp-/- Zpl cells were more sensitive to all four metals than PrP-expressing Zw cells. However, when we introduced PrP or only the empty vector into Zpl cells, we could not discern any protective effect associated with the presence of PrP. This observation was further corroborated when assessing the toxic effect of metals by propidium-iodide staining and fluorescence activated cell sorting analysis. Thus, our results on this mouse cell culture model do not seem to support a strong protective role for PrP against transition metal toxicity and also emphasize the necessity of extreme care when comparing cells derived from PrP knock-out and wild type mice.

No MeSH data available.


Related in: MedlinePlus

The effect of the presence of PrPC on transition metals induced cell death.Cells were tested for cell death after treatment with transition metals for 24 h, cell death was measured by propidium iodide (PI) exclusion assay. (A) Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM of Cu2+-Gly (1:4 mol/mol), or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+. Dead cells were stained with PI, and histograms were obtained by flow cytometric analysis. M1 represents the population of PI positive cells. The bar graphs on the panels b through e indicate the average of PI positive cells in case of 750 μM of Cu2+ treatment (B), 100 μM of Zn2+ treatment (C), 500 μM of Mn2+ treatment (D) and 750 μM of Co2+ treatment (E), respectively. Experiments were performed three times in duplicates, and data represent the mean ± standard deviation (S.D.). *p<0.05, **p<0.01 and ***p<0.001 indicate significant differences between treated (+) and untreated (-) cells; +p<0.05, ++p<0.01 and +++p<0.001 indicate significant differences between the ratios obtained for treated ZW 13–2 cells to treated Zpl 2–1 cells, and treated Zpl 2-1-vector cells to treated Zpl 2-1-PrP cells, respectively.
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pone.0139219.g005: The effect of the presence of PrPC on transition metals induced cell death.Cells were tested for cell death after treatment with transition metals for 24 h, cell death was measured by propidium iodide (PI) exclusion assay. (A) Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM of Cu2+-Gly (1:4 mol/mol), or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+. Dead cells were stained with PI, and histograms were obtained by flow cytometric analysis. M1 represents the population of PI positive cells. The bar graphs on the panels b through e indicate the average of PI positive cells in case of 750 μM of Cu2+ treatment (B), 100 μM of Zn2+ treatment (C), 500 μM of Mn2+ treatment (D) and 750 μM of Co2+ treatment (E), respectively. Experiments were performed three times in duplicates, and data represent the mean ± standard deviation (S.D.). *p<0.05, **p<0.01 and ***p<0.001 indicate significant differences between treated (+) and untreated (-) cells; +p<0.05, ++p<0.01 and +++p<0.001 indicate significant differences between the ratios obtained for treated ZW 13–2 cells to treated Zpl 2–1 cells, and treated Zpl 2-1-vector cells to treated Zpl 2-1-PrP cells, respectively.

Mentions: Next, we attempted to check the effect of the presence of PrPC on transition metal-toxicity by assessing the cell death as a complementary approach. ZW 13–2, Zpl 2–1, Zpl 2-1-PrP and Zpl 2-1-vector cells were treated with the four transition metal ions Cu2+, Zn2+, Mn2+ and Co2+ for a period of 24 h, after which the cell death was measured by propidium iodide (PI) exclusion assay. Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM Cu2+, or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+, respectively. Dead cells were stained with PI and the histograms obtained by flow cytometry analysis were compared (Fig 5A). Exposure to Cu2+, Zn2+, and Mn2+ for 24 h resulted in increase of PI positive cells in most cell lines and conditions (Fig 5B, 5C, 5D and 5E). However, the number of PI positive cells was significantly less in ZW 13–2 as compared to Zpl 2–1 lines in case of Cu2+, Zn2+ and Mn2+ (Fig 5B, 5C and 5D) but not in case of Co2+ treatments. Co2+ was not significantly toxic to either Zpl 2–1 or ZW 13–2 cells compared to untreated control cells (Fig 5E). In case of Zpl 2-1-vector and Zpl 2-1-PrP cells, however, such results could not be observed; the Zpl 2-1-PrP cell population had significantly more PI positive cells compared to Zpl 2-1-vector cells when treated by Cu2+, Zn2+ and Co2+ (Fig 5B, 5C and 5E), with both lines being unresponsive to manganese treatment. These data indicate that PrPC-expressing Zpl 2-1-PrP cells did not gain more resistance to any of the metals tested and at the concentrations applied, compared to PrP-lacking Zpl 2-1-vector cells.


Prion Protein Does Not Confer Resistance to Hippocampus-Derived Zpl Cells against the Toxic Effects of Cu2+, Mn2+, Zn2+ and Co2+ Not Supporting a General Protective Role for PrP in Transition Metal Induced Toxicity.

Cingaram PK, Nyeste A, Dondapati DT, Fodor E, Welker E - PLoS ONE (2015)

The effect of the presence of PrPC on transition metals induced cell death.Cells were tested for cell death after treatment with transition metals for 24 h, cell death was measured by propidium iodide (PI) exclusion assay. (A) Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM of Cu2+-Gly (1:4 mol/mol), or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+. Dead cells were stained with PI, and histograms were obtained by flow cytometric analysis. M1 represents the population of PI positive cells. The bar graphs on the panels b through e indicate the average of PI positive cells in case of 750 μM of Cu2+ treatment (B), 100 μM of Zn2+ treatment (C), 500 μM of Mn2+ treatment (D) and 750 μM of Co2+ treatment (E), respectively. Experiments were performed three times in duplicates, and data represent the mean ± standard deviation (S.D.). *p<0.05, **p<0.01 and ***p<0.001 indicate significant differences between treated (+) and untreated (-) cells; +p<0.05, ++p<0.01 and +++p<0.001 indicate significant differences between the ratios obtained for treated ZW 13–2 cells to treated Zpl 2–1 cells, and treated Zpl 2-1-vector cells to treated Zpl 2-1-PrP cells, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591282&req=5

pone.0139219.g005: The effect of the presence of PrPC on transition metals induced cell death.Cells were tested for cell death after treatment with transition metals for 24 h, cell death was measured by propidium iodide (PI) exclusion assay. (A) Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM of Cu2+-Gly (1:4 mol/mol), or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+. Dead cells were stained with PI, and histograms were obtained by flow cytometric analysis. M1 represents the population of PI positive cells. The bar graphs on the panels b through e indicate the average of PI positive cells in case of 750 μM of Cu2+ treatment (B), 100 μM of Zn2+ treatment (C), 500 μM of Mn2+ treatment (D) and 750 μM of Co2+ treatment (E), respectively. Experiments were performed three times in duplicates, and data represent the mean ± standard deviation (S.D.). *p<0.05, **p<0.01 and ***p<0.001 indicate significant differences between treated (+) and untreated (-) cells; +p<0.05, ++p<0.01 and +++p<0.001 indicate significant differences between the ratios obtained for treated ZW 13–2 cells to treated Zpl 2–1 cells, and treated Zpl 2-1-vector cells to treated Zpl 2-1-PrP cells, respectively.
Mentions: Next, we attempted to check the effect of the presence of PrPC on transition metal-toxicity by assessing the cell death as a complementary approach. ZW 13–2, Zpl 2–1, Zpl 2-1-PrP and Zpl 2-1-vector cells were treated with the four transition metal ions Cu2+, Zn2+, Mn2+ and Co2+ for a period of 24 h, after which the cell death was measured by propidium iodide (PI) exclusion assay. Zpl 2–1, ZW 13–2, Zpl 2-1-vector and Zpl 2-1-PrP cells were treated either with 750 μM Cu2+, or with 100 μM of Zn2+, or 500 μM of Mn2+ or 750 μM of Co2+, respectively. Dead cells were stained with PI and the histograms obtained by flow cytometry analysis were compared (Fig 5A). Exposure to Cu2+, Zn2+, and Mn2+ for 24 h resulted in increase of PI positive cells in most cell lines and conditions (Fig 5B, 5C, 5D and 5E). However, the number of PI positive cells was significantly less in ZW 13–2 as compared to Zpl 2–1 lines in case of Cu2+, Zn2+ and Mn2+ (Fig 5B, 5C and 5D) but not in case of Co2+ treatments. Co2+ was not significantly toxic to either Zpl 2–1 or ZW 13–2 cells compared to untreated control cells (Fig 5E). In case of Zpl 2-1-vector and Zpl 2-1-PrP cells, however, such results could not be observed; the Zpl 2-1-PrP cell population had significantly more PI positive cells compared to Zpl 2-1-vector cells when treated by Cu2+, Zn2+ and Co2+ (Fig 5B, 5C and 5E), with both lines being unresponsive to manganese treatment. These data indicate that PrPC-expressing Zpl 2-1-PrP cells did not gain more resistance to any of the metals tested and at the concentrations applied, compared to PrP-lacking Zpl 2-1-vector cells.

Bottom Line: By employing a cell viability assay, we examined the effects of various concentrations of Cu2+, Zn2+, Mn2+, and Co2+ on Zpl (Prnp-/-) and ZW (Prnp+/+) hippocampus-derived mouse neuronal cells.However, when we introduced PrP or only the empty vector into Zpl cells, we could not discern any protective effect associated with the presence of PrP.Thus, our results on this mouse cell culture model do not seem to support a strong protective role for PrP against transition metal toxicity and also emphasize the necessity of extreme care when comparing cells derived from PrP knock-out and wild type mice.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biochemistry, Biological Research Centre, Hungarian Academy of Sciences, Szeged, Hungary.

ABSTRACT
The interactions of transition metals with the prion protein (PrP) are well-documented and characterized, however, there is no consensus on their role in either the physiology of PrP or PrP-related neurodegenerative disorders. PrP has been reported to protect cells from the toxic stimuli of metals. By employing a cell viability assay, we examined the effects of various concentrations of Cu2+, Zn2+, Mn2+, and Co2+ on Zpl (Prnp-/-) and ZW (Prnp+/+) hippocampus-derived mouse neuronal cells. Prnp-/- Zpl cells were more sensitive to all four metals than PrP-expressing Zw cells. However, when we introduced PrP or only the empty vector into Zpl cells, we could not discern any protective effect associated with the presence of PrP. This observation was further corroborated when assessing the toxic effect of metals by propidium-iodide staining and fluorescence activated cell sorting analysis. Thus, our results on this mouse cell culture model do not seem to support a strong protective role for PrP against transition metal toxicity and also emphasize the necessity of extreme care when comparing cells derived from PrP knock-out and wild type mice.

No MeSH data available.


Related in: MedlinePlus