The hematopoietic tissue of the freshwater crayfish, Pacifastacus leniusculus: organization and expression analysis

The hematopoietic tissue (HPT) and anterior proliferation center (APC) are the main hemocyte-producing organs of the freshwater crayfish, Pacifastacus leniusculus. To deepen our understanding of immune responses to various pathogens, it is essential to identify distinct hemocyte subpopulations with specific functions and to further explore how these cells are generated. Here we provide an in-depth histological study of the HPT and APC in order to localize cell types in different developmental stages, and to provide some information regarding the hemocyte differentiation in the crayfish. We localized mRNA expression of previously identified genes in the HPT/APC and hemocytes by RNA-FISH. The expression of hemolectin and transglutaminase 1 was shown to be co-localized in a high number of the HPT cells, while transglutaminase 2 was expressed in different cell types mainly associated with epithelium or endothelium. Furthermore, by double RNA-FISH for hemolectin and a previously unidentified PDGF-like factor, combined with immunostaining for prophenoloxidase, we could identify several different subtypes of hemocytes, indicating that the immune function of hemocytes in crayfish is more diversified and complex than previously appreciated.