兔骨髓间充质干细胞复合带部分松质骨的小牛皮质骨成骨实验研究

Experimental study of rabbit bone mesenchymal stem cells seeding onto xenogenic calf cortical bone with partial trabecular bone

目的:探讨带部分松质骨的小牛皮质骨材料作为组织工程材料的可能,为该材料的临床应用提供实验依据。方法:分离兔骨髓间充质干细胞(bone mesenchymal stem cells,BMSCs);BMSCs与带部分松质骨的小牛皮质骨复合培养,测定细胞黏附率及细胞毒性,应用电镜观察细胞在骨材料表面生长情况。并饲养3月龄新西兰大白兔60只,随机分为3组(n=20),即BMSCs/异种骨组、单纯异种骨组和自体髂骨组,在髂骨翼分别植入BMSCs成骨诱导后复合的异种骨、单纯异种骨和自体髂骨。术后4,8,12,24周取材并应用RT-PCR检测骨形态诱导蛋白-2(bone morphogenetic protein-2,BMP-2)mRNA。结果:BMSCs对小牛骨黏附率良好,材料毒性评级0-1级。电镜观察细胞均在小牛骨表面生长,并可延入材料孔隙之中,有基质分泌。在各时间点,单纯异种骨组BMP-2mRNA的表达低于BMSCs/异种骨组和自体髂骨组(P<0.05),BMSCs/异种骨组低于自体髂骨组(P<0.05)。结论:复合兔BMSCs的带部分松质骨的小牛皮质骨在新西兰大白兔体内具有良好的成骨能力。

Objective To explore the possibility of xenogenic calf cortical bone with partial trabecular bone as cell carriers in tissue engineering,and to provide direct evidence for clinical application of matrix scaffold.Methods Rabbit bone mesenchymal stem cells（BMSCs） were seperated and cultured on the surface of xenogenic calf cortical bone with partial trabecular bone.Relative growth rate was measured and cytotoxicity was graded.The growth,proliferation and matrix secretion of BMSCs in calf cortical strut were observed by scanning electron microscope.Sixty 3-month-old New Zealand rabbits were divided into 3 groups（20 in each group）： a BMSCs/bone xenograft group,a simple bone xenograft group and an autogenous iliac bone group.Each group was implanted with the BMSCs induced osteogenic composite calf bone,simple bone xenograft and autogenous iliac bone into the ala of ilium.The expressions of bone morphogenetic protein-2（BMP-2） mRNA were detected by reverse transcription polymerase chain reaction in 4,8,12,24 weeks after the operation.Results The toxicity gradation was 0 to 1.BMSCs adhered the surface of calf cortical strut and extended in the cancellous bone,and extracellular matrices were found.The BMP-2 mRNA expression in the simple bone xenograft group was lower than that in the BMSCs/bone xenograft group and the autogenous iliac bone group at all time points（P0.05）,which was lower in the BMSCs/bone xenograft group than that in the autogenous iliac bone group（P0.05）.Conclusion Rabbit BMSCs combined with calf cortical bonescaffold with partial cancellous bone have good osteogenic capacity in the New Zealand rabbits.