Immunomodulatory Effects of Pure Cylindrospermopsin in Rats Orally Exposed for 28 Days

Abstract

Cylindrospermopsin (CYN) is a ubiquitous cyanotoxin showing increasing incidence worldwide. CYN has been classified as a cytotoxin and, among its toxic effects, its immunotoxicity is scarcely studied. This work investigates for the first time the influence of oral CYN exposure (18.75; 37.5 and 75 µg/kg b.w./day, for 28 days) on the mRNA expression of selected interleukin (IL) genes (IL-1β, IL-2, IL-6, Tumor Necrosis Factor alpha (TNF-α), Interferon gamma (IFN-γ)) in the thymus and the spleen of male and female rats, by quantitative real-time polymerase chain reaction (RT-qPCR). Moreover, their serum levels were also measured by a multiplex-bead-based immunoassay, and a histopathological study was performed. CYN produced immunomodulation mainly in the thymus of rats exposed to 75 μg CYN/kg b.w./day in both sexes. However, in the spleen only IL-1β and IL-2 (males), and TNF-α and IFN-γ (females) expression was modified after CYN exposure. Only female rats exposed to 18.75 μg CYN/kg b.w./day showed a significant decrease in TNF-α serum levels. There were no significant differences in the weight or histopathology in the organs studied. Further research is needed to obtain a deeper view of the molecular mechanisms involved in CYN immunotoxicity and its consequences on long-term exposures.

Keywords: cylindrospermopsin; immunotoxicity; mRNA expression; rats; spleen; thymus.

Figure 1

Interleukins (IL) IL-1β (a), IL-2 (b), IL-6 (c), tumour necrosis factor alpha (TNF-α) (d), and interferon-gamma (IFN-γ) (e) gene expression expressed as fold of control in the thymus of female and male rats in the different exposure scenarios (0, 18.75, 37.5, and 75 µg CYN/kg b.w./day, for 28 days). Mean values (±SEM) of N = 6 are calculated (* p < 0.05, ** p < 0.01, *** p < 0.001). The coded symbols represent the following: *—statistical differences compared with respective control group; ^#—statistical differences compared with the 18.75 µg CYN/kg b.w./day dose group; ^$—statistical differences compared with the 37.5 µg CYN/kg b.w./day dose group; ^&—statistical differences between female and male rats exposed to the same dose.

Figure 2

IL-1β (a), IL-2 (b), IL-6 (c), TNF-α (d), and IFN-γ (e) gene expression, expressed as fold of control in the spleen of female and male rats in the different exposure scenarios (0, 18.75, 37.5, and 75 µg CYN/kg b.w./day, for 28 days). Mean values (±SEM) of N = 6 are calculated (* p < 0.05, ** p < 0.01, *** p < 0.001). The coded symbols represent the following: *—statistical differences compared with respective control group; ^#—statistical differences compared with the 18.75 µg CYN/kg b.w./day dose group; ^$—statistical differences compared with the 37.5 µg CYN/kg b.w./day dose group; ^&—statistical differences between female and male rats exposed to the same dose.

Figure 3

Representative photomicrographs of the thymus from control (a,b) and 75 μg CYN/kg b.w./day groups (c,d). b and d images are higher magnifications of the fields framed in black (a and b, respectively). Notice the frequent presence of tingible body macrophages (arrow) in the CYN group. me—medulla; ct—cortex.

Figure 4

Representative photomicrographs of the spleen from control (a,b) and 75 μg CYN/kg b.w./day groups (c,d). b and d images are higher magnifications of the red pulp from the images a and b, respectively. Notice the normal presence of megakaryocytes (arrowhead) in both the control and CYN group. wp—white pulp; rp—red pulp.