目的 观察谷红注射液(guhong injection,GHI)对大鼠胫骨骨折愈合的促进作用,并探讨其作用的相关机制。方法 雄性SD大鼠180只随机分成6组,每组30只:假手术组、模型组、阳性药组(复方骨肽注射液,5 mL·kg-1)和GHI低、中、高剂量组(2.5、5、10 mL·kg-1)。除假手术组外,其余各组建立胫骨骨折大鼠模型。造模后各组行每日1次腹腔注射给药并于1、2、4和6周末取样,心脏取血行血液生化分析和ELISA试剂盒检测;完整取出胫骨行X线摄片、骨生物力学检测、免疫组化法和RT-PCR法。结果 ①给药1、2和4周末,GHI中、高剂量组的血清钙(Ca)、磷(P)含量高于模型组(P<0.05)。②X线摄片显示,GHI各剂量组的外骨痂增长和骨折线消失较模型组快。③与模型组相比,GHI中、高剂量组的最大抗折力、刚性系数均上升(P<0.05)且受力折线图趋势改善明显。④与模型组相比,GHI中、高剂量组的碱性磷酸酶(ALP)含量较高(P<0.05);GHI各剂量组血清中血小板衍生因子(PDGF)有不同程度的上升(P<0.05或P<0.01);给药1、2和4周末,GHI各剂量组血清中骨形态发生蛋白(BMP)-2上升(P<0.05或P<0.01)。⑤与模型组相比,GHI中、高剂量组大鼠骨痂中Runx2 mRNA表达上升且Smad5蛋白表达上升(P<0.05或P<0.01)。结论 GHI能明显改善骨折大鼠骨骼的生物力学性能,其促进骨折愈合的作用可能是通过上调PDGF、BMP-2在骨愈合不同阶段的表达,调控BMP/Smad5/Runx2信号通路可能是其促骨折愈合的机制之一。
Abstract
OBJECTIVE To observe the effect of guhong injection(GHI) on tibial fracture healing in rats and to explore the mechanism of the action of GHI. METHODS One hundred and eighty male SD rats were randomly divided into 6 groups with 30 rats in each group: sham operation group, model group, positive drug group(compound ossotide injection, 5 mL·kg-1), low, medium and high dose of GHI groups(2.5, 5, 10 mL·kg-1). In addition to the sham operation group, the other groups established the rat model of tibial fracture. All were given once daily intraperitoneal injections and samples were taken at 1st, 2nd, 4th and 6th week. Blood biochemical analysis and Elisa kit detection were performed on blood samples. X-rays, biomechanical tests, immunohistochemistry and RT-PCR were performed on the tibial samples. RESULTS ①After administration for one, two and four weeks, the levels of serum calcium(Ca) and phosphorus(P) in medium and high dose of GHI groups were higher than those in model group(P<0.05). ②X-ray showed that the outer callus growth and the disappearance of fracture line in all dose groups of GHI were faster than those in model group. ③Compared with model group, the maximum load and rigidity of medium and high dose of GHI groups were increased at each time point(P<0.05), and the trend of stress line graph were improved obviously. ④The content of alkaline phosphatase(ALP) in medium and high dose of GHI groups were higher than that in model group at each time point(P<0.05). Compared with model group, the serum levels of PDGF were increased in all dose groups of GHI(P<0.05 or P<0.01). After administration for one, two and four weeks, the serum BMP-2 in all dose groups of GHI were higher than those in model group(P<0.05 or P<0.01). ⑤Compared with model group, the expression of Runx2 mRNA were increased in medium and high dose of GHI groups, as well as Smad5 protein expression(P<0.05 or P<0.01). CONCLUSION GHI could significantly improve the biomechanical properties of bone in fracture rats. The promotion of fracture healing might be through the upregulation of PDGF and BMP-2 expression in different stages of bone healing, and the regulation of BMP/Smad5/ Runx2 signaling may be one of the mechanisms of promoting fracture healing.
关键词
谷红注射液 /
生物力学 /
血小板衍生因子 /
BMP/Smad5/Runx2
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Key words
guhong injection /
biomechanics /
PDGF /
BMP/Smad5/Runx2
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中图分类号：
R965
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参考文献
[1] HUANG J J, QI K Z. Traditional Chinese medicine treatment of fracture experimental research progress . Hunan J Tradit Chin Med(湖南中医杂志), 2010, 26(2): 127-128.
[2] O'CONNOR J P, MANIGRASSO M B, KIM B D, et al. Fracture healing and lipid mediators. Bonekey Reports, 2014, 3:517
[3] SCHINDELER A, MCDONALD M M, BOKKO P, et al. Bone remodeling during fracture repair: the cellular picture. Seminars Cell Develop Biol, 2008, 19(5):459-466.
[4] DAI L L, WAN H T, ZHOU H F, et al. Protective effect of Guhong Injection on rat cerebral microvascular endothelial cells injured by oxygen-glucose deprivation. Chin Pharm J(中国药学杂志), 2014, 49(21): 1897-1902.
[5] ARNAUD A, RAMIREZ M, BAXTER J H, et al. Absorption of enterally administered N-acetyl-L-glutamine versus glutamine in pigs. Clin Nutr, 2004, 23(6):1303-1312.
[6] YIN D M, GU J L, SHA B X. Effect of carthamus on the change of nitric oxide content in neonatal rats brain by perinatal asphyxia. Acta Acad Med Nantong(南通医学院学报), 2004, 24(1):21-22.
[7] MA C, GOU S H, HE F, et al. Expressions of type Ⅰ and type Ⅱ collagen detected by Western blot analysis during tibial fracture healing in denervate rats. J Chin Rehabil Tis Eng Res(中国组织工程研究), 2007, 12(10):9978-9981.
[8] TIAN F M, ZHANG L, LUO Y, et al. Effect of simvastatin on fracture healing in osteoporotic rats. Chin Pharm J(中国药学杂志), 2012, 47(21): 1719-1723.
[9] QING Y W U, XIONG X T, BI X U, et al. Effects of prednisone on trabecular microstructure and biomechanical properties of femur in a rat model of type II collagen-induced arthritis . Chin Pharmcol Bull(中国药理学通报), 2014, 30(7):1018-1022.
[10] CAO X S, ZHANG L N, WU X Y, et al. Intermittent+45°head-up-tilt improves mechanical parameters of hindlimb unweightling in rats. J Fourth Mil Med Univ(第四军医大学学报), 2000, 21(6): 655-657.
[11] SUN S L, YE X L, ZHANG H J, et al. Research on the fracture repair of rats with bone fragments and remediation. J Southwest Univ(Nat Sci Ed)(西南大学学报:自然科学版), 2011, 33(6):94-98.
[12] SHI L. Chinese medicine to accelerate the mechanism of fracture healing. J Changchun Univ Tradit Chin Med(长春中医药大学学报), 2010, 26(4): 600-602.
[13] CHEN Z Y. Combined detection of plasma alkaline phosphatase and fibrinogen in patients with fractures. Int J Lab Med(国际检验医学杂志), 2012, 33(23): 2922-2923.
[14] CUI Q. Fractures in patients with plasma fibrinogen and alkaline phosphatase combined detection of clinical significance . Orthop J China(中国矫形外科杂志), 2012, 20(6): 489-489.
[15] MARCOPOULOU C E, VAVOURAKI H N, DEREKA X E, et al. Proliferative effect of growth factors TGF-beta1, PDGF and rhBMP-2 on human gingival fibroblasts and periodontal ligament cells. J Int Acad Periodontol, 2003, 5(3):63-70.
[16] XU Y Q, JIANG X H, WU Y. Expression of platelet-derived growth factor in fracture healing process of rats with femoral fracture. Chin J Bone Tumor Bone Dis(中国骨与关节杂志), 2011, 10(1): 46-49.
[17] CHEN X H, LI S C, SUN P, et al. Downhill running up regulates the BMP-2, Smad1/5 and Runx2 expressions in the bone of growing ovariectomized mice . Chin J Sports Med(中国运动医学杂志), 2013, 32(7): 609-614.
[18] RONG L, LIU J, QI Y, et al. GATA-6 promotes cell survival by up-regulating BMP-2 expression during embryonic stem cell differentiation. Mol Biol Cell, 2012, 23(18): 3754-3763.
[19] UMEHARA K, IIMURA T, SAKAMOTO K, et al. Canine oral mucosal fibroblasts differentiate into osteoblastic cells in response to BMP-2. Anat Rec, 2012, 295(8):1327-1335.
[20] LIU T, GAO Y, SAKAMOTO K, et al. BMP-2 promotes differentiation of osteoblasts and chondroblasts in Runx2-deficient cell lines. J Cell Physiol, 2007, 211(3):728-735.
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脚注
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基金
国家自然科学基金项目资助(81373898,81630105);浙江省卫生高层次创新人才培养工程项目资助(2014-108-24)
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