Experimental study of the oriented differentiation of bone marrow derived mesenchymal stem cells into chondrogenic phenotype in a specific culture fluid

Xiaofei DING; Jingmin ZHAO; Weiping CHEN; Zhi YANG; Wei SU

ABSTRACT

BACKGROUND:

To construct tissue engineering cartilage would open up a novel way for the repair of cartilage damage in avoidance of the disadvantages of traditional therapeutic method.

OBJECTIVE:

To probe the techniques for the isolation of mesenchymal stem cells (MSCs) from bone marrow, as well as the in vitro differentiation into chondrocytic phenotype in a specific culture fluid.

DESIGN:

A complete randomized experiment

SETTING:

The Department of Traumatic Orthopedics and Hand Surgery of the First Affiliated Hospital of Guangxi Medical University, and Teaching and Research Faculty of Histology and Embryology of Guangxi Medical University.

METHODS:

The experiment was carried out at Guangxi Medical University between August 2002 and April 2003. Twenty SD neonatal weaning rats were selected. Bone marrow was aspirated from the bones of rat limbs and was isolated by gradient centrifugation in Percoll, and MSCs could be obtained in combination with adherent screening method, which were then cultured in DMEM-LG with 15% fatal bovine serum (FBS) in the incubator of 37℃ with 5% CO2 for 10-14 days. The passage cells were induced in DMEM-HG with 15% FBS (containing TGF-β1 10 μg/L, 10-7 mol/L dexamethasone, 50 mg/L VitC).MAIN OUTCOME

MEASURES:

The morphology, growth, as well as proliferation and specific expression of chondrogenic matrix of in vitro cultured MSCs due to specific induction.

RESULTS:

Totally 20 SD rats were observed and analyzed with no loss SCs grew in visible symmetric colonies, displaying a long-spindle shape,and the morphological characteristics of marrow-derived MSCs had no obvious changes during passage-culture, but its proliferation time was found from a shuttle fibroblastic appearance to polygonal shape, displaying posiHC staining of type Ⅱ collagen of cartilage specific matrix.bronectin adherent screening technique is a convenient, effective and practical method to separate and collect MSCs from rat bone marrows in chondrogenic phenotype when induced by a specific medium and can secrete cartilage specific matrix, and they can be the optimal seed cells for cartilage tissue engineering.