Chinese Journal of Tissue Engineering Research ›› 2013, Vol. 17 ›› Issue (25): 4629-4636.doi: 10.3969/j.issn.2095-4344.2013.25.010
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Xing Zhi-yuan1, 2, Zhang Ji-bo2, Kong Ling-ju2, Liu Jian-sheng2, Zheng De-yu2
Received:
2013-02-18
Revised:
2013-05-17
Online:
2013-06-18
Published:
2013-06-18
Contact:
Zheng De-yu, M.D., Professor, Master’s supervisor, Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
zdy4673349@163.com
About author:
Xing Zhi-yuan★, Master, Associate chief physician, Department of Orthopedics, Municipal Central Hospital of Panjin, Panjin 124000, Liaoning Province, China; Department of Anatomy, Liaoning Medical University, Jinzhou 121001, Liaoning Province, China
Supported by:
the Educational Bureau of Liaoning Province, No. L2011144
CLC Number:
Xing Zhi-yuan, Zhang Ji-bo, Kong Ling-ju, Liu Jian-sheng, Zheng De-yu. Combination of cryopreserved hydroxyapatite/bone marrow mesenchymal stem cells repairs rabbit radial defects[J]. Chinese Journal of Tissue Engineering Research, 2013, 17(25): 4629-4636.
细胞传代后生长速度较原代培养快,约8 d能达到80%融合。流式细胞仪检测结果显示:培养的第3代兔骨髓间充质干细胞均一表达CD44、CD90及CD105,阳性率分别为94.23%,86.23%及96.24%;而CD34、CD45呈阴性,阳性率分别为1.36%及1.45%,结合他人文献可以认为实验获得的细胞主要为间充质干细胞。 2.3 羟基磷灰石复合成骨诱导后骨髓间充质干细胞构成的复合植骨材料 见图2。成骨诱导的骨髓间充质干细胞与多孔羟基磷灰石共培养7 d后,扫描电镜可见骨髓间充质干细胞在多孔羟基磷灰石表面伸出了突起,相互接触,彼此连接成片,有的细胞已经伸入到羟基磷灰石的孔隙内。在部分骨髓间充质干细胞之间有丝状纤维连接,在细胞表面也可见到有大小不等的白色钙质颗粒,见图2B。复合植骨材料冻存复苏后,多孔羟基磷灰石的表面及孔隙内骨髓间充质干细胞状态与冻存前并没有明显区别,细胞也没有从羟基磷灰石表面脱落的趋势,见图2C。"
2.4 复合材料原位成骨结果 术后所有动物全身状况良好,正常进食。切口无红肿及渗出液,部分动物术区周围有一定程度肿胀,虽未进行特殊处理这些肿胀也逐渐自行消退。无动物死亡,所有动物均计入实验统计。 大体观察:术后8周,冻存复合材料组植骨材料两端均有骨痂生长,断端仍然清晰可见,桡骨及支架材料周围软组织未见充血、变性及坏死表现;其中冻存复合材料组和新鲜复合材料组的支架材料均无移位;羟基磷灰石组有部分标本支架材料可轻微活动。术后12周,冻存复合材料组和新鲜复合材料组植骨材料完全被骨样组织包裹,并形成连续性骨痂,塑形较好;羟基磷灰石组支架材料与自体骨间界限模糊,结合处被骨痂包绕,在某些植骨材料的表面多为纤维样组织包裹。 X射线观察:术后8周,冻存复合材料组和新鲜复合材料组复合植骨材料与自体骨两端交错,可见少量骨痂,断端可见;羟基磷灰石组几乎未见骨痂,断端清晰,见图3。术后12周,冻存复合材料组和新鲜复合材料组可见支架材料与自体骨融合,材料的密度与正常骨相似,有连续性骨痂通过断端,皮质骨形成,髓腔通畅,塑形良好;羟基磷灰石组断端变模糊,骨痂量少,塑形欠佳,仍可见部分支架,见图4。"
组织学观察:术后所有时间点在材料周围均未发现明显的炎性细胞浸润情况。术后8周,冻存复合材料组和新鲜复合材料组复合植骨材料开始降解,在材料边缘有大量类骨质形成,材料内部可见新生的骨岛形成,偶可见小血管,新骨形成增多;而羟基磷灰石组的复合材料内很少见到骨岛形成,仅在材料的边缘形成一薄层的骨组织,大部分为纤维结缔组织,见图5。术后12周,冻存复合材料组和新鲜复合材料组支架材料与自体骨之间可见大量新生骨组织,有规则骨小梁通过,局部可见有少量皮质骨形成,内部可见小血管;羟基磷灰石组支架材料与自体骨间也见新生骨组织,但较少且骨质薄,中间的空隙多数为纤维结缔组织相连,近骨断端侧有骨小梁形成,但尚没有与羟基磷灰石表面组织形成有效连接、贯通,见图6。"
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