关节腔注射移植自体骨髓间充质干细胞对家兔氟骨症疗效观察

Effects of transplanting autologous bone marrow mesenchymal stem cells via articular cavity on skeletal fluorosis rabbits-

目的观察关节腔注射移植自体骨髓间充质干细胞（BMSCs）对家兔氟骨症治疗的有效性和可行性。方法取家兔30只，雌雄各半，按体质量采用随机数字表法分为对照组（6只）和氟骨症模型组（24只），分别饮用含氟化钠0、300mg／L的自来水。90d后将模型组家兔分为单、多次关节腔注射移植组，自然恢复组，溶剂对照组（每组6只，雌雄各半）。分离、培养、鉴定氟骨症家兔BMSCs后，单、多次关节腔注射移植组分别以单次及多次关节腔注射方法移植回氟中毒家兔体内。60d后，X线观察治疗前后骨骼影像学变化，定量CT（QCT）测定家兔活体股骨骨密度，酶联免疫吸附法（ELISA）测定家兔血清碱性磷酸酶（ALP）及骨钙素（BGP），氟离子选择电极法测定家兔股骨骨氟含量．HE染色观察股骨组织病理学改变并计算骨小梁面积。结果X线显示，与治疗前比较，多次关节腔注射移植后家兔股骨上斑片状高密度影淡化，紊乱骨纹有所恢复。多次关节腔注射移植组家兔骨密度[（536．91±25．51）g／cm3]，血清ALP[（20．06±6．25）U／L]、BGP含量[（1230．01±119．50）μg／L]、骨氟含量[（1442．40土458．54）mg／kg]、骨小梁面积[（27．81±2．90）Tb．Ar]均较自然恢复组明显下降[（635．11±93．42）g／cm3、（43．08±2．82）U／L、（3207．73±788．80）μg／L、（2557．65±173．90）mg／kg、（38．52±2．81）Tb．Ar。P均〈0．05]。HE染色显示，与治疗前比较，多次关节腔注射移植组骨髓腔间隙增大，骨小梁数目减少，同时伴有部分新生成的粗细不均、排列整齐、骨髓间隙较均匀的正常骨小梁。单次关节腔注射移植BMSCs未观察到有治疗效果。结论多次关节腔注射移植自体BMSCs可有效修复氟骨症家兔受损骨组织，改善硬化型氟骨症病理损害。

Objective To observe the efficacy and feasibility of transplanting autologous bone marrow mesenchymal stem cells （BMSCs） via articular cavity on skeletal fluorosis rabbits. Methods A total of 30 rabbits, half male and half female, were divided into control group （n = 6） and experimental group （n = 24） by random number table method. The two groups of rabbits were given drinking water containing NaF 0 or 300 mg/L, respectively. After 90 days, 24 rabbits were divided into single treatment group, multiple treatment group, spontaneous recovery group and treatment solvent control group （6 rabbits in each group, half male and half female）. After isolation, the BMSCs from skeletal fluorosis rabbits were cultured and identified, autologous BMSCs were transplanted into rabbit bodies via articular cavity at once or at three different other times, respectively. After 60 days, femur image was observed through X-ray. Femur bone mineral density was measured with quantitative CT （QCT）. Serum alkaline phosphatase （ALP） and osteocalcin （BGP） were also measured using enzyme linked immunosorbent assay （ELISA）. Bone fluoride content was determined via the fluorine ion-selective electrode method. Histopathology changes of femur were observed through HE staining and the trabecular area was calculated. Results In the multiple treatment group, patchy high-density images of femur were disappeared and abnormal bone texture was recovered compared with that of before transplantation. Bone density [（536.91 ± 25.51） g/cm3], ALP concentration [（20.06 ± 6.25） U/L], BGPconcentration [（1 230.01 ± 119.50） μg/L], bone fluoride content [（1 442.40 ± 458.54） mg/kg] and trabecular area [（27.81 ±2.90）Tb.Ar] of the multiple treatment group were lower than those of spontaneous recovery group [（635.11 ± 93.42） g/cm3, （43.08 ± 2.82） U/L, （3 207.73 ± 788.80） μg/L, （2 557.65 ± 173.90） mg/kg, （38.52 ± 2.81） Tb.Ar], and the differences were statistieaUy significant （P〈 0.05）. HE staining in the multiple treatment group showed that bone marrow cavity was enlarged, and the number of trabeculae was decreased, accompanied by some new generated, neatly arranged normal trabeculae. But similar results were not observed in the single treatment group. Conclusion After multiple transplantation via articular cavity, autologous BMSCs from skeletal fluorosis rabbits could repair the damaged bone tissue and improve the pathological damage of skeletal fluorosis with osteosclerosis.