Functionalization impacts the effects of carbon nanotubes on the immune system of rainbow trout, Oncorhynchus mykiss

Abstract

Recent studies have demonstrated the potential for manufactured nanomaterials to reach the aquatic environment. There is a need to determine if these materials will have an impact on aquatic species and at what level of exposure. In addition there is a need to develop models to test the potential effects of the multitude of particle types in production on aquatic vertebrates. The purpose of this research was to determine the impact of manufactured nanomaterials on the immune system of an aquatic vertebrate model, the rainbow trout (Oncorhynchus mykiss). We investigated how structure and type of functionalization of manufactured nanomaterials could affect immunotoxicity. To assess immunotoxicity, we used a well-studied trout macrophage primary cell culture system in conjunction with the expression of IL-1β and IFNα for proinflammatory and antiviral gene expression. There was a significant difference among the different carbon nanotube-based nanomaterials in their level of stimulation of IL-1β in macrophage cells and the dose at which they became stimulatory. At concentrations that were sublethal to cells, almost all nanomaterials were stimulatory at some concentration. Single-walled nanotubes and multi-walled nanotubes that were differentially functionalized to be water-soluble, varied in their effects; specifically the concentrations at which they were stimulatory and they were more stimulatory to IL-1β expression compared with unfunctionalized nanotubes. Each functionalized nanotube type caused a dose-dependent response with the lowest exposures (0.05-1.0 μg/ml) having no stimulatory response and at the highest concentrations (5 μg/ml and 10 μg/ml) stimulating a response similar to the positive LPS positive control. Anionic functionalized multi-walled nanotubes and zwitterionic single-walled nanotubes were stimulatory at the lowest dose (0.5 μg/ml). Sodium deoxycholate, often used to suspend nanomaterials, was also tested and was as stimulatory to the immune cells as the nanomaterials. This study is the first report of the effects of nanomaterials on the function of the immune system in a nonmammalian vertebrate. Since the innate immune system is the first to respond to the intrusion of foreign material, analysis of the effects of nanomaterials on cells of the innate immune system should provide valuable information on how these materials are perceived and affect an animal. Ultimately such research will provide the means to determine which nanomaterials are most harmful to aquatic species and how particles may be altered or functionalized to decrease their toxicity.