骨形态发生蛋白在大段同种异体骨移植中的表达

Expression of bone morphogenetic protein in transplantation with massive bone allograft

目的:通过动物实验来探讨骨形态发生蛋白在大段同种异体骨移植愈合中的表达及其意义。方法:实验于2005-10/2006-01在南京军区总院动物实验中心完成。①24只新西兰大白兔随机分为实验组和对照组各12只。实验组,手术移植用梯度降温深低温保存的同种异体兔股骨半髁植入,以金属螺钉固定;对照组将取下的自体股骨半髁原位植入,用与实验组相同的内固定方法固定。②术后4,8,12周取标本行大体标本观察,苏木精-伊红及骨形态发生蛋白免疫组织化学染色,观察骨形态发生蛋白在大段同种异体骨愈合过程的表达情况。结果:24只兔均进入结果分析。①两组移植骨段大体观察:第12周,实验组与对照组移植骨段已与宿主完全连接成一整体,外周骨痂已塑形。②两组自体骨与异体骨苏木精-伊红染色结果:第12周,实验组自体骨与异体骨间见较多成熟骨小梁,自体骨与异体骨连接处髓腔基本融合,于异体骨外侧新形成皮质骨结构。对照组已形成新皮质结构,有大量成熟的骨小梁结构,髓腔基本融合。③骨形态发生蛋白免疫组织化学染色结果:第4周:实验组皮质骨外周见较强的骨形态发生蛋白表达,阳性染色主要位于不成熟的骨细胞、成骨细胞、软骨细胞的胞浆及其分泌的基质中,表明骨形态发生蛋白通过“旁分泌”和“自分泌”的形式使得间质细胞分化为软骨细胞和骨细胞。对照组骨折断端两侧均有骨形态发生蛋白表达。第8周:异体皮质骨周围的骨痂中可见阳性表达,新形成的管状结构中呈“镶边”样排列的成骨细胞为阳性表达,其周围尚有一些软骨细胞核阳性表达。对照组与实验组相似,新生血管和新骨生成较实验组为多。第12周:实验组及对照组骨形态发生蛋白表达均较少。深低温保存的同种异体骨移植过程中骨形态发生蛋白表达与自体骨相似。骨形态发生蛋白染色在新生骨及其周围类基质表达阳性。结论:①骨形态发生蛋白在同种异体骨移植愈合早期起重要作用,其通过“旁分泌”和“自分泌”的形式使得间质细胞分化为软骨细胞和骨细胞,从而促使新骨形成。②大段同种异体骨移植愈合早期(4周)过程中骨诱导与骨吸收同时存在时,骨形态发生蛋白发生重要作用。

AIM： To explore the expression of bone morphogenetic protein （BMP） in transplantation healing with massive bone allograft and the significance by animal experiment. METHODS： The experiment Area Command from October was performed at the Anima 2005 to January 2006. ①A Experimental Center, General total of 24 New Zealand rabbits Hospital of Nanjing Military were randomly divided into experimental group and control group with 12 in each group. In the experimental group, femoral condyle of allogenic rabbits after gradient cooling and cryopraservation was implanted, fixed with metal screw. In the control group, autoallergic femoral condyle was in situ implanted and fixed the same as that in the experimental group. ②Samples received general sample observation, haematoxylin-eosin （HE） staining and BMP immunohistochemical staining at weeks 4, 8 and 12 after operation so as to observe the expression of BMP in transplantation healing with massive bone allograft. RESULTS： Totally 24 rabbits were involved in the result analysis. ①General observation of allograft of the two groups： At week 12, the allograft in the experimental group and control group had connected with host completely, and peripheral osteotyius had molding. ②Result of HE staining in autogenous bone and allogenic bone of the two groups： At week 12, many mature bone trabecula appeared between autogenous bone and allogenic bone, and medullary cavity connected with autogenous bone and allogenic bone was fused mostly, and formed cortical bone structure at lateral allogenic bone in the experimental group. Neocortex structure had formed and medullary cavity was fused mostly in the control group with many mature bone trabecula. ③Result of BMP immunohistochemical staining： At week 4, strong BMP appeared in peripheral cortical bone, and positive staining mainly located at plasma of immature osteocytes, osteoblasts and cartilage cells and matrixes in the experimental group. It showed that interstitial cell differentiated into cartilage cells and osteocytes by paracrine and autocrine of BMP. BMP appeared at both sides of broken parts in the control group. At week 8, positive expression appeared at callus around allogenic cortical bone. Selvage-like osteoblasts in new-formed channel-shape structure showed positive expression, and around the osteoblasts had some positive cartilage cells. The control group was similar to the experimental group. Mostly angiogenesis and new bone formation appeared in the experimental group. At week 12, BMP rarely appeared in the experimental group and control group. BMP expression of aUogenic bone in cryopreservation was similar to that in the autogenous bone during transplantation. BMP staining was positive in new bone formation and surrounding matrix. CONCLUSION; ①BMP plays an important role in the early allogenic bone transplantation healing. By paracrine and autocrine interstitial cells differentiate into cartilage cells and osteocytes, which can promote new bone formation. ②BMP plays an important role when bone induction and bone resorption exist together in the early （4 weeks） transplantation healing with massive bone allograft.