Résumé
BACKGROUND: Intra-articular injection of cells targeting to change the microenvironment in lesions can act on early osteoarthritis of inflammatory chondrocytes. Implanted cells affect the progress of the disease by the cell characteristics. OBJECTIVE: To explore the synergistic effect of mesenchymal stem cells from human knee adipose (ADMSCs) and synovial tissues (SDMSCs) to inhibit the degeneration of inflammatory chondrocytes. METHODS: ADMSCs, SDMSCs and inflammatory chondrocytes were primary cultured. Under in vitro two-dimensional culture conditions, cell proliferation assay (MTS) was performed to detect the proliferation of three kinds of cells. Differences in chondrogenic markers at mRNA and protein levels between three kinds of adherent cells were detected by quantitative PCR and immunofluorescence. Under in vitro three-dimensional mixed culture conditions, three groups were set up: (1) ADMSCs+inflammatory chondrocytes (A+C group), (2) SDMSCs+inflammatory chondrocytes (S+C group), and (3) ADMSCs-SDMSCs+inflammatory chondrocytes (A+S+C group). Alcian blue staining, safranin O staining and type Ⅱ collagen immunohistochemistry staining were performed on the mixed-cultured cell mass paraffin sections followed by quantitative analysis. Chondrogenic differentiation in each group was detected by quantitative PCR. Culture supernatants were collected to detect the secretion of pro-inflammatory and anti-inflammatory factors by enzyme-linked immunosorbent assay. RESULTS AND CONCLUSION: Under the two-dimensional culture, the proliferative rate of ADMSCs was significantly higher than that of inflammatory chondrocytes and SDMSCs (P < 0.05). The expression of type Ⅱ collagen mRNA and protein and proteoglycan protein in inflammatory chondrocytes was significantly higher than that in the other two kinds of cells (P < 0.01). Under the three-dimensional culture, the percentage of chondrogenic area per total area was significantly higher in the A+S+C group than the S+C and A+C groups (P < 0.05). The expression of type Ⅱ collagen and proteoglycan was significantly higher in the A+S+C group than the S+C and A+C groups (P < 0.05). Compared with the other two groups, the S+C group showed higher levels of interleukin 1, interleukin 6, and tumor necrosis factor α, but lower level of interleukin 10 (P < 0.05). To conclude, the combined use of ADMSCs and SDMSCs synergistically inhibits the degeneration of inflammatory chondrocytes.
Résumé
BACKGROUND: Muscle derived cells in the skeletal muscle have the ability to differentiate into chondrocytes under the induction of bone morphogenic protein 4 and transforming growth factor β3. However, due to the complexity of the skeletal muscle (consisting of muscle fiber, fascia, blood vessel and nerves), the origin of cells has not been identified. OBJECTIVE: To observe the potential for chondrogenic differentiation of fascia-derived cells (FDCs) in order to clarify the chondrogenic ability of the cells derived from the skeletal muscle. METHODS: The fascia was isolated from the rat gluteus maximus, and was then cultured in chondrogenic medium containing bone morphogenic protein 4 and transforming growth factor β3. Samples were harvested on day 14. The chondrogenic differentiation of FDCs was identified by alcian blue, hematoxylin-eosin and safranin O stainings. Cell markers were investigated by immunostaining using the antibodies for desmin, CD34, vimentin, vWF and a-SMA. Immunostaining was used to assay the chondrogenic potential of FDCs cultured with L6 rat myoblasts at varying ratios (1:0, 4:1; 1:1, 1:4 and 0:1). RESULTS AND CONCLUSION: FDCs harvested from the skeletal muscle displayed chondrogenic differentiation and formed cartilaginous tissue when cultured for 14 days in chondrogenic medium after alcian blue and safranin O stainings. Cells isolated from the fascia were negative for desmin, CD34, vWF and a-SMA, but over 90% of the cells were positive for vimentin. Mixed FDC and L6 myoblast pellets showed chondrogenic potential decreased with the increasing ratio of L6 myoblasts. From the analysis of cell surface marker of FDCs in the skeletal muscle, immunostaining, and the comparison with L6 rat myoblasts, it shows that FDCs are the seed cells with chondrogenic potential in the skeletal muscle.
Résumé
<p><b>OBJECTIVE</b>To construct a reasonable substitute for the autograft bone in vitro and transplant it back into the rabbit models to induce the spine fusion.</p><p><b>METHODS</b>The bone marrow stem cell from the seven New Zealand rabbits were cultured. Recombinant human bone morphogenetic protein-4 (rhBMP-4) that has been proved to be bioactive was obtained by the way of genetic engineering. Using the vacuum freezing machine to mix a certain quantity of rhBMP-4 into type I collagen to form a new kind of carrier. Animal model of spine facet process fusion was used. Bone marrow stem cells combined with rhBMP-4 and I type collagen were implanted between the facet process to induce the spine fusion. type I collagen and bone marrow stem cell was used in the controlled group.</p><p><b>RESULTS</b>New bone formation was obvious in the test group. The facet joint was fused very well in this side. No bone formation was present on the other side.</p><p><b>CONCLUSIONS</b>The new composite: bone marrow stem cells, rhBMP-4 and type I collagen was an ideal kind of substitute for the autograft bone.</p>