This dissertation aims to explore the effects of environmental and biological factors on the biological effects of carbon nanomaterials. Part 1: Biophysical assessment of pulmonary surfactant PS predicts the lung toxicity of nanomaterials; Part 2: Oxygen content determines the bio-reactivity and toxicity profiles of carbon black (CB) particles; Part 3: Reduction of graphene oxide (GO) by Na2S differentially alters its cyto-compatibility towards primary macrophages and cell line macrophages; Part 4: Epigenetic toxicity of nano-GO on mouse hepatic parenchymal cells.
Part 1: Biophysical assessment of pulmonary surfactant predicts the lung toxicity of nanomaterials
As a natural surface-active substance, PS is the initial barrier in the lungs against the invasion of nanomaterials and the interaction with nanomaterials. Together with our team at the University of Hawaii, we have implemented an in vitro experimental method called the Constrained Drop Surfactometry (CDS) method by which the effect of ENM on the surface tension of the lung surfactant could real-time observation and quantitative assessment of tension changes. At very low concentrations, four kinds of representative ENMs, including carbon nanotubes (CNTs), GO, zinc oxide (ZnO), all increase the surface tension of surfactant extract from calf lungs. The results of these in vitro experiments correlate well with the extensive alveolar collapse and inflammatory responses observed in mice exposed to these ENMs in the tracheal instillation model. Therefore, there may be a direct correlation between increased surface tension of PS in vitro and pulmonary toxicity of ENM in vivo due to inhibition of ENM. Compared with the general use of animal models, the CDS method has great potential in the development of a method for the determination of acute pulmonary toxicity for rapid and low-cost predictive inhalation of ENMs.
Part 2: Oxygen content determines the bio-reactivity and toxicity profiles of CB particles
Although a large number of studies have explored the impact of fine particles (such as PM2.5) on environmental health and safety (EHS), significant gaps still exists. There is no exact understanding of how different physical structures and chemical properties determine the toxic effects of fine particulate matter. In this article, we compared the cytotoxicity of four CB particles that had similar physicochemical properties except for their oxygen content (C824455 C1864> Printex U> SB4A. Among these CB particles, C824455 and C1864 showed the most prominent lung injury (such as alveolar collapse and local lung inflammation) and macrophage activation, especially C824455. All of these differences in toxic effects, including in vitro and in vivo cytotoxicity, proinflammatory effects, and direct damage to the lung epithelium should be attributed, at least in part, to differences in oxygen content in these CB particles, which determine the different biological transformation of them, that is, different polymerization state. At the same time, PM2.5 also causes severe in vivo and in vitro toxicity to lung cells and macrophages. Taken together, this study provides additional insight into understanding the structure and reactivity (SAR) of particulates and opens up new avenues for elucidation of the physico-chemical determinants of air-lung particle-induced lung injury.
Part 3: Reduction of GO by Na2S differentially alters its cyto-compatibility towards Primary macrophages and cell line macrophages
We compared the differences in biocompatibility (eg, cytotoxicity, proinflammatory effects, and cell morphology, etc.) of sodium sulphite (Na2S) pre-reduction and post-reduction of GO on bone marrow-derived macrophages (BMDMs) and J774A.1 macrophage cell line. We used Na2S as a reducing agent to prepare two RGO (RGO1 and RGO2) with different reduction levels relative to the parent GO. Interestingly, the reduction of Na2S resulted in significant morphological changes of GO, from a flatter GO structure before reduction to the bilayer concave RGO structure after reduction, and the aggregation status did not change significantly. Cytotoxicity assessment showed that RGO was more toxic than GO to both cell types and that J774A.1 cell was more susceptible to the toxicity induced by these materials. RGO induced more pronounced pro-inflammatory responses than GO, especially in BMDM. In addition, GO and RGO exerted different effects on the morphological changes of BMDM cells. The synapses of GO-treated BMDM cells were longer and the synapses of BMDM cells exposed to RGO were shortened and flatter. Mechanistic studies showed that the impaired cell compatibility of RGO may be the result of a combination of oxygen content and changes in functional groups and edge morphology, resulting in RGO stimulating a stronger oxidative stress response in macrophages. Taken together, this study reveals an important contribution of GO and RGO to macrophage cytotoxicity mediated by reductive-mediated deleterious cellular effects.
Part 4: Epigenetic toxicity of nano-GO on mouse hepatic parenchymal cells
As mentioned above, GO shows superior properties in nanomedicine. Such as large specific surface area, photothermal conversion effect, etc., which provide broad potential for application in the fields of drug-carrying and photothermal therapy of tumors. However, the biosafety of GO, especially its in vivo metabolism and apparent toxicity mechanisms, remains poorly understood. In this paper, nano-sized GO, which is used in the recommended biomedicals in current literatures, was selected as the research object. The mice were exposed through the tail vein to analyze the distribution and metabolic conditions of nano-GO in mice, especially in the liver. Through the isolation and purification of liver parenchymal cells, we further studied the effects of nano-GO exposure on the transcriptome and genome-wide methylation of mouse hepatic parenchymal cells. The results show that nano-GO accumulates in the liver parenchyma cells, accompanied by metabolic elimination during the exposure time. During this process, the experimental animals did not show obvious pathological changes. However, whole-genome DNA methylation and methylation levels of liver parenchymal cells have undergone significant changes, and metabolic-related gene expression levels have also changed correspondingly. In summary, this article provides new insights and evidence for revealing the in vivo metabolic pathways and mechanisms of nano-GO and its application in biomedicine.