Regulation of immune functions by microcystins and cannabinoids

Abstract

Contamination of water by cyanobacteria (blue-green algae) is a health threat because of their production of toxic peptides, termed microcystins. The primary target organ for microcystins is liver, but other cellular targets have not been elucidated. In the present study, the effect of microcystins on immune functions was investigated. Representative microcystin compounds, microcystin-LR (MC-LR), microcystin-YR (MC-YR) and nodularin, produced dose-dependent inhibition of in vitro polyclonal antibody response and lymphoproliferative activity to LPS. MC-YR and nodularin decreased lymphoproliferative response to ConA, whereas MC-LR showed no significant effect. Intraperitoneal administration of nodularin produced an inhibition of primary humoral immune responses to the T-cell dependent antigen, sRBC. Interleukin-2 (IL-2) protein secretion in phorbol-12-myristate-13-acetate (PMA) plus ionomycin (Io)-induced splenocytes was measured by enzyme-linked immunosorbent assay (ELISA), which was decreased by the microcystins treatment. When IL-2 mRNA expression was evaluated by quantitative/competitive RT-PCR, microcystins (MC-LR, MC-YR, and nodularin) decreased IL-2 mRNA expression in murine splenocytes and thymocytes in time- and dose-related manner. But no apparent effect by microcystins was observed in EL-4 T-cells. To further characterize the inhibitory mechanism of IL-2 expression by microcystins, IL-2 mRNA stability and the binding activities of transcription factors were evaluated. MC-LR, MC-YR and nodularin led to more rapid destabilization of the expressed IL-2 mRNA. Electrophoretic mobility shift assays (EMSA) showed that nodularin reduced the NF-AT binding activity in PMA/Io-stimulated mouse splenocytes, but no significant effect was observed on AP-1, NF-kB, and Oct binding activity. Based on the fact that IL-4 gene expression is controlled by NF-AT, the effect of nodularin on IL-4 mRNA expression was measured. Nodularin also inhibited IL-4 mRNA expression in PM...