Characterization And Role Of Cellular M-CSF And Its Receptor In Leukemic Cells.

Macrophage colony-stimulating factor (M-CSF) is an important member of cytokine -regulating network in hematopoietic system. Encoded by proto-oncogene c-fms, M-CSF receptor (M-CSF-R) has intrinsic tyrosine kinase activity. M-CSF is characterized by the diversity of expression. Due to alternative splicing of M-CSF mRNA, M-CSF can exist in at least three forms: soluble M-CSF (s-M-CSF), membrane-bound M-CSF (m-M-CSF) and extracellular matrix or proteglycan-bound M-CSF (PG-M-CSF). And then it was found that there were M-CSF and its receptor in the cytoplasma and nucleus of some cells (referred to as cellular M-CSF). To explore the roles of cellular M-CSF and its receptor in leukemic cells, the expression and distribution of cellular M-CSF and its receptor in leukemic cells were studied by immunohistochemical staining and flow cytometry. The results of this report showed that cellular M-CSF and its receptor failed to be detected in normal human peripheral blood mononuclear cells (PBMCs), but a low level of expression was detected in PBMCs after PHA stimulation. Four human leukemic cell lines, including J6-1, J6-2, K562 and HL-60 cells expressed a high level of M-CSF and its receptor. Compared with stimulated PBMCs, the positve rates of m-M-CSF and its receptor in the 4 leukemic cell lines were significantly higher, and the expression level of intracellular M-CSF(cytoplasmic and nuclear M-CSF, cn-M-CSF) and its receptor (M-CSF-cnR) were markedly increased, except for K562. The cytoplasmic and nuclear proteins from HL-60 cells were isolated by differential centrifuge and followed by Western blot analysis. The results showed that three kinds of isoforms of M-CSF with molecular weight of 14 000, 16 000 and 20 000 respectively existed in the cytoplasma and nucleus of HL-60 cells. One form of M-CSF-R with molecular weight of 120 000 was detected inside HL-60 cells. These results suggested that expression of M-CSF in leukemic cells was diverse and heterogeneous.To further analyze the status and interaction between intracellular M-CSF and its receptor in leukemic cells, the intracellular M-CSF-R from HL-60 cells was precipitated with anti-M-CSF-R polyclonal antibody and followed by protein...