目的 构建人骨形态生发蛋白2(hBMP-2)基因真核表达载体,转染脂肪干细胞(ADSCs)后,将其转入去卵巢骨质疏松性大鼠骨缺损模型体内,观察其成骨、修复骨缺损能力.方法 ①选择雌性未孕Wistar 大鼠40只,行双侧卵巢切除,术后定期检测骨密度、骨形态计量学指标及血、尿生化指标,以监测去卵巢骨质疏松性大鼠模型的成功建立;②通过噬菌斑原位杂交筛选人混合细胞cDNA文库获得人骨形态生发蛋白2基因,与真核表达载体pcDNA3.1-连接,构建重组质粒pcDNA3.1-hBMP-2,利用脂质体LipofectamineTM2000介导hBMP-2基因转染第4代脂肪干细胞,并经G418筛选;③将40只模型兔随机分成4组后,于股骨中下段钻孔造成骨缺损,后分别将hBMP2+ pcDNA3.1、ADSCs+ pcDNA3.1、pcDNA3.1-hBMP-2+ ADSCs注入前3组兔的骨缺损处,最后1组为对照组;④定期抽静脉血检测Ca+、P及碱性磷酸酶(ALP)等生化指标,X线观察成骨情况;8周后处死动物取骨缺损区组织,HE染色观察成骨情况及炎症反应.结果术后实验组的检测指标较对照组有明显不同,pcDNA3.1-hBMP-2+ ADSCs组的血清Ca+、P及ALP水平明显升高,X线检查可看到有明显的成骨、骨缺损得到修复,HE染色可见到大量的成骨细胞生成,较对照组有明显的差异,有统计学意义(P<0.05).结论 hBMP-2基因修饰的脂肪干细胞对去卵巢大鼠骨质疏松性骨缺损有很好的修复作用,为绝经后所致的骨质疏松性骨缺损提示了新的治疗思路.%We built a eukaryolie expression vector wilh hBMP-2 gene, then we transfeeled it into ADSCs. Finally we Iransplanled the cells to the hone defect model of oy arieelomized rats to observe the osleogenesis and the eapaeily of hone defect repair. Methods 1 ) Fort) female wislar rats were undergonernhilaleral ovarieelomy. Bone mineral density , hone hislomorphomelry indexes, and blood and urinernbiochemical indicators were monitored to establish the o arieclomized rat model. 2) Cerminal plaque in situ hjhridizalion was used to screen the hBMP2 gene in the human hjhrid cell cDDA librarj. The genewas connected to the eukarjolic expression lector pcU.NA3. 1. hBMP-2 gene was transfecled into the 4th generation of fat stem cells using liposome Lipofeclamine TM2000, and it was selected ivilh C4I8. 3 ) Fort) rabbits randomly diiided into 4 groups. The distal femurs of all the rabbits were drilled to make bone defects. Then hBMP2 + peDNA3. 1, ADSCs + poD_NA3. 1, or peDNA3. 1 - hBMP - 2 + ADSCs were injected into the bone defects of the rabbits in the first three groups, respeclii el). The last group was control group. 4 ) Serum Ca, P, ALP, and other biochemical indicators were delected regularly. Bone Ill-million was obsened using X-ray. The animals were sacrificed 8 iveeks laler. Bone Ill-million and inflammalion were obsened using 11F staining. Results Indicalors in the experimenlal group ivas significanlK differenl comparing to ihose in ihe conlrol group posloperalively. Ca, P, and ALP leiels in ihe serum of ihe pcDNA3. I - hBMP - 2 + ADSCs group increased significanty. Osleogenesis was seen in the X - ray
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