Amniotic fluid stem cells(AFSC) are noncontroversial and abundant potentially transplantable cells. They are derived from fetal and amniotic sources and show osteogenic, neuronal, and adipogenic differentiation in culture. Clinical material must be expanded in culture and sorted if clinical use is desired. Cellular senescence and replicative potential for AFSC cultures has had limited study, with scant data on gene expression over time. We report changes in samples from 17 patients over multiple passages form 10 to 81 population doublings. Longevity was unrelated to telomere length in these cells. It was related to upregulation of TWIST1, which is highly expressed in stem cells, and downregulation of genes associated with apoptosis.