Influence of acidity microenvironment of degenerative intervertebral disc on nucleus pulpous - derived mesenchymal stem cells and adipose- derived mesenchymal stem cells

Abstract

Objective To compare the viability, proliferation and matrix metabolism of nucleus pulpous-derived mesenchymal stem cells (NPMSCs) compared with adipose-derived mesenchymal stem cells (ADMSCs) under intervertebral disc (IVD)like acidic conditions. Methods In our vivo experiment, ADMSCs and NPMSCs from Sprague Dawley rats were cultured under four different pH levels representing the standard condition (pH 7.4) and the normal, mildly degenerated and severely degenerated IVD (pH 7.1, 6.8 and 6.5, separately). By using annexin-V-FITC/propidium iodide staining and a CCK-8 assay, we examined cell viability and cell proliferation respectively. The expression of aggrecan, collagen-Ⅰ , collagen-Ⅱ, MMP-2, ADAMTS4 and TIMP-3 at mRNA and protein levels were measured by RT-PCR and Western blotting. Results Flow cytometry and CCK-8 assays revealed that the viability and proliferation of ADMSCs and NPMSCs decreased with increasing acidity from pH 7.4 to 6.5. Compared with ADMSCs, NPMSCs had significantly fewer apoptotic and necrotic cells under acidic conditions (P<0.05). Cell proliferation of NPMSCs was significantly greater than that of ADMSCs under IVD-like acidic conditions (P<0.05). RT-PCR and western blotting indicated that, in both cell types with increasing acidity, the expression of aggrecan, collagen-Ⅰ , collagen-Ⅱ and TIMP-3 at mRNA and protein levels decreased, while that of MMP-2 and ADAMTS4 increased. Compared with ADMSCs, NPMSCs expressed significantly higher levels of aggrecan and collagen-Ⅱ, and lower levels of collagen-Ⅰ , MMP-2 and ADAMTS4. Conclusion An acidic environment can significantly restrain the IVD regeneration by ADMSCs or NPMSCs. NPMSCs appeared less sensitive to inhibition by acidic pH, and might be promising candidates for cell-based IVD regeneration. Key words: Intervertebral disc degeneration; Mesenchymal stem cells; Cell proliferation