PRODUCTION OF A MACROPHAGE GROWTH FACTOR(S) BY A GOLDFISH MACROPHAGE CELL-LINE AND MACROPHAGES DERIVED FROM GOLDFISH KIDNEY LEUKOCYTES

We recently established a spontaneously proliferating macrophage cell line from the goldfish (GMCL), and in this report demonstrate the prod uction of a macrophage-specific growth factor(s) (MGFs) by these cells . The supernatants from GMCL cultures induced proliferation and differ entiation of macrophage-like cells from kidney hematopoietic tissues o f goldfish. Kidney leukocytes cultured at 6.25 x 10(4) cells/ml in the presence of GMCL-derived derived MGFs proliferated during two weeks o f cultivation, whereas those cultured without the MGFs did not. Leukoc ytes cultured at higher densities (2.5 x 10(5) cells/ml) proliferated in the absence of exogenous growth factor, but not to the same extent as those stimulated with GMCL-derived MGFs, suggesting that kidney leu kocytes may produce endogenous MGFs. At higher cell density (1 x 10(6) cells/ml), kidney leukocytes multiplied extensively over a two-week c ultivation period in the absence of exogenous GMCL-derived MGFs. The s upernatants from these cultures restored the proliferative ability of leukocytes cultured at low densities, providing direct evidence of MGF s production by kidney leukocytes, The predominant cell-type in cultur es grown in the presence of GMCL or kidney leukocyte-MGFs was the macr ophage based on the following criteria: (1) non-specific esterase stai ning; (2) morphologic similarity to GMCL; (3) phagocytosis of the bact erium, A. salmonicida; (4) production of reactive oxygen and nitrogen intermediates in response to stimulation with macrophage activating fa ctors and/or bacterial lipopolysaccharide; and (5) flow cytometric ana lyses. Both in vitro-derived kidney macrophage (IVDKM) and GMCL cultur es contained three distinct populations of cells, (determined by bow c ytometry), suggesting that these macrophage cultures are comprised of cells arrested at distinct differentiation junctures in macrophage dev elopment, Production of MGFs by macrophages and kidney leukocytes may play an important role in regulating macrophage hematopoiesis in fish. (C) 1998 Elsevier Science Ltd. All rights reserved.