Abstract
AbstractThe aim of this study was to analyze adipose tissue-derived mesenchymal stem cells (AT-MSCs) using the Pap test as a first screening step to evaluate genetic stability. Human adipose tissue from six healthy female donors was obtained from elective liposuction procedures. The cells were isolated, cultivated at P2/P3, characterized by flow cytometric analysis, and differentiation induced. The AT-MSCs were stained by Papanicolaou staining and analyzed according to the Bethesda classification, and viability-apoptosis relationships were evaluated. The results of the Pap test for Sample I indicated high-grade alterations consistent with genetic instability; for Samples II-V, atypical cells of undetermined significance; and for Sample VI, normal cells. These results demonstrate the potential of using the Pap test as an initial screening step to evaluate the genetic stability of cultured AT-MSCs and also suggest its use for other adherent cells such as embryonic stem cells or induced pluripotent stem cells.
Highlights
For new therapies, it is necessary to ensure the safety of processes and products in an efficient manner
The obtained cells were demonstrated to be MSCs from adipose tissue why the cells were marked with antibodies that are in consensus of adipose origin as well the cells were integrity preservation demonstrates by great viability and small apoptosis fraction; all samples have demonstrated the normal profile of undifferentiated adipose derived stem cells (Fig. 1), the cells were adherents and all samples were capable of differentiation in lipid cells demonstrated by oil Sudan red and bone by alizarin osseous matrix identification, respectively (Fig. 2)
The Pap test for Sample I showed high-grade alterations consistent with genetic instability (Fig. 3,4), Samples II-V contained atypical cells of undetermined significance (Figs. 5,6) and Sample VI comprised normal cells (Fig. 7). These results demonstrate the potential for using the Pap test as the initial step of a screening methodology to ensure the genetic stability of cultured MSCs
Summary
For new therapies, it is necessary to ensure the safety of processes and products in an efficient manner. When stem cell therapy is used for oncohematological diseases with bone marrow-derived cell transplantation, the protocol is well established for using fresh bone marrow cells or mononuclear cells from cryopreserved umbilical cord blood for allogeneic transplantation or cryopreserved mononuclear cells from peripheral blood for autologous procedures. All these procedures are conducted without cell cultivation to avoid modified cells. IPSCs must undergo expansion under exvivo conditions These considerations highlight the need for the in vitro expansion of stem cells prior to their commitment into tissue-specific applications. Before bioreactors can be used, several factors must be considered, including the need to increase the number of cells, the capacity to support high cell densities in relatively small volumes, the ability to scale up the design, and standardization of the bioprocess, including the safety of the products[11]
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