Subpopulations of mouse resident peritoneal macrophages fractionated on Percoll gradients show differences in cell size, lectin binding and antigen expression suggestive of different stages of maturation.
Aiming the identification of macrophage heterogeneity, mouse resident peritoneal cells were fractionated on discontinuous Percoll gradients into six discrete fractions (0, 1, 2, 3, 4 and 5 in order of increasing density). All six fractions and the total population were characterized by light and electron microscopy, and flow cytometry. The least dense fraction (0) had a low viability (44%); fractions 4 and 5 had a low percentage of macrophages. Light microscopy and flow cytometry of macrophage-enriched fractions 1, 2 and 3 showed an inverse correlation between diameter and cell density, as well as suitable differences in lectin binding to the macrophages of each fraction. The surface of macrophages from fraction 1 had more sialyl residues (higher binding of the lectin LFA), less galactosyl residues (lower binding of the lectin PNA) and expressed more receptors for the antibodies M1/70 (Mac-1) and F4/80; fraction 3 had an opposite binding behavior for the lectins and expressed fewer receptors for both antibodies; fraction 2 had an intermediate behavior for both parameters. Binding of the lectins Con A and HPA showed slight differences, whereas UEA I did not present a detectable difference among the fractions analyzed. These findings suggest that the macrophage heterogeneity achieved by the gradient separation of resident peritoneal cells could be explained by different stages of macrophage maturation.
D000818:Animals; D000954:Antigens, Surface / Q000096:biosynthesis; D002454:Cell Differentiation; D002469:Cell Separation; D048429:Cell Size; D002499:Centrifugation, Density Gradient; D005434:Flow Cytometry; D037102:Lectins / Q000378:metabolism; D017737:Macrophages, Peritoneal / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D008297:Male; D051379:Mice; D008854:Microscopy, Electron; D011205:Povidone; D012822:Silicon Dioxide
D000954:Antigens, Surface; D037102:Lectins; C016039:Percoll; D012822:Silicon Dioxide; D011205:Povidone