Properties of tissue specific macrophages before and after in vitro activation (original) (raw)
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Macrophage activation in vivo and in vitro
Experimental Cell Research, 1977
Morphology, lysosomal enzyme activity and phagocytic ability were tested in peritoneal macrophage cultures after stimulation in vivo or in vitro with endotoxin, mineral oil or latex particles, and compared to the same parameters in normal peritoneal macrophages. Treatment with latex did not give changes in the parameters tested after in vivo or in vitro stimulation. In all other types of stimulation the cells displayed varying degrees of spreading and changes in granule content. Extensive ruffling of cell membrane was obvious in endotoxin-stimulated cells. The pattern of lysosomal enzyme activity was complex and depended on the means of stimulation. Acid phosphatase showed the greatest increase after both in vivo and in vitro stimulation, Nacetyl-glucosaminidase could not be increased in vitro. Internalization of opsonized red cells mediated by the Fc receptor increased after in vivo stimulation. No such change was observed after in vitro stimulation. Normal peritoneal macrophages do not internalize significantly via the C3 receptor. In vivo stimulation triggered the capacity to internalize up to 45 % of the attached red cells. A similar reaction was obtained in vitro when both endotoxin and FCS were added to the culture medium, but not when endotoxin or FCS were used alone. We conclude that the use of the term activation of macrophages should always be based on quantitative changes in well defined parameters. Changes in one parameter will not necessarily be accompanied by the whole range of biochemical and morphological perturbations. The capacity to ingest via the C3 receptor may be the most useful parameter.
Research in Immunology, 1992
Macrophages (MAC) are important effector cells of the immune system. They arise from circulating blood monocytes (MO), which undergo further maturation upon leaving the vasculature and migrating into the various tissues and body cavities. A similar differentiation process can be followed in vitro when monocytes are cultured in the presence of serum. In this study, different factors and serum proteins, either alone or in combination, were tested for their ability to promote the survival and/or maturation of blood MO in the absence of serum. Elutriation-purified MO cultured for 8 days on hydrophobic teflon foils in the presence of 5 % human serum differentiated into large, well-spread MAC, whereas in the absence of serum, MO rapidly died. The serum-induced maturation of MAC was accompanied by a strong expression of CD1 6, CD14 and MAX antigens. Secretion of TNF-alpha and neopterin increased about IO-fold as compared with freshly isolated MO. The replacement of serum by either M-CSF (100 nglml) or immunoglobulin (0.5-5 mglmll had a marked effect on MO survival (about 50 % of serum-cultured MO), but cells were smaller, less spread out and had low expression of CD1 6, CD14 and MAX antigens. Their functional competence in terms of TNF-alpha and neopterin release was reduced to IO-20 % as compared with MAC cultured in the presence of serum. Both albumin (0.5-5 mglml) and 1,25-dihydroxyvitamin D, (1,25(OH),D,) (10e8 M) as substitutes for serum were less effective in terms of MO survival (20-30 % of serum-cultured MO), but were comparable to serum with respect to morphology and phenotype; however, they induced MAC with lower secretory activity. A combination of 1,25(OHI,D, or albumin (0.5 mglml) with immunoglobulin (0.5 mg/ml) resulted in MAC showing serum-cultured characteristics in terms of phenotype, but lower secretory capacity and survival rate. However, the combination of the three factors together could substitute for serum in all parameters tested. The same result was obtained by cultivation of MO with high concentrations of albumin (5 mglml) and immunoglobulin (5 mglml). Other factors tested had no effect on the MO into MAC differentiation process (transferrin, vitamin A, fibronectin, vitamin D,). In summary, we describe defined serum-free culture conditions for the generation of distinct types of MAC, which differ in terms of phenotype, morphology and function.
Journal of Leukocyte Biology, 1984
Rabbit alveolar macrophages (AM) were separated into four subpopulations by centrifugation on discontinuous density gradients of Percoll. The subpopulation s were compared to unseparated AM populations for their ability to provide accessory function to ad herent cell-depleted splenocytes for antigen-stimulated Iymphoproliferation and for the production of Iymphokine. They were also tested for their ability to modulate in vitro plaque-forming (PFC) responses. AM subpopulations that provided accessory function for the production of migration inhibitory factor (M IF)-con taini ng cult ure supernantants were recovered from the least dense fractions of the Percoll gradients. These cells were cytoc hemically characterized as mature cells. AM that suppressed the in vitro PFC response and augmented th e antigen-stimulated Iymphoproliferative response to the greatest degree were recovered from the most dense fractions of the Percoll gradients and were characterized as immature cells. These results suggest that there are distinct subpopulatins of AM , the function of which may represe nt different stages of maturation (or differentiation).
Isolation and Long-term Cultivation of Mouse Alveolar Macrophages
Bio Protoc., 2019
Alveolar macrophages (AM) are tissue-resident macrophages that colonize the lung around birth and can self-maintain long-term in an adult organism without contribution of monocytes. AM are located in the pulmonary alveoli and can be harvested by washing the lungs using the method of bronchoalveolar lavage (BAL). Here, we compared different conditions of BAL to obtain high yields of murine AM for in vitro culture and expansion of AM. In addition, we describe specific culture conditions, under which AM proliferate long-term in liquid culture in the presence of granulocyte-macrophage colony-stimulating factor. This method can be used to obtain large numbers of AM for in vivo transplantation or for in vitro experiments with primary mouse macrophages.
Fems Immunology and Medical Microbiology, 1994
Abstract In the present study, we compared four macrophage (Mφ) cell lines from different anatomical origins for functional and secretory activities against the two morphogenetic forms of the fungus Candida albicans. We show that all the cell lines actively phagocytize the yeast and exert antimicrobial activity against both forms o3 Candida, although Mφ of microglial origin are the most effective. When assessed for secretory properties, microglial Mφ exhibit a peculiar patten with respect to other Mφ populations under either basal or stimulated conditions. In particular, only microglial Mφ fail to respond to the hyphal form of the fungus (H-Candida), which instead acts as a potent tumor necrosis factor inducer in the other Mφ cell lines. When exposed to H-Candida, microglial Mφ are indistinguishable from other Mφ in their ability to modulate specific surface adhesion molecules. In addition to strengthening the knowledge on functional heterogeneity among Mφ, our data provide evidence on the peculiar behavior of microglial Mφ. To what extent Mφ heterogeneity may be related to tissue homeostasis is discussed.
Influence of Various Detachment Procedures on the Functional State of Cultured Murine Macrophages
Immunobiology, 1981
Macrophages were obtained from seven-day-old bone marrow liquid cultures to which a colony-stimulating factor from L-cell-conditioned medium had been added. It was found that the yield of macrophages from the bone marrow liquid culture was dependent on the type and brand of culture dishes. Highest yields were obtained in teflon membrane bag cultures. The cumulative yield after serial passage of macrophages was 700-fold of the cell input after tWO months. After mechanical detachment from plastic culture dishes, the survival rate of cdls was related to the brand of the plastic dish. Viability also varied greatly after pretreatment of cells with scandicain (28 %), proteinase K (78 %), or pronase (99 %). After mechanical detachment of macrophages or pretreatment with scandicain, adherence to plastic, latex phagocytosis and chemotaxis was not or only slightly impaired, whereas the same functions after protease treatment were greatly reduced or even abolished and were only recovered after another culture period. Scandicain, proteinase K, and pronase treatment was mitogenic to macrophages in contrast to mechanical detachment. Pronase treatment also induced release of plasminogen activator activity. The results demonstrate that macrophages respond extremely sensitive to environmental conditions and to various insults by changing their functional state.
Journal of Experimental Medicine, 1975
Culture fluids rich in mononuclear phagocytes have a powerful effect on various in vitro reactions of lymphocytes (1-9). The culture fluids contain a mitogen for thymocytes and mature lymphocytes . Also found are active principles which differentiate memory B cells into antibody-secreting cells and which increase helper activities of T lymphocytes (2, 3). In a previous report we obtained from experiments in which lymphocytes were depleted from the cell preparations, strong evidence that the cellular source of the active component (s) was the macrophage (3). The activity secreted into the medium, however, was variable from experiment to experiment, perhaps related to the state of macrophage activation. In this paper we present results of experiments in which we studied the relation between macrophage activation and/or stimulation with the secretion of the various activities. We have found two ways in which to generate high levels of lymphostimulatory activities in macrophage cultures: one is to add a series of materials that are readily taken up by the macrophages; the second is to add a small number of activated T cells to the macrophage-rich cultures.