Flow cytometry as a tool to monitor the disturbance of phagocytosis in the clam Mya arenaria hemocytes following in vitro exposure to heavy metals.

Abstract

The effectiveness of toxicology biomonitoring programs could be improved by the addition of sensitive biomarkers. In this study the cell viability and sensitivity of phagocytic function of phagocytes from bivalves ( Mya arenaria) to selected heavy metals were measured by flow cytometry, a novel approach. Hemocytes (phagocytes) collected from bivalves by puncture of the posterior adductor muscle were incubated in vitro for 18 h in hemolymph containing 10 −9–10 −3 M of cadmium chloride, zinc chloride, mercuric chloride, methylmercury chloride or silver nitrate, before determining their capacity to phagocytose fluorescent latex beads by flow cytometry. Heterogeneity of the hemocyte cell population was determined by forward scatter (FSC) and side scatter (SSC) cytometric profile which showed two distinct cell populations. At low doses (10 −9, 10 −8 M), all the metal compounds studied stimulated phagocytic activity except silver nitrate. At higher levels of exposure (10 −6, 10 7 M), all metals caused a significant concentration-related decrease in hemocyte phagocytosis activity. From the concentration of each metal inducing 50% suppression (IC50) of the phagocytic activity, the immunotoxic potential of metals with respect to phagocytic function can be ranked in the following increasing order: ZnCl 2 < CdCl 2 < AgNO 3 < HgCl 2 < CH 3HgCl. Parallel analysis of hemocyte viability showed that suppression of phagocytosis by heavy metals was not solely related to a decreased cell viability. These results reveal the high but different degree of sensitivity of the phagocytosis activity of bivalves with respect to heavy metals, as measured by flow cytometry, and demonstrate that flow cytometry is a potentially useful tool in ecotoxicological monitoring.