Paper Title
Senescence Characteristics Emerge Early in Human Peripheral Blood-Derived Mesenchymal Stem Cells in the Culture

Abstract
One of the cell types which is known to be useful for using in cell therapy and regenerative medicine is mesenchymal stem cell (MSC). Since MSCs are exist in almost all body tissues, selecting a suitable source for isolation of MSCs and using for cell therapy has been a great debate in recent years. In this study expression of the senescence characteristics in peripheral blood (PB)-derived and bone marrow (BM)-derived MSCs has been investigated and compared between four different passages of the two cell types. After isolation of MSCs form human PB and BM samples, the cells were characterized by evaluation of their morphological and behavioral characteristics, evaluation of the expression of surface CD markers and differentiation test. At about 90% confluency, the cells were subcultured by teypsinization with a subcultivation ratio of 1:5. Next, passage 1 (P1), P3, P5 and P6 of the two cell types were evaluated in terms of their morphology and behavior, growth rate and expression levels of two proliferation and pluripotency markers (hTERT, Oct4), an adhesion and migration marker (Itgb1) and also an apoptosis marker (P53) by real time RT-PCR. Our results showed that all tested cell passages of BM-MSCs retained their normal morphology and growth rate. For P1, P3 and P5 PB-MSCs morphology and proliferation rate was normal, but in P6 PB-MSCs both morphology and growth rate were changed. Elongated and spindle-shaped MSCs changed to enlarged and flattened elongated, oval and round-shaped cells. Furthermore, molecular analysis showed that all of the tested cells did not express hTERT, while all of them expressed Oct4 and Itgb1 at almost high levels and their level of expression were not different between different cell passages of the two cell types. The interesting finding was the induction of P53 expression in P6 PB-MSCs, which together with changes in morphology and growth rate confirmed the emerging of senescence in these cells. From the results is could be concluded that PB-MSCs showed senescence signs early in the culture and loss their proliferation capacity. This can cause problem in using PB-MSCs in cell therapy protocols. More detailed studies should be performed to find a way to solve this problem and help to establish standard cell therapy systems using PB-MSCs. Keywords - Human, Bone Marrow Mesenchymal Stem Cells, Peripheral Blood Mesenchymal Stem Cells, In Vitro, Senescence, Early