Macrophage-melanocyte heterokaryons. I. Preparation and properties.
01 May 1970-Journal of Experimental Medicine (The Rockefeller University Press)-Vol. 131, Iss: 5, pp 981-1003
TL;DR: Observations indicate that typical macrophage properties cease to be expressed in heterokaryons, and melanocyte functions presumably predominate in heteroksaryons and hybrids.
Abstract: Mouse peritoneal macrophages, which do not synthesize DNA in vitro, were fused with melanocytes, a mouse cell strain which proliferates rapidly in vitro. DNA synthesis was induced in macrophage nuclei 2–3 hr after fusion and occurred irrespective of the number of macrophage nuclei present per melanocyte nucleus in each heterokaryon. 50–80% of macrophage nuclei initiated DNA synthesis in the 3–7 hr period after fusion. The activation of most 11–12-day chick red cell nuclei in melanocyte cytoplasm took longer than 10 hr. The lag before DNA synthesis may reflect the heterochromatin content of each nucleus.
Studies with actinomycin showed that heterokaryon RNA synthesis was essential for subsequent macrophage DNA synthesis. This RNA was synthesized 1–4 hr before the DNA and was unlikely to be ribosomal RNA, since it was insensitive to <0.1 µg/ml actinomycin.
Melanocytes and macrophages were treated before fusion with actinomycin and bromotubercidin to bring about a more selective inhibition of RNA synthesis. Macrophages pretreated for 1 hr with 5 µg/ml of actinomycin showed less than 20% of control RNA synthesis in the first 4 hr after fusion, but a normal activation of macrophage DNA synthesis. Pretreatment of melanocytes for 3–7 hr with 5 µg/ml bromotubercidin, a reversible inhibitor of RNA synthesis, prevented macrophage DNA synthesis without affecting macrophage RNA synthesis in the heterokaryons (81% of control). These studies showed that only melanocyte RNA synthesis was essential for the production of macrophage DNA.
The exposure of one cell partner to actinomycin before fusion caused cross-toxicity of the untreated nucleus after fusion. Bromotubercidin, an adenosine analogue which is incorporated into RNA, did not give rise to such cross-toxicity after fusion.
Once the macrophage nucleus becomes activated in the heterokaryon it becomes less sensitive to the action of actinomycin.
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TL;DR: Dendritic cells represent a novel cell type on both functional and morphological grounds and do not possess the functional properties of other types of reticular cells proposed to exist in lymphoid organs.
Abstract: A novel cell type has been identified in adherent cell populations prepared from mouse peripheral lymphoid organs (spleen, lymph node, Peyer's patch). Though present in small numbers (0.1–1.6% of the total nucleated cells) the cells have distinct morphological features. The nucleus is large, retractile, contorted in shape, and contains small nucleoli (usually two). The abundant cytoplasm is arranged in processes of varying length and width and contains many large spherical mitochondria. In the living state, the cells undergo characteristic movements, and unlike macrophages, do not appear to engage in active endocytosis. The term, dendritic cell, is proposed for this novel cell type.
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Cites background or methods from "Macrophage-melanocyte heterokaryons..."
...Peritoneal macrophages were harvested, without anticoagulants, by standard procedures ( 9 ); the cells were obtained either 4 days after stimulation by intraperitoneai injection of 0.75 ml thioglycollate medium (I0) or from control, unstimulated mice....
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...The morphologic appearance of unsfimulated ( 9 ) and thioglycollatestimulated (20) mouse peritoneal macrophages in culture has been described previously....
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TL;DR: This chapter discusses membrane and cytoplasmic changes in B lymphocytes induced by ligand–surface immunoglobulin (Ig) interaction by using ultrastructural analysis of regular thin section of cells exposed to antibodies conjugated to a large visible molecule.
Abstract: Publisher Summary This chapter discusses membrane and cytoplasmic changes in B lymphocytes induced by ligand–surface immunoglobulin (Ig) interaction. Surface Ig is the receptor for antigen molecules on B lymphocytes. Surface Ig has been identified on the B-cell surface by using immunocytochemistry or direct biochemical analysis. The number of Ig molecules presumably serving as antigen receptors on the B cell surface is on the average of 105 molecules per B cell. The amount of surface-bound Ig is large comparing to the number of hormone receptors needed to trigger certain cellular responses. One could speculate that many of the surface Ig–antigen interactions result in nonfunctional events; therefore, many receptors are needed. The B cell requires such a high number of receptors to effectively bind antigen molecules of large size and variable epitope densities or a number of successive hits are needed in these cells to reach an effective threshold of stimulation. The distribution of surface Ig and other surface macromolecules has been studied at the ultrastructural level by using various methods. One method was the ultrastructural analysis of regular thin section of cells exposed to antibodies conjugated to a large visible molecule.
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