Expression of antimicrobial peptides from Hyas araneus haemocytes following bacterial challenge in vitro

Circulating haemocytes play major roles in the host defense reactions of decapods, including the synthesis and release of antimicrobial peptides (AMPs). Unlike the AMPs from insects, those in decapods are constitutively expressed. This study aims to establish primary cell cultures of the three main haemocyte types in Hyas araneus haemocytes, and to measure the in vitro expression of AMP genes in the cells following microbial challenge. The haemocyte populations were separated on Percoll gradients and cultured in modified L-15 medium. Expression analysis by real-time RT-PCR showed that the granular cells are the main producers of crustin, hyastatin and arasin 1 AMPs, but the hyaline cells and semigranular cells also show some expression of these genes. Incubating the cell populations with Aerococcus viridans var. homari (a Gram-positive bacterium) or Listonella anguillarum (a Gram-negative pathogen) provoked no dramatic changes in the gene expression of any of the AMP, and although there was a small (single doubling) significant increase in expression of the crustin gene in granular cells 24 h after exposure to L. anguillarum, it is unclear if this is biologically relevant under in vitro conditions. The results presented in this study are in accordance with several in vivo studies.