Multixenobiotic resistance in coelomocytes from three echinoderm species

Abstract

Multixenobiotic resistance (MXR) proteins are known to be present in most living organisms, but only a few studies have been conducted on echinoderms and especially on their cir- culating cells, the coelomocytes. The objective of the present study was to investigate the presence of MXR activity in coelomocytes of the sea urchin Strongylocentrotus droebachiensis, the sea star Lept - asterias polaris and the sea cucumber Cucumaria frondosa. Cells were exposed to fluorescent sub- strates (1 µM Rhodamine B (RB) or 0.5 µM calcein-AM (CAM)), with or without inhibitors (50 µM ver- apamil (Ver), 5 µM cyclosporin-A (CsA) and 5 µM MK571 (MK)), and single-cell fluorescence was measured by flow cytometry. The combinations RB + CsA and RB + Ver induced a fluorescence in - crease in S. droebachiensis and L. polaris coelomocytes, as well as in S. droebachiensis vibratile cells. The combination RB + MK induced a fluorescence increase in S. droebachiensis and C. frondosa coelomocytes. Finally, the combination CAM + MK induced a fluorescence diminution in L. polaris coelomocytes and S. droebachiensis vibratile cells. This difference in fluorescence incorporation indi- cated an MXR-like activity in coelomocytes, probably due to the presence of a P-glycoprotein (Pgp) and a multidrug resistance-associated protein (MRP)-like transporter. Western blot analysis was also carried out (Ab C219 and Ab C9) in order to detect potential MXR proteins using anti-MXR anti - bodies. Both Pgp and MRP were detected, but could not be further discriminated. MXR activity was clearly demonstrated in coelomocytes of S. droebachiensis, L. polaris and C. frondosa, although the identity of proteins responsible for this activity needs to be confirmed.