Transferrin as a growth factor for rat bladder carcinoma cells in culture.

Using the heterotopically transplanted rat urinary bladder system, we previously showed that normal urine has a tumor-enhancing effect on carcinogen-initiated urothelium. In an attempt to isolate a urinary growth-stimulating (tumor-enhancing) factor(s), urine was first fractionated by Bio-Gel P-100 column chromatography, and each fraction was tested for inducibility of ornithine decarboxylase (ODC) and growth-stimulatory activity in a target rat bladder carcinoma cell line, 804G. ODC inducibility was chosen as a marker for tumor-enhancing effect because it is a key characteristic of tumor promoters. There was a single peak demonstrating a strong growth-stimulatory activity as measured by [3H]thymidine incorporation. There were two ODC-inducible peaks, one located at a high molecular weight region and partly overlapped with the growth-stimulatory peak. The other was located at a lower molecular weight region. CM-Sephadex chromatography and subsequent high performance liquid chromatography successfully separated the high molecular weight-ODC activity from the growth-stimulatory activity. The latter component was found to contain transferrin (TF) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunodiffusion with anti-rat TF antibody and was designated as urinary transferrin fraction. The urinary TF fraction and authentic rat TF stimulated growth of several rat bladder carcinoma cells maintained in a serum-free as well as a serum-deficient medium. The response was proportional to the concentration of TF ranging from 0.2 to 5 microgram/ml. Preincubation of the urinary TF fraction or TF with an anti-rat TF significantly reduced their growth-stimulatory effects in 804G cells. The high molecular weight-ODC also stimulated cell growth but to a lesser extent. These results when combined with our previous observations suggest that TF and possibly also ODC-inducible substances may be important urinary components participating in the tumor promotion by urine.