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Silver Nanoparticles Induced RNA Polymerase-Silver Binding and RNA Transcription Inhibition in Erythroid Progenitor Cells Zhe Wang, Sijin Liu, Juan Ma, Guangbo Qu, Xiaoyan Wang, Suquan Yu, Jiuyang He, Jingfu Liu, Tian Xia, and Gui-Bin Jiang ACS Nano, Just Accepted Manuscript • DOI: 10.1021/nn400594s • Publication Date (Web): 09 Apr 2013 Downloaded from http://pubs.acs.org on April 14, 2013
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ACS Nano
Silver Nanoparticles Induced RNA Polymerase-Silver Binding and RNA Transcription Inhibition in Erythroid Progenitor Cells
Zhe Wang1, Sijin Liu1,*, Juan Ma1, Guangbo Qu1, Xiaoyan Wang1, Sujuan Yu1, Jiuyang He2, Jingfu Liu1, Tian Xia3, Gui-Bin Jiang1
1. State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China. 2. Department of Biomedical Engineering, Tufts University, Medford, Massachusetts Medford 02155, USA. 3. Division of NanoMedicine, Department of Medicine, UCLA, Los Angeles, California 90095, USA.
*Correspondence to Sijin Liu Sijin Liu, Ph.D, Email:
[email protected] Tel/Fax: 8610-62849330
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Abstract Due to its antimicrobial activity, nanosilver (nAg) has become the most widely used nanomaterial. Thus far, the mechanisms responsible for nAg-induced antimicrobial properties and nAg-mediated toxicity to organisms have not been clearly recognized. Silver (Ag) ions certainly play a crucial role, and the form of nanoparticles can change the dissolution rate, bioavailability, biodistribution, and cellular uptake of Ag. However, whether nAg exerts direct “particle-specific” effects has been under debate. Here, we demonstrated that nAg exhibited a robust inhibition on RNA polymerase activity and overall RNA transcription through direct Ag binding to RNA polymerase, which is separated from the cytotoxicity pathway induced by Ag ions. nAg treatment in vitro resulted in reduced hemoglobin concentration in erythroid cells; in vivo administration of nAg in mice caused profound reduction of hemoglobin content in embryonic erythrocytes, associated with anemia in the embryos. Embryonic anemia and general proliferation deficit due to the significant inhibition on RNA synthesis, at least partially, accounted for embryonic developmental retardation upon nAg administration. To date, there is no conclusive answer to the sources of nAg-mediated toxicity: Ag ions or “particle-specific” effects, or both. We here demonstrated that both Ag ions and nAg particles simultaneously existed inside cells, demonstrating the “Trojan horse” effects of nAg particles in posing biological impacts on erythroid cells. Moreover, our results suggested that “particle-specific” effects could be the predominant mediator in eliciting biological influences on erythroid cells under relatively low concentrations of nAg exposure. The combined data highlighted the inhibitory effect of nAg on RNA polymerase activity through a direct reciprocal interaction. Key words: Nanosilver; Trojan horse; erythroid cells; RNA polymerase; Transcription.
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With the explosive growth of nanotechnology, nanosilver (nAg) has become the most widely used nanomaterials.1,2 Due to its novel properties, in particular its intrinsic antimicrobial activity, nAg has been applied to diverse fields with significant advantages, such as medicine and personal care.3-7 To date, the mechanisms underlying nAg-mediated antimicrobial activity have not been fully characterized. Meanwhile, large-scale applications of nAg-related products pose potential risks to the environment and health.8,9 A number of studies have documented the toxicities of nAg in a variety of cells and organisms.10-12 There is also possible interruption of genetic integrity,13-15 since nAg could be readily taken up into cells, and even the nuclei.14,16 However, studies on nAg-mediated genotoxicity are still limited; thus far, there has been scarce investigation on the effect of nAg exposure on RNA polymerase activity and RNA polymerase-conducted transcription. Until now, the sources of nAg-mediated toxicity have not been clearly determined and without conclusive answer. In other words, the mechanism of nAg-mediated toxicity is not clear: ion effects or particle effects, or both. Previous studies have demonstrated that nAg has the potential to release Ag ions, and Ag ions could surely exert significant toxicity to cells and organisms.3,17,18 However, whether nAg conducts direct particle effects has been in issue. In the current report, we employed a panel of nAg of different sizes and shapes, and their detailed properties are described in Table S1 with the representative TEM images shown in Figure 1. We studied the mechanisms responsible for nAg-induced cytotoxicity from the perspective of nAg’s interference on RNA polymerase activity and the interaction between Ag and RNA polymerase in erythroid cells. We here chose erythroid cells because hemoglobins are the predominant proteins produced by these cells, and thus it is relatively easy to study the regulation of mRNA transcription for globin genes. We demonstrated that nAg conduced a significant suppression on RNA polymerase activity and overall RNA transcription through direct Ag binding to RNA polymerase, and this effect was separated from the 3
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cytotoxicity pathway induced by Ag ions. Our results suggested that the particle effects could be the predominant mediator in inhibiting RNA transcription in erythroid cells under relatively low concentrations of nAg exposure. Our combined data confirmed the “Trojan horse” effects of nAg in exerting biological effects on erythroid cells. RESULTS AND DISCUSSION We first surveyed the potential influence of nAg treatment on globin mRNA transcription by exposing mouse erythroleukemia (MEL) cells to nAg. MEL cells are erythroid progenitor cells at the stage of proerythroblast, and these cells continue to differentiate upon induction with DMSO.19 As shown in Figure S1, the qRT-PCR analysis reflected a large reduction for both α-globin and β-globin mRNA concentrations in MEL cells treated with 10 nm, 25 nm, 40 nm or 110 nm spherical nAg, or 45 nm plate-like nAg, at a concentration of 1 µg/ml (P
