Comparative immunological study on granulocyte populations in the Leydig organ of two elasmobranch species

Elasmobranchs are the most primitive jawed vertebrates, and their immune system is similar to those of amniotes. Therefore, sharks and rays represent excellent models for investigating the evolution of immune system components and functions. In addition to the typical lymphoid tissues, namely the thymus and spleen, elasmobranchs have Leydig (LO) and epigonal organs instead of bone marrow. The LO is located between the submucosa and muscular layers of the esophagus. Here, we report the cell composition of the LO in two elasmobranch species, the thornback ray (Raja clavata) and the small-spotted catshark (Scyliorhinus canicula). In February 2021, ten specimens of each species were collected from North Sardinia (Italy), and the esophagus was excised and studied using immunohistochemistry to characterize the immune cells in the LO. The anti-proliferating cell nuclear antigen antibody showed that approximately 44 % of cells were in DNA synthesis activity, and approximately 14 % were myeloid progenitors positive for the anti-c-kit antibody. A panel of antibodies targeting the main molecules of immune system cells was employed, showing that the LO of R. clavata and S. canicula had similar percentages of areas occupied by granulocyte types containing lysozyme, serotonin, histamine, tumor necrosis factor-like peptide, and inducible nitric oxide synthase. A comparison of the current data on these two species with previous results on the blackmouth catshark, Galeus melastomus, revealed some differences in the percentages of areas with granulocyte populations that express histamine, serotonin, and tumor necrosis factor-like peptide.