BACKGROUND: Fibroblasts are considered as seed cells necessary for the construction of tissue engineering skin. The ultrastructure of cells of various generations was observed under the electron microscope in the hope of providing foundation for proper selection of seed cells for tissue engineering skin.OBJECTIVE: To observe the ultrastructural changes of normal human fibroblasts during in vitro culture.DESIGN: Self-control observation.SETTING: Institute of Plastic Surgery, Weifang Medical College; Department of General Surgery affiliated to Weifang Medical College.MATERIALS: This experiment was carried out in the Institute of Plastic Surgery, Weifang Medical College, between September 2000 and September 2002. Healthy prepuce specimens were collected during posthetomy from normal boys aged 6-8 years after the informed consent was obtained from their guardians.METHODS: The normal human diploid fibroblasts were used to carry out consecutive subculture; cells were collected from different generations for morphological and ultrastructural observation under the inverted phase contrast microscope and transmission electron microscope.ture under the transmission electron microscope.microscope: Cells could pass on for 65-70 generations and survive for 280-300 days. Cells within 45 generations could grow rapidly, but gradually grew slowly after the 45th generation, and even displayed no proliferaunder the transmission electron microscope: There were no obvious changes in cell ultrastructure within 40 generations, but cells presented inward tolds of nucleus membrane from the onset of generations 41-65, with the ratio of cell nuclear/plasma reduced as well as cell surface process and microvilli also reduced.CONCLUSION: The ultrastructural change of in vitro cultured fibroblasts varied between different generations, which became obvious after the 41st generation, suggesting that fibroblasts within 40 generations are considered preferable seed cells for the construction of tissue engineering skin.