Culturing mesenchymal stem cells from canine umbilical cord blood in endothelial basal medium using different methods

Jinben MA; Genfa SHAN

ABSTRACT

BACKGRoUND:

Endothelial basal medium is mainly used to culture endothelial progenitor cells.Studies on mesenchymal stem cells (MSCs) cultured in this medium are few.

OBJECTIVE:

To compare the outcome of MSCs cultured in different mediums including endothelial basal medium. DESiGN.TIME AND

SETTING:

The control eell experiment was performed at the Muniopal Key Laboratory of Xinhua Hospital of Shanghai.China from September 2005 to May 2006. MATERIALSEight pregnant mongrel dogs were selected to obtain umbilical cord blood for isolation and culture of stem cells. Endothelial eell basal medium and endothelial cell medium were bought from Clonetics.USA.Mouse anti-CDlla monoclonal andbody,mouse anti-CDllb monoclonal antibody.mouse anti-CD29 monoclenal antibody and mouse anti-CD7l monoclonal antibody Were purchased from Antibody diagnostica,USA.Mouse anti-CD34 monoclonal antibody was obtained from Lab Vision Corporation.USA.

METHODS:

Umbilical cord blood stem cells were divided into four groups.Umbilical cord blood steTn cells in the group A were incubated in the endothellal basal medium.Umbilical cord blood steHl cells in the group B were incubated in the endothelial basal medium containing microvascular endothelial cells in a 6-well plate.Umbilical cord blood stem cells in the group C were incubated in the endothelial basal medium containing endotheliaI medium in a 6-well plate coated with fibronectin.Umbilical cord blood stelrn cells in the group D were incubated in the endothelial basal medium containing endothelial medium in a 25 cm2 ctdturing flask.MAIN OUTCOME

MEASURES:

Cell morphology and population doublings were observed.CDlla,CDllb,CD34,CD29 and CD71 expression was detected by immunohistochemistry.

RESULTS:

Fibroblast-like cells were measured in each group.The celIs grew badly in morphology and proliferated slowly in the group A,while cells proliferated rapidlyinthe group B.The cell clones were instable and inclined to aging in the group C,with a new cell clone was found.The cells in the group D were similar to those in the group C.Surface antigens detected by immunohistochemistry showed CDlla(-),CDllb(-),CD34(-),CD29(+)and CD7l(+).

CONCLUSION:

MSCs witll well growth and rapid proliferation can be detected in the endothelial basal medium containing endothellal medium in an intact 6-well plate.The outcome is bad in fibronectm-coated plate or 25 cm2 culture flask.