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The role of hypoxia-inducible factor-1 α in zinc oxide nanoparticle-induced nephrotoxicity in vitro and in vivo

View Article: PubMed Central - PubMed

ABSTRACT

Background: Zinc oxide nanoparticles (ZnO NPs) are used in an increasing number of products, including rubber manufacture, cosmetics, pigments, food additives, medicine, chemical fibers and electronics. However, the molecular mechanisms underlying ZnO NP nephrotoxicity remain unclear. In this study, we evaluated the potential toxicity of ZnO NPs in kidney cells in vitro and in vivo.

Results: We found that ZnO NPs were apparently engulfed by the HEK-293 human embryonic kidney cells and then induced reactive oxygen species (ROS) generation. Furthermore, exposure to ZnO NPs led to a reduction in cell viability and induction of apoptosis and autophagy. Interestingly, the ROS-induced hypoxia-inducible factor-1α (HIF-1α) signaling pathway was significantly increased following ZnO NPs exposure. Additionally, connective tissue growth factor (CTGF) and plasminogen activator inhibitor-1 (PAI-1), which are directly regulated by HIF-1 and are involved in the pathogenesis of kidney diseases, displayed significantly increased levels following ZnO NPs exposure in HEK-293 cells. HIF-1α knockdown resulted in significantly decreased levels of autophagy and increased cytotoxicity. Therefore, our results suggest that HIF-1α may have a protective role in adaptation to the toxicity of ZnO NPs in kidney cells. In an animal study, fluorescent ZnO NPs were clearly observed in the liver, lungs, kidneys, spleen and heart. ZnO NPs caused histopathological lesions in the kidney and increase in serum creatinine and blood urea nitrogen (BUN) which indicate possible renal possible damage. Moreover, ZnO NPs enhanced the HIF-1α signaling pathway, apoptosis and autophagy in mouse kidney tissues.

Conclusions: ZnO NPs may cause nephrotoxicity, and the results demonstrate the importance of considering the toxicological hazards of ZnO NP production and application, especially for medicinal use.

Electronic supplementary material: The online version of this article (doi:10.1186/s12989-016-0163-3) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

HIF-1α knockdown by siRNA affects the cytotoxicity and autophagy of HEK-293 cells. a Transfection efficacy was verified by western blot analysis. HIF-1α protein expression in HEK-293 cells transfected with control or HIF-1α siRNA for 24 h. b Cell viability in the absence or presence of HIF-1α siRNA in HEK-293 cells. c Quantification of apoptotic cells with Annexin V-stained cells using flow cytometry. d Quantification of AVOs with AO-stained cells transfected with control or HIF-1α siRNA using flow cytometry. The cells were transfected with control or HIF-1α siRNA for 24 h and treated with medium alone or 20 μg/ml ZnO NPs for 24 h. *p < 0.05, control siRNA + ZnO NPs versus HIF-1α siRNA + ZnO NPs
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Fig5: HIF-1α knockdown by siRNA affects the cytotoxicity and autophagy of HEK-293 cells. a Transfection efficacy was verified by western blot analysis. HIF-1α protein expression in HEK-293 cells transfected with control or HIF-1α siRNA for 24 h. b Cell viability in the absence or presence of HIF-1α siRNA in HEK-293 cells. c Quantification of apoptotic cells with Annexin V-stained cells using flow cytometry. d Quantification of AVOs with AO-stained cells transfected with control or HIF-1α siRNA using flow cytometry. The cells were transfected with control or HIF-1α siRNA for 24 h and treated with medium alone or 20 μg/ml ZnO NPs for 24 h. *p < 0.05, control siRNA + ZnO NPs versus HIF-1α siRNA + ZnO NPs

Mentions: To further define the role of HIF-1α, we silenced HIF-1α expression using HIF-1α siRNA in HEK-293 cells. The expression of the HIF-1α proteins was markedly decreased in the cells treated with the HIF-1α siRNA compared with the control siRNA (Fig. 5a). The viability of the cells after siRNA transfection and ZnO NP treatment was determined by MTS assays. As shown in Fig. 5b, transfection with HIF-1α siRNA significantly enhanced the cytotoxicity of ZnO NPs in HEK-293 cells. Furthermore, we used HIF-1α siRNA to determine whether inhibition of HIF-1α alters ZnO NP treatment-induced apoptosis and autophagy. The results indicated that ZnO NP treatment with HIF-1α siRNA let to a significant decrease in autophagic cells compared to ZnO NPs alone (Fig. 5d). However, HIF-1α siRNA alone showed no noticeable change in apoptotic cells (Fig. 5c). These results suggest that HIF-1α-knockdown enhances the ZnO NP-induced cytotoxicity and suppresses the ZnO NP-induced autophagy.Fig. 5


The role of hypoxia-inducible factor-1 α in zinc oxide nanoparticle-induced nephrotoxicity in vitro and in vivo
HIF-1α knockdown by siRNA affects the cytotoxicity and autophagy of HEK-293 cells. a Transfection efficacy was verified by western blot analysis. HIF-1α protein expression in HEK-293 cells transfected with control or HIF-1α siRNA for 24 h. b Cell viability in the absence or presence of HIF-1α siRNA in HEK-293 cells. c Quantification of apoptotic cells with Annexin V-stained cells using flow cytometry. d Quantification of AVOs with AO-stained cells transfected with control or HIF-1α siRNA using flow cytometry. The cells were transfected with control or HIF-1α siRNA for 24 h and treated with medium alone or 20 μg/ml ZnO NPs for 24 h. *p < 0.05, control siRNA + ZnO NPs versus HIF-1α siRNA + ZnO NPs
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig5: HIF-1α knockdown by siRNA affects the cytotoxicity and autophagy of HEK-293 cells. a Transfection efficacy was verified by western blot analysis. HIF-1α protein expression in HEK-293 cells transfected with control or HIF-1α siRNA for 24 h. b Cell viability in the absence or presence of HIF-1α siRNA in HEK-293 cells. c Quantification of apoptotic cells with Annexin V-stained cells using flow cytometry. d Quantification of AVOs with AO-stained cells transfected with control or HIF-1α siRNA using flow cytometry. The cells were transfected with control or HIF-1α siRNA for 24 h and treated with medium alone or 20 μg/ml ZnO NPs for 24 h. *p < 0.05, control siRNA + ZnO NPs versus HIF-1α siRNA + ZnO NPs
Mentions: To further define the role of HIF-1α, we silenced HIF-1α expression using HIF-1α siRNA in HEK-293 cells. The expression of the HIF-1α proteins was markedly decreased in the cells treated with the HIF-1α siRNA compared with the control siRNA (Fig. 5a). The viability of the cells after siRNA transfection and ZnO NP treatment was determined by MTS assays. As shown in Fig. 5b, transfection with HIF-1α siRNA significantly enhanced the cytotoxicity of ZnO NPs in HEK-293 cells. Furthermore, we used HIF-1α siRNA to determine whether inhibition of HIF-1α alters ZnO NP treatment-induced apoptosis and autophagy. The results indicated that ZnO NP treatment with HIF-1α siRNA let to a significant decrease in autophagic cells compared to ZnO NPs alone (Fig. 5d). However, HIF-1α siRNA alone showed no noticeable change in apoptotic cells (Fig. 5c). These results suggest that HIF-1α-knockdown enhances the ZnO NP-induced cytotoxicity and suppresses the ZnO NP-induced autophagy.Fig. 5

View Article: PubMed Central - PubMed

ABSTRACT

Background: Zinc oxide nanoparticles (ZnO NPs) are used in an increasing number of products, including rubber manufacture, cosmetics, pigments, food additives, medicine, chemical fibers and electronics. However, the molecular mechanisms underlying ZnO NP nephrotoxicity remain unclear. In this study, we evaluated the potential toxicity of ZnO NPs in kidney cells in vitro and in vivo.

Results: We found that ZnO NPs were apparently engulfed by the HEK-293 human embryonic kidney cells and then induced reactive oxygen species (ROS) generation. Furthermore, exposure to ZnO NPs led to a reduction in cell viability and induction of apoptosis and autophagy. Interestingly, the ROS-induced hypoxia-inducible factor-1&alpha; (HIF-1&alpha;) signaling pathway was significantly increased following ZnO NPs exposure. Additionally, connective tissue growth factor (CTGF) and plasminogen activator inhibitor-1 (PAI-1), which are directly regulated by HIF-1 and are involved in the pathogenesis of kidney diseases, displayed significantly increased levels following ZnO NPs exposure in HEK-293 cells. HIF-1&alpha; knockdown resulted in significantly decreased levels of autophagy and increased cytotoxicity. Therefore, our results suggest that HIF-1&alpha; may have a protective role in adaptation to the toxicity of ZnO NPs in kidney cells. In an animal study, fluorescent ZnO NPs were clearly observed in the liver, lungs, kidneys, spleen and heart. ZnO NPs caused histopathological lesions in the kidney and increase in serum creatinine and blood urea nitrogen (BUN) which indicate possible renal possible damage. Moreover, ZnO NPs enhanced the HIF-1&alpha; signaling pathway, apoptosis and autophagy in mouse kidney tissues.

Conclusions: ZnO NPs may cause nephrotoxicity, and the results demonstrate the importance of considering the toxicological hazards of ZnO NP production and application, especially for medicinal use.

Electronic supplementary material: The online version of this article (doi:10.1186/s12989-016-0163-3) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus