Specific inhibitors of mitogen-activated protein kinase and PI3-K pathways impair immune responses by hemocytes of trematode intermediate host snails

Abstract

To characterize molecular mechanisms regulating snail cellular immune responses, the contributions of mitogen-activated protein kinases (MAPKs) and phosphatidylinositol 3-kinase (PI3-K) were examined in hemocytes of the trematode intermediate host snails Biomphalaria glabrata and Lymnaea stagnalis. Simultaneous measurement of phagocytosis/encapsulation and H2O2 production by hemocytes in the presence or absence of specific signal transduction inhibitors was used to assess the role of extracellular-signal regulated kinases 1 and 2 (ERK1/2), p38, JNK and PI3-K. Hemocyte spreading was significantly reduced in a dose-dependent manner by the ERK inhibitor, PD098059, and by wortmannin, a potent PI3-K inhibitor. The JNK inhibitor, SP600125, and the p38 kinase inhibitor, SB203580, had no effect on hemocyte spreading. Sheep red blood cell phagocytosis was significantly impaired by PD098059, SP600125, and SB203580. Hydrogen peroxide production during phagocytosis was severely inhibited by PD098059. Additionally, PD098059, but not the other inhibitors, significantly impaired the cellular encapsulation of trematode larvae and H2O2 production during encapsulation. These results suggest that MAPK and PI3-K signal transduction pathways play a pivotal role in the immune responses of snail hemocytes. PI3-K and ERK appear to strongly regulate cell motility. ERK, JNK and p38 contribute to phagocytosis-mediated signal transduction. ERK also play a major role in oxidative burst activation and the encapsulation of trematode larvae by snail hemocytes.