骨髓间充质干细胞对大鼠BRONJ治疗的研究

doi:10.13591/j.cnki.kqyx.2022.08.004

摘要/Abstract

摘要： 目的评估骨髓间充质干细胞(bone mesenchymal stem cells,BMSCs)对大鼠双膦酸盐相关颌骨坏死(bisphosphonate-related osteonecrosis of the jaw, BRONJ)的治疗效果。方法 SD大鼠每周给予尾静脉注射唑来膦酸(80 μg/kg),用药2周后,在全麻下拔除两侧上颌第一磨牙,之后继续用药至12周,构建大鼠BRONJ模型。取第3代大鼠外周骨BMSCs与骨粉支架在体外构建细胞支架复合体以备用。将BRONJ大鼠随机分为4组,在全麻下进行局部去骨清创,分别移植入细胞支架、支架,并设立清创组和对照组,最后缝合创口。移植术后4周和8周分批处死,分离并收集骨组织样本。Micro-CT和组织学分析各组骨缺损区新骨形成情况。ELISA法检测信号通路相关因子NF-kB受体活化因子配体(receptor activator of nuclear factor -κB,RANKL)和骨保护素(osteoprotegerin,OPG)的表达。结果 Micro-CT和组织学染色显示,细胞支架组成骨修复效果最佳,支架降解快,新骨生成多,骨矿化密度最高(P<0.05)。支架组成骨修复效果居其次,支架降解较慢,类骨质和矿化骨基质生成较慢,骨矿化密度较低。清创组成骨修复效果最差,凹坑状骨缺损明显,骨面暴露,死骨形成,且缺损范围较清创术后呈现扩大趋势。对照组拔牙创黏膜未愈合或延迟愈合,未愈合的黏膜下方可见暴露坏死骨,结缔组织内有明显的炎症浸润,骨质出现不同程度的硬化,为典型的BRONJ表现。细胞因子检测显示,术后4周和8周,细胞支架组、支架组和清创组RANKL/OPG含量比值依次增高,细胞支架组和支架组较对照组下降(P<0.05),清创组较对照组明显增高(P<0.05)。结论外周骨BMSCs对BRONJ具有一定的治疗效果,促进BRONJ大鼠清创后的骨缺损修复。RANKL/RANK/OPG信号通路可能在BRONJ病程发展中起重要作用。

关键词: 骨髓间充质干细胞, 双膦酸盐相关颌骨坏死, 治疗

Abstract: Objective To evaluate therapeutic effects of BMSCs on BRONJ in rats. Methods SD rats were injected intravenously with zoledronate (80 μg/kg/week) via the tail vein. After two weeks of administration, under general anesthesia, the first molars from bilateral maxillae were extracted. Drugs were continuously injected for 12 weeks to establish a rat BRONJ model. The 3rd generation of isolated BMSCs from peripheral bones were cultivated in the Bio-Oss scaffold for later use. BRONJ rats were randomly divided into four groups, and treated with surgical debridement under general anesthesia. Then, BMSCs/scaffold and scaffold were transplanted into operation sites of BRONJ rats under aseptic conditions, respectively. Rats with or without surgical debridement were used as negative control and blank control. Thereafter, rats were sacrificed at 4 and 8 weeks in batches, and bone samples were collected. New bone formation was observed by Micro-CT scanning and histological analysis. Expressions of signaling pathway related factors including RANKL and OPG were detected using ELISA testing. Results Micro-CT and histological analysis showed that bone repair effect of the BMSCs/scaffold group was optimal, presenting rapid scaffold degradation, active new bone formation and high bone mineral density (P<0.05). Bone repair effect of the scaffold group was the second, presenting slower degradation, less new bone formation and lower bone mineral density compared with the BMSCs/scaffold group. Bone repair effect of the debridement group was the worst, showing pit-like bone defect and exposed dead bone. Even worse, the bone defect was enlarged after surgical debridement. The control group presented major histopathological manifestations of human BRONJ, including non-healed or delay-healed mucosa, exposed necrotic bone, osseous sclerosis and inflammatory infiltration. Cytokines results showed that RANKL/OPG ratio was the lowest in BMSCs/scaffold group and the highest in debridement group at 4 and 8 weeks. The RANKL/OPG ratio of BMSCs/scaffold group and scaffold group significantly decreased than that of control group (P<0.05), but the ratio of debridement group significantly increased compared with the control group (P<0.05). Conclusion BMSCs from peripheral bones have a therapeutic effect on BRONJ, which can promote the repair of bone defect after debridement in BRONJ rats. RANKL/RANK/OPG signaling pathway may play an essential role in the development of BRONJ.

Key words: bone mesenchymal stem cell, bisphosphonate-related jaw osteonecrosis, therapy

中图分类号:

R456.9

龚雪, 钱文昊, 苏俭生. 骨髓间充质干细胞对大鼠BRONJ治疗的研究[J]. 口腔医学, 2022, 42(8): 694-700.

GONG Xue, QIAN Wenhao, SU Jiansheng. Bonemesenchymal stem cell therapy for bisphosphonate-related jaw osteonecrosis in rats[J]. Stomatology, 2022, 42(8): 694-700.

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