Growth and metastases of KGN cells in nude mice. KGN cells at the early (P7) and advanced (P58) passages were transplanted in the flanks of nude mice at a concentration of 5 × 106 cells. After 3 months, the growth of KGN cells at the injected region and metastases in the bowel were examined. (A) Growth of KGN cells (P7 and P59) at the injected regions shown by arrowheads. Tumour growth under the skin is shown in the enlarged windows. (B) Metastases of KGN cells in the bowel of nude mice. Arrowheads indicate metastases (nodules). The metastases were about 3 mm × 3 mm in size and were observed in 7 out of 9 mice. (C) The number of tumour nodules formed in the bowel was counted from 7 nude mice. Data are shown as the mean number of nodules with SEM. Statistical difference was analysed by Student's t-test (*P < 0.05). These experiments were quadruplicated. (D) Semi-quantitative RT-PCR analysis of human p53 in intestines with or without tumour nodules. Total RNA was isolated from the small intestine with or without nodules in each experimental group, and PCR analysis was performed using human-specific primers for p53. The upper panel represents a typical digital photograph taken on a transilluminator. Lower panel represents the averaged band intensity of p53 with SEM from four independent experiments.