Viability and Expression Pattern of Cryopreserved Mesenchymal Stem Cells derived From Buffalo Bone Marrow
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Date
2016
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Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana
Abstract
Cryopreservation of mesenchymal stem cells is necessary, because of their tremendous potential for cellular therapy and regenerative medicine so that they can be used at the time of treatment of clinical cases. The present investigation was carried out to compare the viability and expression pattern of cryopreserved mesenchymal stem cells from fresh MSCs derived from buffalo bone marrow. Bone marrow aspirates were collected from iliac crest of pelvis from buffalo calves immediately after slaughtering of animals. MSCs were separated by density gradient method and cultured in high glucose DMEM supplemented with 15% FBS. Population doubling time and characterization of MSCs was done at 4th passage. Population doubling time for fourth passage cells was 38.63±1.70 hours. Buffalo MSCs showed positive alkaline phosphatase (AP) activity and positive expression of surfacemarkers (CD73 and CD105). MSCs were cryopreserved by slow freezing and fast freezing method for three months. DMSO and Glycerol were used as cryoprotectants in five different combinations for freezing MSCs. Post-cryopreserved MSCs were analyzed for viability, population doubling time and their expression pattern. The results of present study revealed that population doubling time was significantly lower in slow freezing as compared to fast freezing with cryopreservation media II and V. Similarly, viability percentage was significantly higher in slow freezing as compare to fast freezing with cryopreservation media II and V. Cryopreserved MSCs maintained the stemness and expressed CD73 and CD105 markers which was similar to fresh MSCs. On the basis of results of present investigation, it can be concluded that MSCs derived from buffalo bone marrow are better cryopreserved by slow freezing protocol with cryomedia II and V as compare to fast freezing
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