Influence Of Simvastatin On The Proliferation And Secretion Functions Of Nucleus Pulposus-derived Mesenchymal Stem Cells

ObjectiveTo investigate the effect of simvastatin on the proliferation and secretion functions of nucleus pulposus-derived mesenchymal stem cell(NPMSC)in vitro and explore its possible mechanism.MethodsNPMSC was isolated and cultured with collagenase and sequential trypsin digestion from the caudal spinal nucleus pulposus of 8-week-old Sprague-Dawley rats.Cellular morphology was observed and cellular proliferation was detected by MTT assay.The mRNA expression of CD11b,CD45,CD73,CD90 and CD105 was detected using flow cytometry.The stem cell gene Sox2,Oct-4 and Nanog expression of NPMSC was measured and compared with bone marrow derived mesenchymal stem cell.The third generation NPMSC was incubated with various concentrations of simvastatin(Omol/L,0.01?mol/L,O.1?mol/L and 1?mol/L)for 1,3,7,14 and 21 days.The influence of simvastatin on proliferation was assayed with CCK-8 assay,and mRNA expressions of type II collagen,aggrecan,hypoxia-inducible factor-1?(HIF-la),glucose transporter 1(GLUT-1)and vascular endothelial growth factor(VEGF)were detected by semi-quantitative reverse transcription polymerase chain reaction(RT-PCR).ResultsNPMSC isolated from rat nucleus pulposus could form sunflower-like colonies.NPMSC of third passage became homogeneous and exhibited spindle-like morphology.Meanwhile,high expression levels of stem cell-related positive antigen molecules(CD73,CD90,CD 105)and low expression levels of negative antigen molecules(CD45,CD11b)were observed.Moreover,NPMSC expressed comparable stem cell-related genes(Sox2,Oct-4,Nanog)with bone marrow mesenchymal stem cells.Simvastatin at a certain range of concentration(0.01?mol/L-0.1?mol/L)significantly promote proliferation and secretion(HIF-1?,GLUT-1,VEGF)functions(P<0.05).Simvastatin at 0.1?mol/L concentration reached the maximum effects on the proliferation and secretion functions(P<0.05),while simvastatin at 1?mol/L concentration showed descended proliferation and secretion functions(P<0.05).Simvastatin promote the mRNA expression of type II collagen and aggrecan,of which 0.1?mol/L concentration reached the maximum effects.ConclusionSimvastatin at a certain concentration(0.01 pmol/L-01?mol/L)range could improve proliferation capability and promote mRNA expressions of type collagen II and aggrecan of NPMSC,and the HIF-la mediated pathway involved in the mechanism.