Title: Viability and expression pattern of cryopreserved mesenchymal stem cells derived from buffalo bone marrow

Authors: Pooja Devi, Manjinder Sharma, DV Singh, MK Lonare and Rahul Udehiya

Source: Ruminant Science (2017)-6(1):7-12.

Cite this reference as: Devi Pooja, Sharma Manjinder, Singh DV, Lonare MK and Udehiya Rahul (2017).Viability and expression pattern of cryopreserved mesenchymal stem cells derived from buffalo bone marrow. Ruminant Science 6(1):7-12.

Abstract

The present investigation was carried out to compare the viability and expression pattern of cryopreserved mesenchymal stem cells (MSCs) with fresh MSCs derived from buffalo bone marrow. Bone marrow aspirates were collected from iliac crest of pelvis of 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 passaged fresh MSCs was 38.63±1.70 hrs. Buffalo MSCs showed positive alkaline phosphatase (AP) activity and positive expression of surface markers (CD73 and CD105) before cryopreservation. MSCs were cryopreserved by slow freezing and fast freezing method for three months. Me2SO and Glycerol were used as cryoprotectants in five different combinations viz. medium I (10% Me2SO), II (7% Me2SO and 3% Glycerol), III (5% Me2SO and 5% Glycerol), IV (3% Me2SO and 7% Glycerol) and V (Commercial serum free medium).for freezing MSCs. Post-cryopreserved MSCs were analyzed for viability, population doubling time and their expression pattern. The results of present study revealed significantly lower population doubling time and higher viability percentage after slow freezing as compared to fast freezing with cryopreservation media II and V. Cryopreserved MSCs maintained the expression pattern of CD73 and CD105 markers similar to fresh MSCs concluding the study that MSCs can be better cryopreserved with cryomedia II and V by slow freezing protocol.

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