两种壳聚糖载rhBMP-2纳米微球对SD大鼠体内异位成骨的影响

doi:10.13418/j.issn.1001-165x.2016.04.012

中国临床解剖学杂志 ›› 2016, Vol. 34 ›› Issue (4): 412-418.doi: 10.13418/j.issn.1001-165x.2016.04.012

两种壳聚糖载rhBMP-2纳米微球对SD大鼠体内异位成骨的影响

余翔¹，　夏远军²，　郑晓辉³，　章莹²，　陈泽鹏²，　夏虹²，　尹庆水²

1.广州中医药大学第一临床医学院，广州 510405；　2.广州军区广州总医院创伤骨科，广州 510010；
3.广州中医药大学第一附属医院创伤骨科, 广州 510405

收稿日期:2016-04-22 出版日期:2016-07-25 发布日期:2016-08-03
通讯作者: 章莹，主任医师，硕士生导师，E-mail: zhangying-doc@aliyun.com
作者简介:余翔(1990-), 男，云南宣威人，硕士研究生，主要从事骨材料学方面的研究，E-mail:879925091@qq.com
基金资助:
国家自然科学基金青年基金资助项目（51303031）；广东省自然科学基金资助项目（S2012010009743）；广东省应用基础研究计划项目（2013J4100120）

Ectopic osteogenesis evaluation of two kind of chitosan rhBMP-2 loaded microspheres in SD rats

YU Xiang¹, XIA Yuan-jun²，ZHENG Xiao-hui³, ZHANG Ying², CHEN Ze-peng², XIA Hong²,YIN Qing-shui²

1. The First Clinical Medical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, China;　2. Department of Orthopaedics & Traumatology, General Hospital of Guangzhou Military Command, Guangzhou 510010, China; 3.Department of Traumatic Orthopedics, The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, Guangdong 510405, China

Received:2016-04-22 Online:2016-07-25 Published:2016-08-03
Contact: ZHANG Ying, E-mail: zhangying-doc@aliyun.com

摘要/Abstract

摘要：

目的　通过SD大鼠异位成骨实验来探究重组人骨形态发生蛋白-2/壳聚糖/硫酸葡聚糖(rhBMP-2/CS/DS) 复合微球和重组人骨形态发生蛋白-2/壳聚糖 (rhBMP-2/CS) 微球对SD大鼠体内异位成骨的影响。方法　随机将36只SD大鼠平均分为三组（n=12），分别为A组 (rhBMP-2), B组(rhBMP-2/CS), C组(rhBMP-2/CS/DS)。制备股四头肌肌袋模型后，分别将三种材料植入股四头肌肌袋肌间隙中。分别在4，8和12周时大体观察植入区组织硬度，每组处死4只大鼠后取出异位骨块，并切取异位骨化的组织行micro-CT扫描及 Mimics软件三维重建；检测各组织块骨体积分数（bone volume fraction,BVF）、骨小梁厚度（trabecular thichness,Tb.Th）、骨密度（bone mineral density,BMD）；并行组织学观察和ALP活性、钙含量检测。结果 4周时，A、B、C三组植入区周围组织质地均稍硬，三者并无明显区别；8周和12周时，三组植入区硬度明显增加，且C组比A、B组质地更硬。4周时，HE染色可见三组有少量骨组织形成，但不明显；B、C两组BVF、Tb.Th、BMD，碱性磷酸酶（ALP）活性、钙含量均高于A组；B、C两组以上指标差异无统计学意义。8、12周时，HE染色可见到三组骨组织逐渐增多，并逐渐成熟，且B、C两组可见到比A组更成熟的骨组织，C组骨组织比B组更成熟；B、C两组BVF、Tb.Th、BMD，ALP活性、钙含量均高于A组，C组以上指标均高于B组。结论　rhBMP-2/CS/DS纳米缓释微球的成骨效果明显强于rhBMP-2/CS纳米微球和单独rhBMP-2，其可能在骨组织工程领域有较好的运用前景。

关键词: 重组人骨形态发生蛋白-2, 壳聚糖, 硫酸葡聚糖, 异位成骨, Micro-CT, 三维重建

Abstract:

Objective　To explore the ectopic osteogenesis introducing ability of recombinant human bone morphogenetic protein-2/chitosan/dextran sulfate composite microspheres and recombinant human bone morphogenetic protein-2/chitosan composite microspheres on SD rats.　Method　36 SD rats were randomly divided into three groups, in which the quadriceps muscle bag model of the rats were established; the bags were implanted respectively with rhBMP-2(group A), CS/DS microspheres (group B), and rhBMP-2/CS/DS microspheres (group C). Hardness of the implanted regions tissuesat the 4th,8th,12th weekend were observed, and four rats were sacrificed in each group, from which the ectopic bone tissues were harvested. The bone formation situation of plantation areas were generally observed, and the heterotopic ossification organization tissues were scanned by micro-CT, and were three-dimensionally reconstructed by using the Mimics software；parameters such astissue bone volume fraction (BVF), trabecular thickness (Tb.Th), bone mineral density (BMD) were detected; Some heterotopic ossification organization tissues were taken for histological observation and analysis of ALP activity and calcium content.　Results　At the 4th weekend, tissues around implanted regions of each group had slightly harder texture, and there was no significant difference among the three groups; at the 8th and 12th weekend, hardness of tissues around implanted regions of each group increased significantly, and the group C had harder tissues formed than group A and B. At the 4th weekend, HE staining showed that the three groups had a small amount of bone tissue formation, but not obviously; the BVF, Tb.Th, BMD, ALP activity and calcium levels of B, C groups were significantly higher than group A; there were no statistically significance on the above indicators of group B and C. At the 8th and 12th weekend, gradually-increased and marture bone tissues could be seen in the three groups under HE staining. However,there were more mature ectopic bone tissues in group B and C than group A; And the bone tissues of group C were more mature than group B. Meanwhile, the BVF, Tb.Th, BMD, ALP activity and calcium levels of group B and C were significantly higher than group A, and the above-mentioned indicators of group C were higher than that of group B.　Conclusion　rhBMP-2/CS/DS microspheres have stronger ability of inducing ectopic bone than rhBMP-2/CS microspheres and single rhBMP-2, and it mayhave good prospects for applicationin bone tissue engineering.

Key words: Bone morphogenetic proteins-2, Chitosan, Dextran sulfate, Ectopic osteogenesis, Micro-CT, Three-dimensional reconstruction

余翔,夏远军,郑晓辉,章莹,陈泽鹏,夏虹,尹庆水. 两种壳聚糖载rhBMP-2纳米微球对SD大鼠体内异位成骨的影响[J]. 中国临床解剖学杂志, 2016, 34(4): 412-418.

YU Xiang, XIA Yuan-jun，ZHENG Xiao-hui, ZHANG Ying, CHEN Ze-peng, XIA Hong,YIN Qing-shui. Ectopic osteogenesis evaluation of two kind of chitosan rhBMP-2 loaded microspheres in SD rats[J]. Chinese Journal Of Clinical Anatomy, 2016, 34(4): 412-418.

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两种壳聚糖载rhBMP-2纳米微球对SD大鼠体内异位成骨的影响

Ectopic osteogenesis evaluation of two kind of chitosan rhBMP-2 loaded microspheres in SD rats

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