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Zinc supplementation protects against cadmium accumulation and cytotoxicity in Madin-Darby bovine kidney cells.

Zhang D, Liu J, Gao J, Shahzad M, Han Z, Wang Z, Li J, Sjölinder H - PLoS ONE (2014)

Bottom Line: Metallothioneins (MTs) are metal-binding proteins that play an essential role in heavy metal ion detoxification.We found that co-exposure to Zn2+ and Cd2+ synergistically enhanced RNA and protein expression of MT-1, MT-2, and the metal-regulatory transcription factor 1 in MDBK cells.Notably, addition of Zn2+ reduced the amounts of cytosolic Cd2+ detected following MDBK exposure to 10 μM Cd2+.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, PR China.

ABSTRACT
Cadmium ions (Cd2+) have been reported to accumulate in bovine tissues, although Cd2+ cytotoxicity has not been investigated thoroughly in this species. Zinc ions (Zn2+) have been shown to antagonize the toxic effects of heavy metals such as Cd2+ in some systems. The present study investigated Cd2+ cytotoxicity in Madin-Darby bovine kidney (MDBK) epithelial cells, and explored whether this was modified by Zn2+. Exposure to Cd2+ led to a dose- and time-dependent increase in apoptotic cell death, with increased intracellular levels of reactive oxygen species and mitochondrial damage. Zn2+ supplementation alleviated Cd2+-induced cytotoxicity and this protective effect was more obvious when cells were exposed to a lower concentration of Cd2+ (10 μM), as compared to 50 μM Cd2+. This indicated that high levels of Cd2+ accumulation might induce irreversible damage in bovine kidney cells. Metallothioneins (MTs) are metal-binding proteins that play an essential role in heavy metal ion detoxification. We found that co-exposure to Zn2+ and Cd2+ synergistically enhanced RNA and protein expression of MT-1, MT-2, and the metal-regulatory transcription factor 1 in MDBK cells. Notably, addition of Zn2+ reduced the amounts of cytosolic Cd2+ detected following MDBK exposure to 10 μM Cd2+. These findings revealed a protective role of Zn2+ in counteracting Cd2+ uptake and toxicity in MDBK cells, indicating that this approach may provide a means to protect livestock from excessive Cd2+ accumulation.

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Effects of Zn2+ and Cd2+ on MDBK apoptosis.(A) MDBK cells were treated with 10 μM CdCl2, alone or in combination with ZnCl2 (0, 10, or 50 μM), for 12 h. Apoptotic cell death was quantified by flow cytometry following double staining with propidium iodide (PI) and a fluorescein isothiocyanate (FITC)-conjugated annexin V antibody. Control cells were treated with medium. (B) MDBK cells were treated with CdCl2 alone (0, 10, or 50 μM) or in combination with ZnCl2 (0, 10, or 50 μM) for the indicated time periods. The percentage of PI- or annexin V-positive apoptotic cells was quantified by flow cytometry. The data were expressed as the mean ± SD (n = 4). #P<0.05, ##P<0.01, and ###P<0.001 for the comparison with the medium-treated control group; *P<0.05 and **P<0.01 compared to cells exposed to Cd2+ only (10 or 50 μM).
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pone-0103427-g001: Effects of Zn2+ and Cd2+ on MDBK apoptosis.(A) MDBK cells were treated with 10 μM CdCl2, alone or in combination with ZnCl2 (0, 10, or 50 μM), for 12 h. Apoptotic cell death was quantified by flow cytometry following double staining with propidium iodide (PI) and a fluorescein isothiocyanate (FITC)-conjugated annexin V antibody. Control cells were treated with medium. (B) MDBK cells were treated with CdCl2 alone (0, 10, or 50 μM) or in combination with ZnCl2 (0, 10, or 50 μM) for the indicated time periods. The percentage of PI- or annexin V-positive apoptotic cells was quantified by flow cytometry. The data were expressed as the mean ± SD (n = 4). #P<0.05, ##P<0.01, and ###P<0.001 for the comparison with the medium-treated control group; *P<0.05 and **P<0.01 compared to cells exposed to Cd2+ only (10 or 50 μM).

Mentions: Cd2+ and Zn2+ have both been shown to accelerate apoptosis in dose- and species-dependent manners [16], [19], [20] and we therefore determined the concentrations of these ions that induced toxic effects on MDBK cells. Cells were treated with 10–100 μM of CdCl2 or 10–200 μM of ZnCl2 for 12 h and cell survival was analyzed by flow cytometry with staining of annexin V and PI, to detect early and late apoptosis. We observed that 10 μM Cd2+ induced mild apoptotic cell damage (Fig. 1A), whereas 50 μM Cd2+ caused severe cell death (Fig. 1B). Nearly all cells treated with 100 μM CdCl2 underwent apoptosis (data not shown). We found that Cd2+-triggered cytotoxicity was both time- and dose-dependent. Compared to these cytotoxic effects of Cd2+, addition of 10 or 50 μM Zn2+ did not induce apoptosis (Fig. 1B). Based on these findings, 10 μM or 50 μM Cd2+ was used subsequently to induce mild or severe toxicity, respectively, for investigation of the possible protective role of Zn2+ supplementation (10 or 50 μM).


Zinc supplementation protects against cadmium accumulation and cytotoxicity in Madin-Darby bovine kidney cells.

Zhang D, Liu J, Gao J, Shahzad M, Han Z, Wang Z, Li J, Sjölinder H - PLoS ONE (2014)

Effects of Zn2+ and Cd2+ on MDBK apoptosis.(A) MDBK cells were treated with 10 μM CdCl2, alone or in combination with ZnCl2 (0, 10, or 50 μM), for 12 h. Apoptotic cell death was quantified by flow cytometry following double staining with propidium iodide (PI) and a fluorescein isothiocyanate (FITC)-conjugated annexin V antibody. Control cells were treated with medium. (B) MDBK cells were treated with CdCl2 alone (0, 10, or 50 μM) or in combination with ZnCl2 (0, 10, or 50 μM) for the indicated time periods. The percentage of PI- or annexin V-positive apoptotic cells was quantified by flow cytometry. The data were expressed as the mean ± SD (n = 4). #P<0.05, ##P<0.01, and ###P<0.001 for the comparison with the medium-treated control group; *P<0.05 and **P<0.01 compared to cells exposed to Cd2+ only (10 or 50 μM).
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Related In: Results  -  Collection

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pone-0103427-g001: Effects of Zn2+ and Cd2+ on MDBK apoptosis.(A) MDBK cells were treated with 10 μM CdCl2, alone or in combination with ZnCl2 (0, 10, or 50 μM), for 12 h. Apoptotic cell death was quantified by flow cytometry following double staining with propidium iodide (PI) and a fluorescein isothiocyanate (FITC)-conjugated annexin V antibody. Control cells were treated with medium. (B) MDBK cells were treated with CdCl2 alone (0, 10, or 50 μM) or in combination with ZnCl2 (0, 10, or 50 μM) for the indicated time periods. The percentage of PI- or annexin V-positive apoptotic cells was quantified by flow cytometry. The data were expressed as the mean ± SD (n = 4). #P<0.05, ##P<0.01, and ###P<0.001 for the comparison with the medium-treated control group; *P<0.05 and **P<0.01 compared to cells exposed to Cd2+ only (10 or 50 μM).
Mentions: Cd2+ and Zn2+ have both been shown to accelerate apoptosis in dose- and species-dependent manners [16], [19], [20] and we therefore determined the concentrations of these ions that induced toxic effects on MDBK cells. Cells were treated with 10–100 μM of CdCl2 or 10–200 μM of ZnCl2 for 12 h and cell survival was analyzed by flow cytometry with staining of annexin V and PI, to detect early and late apoptosis. We observed that 10 μM Cd2+ induced mild apoptotic cell damage (Fig. 1A), whereas 50 μM Cd2+ caused severe cell death (Fig. 1B). Nearly all cells treated with 100 μM CdCl2 underwent apoptosis (data not shown). We found that Cd2+-triggered cytotoxicity was both time- and dose-dependent. Compared to these cytotoxic effects of Cd2+, addition of 10 or 50 μM Zn2+ did not induce apoptosis (Fig. 1B). Based on these findings, 10 μM or 50 μM Cd2+ was used subsequently to induce mild or severe toxicity, respectively, for investigation of the possible protective role of Zn2+ supplementation (10 or 50 μM).

Bottom Line: Metallothioneins (MTs) are metal-binding proteins that play an essential role in heavy metal ion detoxification.We found that co-exposure to Zn2+ and Cd2+ synergistically enhanced RNA and protein expression of MT-1, MT-2, and the metal-regulatory transcription factor 1 in MDBK cells.Notably, addition of Zn2+ reduced the amounts of cytosolic Cd2+ detected following MDBK exposure to 10 μM Cd2+.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, PR China.

ABSTRACT
Cadmium ions (Cd2+) have been reported to accumulate in bovine tissues, although Cd2+ cytotoxicity has not been investigated thoroughly in this species. Zinc ions (Zn2+) have been shown to antagonize the toxic effects of heavy metals such as Cd2+ in some systems. The present study investigated Cd2+ cytotoxicity in Madin-Darby bovine kidney (MDBK) epithelial cells, and explored whether this was modified by Zn2+. Exposure to Cd2+ led to a dose- and time-dependent increase in apoptotic cell death, with increased intracellular levels of reactive oxygen species and mitochondrial damage. Zn2+ supplementation alleviated Cd2+-induced cytotoxicity and this protective effect was more obvious when cells were exposed to a lower concentration of Cd2+ (10 μM), as compared to 50 μM Cd2+. This indicated that high levels of Cd2+ accumulation might induce irreversible damage in bovine kidney cells. Metallothioneins (MTs) are metal-binding proteins that play an essential role in heavy metal ion detoxification. We found that co-exposure to Zn2+ and Cd2+ synergistically enhanced RNA and protein expression of MT-1, MT-2, and the metal-regulatory transcription factor 1 in MDBK cells. Notably, addition of Zn2+ reduced the amounts of cytosolic Cd2+ detected following MDBK exposure to 10 μM Cd2+. These findings revealed a protective role of Zn2+ in counteracting Cd2+ uptake and toxicity in MDBK cells, indicating that this approach may provide a means to protect livestock from excessive Cd2+ accumulation.

Show MeSH
Related in: MedlinePlus