The hemolymph coagulation mechanism of the spiny lobster, Panulirus japonicus, was investigated in vitro focusing on transglutaminase (electing enzyme), cytolysis and plasma gelation. Hemocytes and plasma were prepared separately from hemolymph sampled into anticoagulant saline. Transglutaminase activity of hemolymph was detected only in hemocytes, and most of the activity in hemolymph was derived from hyaline and semigranular cells (H/SG cells). Plasma showed strong cytolytic activity toward H/SG cells, and the activity was calcium-dependent. Plasma gelled after being mixed with unseparated whole hemocytes. Plasma also gelled when mixed with separated H/SG cells, while no gelation was detected in plasma mixed with granular cells. When freshly drawn hemolymph was sonicated to disrupt hemocytes, gelation time of the hemolymph was remarkably shortened compared with unsonicated hemolymph. From these results, the reaction sequence in hemolymph coagulation system was outlined: 1) plasma induces cytolysis in H/SG cells; 2) clotting enzyme (transglutaminase) is released by the lysis of these hemocytes; and) the enzyme catalyzes gelation of plasma. Therefore, the reaction of hemolymph coagulation proceeds by interaction between hemocytes and plasma, and the cytolysis of H/SG cells induced by plasma is a key phenomenon in this system.
