Non-specific Cellular Function of Eisenia Fetida Regulated by Polycyclic Aromatic Hydrocarbons

Exposure to ubiquitous environmental chemicals, such as polycyclic aromatic hydrocarbons (PAH), may affect the earthworm cellular immune-defense system. Historically, the coelomocytes of earthworms were classified into several cell types, including lymphoid cells and granular cells. Those cells revealed a cellular defense mechanism related to phagocytosis, hemolysis and cytolysis. We examined cellular functions of coelomocytes such as phagocytosis and H(2)O(2) activity by flow cytometry after exposure to soil contaminated with 7,1 2-dimethylbenzanthracene (DMBA). In addition, increased synthesis of cytochrome P450 was examined by immunohistochemistry and western blot. Sexually mature Eisenia fetida were kept in dark at 18 degreesC with various doses of DMBA (0-100 mug) contaminated artificial soil (10g each in glass jars) for 7 days. Coelomocytes were harvested from earthworms and processed for in vitro phagocytosis and H(2)O(2) activity. Phagocytosis was assessed by ingestion of fluorescence beads and H(2)O(2) activity examined using 2', 7'-dichloro-fluorescin diacetate that was measured by flowcytometery 2h after incubation. Coelomocytes were classified as small and large cells using flowcytometery. The small cells including lymphoid cells, granular cells and hyaline cells were major effector cells for earthworm non-specific immune response. Results indicated that cell functions were down regulated in a dose dependent manner after exposure to sublethal doses of DMBA. Cytochrome P450 expression increased as revealed by immunohistochemistry and western bolt analysis, These non-specific cellular immune functions (expressions of innate immunity) permit us to evaluate biological activity of earthworms and the effect of soil contamination for future ecotoxicological studies.