Isolation and characterization of inflammatory leukocytes from glomeruli in an in situ model of glomerulonephritis in the rat.
Inflammatory cell populations in glomerulonephritis (GN) are not well characterized. A method is reported for isolating leukocytes from glomeruli. GN was induced in rats by perfusing left kidneys (LKs) with cationized human IgG followed by intravenous rat anti-human IgG serum. Acute GN developed in LKs with proteinuria, deposition of human and rat IgG and C3, leukocyte infiltration, and capillary wall electron-dense deposits. Glomeruli (GL) isolated at 24 hours were digested with collagenase, trypsin, and DNase, and the resulting cells were as follows (mean +/- SEM): LK, 354 +/- 25/GL; RK, 214 +/- 32/GL. Cells were labeled with monoclonal antibody MRCOX1 (anti-rat leukocyte common [LC] antigen) followed by FITC F(ab')2 rabbit anti-mouse Ig: LK, 170 +/- 11 leukocytes/GL;RK, 8 +/- 2 leukocytes/GL (P less than 0.001). Isolated cells were sorted by flow cytometry to 98% pure LC+ cells with greater than 80% viability (Giemsa staining: 86% mononuclear cells, 14% neutrophils); the ultrastructure was that of maturing macrophages and neutrophils. This method quantitates leukocyte infiltration and provides leukocytes from nephritic glomeruli suitable for in vitro studies.
D000818:Animals; D002469:Cell Separation; D005434:Flow Cytometry; D005921:Glomerulonephritis / Q000473:pathology*; D007124:Immunoenzyme Techniques; D007678:Kidney Glomerulus / Q000473:pathology* / Q000648:ultrastructure; D007962:Leukocytes / Q000473:pathology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D051381:Rats; D011917:Rats, Inbred Lew