The Effect Of Adenovirus-mediated Huamn Bone Morphogenetic Protein4Gene On The Human Degenerative Ucleus Pulposus Cells

Objective:To research the transfer of adenovirus human bonemorphogenetic protein4(Ad-hBMP-4) to human degenerative lumbarintervertebral disc cells in vitro and analyze its effect on the proteoglycan,collagen type II, and Sox9of intervertebral disc cells.Methods:(1)Degenerative intervertebral disc were isolated and cultured invitro. Cells were stained by HE and the expressing position of collagen typeII was identified in the intervertebral disc cells through the laser confocalmicroscope(2)The first generation of disc cells were grown to70%-80%confluence for the purpose of cells. DMEM/F12medium containing20%fetal bovine serum cultured for3,6days after transfection by Ad-GFP andAd-hBMP-41h,so grouped P1cell fusion rate of70%-80%, the P1cellstransfected byAd-GFP, Ad-hBMP-4for3days, the P1cells transfected byAd-GFP, Ad-hBMP-4for6days,(3)Reverse transcription-polymerase chain reaction for mRNAexpression of proteoglycan, collagen type II, and Sox9was performed. The expression of proteoglycan, collagen type II detected by Western blot.Non-transfected group as controlsResults:(1)HE staining: P1cells were polygonal, triangle, nucleus were dyedblue and round or circular.Part of the cell aggregates fusion and nucleuswas obvious. Cytoplasm was dyed different shades of pink(2)Collagen type Ⅱ mainly express in the nucleus pulposus cellcytoplasm.(3)Through the RT-PCR and Western blot the extracellular matrix inthese cells after infection3，6days compared with controls have increasedsignificantly（p<0.05）.The difference between after3and6days wasstatistically significant （p<0.05）.Conclusion:Ad-hBMP-4could infect nucleus pulposus cells with highefficiency and promote collagen type Ⅱ，proteoglycan and Sox9geneexpression along with the infected days increasing. BMP-4may play animportant role in the repair process during early disc degeneration.