Effects of dietary chitosan-selenium nanoparticles (CTS-SeNPs) on zebrafish (Danio rerio) immune system

Abstract

Selenium (Se) is an essential micronutrient for human and all animals. There are 25 selenoproteins in human and they are primarily related to antioxidant functions. Effect of Se to the body is complex: it is dependent on the administration dose, the subject's baseline Se status, the administered form of Se and its subsequent metabolism. Recently, there was increasing interest in the bioactivity of selenium nanoparticles (SeNPs). They are small particles (typically <100nm) of elemental Se and are typically stabilized with polysaccharides and/or proteins at the surface to enhance stability and dispersibility in water. In this study, chitosan stabilized selenium nanoparticles (CTS-SeNPs) were synthesized by using controllable reduction methods. immunostimulation activity of CTS-SeNPs was explored in zebrafish (Danio rerio) innate and adaptive immune systems. This thesis aims to explore the immune effects of CTS-SeNPs and its underlying mechanism. In Chapter 2, zebrafish were fed different doses of CTS-SeNP for 3-60d. Immune biomarker assays and bacterial challenge were conducted to evaluate the immunostimulation effects of CTS-SeNP. CTS-SeNP showed immunostimulation effect from 5 to 20 µg/g but with the best outcome at 10 µg/g. Immunostimulation effect were rapidly induced after 3-9d and can sustain to 60d. The zebrafish fed with 10 µg/g CTS-SeNP also had survivalship 26.7 % higher than the control after intraperitoneal injection of common bacterium Aeromonas hydrophila. Our results suggested that CTS-SeNP is an effective immunostimulant to fish and has potential application in aquaculture. In Chapter 3, mechanism of immunostimulation effects of CTS-SeNP was explored with proteomics approach on fish serum. Protein expression profiles of fish fed on CTS-SeNP and control diet under 1) non-infection (normal) condition or 2) under bacterial infection condition were compared. Molecular network analyses indicated in non-infection condition, CTS-SeNP activated free radical scavenging network by increasing the expression of two key regulator, SOD1 and NF-KB. It suggested that SeNP might be anti-inflammatory under normal CTS-SeNP supplementary condition. Under infection condition, complement pathways were more activated in zebrafish supplied with CTS-SeNP. Improvement of complement activation indirectly enhance the downstream immune responses. In Chapter 4, to test the hypothesis that immunomodulation of CTS-SeNP was driven by improvement in redox condition, effects of CTS-SeNP was compared with Trolox at equal antioxidant capacity. CTS-SeNP and Trolox offered similar protection effect on bacterial infection, but CTS-SeNP has better redox regulation and immune biomarker responses. These effects were likely related to functions of selenoproteins, indicating that CTS-SeNP was biologically metabolized to improve both the immune system. In Chapter 5, transcriptome analysis of zebrafish kidney was carried out by RNAseq. Results again demonstrated that CTS-SeNP affect pathways involved in immune system, and also lipid metabolism and environmental adaptation. Findings echoed results in Chapter 2 that many genes involve in T-cell regulation and innate immunity was up-regulated by CTS-SeNP. Finally, in Chapter 6 an overall summary of CTS-SeNPs immunostimulation effects was presented and potential immune mechanism of SeNP was discussed. Limitations of the study, future research need and potential applications of this NP were also presented.