Osteogenic Potential Of Human Intervertebral Disc Cell In Vitro

Background:Most adolescent idiopathic scoliosis patients need surgical therapy eventually.But the Rx is complex and expensive,and necessary fusion materials are not perfect.Spine fusi on may be achieved if the intervertebral disc as an important connect unit can beinduced to be osteogenetic.Intervertebral disc consists of anulus fibrosis(AF)and nucleus pulpous(NP). Both share the common characteristic of fewer cells and more matrix. Homo-cartilage cells exist in NP and the inner layer of AF,fibroblasts exist in the outer layer of AF.Transdifferentiation is defined as cell reverse from one kind of differentiation state to another differentiation state.Differentiation occurs under two situations:one is when the dell includes relevant germ plasm,the other is when the environment is in favor of differentiation reverse.Objective:Observe the cytological characteristics and biological spedfics in vitro.To evaluate the provocative effect of combined recombinant human Bone Morphogenetic Protein-2(rh-BMP₂)and basic Fibroblast Growth Factor(bFGF) on intervertebral disc.Methods:The samples were acquired from young pati ents ranging from 16 to 25 years old,without pathological changes in the intervertebral disc.The samples were delivered to the laboratory in thirty minutes.There AF and NP were separated without other tissues.Human AF call and NP call line system was established by utilizing cell culture technique in vitro.The osteogenic potential and biology changes were observed which were induced by cell factors rh-BMP₂ and bFGF.There are four cell culture groups:GroupsA,B,C and D.GroupA: DMEM/FBS;Group B:DMEM/FBS+rh-BMP₂;Group C:DMEM/ FBS+bFGF;Group D:DMEM/FBS+rh-BMP₂+bFGF.The objects of observation include the changes in cell growth curve and Alkaline Phosphatase (ALP)activity,calcification of the compl exes,secretion of typeâ… ,â…¡collagen and activity of osteocalcin.Results:With the time of cell culture prolonged,the fibroblasts orientated themselves in a radiating pattern,and partial matrix accumulate,rh-BMP_{2 on}with no influence on AF cell proliferation,can enhance ALP activity,the secretion of typeâ… collagen,the expression of osteocalcin,the deposition of calcium salt and formation of calcium nodes,bFGF can considerably stimulate AF cell proliferation and depress ALP activity,while enhancing the secretion of typeâ… collagen,the deposition of calcium salt and formation of calcium nodes. Combined use rh-BMP₂ and bFGF can promote AF cell proliferation with no observable effect on ALP activity.It can notably enhance the secretion of typeâ… collagen,the deposition of calcium salt and formation of calcium nodes.The effect excelled that of using either cell factor alone.Original NP cell isin the shape of a polygon,second generation NP cell is in the shape of a star or a shuttle.Along with the prolonged time of cell culture,the fibroblasts orientated themselves in a radiating pattern,rh-BMP₂ can restrain NP cell proliferation,enhance the expression of osteocalcin,promote the deposition of catcium salt and form calcium nodes,bFGF can enhance the secretion of typeâ… collagen,the deposition of calcium salt and formation of calcium nodes. Combined use of rh-BMP₂ and bFGF can promote AF cell proliferation, enhance the secretion of typeâ… collagen:improve the expression of osteocalcin and the deposition of calcium salt,and form calcium nodes,thus achieving the best effect of osteogenic inducement.The expression of typeâ… collagen was negativein all the four groupsConclusion:rh-BMP₂ can noticeably provocate the osteogenic potential of AF cell in vitro,and bFGF can enhance this effect.rh-BMP₂ can provocate the osteogenic potential of NP cell in vitro,and bFGF can enhance this effect.