Flow cytometric identification and isolation of hypertrophic type II pneumocytes after partial pneumonectomy.
Type II pneumocytes were isolated by either Percoll density gradient centrifugation or by immunoglobulin G (IgG) panning from the lungs of normal rats and the right lung of rats subjected to left pneumonectomy. Cells were studied at 7- (pnx-7) and 15- (pnx-15) days postoperative, times during and after, respectively, rapid compensatory growth of the right lung. Acridine orange staining permitted resolution of type II cells from contaminants on the basis of high red fluorescence (greater than 590 nm). Simultaneous measurement of forward-angle light scatter (FALS) suggested a shift of pnx-7 cells toward greater size, which was reversed in pnx-15 cells. By Percoll gradient isolation, approximately 15% of pnx-7 cells analyzed were above the mean FALS of control cells. In contrast, approximately 30% of the pnx-7 cells isolated by IgG panning were above the mean FALS of corresponding control cells. Biochemical analyses of pnx-7 cells separated by cell sorting into "high FALS" and "low FALS" subgroups revealed that high FALS type II cells contained 50% more protein (P less than 0.05) and 140% more RNA (P less than 0.01) than low FALS cells, with no significant change in cellular DNA content. These data are consistent with previous studies of type II cells isolated from the lungs of pneumonectomized animals and confirm the presence of hypertrophic cells in these preparations. They provide a foundation from which to design further flow cytometric studies of the role of hypertrophic type II pneumocytes in compensatory lung growth.
Department of Cellular and Molecular Physiology, College of Medicine, Pennsylvania State University, Hershey 17033.
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D000818:Animals; D002469:Cell Separation / Q000379:methods; D005434:Flow Cytometry; D006984:Hypertrophy / Q000473:pathology; D008168:Lung / Q000473:pathology*; D008297:Male; D008856:Microscopy, Fluorescence; D011013:Pneumonectomy; D051381:Rats; D011919:Rats, Inbred Strains