本文關(guān)鍵詞： 丹參 股骨骨折 磁性微球 BMP2 BMP7　出處：《蘭州大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

【摘要】：背景：骨損傷后愈合障礙是骨科常見的臨床疾病,迄今在骨修復(fù)中仍存在重大挑戰(zhàn),近年來(lái)對(duì)骨折愈合的骨生長(zhǎng)因子關(guān)于細(xì)胞分子和基因水平作用的研究越來(lái)越多。其中BMP-2、BMP-7在骨折愈合中的作用已得到認(rèn)可。丹參作為活血化瘀類代表性中藥,在促骨愈合的作用機(jī)理已有研究,但傳統(tǒng)中藥給藥方式仍存在局限性,開展新的給藥途徑并用于促骨愈合中的研究較少,本實(shí)驗(yàn)通過(guò)磁靶向給藥系統(tǒng)(Magnetic tageting drug delivery system, MTDDS),觀察磁性丹參微球?qū)钦塾线^(guò)程中BMP-2、BMP-7表達(dá)的影響及其作用。目的：觀察磁性丹參微球(Feromagnetic tanshinol microspheres)在大鼠骨損傷后的體內(nèi)分布特點(diǎn)及對(duì)成骨細(xì)胞的增殖及功能分化的影響,探討磁性微球促骨愈合緩釋誘導(dǎo)的作用機(jī)制。方法：建立大鼠股骨骨折模型,在磁靶向作用下注射磁性丹參微球制劑,篩選最佳濃度,考察其磁響應(yīng)性。選用造模成功健康成年SD大鼠,雌雄不計(jì),體重230±30g,使用隨機(jī)數(shù)字表進(jìn)行隨機(jī)分為四組：空白組；藥物組；磁場(chǎng)組；藥物+磁場(chǎng)組。空白組未做任何干預(yù),藥物組只給予磁性丹參微球未外加磁場(chǎng)。磁場(chǎng)組只給外加磁場(chǎng)未給予磁性丹參微球；藥物+磁場(chǎng)組是指給磁性微球的同時(shí)給予外加磁場(chǎng)。無(wú)菌條件下,取下術(shù)后股骨,用注射器沖取骨髓,離心獲取乳白色單個(gè)核細(xì)胞,成骨誘導(dǎo)培養(yǎng)液進(jìn)行誘導(dǎo)培養(yǎng),得到成骨細(xì)胞。丹參磁性微球分別給藥1天、4天、8天和12天,最后一天給藥后2小時(shí)處死大鼠并提取骨折組織。MTT法檢測(cè)骨折組織中第1天、第4天、第8天及第12天成骨細(xì)胞的增殖情況,Western blotting檢測(cè)BMP-2和BMP-7蛋白各時(shí)間點(diǎn)的表達(dá)情況,RT-PCR檢測(cè)BMP-2和BMP-7 mRNA的表達(dá)。結(jié)果：(1)在35000e外加磁場(chǎng)下給予60mg/L的微球制劑后骨折組織的丹參素含量檢測(cè)達(dá)到最大值(P0.05)。(2)MTT試驗(yàn)顯示在成骨細(xì)胞對(duì)數(shù)生長(zhǎng)期第4天和第8天藥物組+磁場(chǎng)組增殖效果明顯高于其他三組(P0.01),藥物組高于空白組和磁場(chǎng)組(P0.05)。(3)Western blotting檢測(cè)藥物+磁場(chǎng)組BMP-2和BMP-7所有時(shí)間點(diǎn)表達(dá)量均高于對(duì)照組(P0.05),BMP-2、BMP-7藥物組表達(dá)量高于對(duì)照組和磁場(chǎng)組(P0.05)。(4)實(shí)時(shí)熒光定量PCR檢測(cè)第12天時(shí)BMP-2、BMP-7mRNA表達(dá)升高最為明顯(P0.05),：藥物組和藥物+磁場(chǎng)組表達(dá)量高于空白組和磁場(chǎng)組(P0.05)。結(jié)論：磁性丹參微球大鼠體內(nèi)有較好的磁響應(yīng)性,并對(duì)成骨細(xì)胞的增殖以及骨形態(tài)蛋白BMP2和BMP7的表達(dá)有促進(jìn)作用。
[Abstract]:Background: the disorder of healing after bone injury is a common clinical disease in orthopedic department. So far, there are still major challenges in bone repair. In recent years, more and more researches have been made on the effects of bone growth factor on cell molecular and gene levels in fracture healing. The role of BMP-2BMP-7 in fracture healing has been recognized. Salvia miltiorrhiza is a representative Chinese medicine for promoting blood circulation and removing blood stasis. The mechanism of promoting bone healing has been studied, but there are still some limitations in traditional Chinese medicine administration. The effect of magnetic Danshen microspheres on the expression of BMP-2 and BMP-7 during fracture healing was observed by magnetic tageting drug delivery system (MTDDSX). Objective: to observe the effect of magnetic Danshen microspheres Feromagnetic tanshinol microspheres after bone injury in rats. Characteristics of osteoblasts and their effects on proliferation and functional differentiation of osteoblasts, Methods: a model of femur fracture in rats was established. Magnetic Danshen microspheres were injected with magnetic target to screen the best concentration. To investigate its magnetic response, male and female male and female SD rats, weighing 230 鹵30 g, were randomly divided into four groups: blank group, drug group, magnetic field group, drug magnetic field group, and blank group without any intervention. The drug group was only given magnetic Salvia miltiorrhiza microspheres without external magnetic field. The magnetic field group only gave magnetic field not magnetic Salvia miltiorrhiza microspheres. In the drug magnetic field group, the magnetic field was given to the magnetic microspheres at the same time as the external magnetic field. Bone marrow was washed with syringe, milky white mononuclear cells were obtained by centrifugation, osteoblasts were cultured in osteoblast induction medium, and Salvia miltiorrhiza magnetic microspheres were given 1 day, 4 days, 8 days and 12 days, respectively. The rats were killed 2 hours after the last day of administration and the fracture tissue was extracted by MTT method to detect the first and fourth day of fracture tissue. Proliferation of osteoblasts on the 8th and 12th day the expression of BMP-2 and BMP-7 protein was detected by Western blotting and the expression of BMP-2 and BMP-7 mRNA was detected by RT-PCR. Results: the bone marrow cells were treated with 60mg / L of microspheres at 35000e magnetic field to detect the expression of BMP-2 and BMP-7 mRNA. The results of MTT assay showed that the proliferative effect of the magnetic field group on the 4th and 8th day of logarithmic growth of osteoblasts was significantly higher than that of the other three groups, and that of the drug group was higher than that of the blank group and the magnetic field group. The expression of BMP-2 and BMP-7 in the magnetic field group was higher than that in the control group at all time points. The expression of BMP-2BMP-7 in the drug group was higher than that in the control group and the magnetic field group.) the expression of BMP-7 mRNA in the drug group and the drug magnetic field group was significantly higher than that in the control group and the magnetic field group on the 12th day. The expression of P0.05 in the field group was higher than that in the blank group and the magnetic field group. Conclusion: the magnetic salvia miltiorrhiza microspheres have better magnetic response in vivo. It also promotes the proliferation of osteoblasts and the expression of bone morphogenetic protein BMP2 and BMP7.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R683.4

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本文編號(hào)：1500869