Kidney glomerular explants in serum-free media: role of individual medium components in cell outgrowth

Guinea pig glomeruli were grown for 22 days in a serum-free medium composed of Waymouth's MB 752/1 supplemented with sodium pyruvate, nonessential amino acids, and antibiotics. To this basic medium was added insulin, transferrin, selenium (Se), tri-iodothyronine, or fibronectin (FN) - either singly, or in various combinations - and sequential quantitative studies of the glomerular outgrowths were performed. Total cells in glomerular outgrowths, mitotic index, and glomerular attachment rate were determined and compared with values for glomerular outgrowths in media containing either no additions or all of the above components. FN was required for whole glomerular attachment, while transferrin plus FN was required for mitosis in glomerular cell outgrowths. Insulin and tri-iodothyronine slightly increased glomerular cell outgrowth by slightly increasing whole glomerular attachment, but had little effect on mitosis in glomerular outgrowths. The effect of Se was complex. Se did not affect whole glomerular attachment or mitosis in the presence of transferrin plus FN. However, in a medium containing transferrin, FN, and 3-amino-1,2,4-triazole (AT) (an inhibitor of catalase and glutathione peroxidase), Se increased total cell number but had little effect on the glomerular attachment rate or the mitotic index. Morphologic analysis of glomeruli early in culture suggested that Se may act by decreasing the amount of or delaying the time of cell death. In all of the media tested, total DNA was relatively constant over the course of 22 days, suggesting the possibility that glomerular cells cultured in a serum-free medium are part of a cell renewal system.