Immune priming confers a sustained, augmented response of innate immune cells to a secondary challenge, a process that is characteristically reliant on metabolic reprogramming. Recent evidence suggests that histone demethylases play essential roles in the immune priming, while its regulation role in the metabolic reprogramming remains largely unknown. In the present study, the concentration of glucose was significantly down-regulated in the hemocytes of crab Eriocheir sinensis after secondary stimulation with Aeromonas hydrophila, while the expression levels of phosphofructokinase (EsPFK) pyruvate kinase (EsPK), hexokinase-2 (EsHK-2) and Glucose-6-phosphate dehydrogenase (EsG-6-PD), along with the concentrations of lactate and the ratio of NAD⁺/NADH, were elevated. Additionally, the levels of H3K9me3 and its enrichment at the promoters of EsPFK and EsG-6-PD were significantly decreased at 7 days after A. hydrophila stimulation. The lysine Demethylase 4 homologue (EsKDM4) was observed to translocate into the nucleus of crab hemocytes after A. hydrophila stimulation, and its activity markedly increased after secondary stimulation with A. hydrophila. Following RNA interference of EsKDM4, there was a significant increase in H3K9me3 levels, and the enrichment of H3K9me3 at the EsPFK and EsG-6-PD promoters, as well as the concentration of glucose, in the hemocytes of crabs after secondary stimulation with A. hydrophila. Furthermore, mRNA transcripts of EsPFK and EsG-6-PD, as well as the concentration of lactate and ratio of NAD⁺/NADH, significantly decreased after secondary stimulation. These results suggested that EsKDM4 mediates the enrichment of H3K9me3 at the promoters of EsPFK and EsG-6-PD, thereby regulating glycolysis during the immune priming of crabs.

Hematopoiesis is a complex process of creating new hemocytes and releasing them from hematopoietic tissue. In the present study, the hematopoietic site in oyster Crassostrea gigas was successfully identified in the proximal sector (designated G2–G3) of the gill hinge with a substantial number of newborn cells and a minor presence of stem-like cells. The homologues of VEGF (CgVEGF) and its receptor CgVEGFR were characterized, and they interacted with each other. After the oysters received an injection of rCgVEGF, the number of EdU-positive (EdU⁺) cells increased within the G2–G3 sector and the hemolymph. When the expression of CgVEGFR was inhibited by RNAi, the percentage of EdU⁺ cells in the hemolymph declined dramatically, but increased significantly in the G2–G3 sector and EdU⁺ cells aggregated in this region. Meanwhile, the phosphorylation levels of CgErk and CgJNK, mRNA transcripts of cell proliferation-related and cell migration-related genes, reduced significantly. These results indicate that the proximal region of the hinge in gill was the site producing hemocytes, and CgVEGF-VEGFR-MAPK signaling pathway induced the migration of newborn hemocytes from this site to the circulating hemolymph, which provides new clues about hematopoiesis in primary invertebrates.

Staphylococcus aureus is a leading cause of severe infections in humans and animals, and the emergence of multidrug-resistant strains highlights the need to develop effective vaccines to prevent such diseases. Epitope-based vaccines use short antigen-derived peptides corresponding to immune epitopes, which are administered to trigger protective humoral and cellular immune responses. In this study, in silico MHC affinity measurement methods were used to predict possible binding regions, and five 20-mer synthetic TRAP peptides (TRAPP) were synthesized. Epitope-based vaccines, named PT and PTR, incorporating the identified CD4⁺ T and B cell epitopes, were constructed. Peptides TRAP_20–39 and TRAP_94–113 elicited significant peptide-stimulated T-cell proliferation responses in vivo. Additionally, high levels of IFN-γ and IL-17A, along with moderate levels of IL-4, were detected in ex vivo stimulated CD4⁺ T cells isolated from rTRAP- and TRAPP-immunized mice, suggesting that these peptides are classified as Th1 and Th17 epitopes. Immunization with PT or PTR induces robust humoral and cellular immune responses. Moreover, the epitope-based vaccine, PT, exhibited a stronger protective immune response than the intact TRAP in a murine systemic S. aureus infection model. Based on the results presented herein, an epitope-based vaccine is a promising and potentially more effective candidate.