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p53 activation contributes to patulin-induced nephrotoxicity via modulation of reactive oxygen species generation.

Jin H, Yin S, Song X, Zhang E, Fan L, Hu H - Sci Rep (2016)

Bottom Line: Patulin is a major mycotoxin found in fungal contaminated fruits and their derivative products.To the best of our knowledge, this is the first report addressing the functional role of p53 in patulin-induced oxidative stress.The findings of the present study provided novel insights into understanding mechanisms behind oxidative stress in response to patulin exposure.

View Article: PubMed Central - PubMed

Affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, Department of Nutrition and Health, College of Food Science and Nutritional Engineering, China Agricultural University, No17 Qinghua East Road, Haidian District, Beijing 100083, China.

ABSTRACT
Patulin is a major mycotoxin found in fungal contaminated fruits and their derivative products. Previous studies showed that patulin was able to induce increase of reactive oxygen species (ROS) generation and oxidative stress was suggested to play a pivotal role in patulin-induced multiple toxic signaling. The objective of the present study was to investigate the functional role of p53 in patulin-induced oxidative stress. Our study demonstrated that higher levels of ROS generation and DNA damage were induced in wild-type p53 cell lines than that found in either knockdown or knockout p53 cell lines in response to patulin exposure, suggesting p53 activation contributed to patulin-induced ROS generation. Mechanistically, we revealed that the pro-oxidant role of p53 in response to patulin was attributed to its ability to suppress catalase activity through up-regulation of PIG3. Moreover, these in vitro findings were further validated in the p53 wild-type/knockout mouse model. To the best of our knowledge, this is the first report addressing the functional role of p53 in patulin-induced oxidative stress. The findings of the present study provided novel insights into understanding mechanisms behind oxidative stress in response to patulin exposure.

No MeSH data available.


Related in: MedlinePlus

Induction of PIG3 was required for the pro-oxidant function of p53 in patulin-induced oxidative stress.(A). Effects of patulin treatment on PIG3 expression in HEK293 cells. The cells were treated with various concentrations patulin for 24 h. The protein level of PIG3 was analyzed by western blotting. (B). Effects of patulin treatment on PIG3 expression in p53 wild type/knockout MEF cells. The cells were treated with various patulin concentrations for 12 h and then PIG3 expression was measured by western blotting. (C). Effects of p53 knockdown on PIG3 expression by patulin. The cells were transfected with p53 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then PIG3 expression was analyzed by western blotting. (D). Effects of PIG3 knockdown on H2AX phosphorylation by patulin. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then H2AX phosphorylation was measured by western blotting. (E,F). Effects of PIG3 inactivation on patulin induced ROS generation and inhibitory effects of catalase activity. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h. ROS levels were measured by flow cytometry after staining with H2DCFDA (E) and catalase activity was analyzed by catalase activity assay kit (F). n = 3, *P < 0.05, **P < 0.01. (The blots shown are representative of three independent experiments).
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f3: Induction of PIG3 was required for the pro-oxidant function of p53 in patulin-induced oxidative stress.(A). Effects of patulin treatment on PIG3 expression in HEK293 cells. The cells were treated with various concentrations patulin for 24 h. The protein level of PIG3 was analyzed by western blotting. (B). Effects of patulin treatment on PIG3 expression in p53 wild type/knockout MEF cells. The cells were treated with various patulin concentrations for 12 h and then PIG3 expression was measured by western blotting. (C). Effects of p53 knockdown on PIG3 expression by patulin. The cells were transfected with p53 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then PIG3 expression was analyzed by western blotting. (D). Effects of PIG3 knockdown on H2AX phosphorylation by patulin. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then H2AX phosphorylation was measured by western blotting. (E,F). Effects of PIG3 inactivation on patulin induced ROS generation and inhibitory effects of catalase activity. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h. ROS levels were measured by flow cytometry after staining with H2DCFDA (E) and catalase activity was analyzed by catalase activity assay kit (F). n = 3, *P < 0.05, **P < 0.01. (The blots shown are representative of three independent experiments).

Mentions: PIG3 (p53-inducible gene 3), a transcriptional target of p53, has been reported to play a role in the pro-oxidant activity of p53 in some model systems32021. We then asked if up-regulation of PIG3 contributed to patulin-induced p53-mediated ROS generation. The changes of PIG3 expression in response to patulin exposure were analyzed by western blotting and the results are shown in Fig. 3A,B. As expected, exposure to patulin caused a concentration-dependent up-regulation of PIG3 in both HEK293 and p53 wt MEF cells, further supporting transcriptional activation of p53 in response to patulin exposure in these cell lines. Moreover, we confirmed that up-regulation of PIG3 was indeed due to p53 activation by the evidence that knockdown of p53 led to a decreased PIG3 expression in HEK293 cells (Fig. 3C), whereas knockout of p53 resulted in abolishment of PIG3 induction in MEF cells (Fig. 3B). To examine the role of PIG3 induction in patulin-induced ROS generation, we measured the influences of PIG3 inhibition by RNAi on the levels of ROS generation. As shown in Fig. 3D, PIG3 was efficiently suppressed by its siRNA. Under such condition, DNA damage marker H2AX phosphorylation induced by patulin exposure was inhibited partially. Consistent with the decreased DNA damage, the ROS generation by patulin was suppressed under PIG3 silencing condition (Fig. 3E). In line with the decreased ROS generation, the inhibition of catalase activity by patulin was significantly attenuated when PIG3 was inactivated by RNAi (Fig. 3F). Together, the results suggested that p53-dependent up-regulation of PIG3 contributed to ROS generation in response to patulin exposure.


p53 activation contributes to patulin-induced nephrotoxicity via modulation of reactive oxygen species generation.

Jin H, Yin S, Song X, Zhang E, Fan L, Hu H - Sci Rep (2016)

Induction of PIG3 was required for the pro-oxidant function of p53 in patulin-induced oxidative stress.(A). Effects of patulin treatment on PIG3 expression in HEK293 cells. The cells were treated with various concentrations patulin for 24 h. The protein level of PIG3 was analyzed by western blotting. (B). Effects of patulin treatment on PIG3 expression in p53 wild type/knockout MEF cells. The cells were treated with various patulin concentrations for 12 h and then PIG3 expression was measured by western blotting. (C). Effects of p53 knockdown on PIG3 expression by patulin. The cells were transfected with p53 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then PIG3 expression was analyzed by western blotting. (D). Effects of PIG3 knockdown on H2AX phosphorylation by patulin. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then H2AX phosphorylation was measured by western blotting. (E,F). Effects of PIG3 inactivation on patulin induced ROS generation and inhibitory effects of catalase activity. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h. ROS levels were measured by flow cytometry after staining with H2DCFDA (E) and catalase activity was analyzed by catalase activity assay kit (F). n = 3, *P < 0.05, **P < 0.01. (The blots shown are representative of three independent experiments).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4829895&req=5

f3: Induction of PIG3 was required for the pro-oxidant function of p53 in patulin-induced oxidative stress.(A). Effects of patulin treatment on PIG3 expression in HEK293 cells. The cells were treated with various concentrations patulin for 24 h. The protein level of PIG3 was analyzed by western blotting. (B). Effects of patulin treatment on PIG3 expression in p53 wild type/knockout MEF cells. The cells were treated with various patulin concentrations for 12 h and then PIG3 expression was measured by western blotting. (C). Effects of p53 knockdown on PIG3 expression by patulin. The cells were transfected with p53 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then PIG3 expression was analyzed by western blotting. (D). Effects of PIG3 knockdown on H2AX phosphorylation by patulin. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h and then H2AX phosphorylation was measured by western blotting. (E,F). Effects of PIG3 inactivation on patulin induced ROS generation and inhibitory effects of catalase activity. The cells were transfected with PIG3 siRNA using INTERFER siRNA transfection agent. At 24 h post-transfection, the cells were treated with 7 μM patulin for 24 h. ROS levels were measured by flow cytometry after staining with H2DCFDA (E) and catalase activity was analyzed by catalase activity assay kit (F). n = 3, *P < 0.05, **P < 0.01. (The blots shown are representative of three independent experiments).
Mentions: PIG3 (p53-inducible gene 3), a transcriptional target of p53, has been reported to play a role in the pro-oxidant activity of p53 in some model systems32021. We then asked if up-regulation of PIG3 contributed to patulin-induced p53-mediated ROS generation. The changes of PIG3 expression in response to patulin exposure were analyzed by western blotting and the results are shown in Fig. 3A,B. As expected, exposure to patulin caused a concentration-dependent up-regulation of PIG3 in both HEK293 and p53 wt MEF cells, further supporting transcriptional activation of p53 in response to patulin exposure in these cell lines. Moreover, we confirmed that up-regulation of PIG3 was indeed due to p53 activation by the evidence that knockdown of p53 led to a decreased PIG3 expression in HEK293 cells (Fig. 3C), whereas knockout of p53 resulted in abolishment of PIG3 induction in MEF cells (Fig. 3B). To examine the role of PIG3 induction in patulin-induced ROS generation, we measured the influences of PIG3 inhibition by RNAi on the levels of ROS generation. As shown in Fig. 3D, PIG3 was efficiently suppressed by its siRNA. Under such condition, DNA damage marker H2AX phosphorylation induced by patulin exposure was inhibited partially. Consistent with the decreased DNA damage, the ROS generation by patulin was suppressed under PIG3 silencing condition (Fig. 3E). In line with the decreased ROS generation, the inhibition of catalase activity by patulin was significantly attenuated when PIG3 was inactivated by RNAi (Fig. 3F). Together, the results suggested that p53-dependent up-regulation of PIG3 contributed to ROS generation in response to patulin exposure.

Bottom Line: Patulin is a major mycotoxin found in fungal contaminated fruits and their derivative products.To the best of our knowledge, this is the first report addressing the functional role of p53 in patulin-induced oxidative stress.The findings of the present study provided novel insights into understanding mechanisms behind oxidative stress in response to patulin exposure.

View Article: PubMed Central - PubMed

Affiliation: Beijing Advanced Innovation Center for Food Nutrition and Human Health, Department of Nutrition and Health, College of Food Science and Nutritional Engineering, China Agricultural University, No17 Qinghua East Road, Haidian District, Beijing 100083, China.

ABSTRACT
Patulin is a major mycotoxin found in fungal contaminated fruits and their derivative products. Previous studies showed that patulin was able to induce increase of reactive oxygen species (ROS) generation and oxidative stress was suggested to play a pivotal role in patulin-induced multiple toxic signaling. The objective of the present study was to investigate the functional role of p53 in patulin-induced oxidative stress. Our study demonstrated that higher levels of ROS generation and DNA damage were induced in wild-type p53 cell lines than that found in either knockdown or knockout p53 cell lines in response to patulin exposure, suggesting p53 activation contributed to patulin-induced ROS generation. Mechanistically, we revealed that the pro-oxidant role of p53 in response to patulin was attributed to its ability to suppress catalase activity through up-regulation of PIG3. Moreover, these in vitro findings were further validated in the p53 wild-type/knockout mouse model. To the best of our knowledge, this is the first report addressing the functional role of p53 in patulin-induced oxidative stress. The findings of the present study provided novel insights into understanding mechanisms behind oxidative stress in response to patulin exposure.

No MeSH data available.


Related in: MedlinePlus