Biological characteristics and karyotiping of a new isolation method for human adipose mesenchymal stem cells in vitro

ObjectiveA new method was presented to prepare clinical-grade human adipose-derived stromal stem cells (ASCs) and its safety in vitro, such as biological characteristics and genetic features alteration were investigated.MethodsThe morphology of the ASCs which were cultured in vitro using serum-free medium was observed. Cell cycle and CD markers profile were tested by flow cytometry, while karyotype was analyzed by the chromosome G-banding technology. Growth factors expression was tested by ELISA and tumor-related genes were analyzed by the real-time PCR, respectively.ResultsASCs were adult stem cells with spindle shape. The proliferation ratio of ASCs began to slow down after 10 passages, and was significant after 15 passages. Cell cycle analysis revealed that the percentage of G2 phase and S phase cells was stable. There was no obvious missing, translocation or dislocation in terms of karyotype. Expression level of tumor relevant genes and cytokines at different passages had no significant difference.ConclusionsThe clinical-grade ASCs prepared with this new method, less than ten passages, was safe for clinical trials.