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| HU Jinquan,YUAN Wen,CAO Peng.The effect of rat bone marrow mesenchymal stem cells on IL-1β induced degenerative nucleus pulposus cells under the condition of co-culture[J].Chinese Journal of Spine and Spinal Cord,2015,(4):361-366. |
| The effect of rat bone marrow mesenchymal stem cells on IL-1β induced degenerative nucleus pulposus cells under the condition of co-culture |
| Received:November 28, 2014 Revised:March 12, 2015 |
| English Keywords:Mesenchymal stem cells Disc degeneration Nucleus pulposus cell Apoptosis Co-culture system |
| Fund:上海市科委国际合作项目（编号：13430721000） |
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| English Abstract: |
| 【abstract】 Objectives: To investigate the effect of rat bone marrow mesenchymal stem cells(BMSCs) on IL-1β induced degenerative nucleus pulposus cells(NPCs) under the condition of co-culture. Methods: NPCs and BMSCs were isolated from SD rat and cultured to the 3rd passage respectively. BMSCs were taken for flow cytometry to analyze the phenotypes of CD29, CD31, CD45, CD90. Experiments were divided into three groups by using the 3rd passage NPCs. Group A, without IL-1β intervention and without BMSCs co-culture were regarded as control group; group B, IL-1β induced NP cells but without BMSCs co-culture; group C, IL-1β induced NP cells and then co-cultured with BMSCs by indirect co-culture transwell. Both IL-1β intervention time and BMSCs co-culture time were set at 24h, then the transwell was removed. The gene(ADAMTS-4, ADAMTS-5, MMP-13) expressions of NPCs through real-time fluorescent quantitative PCR(RT-PCR) were detected in each group, at the same time the Annexin V-FITC/propidium iodide(PI) apoptosis kit and Caspase 3 kits were used to observe the apoptosis in each group. Results: BMSCs flow results showed that more than 90% of BMSCs presented as CD29, CD90 positive, less than 5% of BMSCs presented as CD31 and CD45 positive, which showed good uniform. ADAMTS-4, ADAMTS-5, MMP-13 gene relative expressions in each group were obtained by RT-PCR, 0.98±0.19, 1.10±0.08, 1.21±0.24 respectively in group A, 5.23±0.25, 6.92±2.33, 23.39±0.09 respectively in group B, 2.31±0.26, 3.33±1.52, 12.68±0.11 respectively in group C. Compared with group A, the NP cell in group B and group C showed significant increase of ADAMTS-4, ADAMTS-5, MMP-13 gene expression(P<0.05), and compared with group B, the rise of ADAMTS-4, ADAMTS-5, MMP-13 gene expression showed obvious inhibition in group C(P<0.05). In detection of apoptosis, the relative expression of Caspase 3 activity was 1.20±0.18 in group A, 5.92±0.93 in group B, 2.33±0.52 in group C. Compared with group A, the relative expression quantity of Caspase 3 in group B and group C showed significant increase(P<0.05), and compared with group B, the rise of Caspase 3 activity was suppressed obviously in group C(P<0.05). Annexin V-FITC/propidium iodide(PI) by flow cytometry indicated the increase of apoptotic cells in group B and group C compared with that in group A(P<0.05). Conculsion: BMSCs effectively decrease the degeneration and apoptosis of NP cells induced by IL-1β under the condition of co-culture. As seed cells, BMSCs have therapeutical effect on intervertebral disc inflammation. |
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