[Differentiation of intervertebral nucleus pulposus-like cells from hBMSCs]

To compare the molecular phenotype of human intervertebral disc cells and articular chondrocytes and to analyze whether hBMSCs can differentiate into both chondrocytes and nucleus pulposus cells after combined induction of TGF-beta3 and BMP-7 in vitro.

The cells with the characteristics of hBMSCs were isolated from marrow aspirates of the volunteer donors' iliac crest. Human bone marrow was removed and fractionated, and adherent cell cultures were established. The 4th passage cells were then translated into an aggregate culture system in a serum-free medium. The pellet cultures of hBMSCs were divided into four groups: 10 ng/mL TGF-beta3 group (group A), 200 ng/mL BMP-7 group (group B), combination group of TGF-beta3 and BMP-7 (group C) and blank group as the control (group D). Histological observation, RT-PCR and RQ-PCR were applied to measure the expressions of collagen type I, II, X, aggrecan and SOX9 on the 4th and 21st day after cell induction, respectively.

As was shown by histological observation, the induced cells expressed the feature of chondrocytes in morphology and ECM in groups A and C on the 21st day after the culture. And the collagen type II was positive after staining in groups A and C. The cell morphology of the induced cells in groups B and C had no obviously changed. PCR detection showed that the expressions of SOX9, aggrecan, collagen type I, II in groups A and C at 21st day were more increased than those at 4th day (P < 0.05). The only expressions of collagen type I in groups B and D at 21st day were more increased than those at 4th day (P < 0.05). The expressions of collagen type X only was positive in group A.

Combination of TGF-beta3 and BMP-7 can make the differentiated cells from hBMSCs much closer to intervertebral disc cells, so it perhaps could provide seed cells for intervertebral disc tissue engineering.