Cell fusion of human and mouse cells as a source for new cells retaining human markers. Analysis of DNA content, membrane and cytoplasmic antigen expression.
Flow cytometry and ultrastructural morphometry were used to study some characteristics of cells obtained by fusion with polyethylene-glycol 4000 between mouse fibroblasts 3T3.4E and normal keratinocytes (3T3.4E x NHK) or hand wart human keratinocytes (3T3.4E x HWK), at late passages. The cell cycle and the expression of human beta 2-microglobulin, human EGF-receptors (EGF-r), vimentin were simultaneously studied by flow cytometry. Epithelial CaSki cells, derived from a human uterine carcinoma, expressing high levels of beta 2-microglobulin, EGF-r and vimentin, were used as a positive control. In mouse fibroblasts 3T3.4E only vimentin was expressed whereas in cells derived from fusion, human beta 2-microglobulin, human EGF-r and vimentin were detected. The cell cycle analysis revealed that the peak position of G0/G1 differed with the cells (channel 11 for 3T3.4E cells, 13 for 3T3.4E x HWK and 15 for 3T3.4E x NHK). The area of the cell compartments from each cell type was also different by quantitative ultrastructural morphometry. The hybrid phenotype was maintained in late passages in cells (3T3.4E x NHK) and (3T3.4E x HWK), as shown by the expression of human antigens, differences in DNA contents and nuclear area. Flow cytometry may be a very accurate and precise tool for studying low antigenic expression. The combination of different methods including analysis of DNA content, antigenic expression and ultrastructural morphometry confirmed that 3T3.4E, 3T3.4E x NHK and 3T3.4E x HWK cells are different cell types. These techniques are complementary to cell phenotype analysis.(ABSTRACT TRUNCATED AT 250 WORDS)
Lizard
G
G
Regroupement de Bio-Technologies Médicales, Dijon, France.
Chardonnet
Y
Y
Chignol
M C
MC
Thivolet
J
J
D016475:3T3 Cells / Q000737:chemistry / Q000648:ultrastructure; D000818:Animals; D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm / Q000096:biosynthesis; D000954:Antigens, Surface / Q000096:biosynthesis; D015415:Biomarkers; D002294:Carcinoma, Squamous Cell / Q000473:pathology*; D002453:Cell Cycle; D002459:Cell Fusion; D002467:Cell Nucleus / Q000648:ultrastructure; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis; D066246:ErbB Receptors / Q000096:biosynthesis; D005260:Female; D005434:Flow Cytometry; D006229:Hand Dermatoses / Q000473:pathology; D006801:Humans; D006822:Hybrid Cells / Q000737:chemistry / Q000648:ultrastructure; D015603:Keratinocytes / Q000737:chemistry / Q000648:ultrastructure; D008565:Membrane Proteins / Q000096:biosynthesis; D051379:Mice; D009363:Neoplasm Proteins / Q000096:biosynthesis; D010641:Phenotype; D011092:Polyethylene Glycols; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000648:ultrastructure; D014594:Uterine Neoplasms / Q000473:pathology*; D014746:Vimentin / Q000096:biosynthesis; D014860:Warts / Q000473:pathology; D001613:beta 2-Microglobulin / Q000096:biosynthesis
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D015415:Biomarkers; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D014746:Vimentin; D001613:beta 2-Microglobulin; D011092:Polyethylene Glycols; D004247:DNA; D066246:ErbB Receptors