Establishment and characterization of two head kidney macrophage cell lines from large yellow croaker (Larimichthys crocea).

Two continuous macrophage cell lines (LCM07 and LCM10) were established for the first time from the head kidney of the marine fish large yellow croaker (Larimichthys crocea). To date, both cell lines have been subcultured for more than 100 passages in 12 months. Notably, the LCM07 and LCM10 cells have distinct morphology and immune function. LCM07 cells showed strong contact inhibition in crowded conditions, while this was not observed in the LCM10 cells because they could grow in an overlapping manner. Correspondingly, LCM10 cells were slenderer than LCM07 cells. LCM07 cells had stronger phagocytic ability than LCM10 cells, while LCM10 cells had stronger respiratory burst activity after incubation with lipopolysaccharide (LPS) and phorbol ester (PMA). LCM07 cells had stronger Escherichia coli killing ability than LCM10 cells. The mRNA of macrophage markers, namely that of CD11b, CD114, CD68, CD86, CD209, and CD163, were all expressed in primary macrophages as well as the two cell lines. The mRNA expression levels of selected inflammatory cytokines, namely interleukin (IL)-1β, IL-8, and tumor necrosis factor (TNF)α, were all upregulated after incubation with LPS. LPS also regulated key components of the mitogen-activated protein kinase (MAPK) signaling pathway, i.e., p38, ERK (extracellular signal-regulated kinase), and JNK (Jun N-terminal kinase) and their phosphorylated forms. Arachidonic acid (ARA) downregulated the LPS-induced upregulation of IL-1β, IL-8, and TNFα, revealing that LCM07 and LCM10 cells are useful for studying nutritional immunity. In conclusion, two distinct macrophage cell lines have been established for the first time from the head kidney of marine fish, which could be useful for studying immunity and nutritional immunity.