DCMTSA对创伤性股骨头坏死大鼠BMP-2和BMP-7表达的影响
发布时间：2018-11-12 14:28
【摘要】：背景骨折愈合是复杂的生物学修复过程,临床上骨折延迟愈合／不愈合的间题仍未解决,传统的中西药物干预治疗时间长,效果不理想。目前,促进骨折愈合最好细胞因子是骨形态发生蛋白。实验研究发现骨形成蛋白具有诱导骨髓基质细胞向成骨细胞分化的作用。海藻酸钠(sodium alginate,SA)是一种从褐藻中提取的天然多糖聚合物,若能使SA部分糖醛酸单元的羟基转变成醛基,即羧甲基海藻酸钠(dicarboxmethyl trisodium alignate,DCMTSA),可改善SA的生物相容性,可作为诱导骨形态发生蛋白的有效药物。目的改善SA的生物学特性,探讨DCMTSA对骨形成蛋白表达的影响,明确DCMTSA对骨髓基质细胞向成骨细胞分化促进作用,提供一种制备方法简单、原料来源广泛的治疗和预防骨折的药物,为应用于临床预防和治疗骨折等疾病提供实验室依据。方法1制备DCMTSA:SA经过氧化氢溶液或高锰酸钾部分氧化,部分糖醛酸单元的羟基转变成醛基,改善SA的生物相容性。2动物模型制备及分组：45只体重(261±12)g,2-3月龄SD大鼠,制成创伤性股骨头坏死模型后,以数字随机法平均分为三组,每组15只。DCMTSA组,大鼠每日以DCMTSA 0.1 g.kg-1.d-1灌胃；SA组,大鼠每日以SA 0.1g.kg-1.d-1灌胃模型组；对照组,大鼠每天0.1g.kg-1.d-1生理盐水灌胃。3实验方法：每组大鼠分别于术后7d、14d、21d各处死5只。取大鼠骨折股骨分为两份：一份经10%甲醛固定后脱钙,HE染色光镜下观察各组骨折处骨痂生长组织结构,免疫组化SP法检测骨形成蛋白一2(bone morphogenetie protein-2,BMP-2)骨形成蛋白-7(bone morphogenetie protein-7,BMP-7)的表达情况；一份脱钙软化后组织匀浆,以RT-PCR检测BMP-2 mRNA和BMP-7 mRNA的表达。结果1 HE染色结果：术后7d、14d、21d,DCMTSA组大鼠骨折断端骨组织的纤维组织增生,骨小梁形成,骨基质形成及骨结构的形成情况明显优于SA组,SA组优于对照组。2免疫组化实验结果：术后7d、14d、21d,三组大鼠BMP-2和BMP-7蛋白表达DCMTSA组高于SA组,SA组高于对照组。3 RT-PCR检测结果：术后21d,BMP-2和BMP-7蛋白mRNA的表达DCMTSA组高于SA组,SA组高于对照组。4股骨骨组织计量学结果：术后21d大鼠平均骨小梁数、平均骨小梁厚度和平均骨小梁面积DCMTSA组大于SA组,SA组大于对照组。且对照组的松骨质和密骨质容积明显降低,骨小梁数目明显减少,骨髓腔明显扩大,骨小梁厚度和骨皮质变薄。结论DCMTSA对骨组织细胞释放BMP-2和BMP-7的促进作用大于SA,促进成骨细胞分化,抑制破骨细胞造成的骨吸收,有利于创伤性股骨头坏死大鼠的骨愈合。
[Abstract]:Background fracture healing is a complex biological repair process, the clinical delayed healing / non-union problem remains unresolved, the traditional Chinese and western medicine intervention treatment time is long, the effect is not ideal. At present, bone morphogenetic protein is the best cytokine to promote fracture healing. It was found that bone morphogenetic protein could induce bone marrow stromal cells to differentiate into osteoblasts. Sodium alginate (sodium alginate,SA) is a kind of natural polysaccharide polymer extracted from brown algae. The biocompatibility of SA can be improved if the hydroxyl group of part of SA uronic acid unit is converted into aldehyde group, that is, carboxymethyl alginate sodium (dicarboxmethyl trisodium alignate,DCMTSA. It can be used as an effective drug to induce bone morphogenetic protein. Objective to improve the biological characteristics of SA, to investigate the effect of DCMTSA on the expression of bone morphogenetic protein, to clarify the effect of DCMTSA on the differentiation of bone marrow stromal cells into osteoblasts, and to provide a simple preparation method. A wide range of raw materials for the treatment and prevention of fracture drugs, for clinical prevention and treatment of fracture and other diseases to provide laboratory basis. Methods (1) DCMTSA:SA was partially oxidized by hydrogen peroxide solution or potassium permanganate, and the hydroxyl groups of some uronic acid units were transformed into aldehydes, which improved the biocompatibility of SA. SD rats aged 2-3 months were made into traumatic femoral head necrosis model and were randomly divided into three groups with 15 rats in each group. The rats in the DCMTSA group were given DCMTSA 0.1 g.kg-1.d-1 daily. SA group, rats were fed with SA 0.1g.kg-1.d-1 daily; Rats in control group were treated with 0.1g.kg-1.d-1 normal saline daily. 3 Experimental methods: 5 rats in each group died at 14 days and 21 days after operation respectively. The femur of rat fracture was divided into two parts: one was decalcified after 10% formaldehyde fixation, the structure of callus growth was observed under light microscope with HE staining, and bone morphogenetic protein-2 (bone morphogenetie protein-2, was detected by immunohistochemical SP method. The expression of bone morphogenetic protein-7 (bone morphogenetie protein-7,BMP-7 (BMP-2); After decalcification and softening, the expression of BMP-2 mRNA and BMP-7 mRNA was detected by RT-PCR. Results 1 the results of HE staining showed that the fibrous tissue proliferation, trabecular formation, bone matrix formation and bone structure formation in the DCMTSA group were significantly better than those in the SA group. SA group was superior to control group. 2 Immunohistochemical results showed that the expression of BMP-2 and BMP-7 protein in DCMTSA group was higher than that in SA group, and that in SA group was higher than that in SA group. 3 the results of RT-PCR detection in SA group were higher than those in control group: 21 days after operation, 21 days after operation, the expression of BMP-2 and BMP-7 protein in three groups was higher than in SA group. The expression of BMP-2 and BMP-7 protein mRNA in DCMTSA group was higher than that in SA group, and that in SA group was higher than that in control group. 4 the average trabecular bone count, mean trabecular thickness and mean trabecular area in DCMTSA group were higher than those in SA group 21 days after operation. The SA group was larger than the control group. In the control group, the volume of cancellous bone and dense bone decreased significantly, the number of trabecular bone decreased, the medullary cavity enlarged, and the thickness of trabecular bone and cortical bone thinned. Conclusion DCMTSA can promote the release of BMP-2 and BMP-7 from bone tissue cells more than SA, can promote osteoblast differentiation and inhibit bone resorption caused by osteoclasts, which is beneficial to bone healing in rats with traumatic osteonecrosis of femoral head.
【学位授予单位】：新乡医学院
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R683
本文编号：2327378
[Abstract]:Background fracture healing is a complex biological repair process, the clinical delayed healing / non-union problem remains unresolved, the traditional Chinese and western medicine intervention treatment time is long, the effect is not ideal. At present, bone morphogenetic protein is the best cytokine to promote fracture healing. It was found that bone morphogenetic protein could induce bone marrow stromal cells to differentiate into osteoblasts. Sodium alginate (sodium alginate,SA) is a kind of natural polysaccharide polymer extracted from brown algae. The biocompatibility of SA can be improved if the hydroxyl group of part of SA uronic acid unit is converted into aldehyde group, that is, carboxymethyl alginate sodium (dicarboxmethyl trisodium alignate,DCMTSA. It can be used as an effective drug to induce bone morphogenetic protein. Objective to improve the biological characteristics of SA, to investigate the effect of DCMTSA on the expression of bone morphogenetic protein, to clarify the effect of DCMTSA on the differentiation of bone marrow stromal cells into osteoblasts, and to provide a simple preparation method. A wide range of raw materials for the treatment and prevention of fracture drugs, for clinical prevention and treatment of fracture and other diseases to provide laboratory basis. Methods (1) DCMTSA:SA was partially oxidized by hydrogen peroxide solution or potassium permanganate, and the hydroxyl groups of some uronic acid units were transformed into aldehydes, which improved the biocompatibility of SA. SD rats aged 2-3 months were made into traumatic femoral head necrosis model and were randomly divided into three groups with 15 rats in each group. The rats in the DCMTSA group were given DCMTSA 0.1 g.kg-1.d-1 daily. SA group, rats were fed with SA 0.1g.kg-1.d-1 daily; Rats in control group were treated with 0.1g.kg-1.d-1 normal saline daily. 3 Experimental methods: 5 rats in each group died at 14 days and 21 days after operation respectively. The femur of rat fracture was divided into two parts: one was decalcified after 10% formaldehyde fixation, the structure of callus growth was observed under light microscope with HE staining, and bone morphogenetic protein-2 (bone morphogenetie protein-2, was detected by immunohistochemical SP method. The expression of bone morphogenetic protein-7 (bone morphogenetie protein-7,BMP-7 (BMP-2); After decalcification and softening, the expression of BMP-2 mRNA and BMP-7 mRNA was detected by RT-PCR. Results 1 the results of HE staining showed that the fibrous tissue proliferation, trabecular formation, bone matrix formation and bone structure formation in the DCMTSA group were significantly better than those in the SA group. SA group was superior to control group. 2 Immunohistochemical results showed that the expression of BMP-2 and BMP-7 protein in DCMTSA group was higher than that in SA group, and that in SA group was higher than that in SA group. 3 the results of RT-PCR detection in SA group were higher than those in control group: 21 days after operation, 21 days after operation, the expression of BMP-2 and BMP-7 protein in three groups was higher than in SA group. The expression of BMP-2 and BMP-7 protein mRNA in DCMTSA group was higher than that in SA group, and that in SA group was higher than that in control group. 4 the average trabecular bone count, mean trabecular thickness and mean trabecular area in DCMTSA group were higher than those in SA group 21 days after operation. The SA group was larger than the control group. In the control group, the volume of cancellous bone and dense bone decreased significantly, the number of trabecular bone decreased, the medullary cavity enlarged, and the thickness of trabecular bone and cortical bone thinned. Conclusion DCMTSA can promote the release of BMP-2 and BMP-7 from bone tissue cells more than SA, can promote osteoblast differentiation and inhibit bone resorption caused by osteoclasts, which is beneficial to bone healing in rats with traumatic osteonecrosis of femoral head.
【学位授予单位】：新乡医学院
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R683
【参考文献】
相关期刊论文 前10条
1 刘泽汉;王立春;张宏颖;公伟勋;荆鹏伟;陈琳;何旭;;动物脂肪干细胞联合双重转化生长因子释放支架修复关节软骨损伤的实验研究[J];中国骨与关节损伤杂志;2012年12期
2 张新宽;赵京生;刘英杰;柴青芬;许佳平;;四法疏经法治疗股骨头缺血性坏死的临床研究[J];光明中医;2014年02期
3 刘国岩;徐琬梨;张世华;;活血补髓汤治疗激素性股骨头坏死的实验研究[J];江西中医药;2014年02期
4 王平,高红艳;辨证分型治疗激素型股骨头坏死117例[J];上海中医药大学学报;2003年01期
5 章恒;曾明珠;冯强;;补肾活血方联合银质针导热治疗早中期股骨头坏死临床研究[J];新中医;2014年02期
6 司艳莉;李冬霞;刘友才;千智斌;;海藻酸多糖衍生物治疗大鼠骨质疏松(英文)[J];中国组织工程研究与临床康复;2011年24期
7 林永城;曾炎辉;张泽玫;;补肾复活汤对中老年股骨头缺血性坏死患者血液流变学的影响[J];中国康复;2007年01期
8 黄自冲;;固附汤治疗股骨头坏死[J];中国民间疗法;2009年07期
9 孙缨;张碧霞;王玲;;硝苯地平-海藻酸钠缓释片的制备和性能研究[J];中南药学;2012年04期
10 顾晓东;李雪;;纳米羟基磷灰石/胶原蛋白复合骨结合血管内皮生长因子/骨形态发生蛋白-7修复骨缺损的实验研究[J];中国医药指南;2013年28期
相关博士学位论文 前1条
1 毕锴;中医骨折分期治疗对骨折愈合bFGF、TGF-β、VEGF、BMP-2基因表达影响的实验研究[D];中国中医科学院;2013年
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