Isolation and Culture of Human Umbilical Cord Vein Mesenchymal Stem Cells and Identification of Their Biological Characteristics

Abstract: To isolate and culture human umbilical cord vein mesenchymal stem cells (MSCs) and identify their biological characteristics， sequential collagenase digestions were adopted to isolate human umbilical cord MSCs. Morphology and cell growth curve were detected in hUVMSC2 cell line. Different induction conditions were used to direct hUVMSC2 cells to differentiate into osteoblasts and adipocytes respectively. Flow cytometry was adopted to analyze expression of surface markers such as CD54， CD105， CD29， CD166， CD44， CD31， CD34， CD49， and CD106， etc. Morphologically hUVMSC2 cells were fibroblast-like， did not express endothelial-specific marker vWF. Under different induction conditions， hUVMSC2 cells could be directed to differentiate into osteoblasts and adipocytes respectively in vitro. Through flow cytometry assay， hUVMSC2 cells express adhesion molecules such as CD54， CD105， CD29， CD166， CD44， but not CD31， CD34， CD49， and CD106 endothelial or hematopoietic markers. During exponential growth period， hUVMSC2 cells doubling time is about 26 h， whereas when mitogen bFGF added， cells grew faster and the doubling time became shorter as16 h. This investigation confirmed that human umbilical cord vein mesenchymal stem cells exist under endothelia layer. And sequential collagenase digestion method could successfully be used to isolate and culture two types of cells as endothelial cells and MSCs simultaneously from a single human umbilical cord vein. These MSCs had bi-directional differentiation potential at least， and also expressed important adhesion molecules such as ICAM-1， β3-integrin， and ALCAM， etc. This study also provided new MSC origin and basic data for stem cell study and clinical MSC application in combination with hematopoietic stem cell transplantation.

CSTR: 32200.14.cjcb.2005.02.0022