摘要
目的:研究中药接骨方在骨折愈合过程中对骨粘连蛋白信使核糖核酸表达的影响,探讨中药接骨方促进骨折愈合的作用机制。方法:将40只2月龄清洁级雌性SD大鼠制成右股骨中段骨折模型,并行髓内固定。造模成功后将40只大鼠分为治疗组和对照组,每组20只。自造模后第1天开始,治疗组以接骨方药液按10.4 mL.kg-1体质量灌胃,对照组以等体积生理盐水灌胃。造模后第14天和第28天分别从2组各取10只大鼠处死,取出骨痂组织。将每个标本的骨痂组织按需要分成2份,1份迅速冰冻切片进行原位杂交处理,测定骨粘连蛋白信使核糖核酸表达情况;另1份制成石蜡切片,染色后观察切片中成骨细胞、破骨细胞、成软骨细胞的情况。结果:①细胞学观察。造模后第14天,治疗组和对照组均可见较多骨痂形成。治疗组骨痂组织中有大量新生骨小梁和软骨组织,并连接成片,其间成骨细胞、成软骨细胞较为活跃,有少量破骨细胞;对照组纤维组织增生活跃,有大量成骨细胞形成,并见有一定的软骨组织。治疗组成骨细胞及破骨细胞数量多于对照组(t=6.520,P=0.000;t=3.670,P=0.002),但成软骨细胞数量少于对照组(t=12.490,P=0.000)。造模后第28天,治疗组骨痂组织中骨小梁数量较多,粗细均匀,排列方向一致,相互连接,折光性强。可见由成骨细胞转化而成并被钙化骨基质包埋的骨细胞,细胞形态正常,有骨小管与基质相连。大量排列较为整齐的成骨细胞和骨细胞出现,破骨细胞数量较少。对照组骨痂中骨小梁排列紊乱,折光强弱不一,并可见小吸收腔,骨折处可见较多纤维细胞、成软骨细胞、成骨细胞和破骨细胞。治疗组成骨细胞、成软骨细胞及破骨细胞数量均少于对照组(t=6.640,P=0.000;t=9.940,P=0.000;t=8.330,P=0.000)。②骨粘连蛋白信使核糖核酸表达情况。造模后第14天,治疗组大鼠骨痂骨粘连蛋白信使核糖核酸的表达水平高于对照组(t=6.700,P=0.000);造模后第28天,2组大鼠骨痂组织骨粘连蛋白信使核糖核酸表达水平比较,差异无统计学意义(t=1.830,P=0.084)。治疗组造模后第28天骨粘连蛋白信使核糖核酸的表达水平低于造模后第14天(t=4.330,P=0.000);对照组造模后第28天骨粘连蛋白信使核糖核酸的表达水平高于造模后第14天(t=4.770,P=0.000)。结论:中药接骨方通过提高骨粘连蛋白信使核糖核酸的表达水平,使软骨修复提早进入骨化及塑形期,从而加速骨折愈合。
Objective ：To study on the etffect of traditional Chinese medicine（ TCM ） JIEGU DECOCTION on the expression of osteonectin（ON） mRNA during fracture healing, and to explore the mechanism of action of TCM JIEGU DECOCTION in promoting fracture healing. Methods ：Forty clean femal SD rats of 2 months old were built models of right mid-femur fi＇actnres with intramedullar7 fixation. After sucessful molding,40 rats were divided into treatment group and control group,20 cases in each group. Since the first day from molding, rats in the treatment group were intragastric administrated with JIEGU DECOCTION （ 10.4 mL · kg ^-1 ） ,while the others in the control group were intragastric administrated with equal volume of normal saline. On the 14th day and 28th day after molding, 10 rats selected from each group were executed,and their bony callus tissues were fetched out. Bony callus tissue of each specimen was divided into 2 parts. Ch,e part was made into frozen section and processed with in-situ hybridization, then ON mRNA expression was evaluated, while another part was made in- to paraffin section,and the morphology and amount of osteoblast, osteoclast and chondroblast in the paraffin sections were observed after staining. Results：On the 14th day after molding,massive bony callus tissues were found in treatment group and control group. In the bony callus tissue of treatment group,there were a large amount of new bone trabeeula and carlilaginous tissue that connected into pieces,among which the osteoblast and chondroblast were active relatively while only a tew number of osteoclasts were found. In the control group,lhe proliferation of fibrous tissue was active, and massive osteoblasts and a certain number of cartilaginous tissues were found. The amonnt of osteoblast and osteoclast in the treatment group was larger than that of control group （ t = 6. 520, P = 0. 000 ; t = 3. 670, P = 0. 002 ）, but the amount of chondroblast in the treatment group was smaller than that of control group（ t = 12. 490,P = O. 000）. On the 28th day after molding, a large amount of of bone trabeculas were found in the bony callus tissue in treatment group, and they were uniform in thickness and arranged in consistent direction and interconnected with strong light refraction. It was showen that osteoeytes transformed from osteoblasts were embedded by calcified bone matrix,the shape of cells were normal,and there were connections between bone eanalicules and matrix. Massive osteoblasts and osteocytes appeared and arranged in regular order,while a few amount of osteoclasts were shown. [n the control group, bone trabeculas in the bony callus were in a disordered arrangement with uniform strength in refraction, and small absorption cavity was fi）und. Massive fibroblasts,chondroblast, osteoblast and osteoclast were found in the broken ends of fractured bone. The amount of osteo- blast,chondroblast and osteoclast in the treatment group were all smaller than those of control group（ t = 6. 640,P = 0. 000 ;t = 9. 940, P = 0. 000 ;t = 8. 330,P = 0. 000）. On the 14th day after molding, ON mRNA expression level in bony callus tissue of rats in the treatment group was higher than that of control group（ t = 6. 700,P = 0. 000）. On the 28th day after molding, there was no statistical ditterence in ON mRNA expression level in bony callus tissue of rats between the 2 groups （t = 1. 830 ,P = 0. 084）. ON mRNA expression level of treatment group on the 28th day after molding was lower than that on the 14th day after molding（ t = 4. 330,P = 0. 000）. ON mRNA expression level in control group on the 28th day after molding was higher than that on the 14th day after molding（ t = 4. 770, P = 0. 000）. Conclusion： JIEGU DECOCTION can raise ON mRNA expression level. As a result, cartilage repair can enter into ossification and remodeling period earlier and the fracture healing is promoted.
出处
《中医正骨》
2012年第9期11-14,共4页
The Journal of Traditional Chinese Orthopedics and Traumatology
关键词
骨折愈合
骨结合素
接骨方
基因表达调控
RNA
信使
动物实验
Fracture healing
Osteonectin
JIEGU DECOCTION
Gene expression regulation
RNA, messenger
Animal experimentation
