Comparison Of Biological Characteristics Of Nucleus Pulposus Mesenchymal Stem Cells From Normal And Degenerative Nucleus Pulposus In Rats

Objective To compare the biological characteristics of nucleus pulposu mesenchymal stem cells(NPMSC)derived from normal and degenerative intervertebral disc(IVD).Methods NPMSC were isolated from normal and degenerative IVD and cultured in vitro.Flow cytometric analysis was performed to detect the expressions of cell surface markers including CD73,CD90,CD105,CD45 and CD11b/c.The expression of the stemness genes sex determining region Y-box 2(Sox-2),octamer-binding transcription factor 4(Oct-4)and Nanog was determined RT-PCR.NPMSC from normal(N-NPMSC)and degenerative IVD(D-NPMSC)were induced to differentiate toward osteogenic,adipogenic and chondrogenic linages.The biological characteristics of the NPMSC,including the cell colony formation,cell proliferation cell apoptosis,caspase-3 activity,mRNA and protein expressions of HIF-1α,GLUT-1,VEGF,SIRT1 and SIRT6 were compared.Results NPMSC from both groups highly expressed CD105,CD90 and CD73(characteristic cell surface marker of MSC),and lower expressed CD45 and CD11b/c.The original cells from both normal and degenerative IVD could form sunflower-like colonies and the third passage NPMSC became homogeneous and exhibited spindle-like morphology.The cell morphology of two groups showed no significant difference.N-NPMSC and D-NPMSC had a comparable capacity for osteogenic,adipogenic and chondrogenic multipotent differentiation,but N-NPMSC showed a lower expression of stemness gene(P < 0.05).D-NPMSC showed lower clonogenic capacity and proliferative activity than N-NPMSC,while an increased rate of cell apoptosis and expression of caspase-3(P < 0.05).D-NPMSC also showed lowerexpression of HIF-1α,GLUT-1,VEGF,SIRT1 and SIRT6 in the mRNA and protein level than N-NPMSC(P < 0.05).Conclusions NPMSC can be isolated from both normal and degenerative IVD.However,NPMSCs derived from degenerative IVD showed different biological behaviors including lower proliferative activity,lower expression of stemness gene and higher cell apoptosis rate.The HIF-1α mediated pathway may be involved in the mechanism of cell proliferation.