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Glutamine starvation enhances PCV2 replication via the phosphorylation of p38 MAPK, as promoted by reducing glutathione levels.

Chen X, Shi X, Gan F, Huang D, Huang K - Vet. Res. (2015)

Bottom Line: The results show that glutamine promoted PK-15 cell viability.Meanwhile, p38 MAPK phosphorylation and PCV2 replication significantly decreased in p38-knockdown PK-15 cells.Our findings may contribute toward interpreting the possible pathogenic mechanism of PCV2 and provide a theoretical reference for application of glutamine in controlling porcine circovirus-associated diseases.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China. cxx@njau.edu.cn.

ABSTRACT
Glutamine has a positive effect on ameliorating reproductive failure caused by porcine circovirus type 2 (PCV2). However, the mechanism by which glutamine affects PCV2 replication remains unclear. This study was conducted to investigate the effects of glutamine on PCV2 replication and its underlying mechanisms in vitro. The results show that glutamine promoted PK-15 cell viability. Surprisingly, glutamine starvation significantly increased PCV2 replication. The promotion of PCV2 replication by glutamine starvation disappeared after fresh media with 4 mM glutamine was added. Likewise, promotion of PCV2 was observed after adding buthionine sulfoximine (BSO). Glutamine starvation or BSO treatment increased the level of p38 MAPK phosphorylation and PCV2 replication in PK-15 cells. Meanwhile, p38 MAPK phosphorylation and PCV2 replication significantly decreased in p38-knockdown PK-15 cells. Promotion of PCV2 replication caused by glutamine starvation could be blocked in p38-knockdown PK-15 cells. Therefore, glutamine starvation increased PCV2 replication by promoting p38 MAPK activation, which was associated with the down regulation of intracellular glutathione levels. Our findings may contribute toward interpreting the possible pathogenic mechanism of PCV2 and provide a theoretical reference for application of glutamine in controlling porcine circovirus-associated diseases.

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Related in: MedlinePlus

Glutamine supplementation blocks the promotion of PCV2 replication by glutamine starvation. PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine. The cells were incubated for another 48 h and then harvested to determine the number of PCV2 log10 DNA copies per 106 cells (A) and the relative proportion of PCV2 infected cells (B and C). Groups were compared by a 1-way ANOVA followed by a least-significant difference test. Significant changes are indicated by *(P < 0.05) in comparison with control.
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Fig3: Glutamine supplementation blocks the promotion of PCV2 replication by glutamine starvation. PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine. The cells were incubated for another 48 h and then harvested to determine the number of PCV2 log10 DNA copies per 106 cells (A) and the relative proportion of PCV2 infected cells (B and C). Groups were compared by a 1-way ANOVA followed by a least-significant difference test. Significant changes are indicated by *(P < 0.05) in comparison with control.

Mentions: Another set of experiments was performed to determine whether glutamine supplementation blocks the positive effect of glutamine starvation on PCV2 replication. First, PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine and the cells were incubated for another 48 h. Finally, the number of PCV2 DNA copies and the relative proportion of PCV2-infected cells were calculated. As shown in Figure 3, the shift to fresh media with 4 mM glutamine at 72 h and the additional incubation for 48 h did not significantly alter the PCV2 replication in all groups (P > 0.05). The results indicate that the addition of fresh media with 4 mM glutamine blocked the promotion of PCV2 replication by glutamine starvation.Figure 3


Glutamine starvation enhances PCV2 replication via the phosphorylation of p38 MAPK, as promoted by reducing glutathione levels.

Chen X, Shi X, Gan F, Huang D, Huang K - Vet. Res. (2015)

Glutamine supplementation blocks the promotion of PCV2 replication by glutamine starvation. PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine. The cells were incubated for another 48 h and then harvested to determine the number of PCV2 log10 DNA copies per 106 cells (A) and the relative proportion of PCV2 infected cells (B and C). Groups were compared by a 1-way ANOVA followed by a least-significant difference test. Significant changes are indicated by *(P < 0.05) in comparison with control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4363047&req=5

Fig3: Glutamine supplementation blocks the promotion of PCV2 replication by glutamine starvation. PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine. The cells were incubated for another 48 h and then harvested to determine the number of PCV2 log10 DNA copies per 106 cells (A) and the relative proportion of PCV2 infected cells (B and C). Groups were compared by a 1-way ANOVA followed by a least-significant difference test. Significant changes are indicated by *(P < 0.05) in comparison with control.
Mentions: Another set of experiments was performed to determine whether glutamine supplementation blocks the positive effect of glutamine starvation on PCV2 replication. First, PK-15 cells were infected with PCV2 at an MOI of 1 in complete medium for 24 h. These PK-15 cells were subsequently grown in medium containing various concentrations of glutamine for 48 h before the media of all treatment groups were changed to fresh media with 4 mM glutamine and the cells were incubated for another 48 h. Finally, the number of PCV2 DNA copies and the relative proportion of PCV2-infected cells were calculated. As shown in Figure 3, the shift to fresh media with 4 mM glutamine at 72 h and the additional incubation for 48 h did not significantly alter the PCV2 replication in all groups (P > 0.05). The results indicate that the addition of fresh media with 4 mM glutamine blocked the promotion of PCV2 replication by glutamine starvation.Figure 3

Bottom Line: The results show that glutamine promoted PK-15 cell viability.Meanwhile, p38 MAPK phosphorylation and PCV2 replication significantly decreased in p38-knockdown PK-15 cells.Our findings may contribute toward interpreting the possible pathogenic mechanism of PCV2 and provide a theoretical reference for application of glutamine in controlling porcine circovirus-associated diseases.

View Article: PubMed Central - PubMed

Affiliation: College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China. cxx@njau.edu.cn.

ABSTRACT
Glutamine has a positive effect on ameliorating reproductive failure caused by porcine circovirus type 2 (PCV2). However, the mechanism by which glutamine affects PCV2 replication remains unclear. This study was conducted to investigate the effects of glutamine on PCV2 replication and its underlying mechanisms in vitro. The results show that glutamine promoted PK-15 cell viability. Surprisingly, glutamine starvation significantly increased PCV2 replication. The promotion of PCV2 replication by glutamine starvation disappeared after fresh media with 4 mM glutamine was added. Likewise, promotion of PCV2 was observed after adding buthionine sulfoximine (BSO). Glutamine starvation or BSO treatment increased the level of p38 MAPK phosphorylation and PCV2 replication in PK-15 cells. Meanwhile, p38 MAPK phosphorylation and PCV2 replication significantly decreased in p38-knockdown PK-15 cells. Promotion of PCV2 replication caused by glutamine starvation could be blocked in p38-knockdown PK-15 cells. Therefore, glutamine starvation increased PCV2 replication by promoting p38 MAPK activation, which was associated with the down regulation of intracellular glutathione levels. Our findings may contribute toward interpreting the possible pathogenic mechanism of PCV2 and provide a theoretical reference for application of glutamine in controlling porcine circovirus-associated diseases.

Show MeSH
Related in: MedlinePlus