Limits...
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.

Show MeSH

Related in: MedlinePlus

Effects of different concentrations of glutamine on PCV2 replication. PK-15 cells were infected with PCV2 at an MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in medium containing various concentrations of glutamine. After 48 h, the PK-15 cells were 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) by real-time PCR and immunofluorescence microscopy, respectively. Values shown are means ± SD mean from three independent experiments. Asterisks indicate groups statistically significantly different from control by a 1-way ANOVA followed by least-significant difference test (**P < 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig2: Effects of different concentrations of glutamine on PCV2 replication. PK-15 cells were infected with PCV2 at an MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in medium containing various concentrations of glutamine. After 48 h, the PK-15 cells were 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) by real-time PCR and immunofluorescence microscopy, respectively. Values shown are means ± SD mean from three independent experiments. Asterisks indicate groups statistically significantly different from control by a 1-way ANOVA followed by least-significant difference test (**P < 0.01).

Mentions: To establish the viral infection and ensure that PK-15 cells were able to survive in glutamine-free medium in our experiment, the cells were infected with PCV2 at a MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in media containing various concentrations of glutamine. After 48 h, the PK-15 cells were harvested to determine the number of PCV2 DNA copies and the relative proportion of PCV2-infected cells by real-time PCR and immunofluorescence microscopy, respectively. The PCV2 log10 DNA copies per 106 cells (Figure 2A) and the proportion of PCV2 infected cells (Figures 2B and 2C) were significantly increased when the glutamine concentration was below 2 mM as compared with cells grown with >4 mM glutamine (P < 0.05). The number of PCV2 log10 DNA copies per 106 cells was significantly increased by 2%, 4%, 5%, and 6% in groups with glutamine starvation at concentrations of 1, 0.5, 0.1, and 0 mM, respectively, as compared with the controls treated with 4 mM glutamine. The relative proportions of the PCV2-infected cells were 116%, 125%, 138%, and 145% when glutamine was supplemented at concentrations of 1, 0.5, 0.1, and 0 mM, respectively. Our results show that glutamine starvation increases PCV2 replication.Figure 2


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)

Effects of different concentrations of glutamine on PCV2 replication. PK-15 cells were infected with PCV2 at an MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in medium containing various concentrations of glutamine. After 48 h, the PK-15 cells were 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) by real-time PCR and immunofluorescence microscopy, respectively. Values shown are means ± SD mean from three independent experiments. Asterisks indicate groups statistically significantly different from control by a 1-way ANOVA followed by least-significant difference test (**P < 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig2: Effects of different concentrations of glutamine on PCV2 replication. PK-15 cells were infected with PCV2 at an MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in medium containing various concentrations of glutamine. After 48 h, the PK-15 cells were 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) by real-time PCR and immunofluorescence microscopy, respectively. Values shown are means ± SD mean from three independent experiments. Asterisks indicate groups statistically significantly different from control by a 1-way ANOVA followed by least-significant difference test (**P < 0.01).
Mentions: To establish the viral infection and ensure that PK-15 cells were able to survive in glutamine-free medium in our experiment, the cells were infected with PCV2 at a MOI of 1 and incubated in the complete medium for 24 h. Subsequently, PK-15 cells were grown in media containing various concentrations of glutamine. After 48 h, the PK-15 cells were harvested to determine the number of PCV2 DNA copies and the relative proportion of PCV2-infected cells by real-time PCR and immunofluorescence microscopy, respectively. The PCV2 log10 DNA copies per 106 cells (Figure 2A) and the proportion of PCV2 infected cells (Figures 2B and 2C) were significantly increased when the glutamine concentration was below 2 mM as compared with cells grown with >4 mM glutamine (P < 0.05). The number of PCV2 log10 DNA copies per 106 cells was significantly increased by 2%, 4%, 5%, and 6% in groups with glutamine starvation at concentrations of 1, 0.5, 0.1, and 0 mM, respectively, as compared with the controls treated with 4 mM glutamine. The relative proportions of the PCV2-infected cells were 116%, 125%, 138%, and 145% when glutamine was supplemented at concentrations of 1, 0.5, 0.1, and 0 mM, respectively. Our results show that glutamine starvation increases PCV2 replication.Figure 2

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