An Experimental Research Into Bone Marrow Mesenchymal Stem Cells And Its' Chondrogenetic Ability Under Growth Factor

Objective To separate bone marrow MSCs and cultivate them in vitro,and demonstrate the role of growth factor-induced in the course of bone marrow mesenchymal stem cell(MSC) differentiation.Material and method All 32 New Zealand white rabbits,male,age from 4 month to 6 month,weight from 2.5 to 3.5 kg.Pick out one of them randomly to aspirate marrow from hibateral ilium,separate bone marrow MSCs and cultivate them in vitro. In experimental group,rhTGF-β₁ and rhIGF-â… were applied into DMEM to induce proliferation and chondrogenic transformation,while in control group only DMEM was used.Contrast the proliferation and chondrogenic transformation between two groups.Allogeneic decalcificated bone matrix(DBM) was prepared by successive defatting and decalcification,observed by scanning electronic microscope.The third passage MSCs induced by rhTGF-β₁ and rhIGF-â… were seeded into DBM to construct tissue engineered cartilage,and then place into rabbits' knee cavity after parceled with fascia.At 4^th w,8^th w,12^th w,culture masses were sectioned and stained for observation.Results Colony forming efficiency(CFE) was 20/10⁶ in experimental group, significantly higher than that of control group,which was 4/10⁶(u=3.326,P＜0.01). Immunohistochemical identification ofâ…¡collagen was positive in experimental groups,while negative in control groups.Allogeneic DBM prepared was a three -dimensional highly-porous structure with good plasticity and suitable mechanical hardness.At 12^th week,culture masses were observated,HE stain show that the chondrocytes have began to proliferate,caryon is blue.Toluidine blue stain show that the chondrocytes aligned,there are plenty of cartilage lacunas,and were surrounded by lots of extracellar matrix.Immunohistochemical identification ofâ…¡collagen was positive,lots of brown-yellow stained particles could be discerned in extracellar matrix.Conclusion Proliferation and expression of chondrocytic phenotype by MSCs can be induced by the synergistic action of rhTGF-β₁ and rhIGF-â… .MSCs combine with DBM can successfully cultivate tissue engineered cartilage.Allogeneic DBM may satisfy the demands of scaffold for tissue-engineered cartilage.