Phenotypic, Functional, and Safety Control at Preimplantation Phase of MSC-Based Therapy

Figure 4

(a) The influence of physioxic 5% O₂ or normoxic 21% O₂ atmosphere on the quality of mesenchymal stem cell populations. AD-MSC cultured in 5% O₂ retain relatively stable proliferation rate, when AD-MSC culture in 21% O₂ atmosphere retain unstable, oscillating growth rate with the estimated population doubling time apparently falling down and rising again successively. (b) Percentage of senescent cells. In 21% O₂ atmosphere the increased number of SA-βgal expressing cells was observed relative to cells cultured in 5% O₂ atmosphere. (c) The morphology of AD-MSC in physioxic and normoxic conditions. Whereas in the physioxic conditions AD-MSC retained spindle-shaped morphology during the whole estimated period, in normoxic conditions they firstly acquired typical feature of aging cells and then their phenotype changed. (d) Flow cytometry analysis. After long-term culture in 21% O₂ atmosphere we have noticed disturbed expression of MSC-specific surface antigen pattern, including complete lack of CD90, CD105, and CD73. AD-MSC growing in 5% O₂ by the same number of passages still presented correct CD90, CD105, and CD73 antigens pattern with typical lack of hematopoietic markers, CD34, CD11B, CD19, CD45, and HLA-DR expression. (e) The comparative chromosome analysis of AD-MSC cultures growing 28 passages in either 5% O₂ or 21% oxygen atmospheres. The cells growing in the physioxic conditions demonstrated proper karyotypes, whereas AD-MSC cultured in the air environment displayed severe karyotyping abnormalities, such as polyploidy of chromosomes (20 in 30 analyzed metaphases) and haploid chromosome numbers (10 in 30 analyzed metaphases); ; ; .