膝骨性关节炎中的血管增生研究

发布时间：2018-04-04 12:29

  本文选题：膝骨关节炎　切入点：微血管侵入　出处：《中国人民解放军医学院》2015年硕士论文

【摘要】：研究背景：膝骨关节炎(knee osteoarthritis, KOA)是老年人多种退行性病变中最常见的一种,膝关节活动性疼痛、骨赘增生、内翻畸形为常见症状。涉及KOA病变的因素有很多,体重超标、外伤、剧烈活动过多都与其有关。膝骨关节炎软骨中可见微血管侵入,血管增生机制尚不清楚,各种调节因子作用机制、分子生物学水平有待进一步研究。缺乏形象、直观、特异性强的观察方法观测OA血管增生的部位与数量。本研究基于显微CT的检测人与大鼠膝骨性关节炎(OA)不同时期的软骨、软骨下骨的血管增生情况,探讨骨关节炎不同阶段、不同部位的血管增生变化,进一步阐明软骨血管侵入的机制,探索关节炎发病的机制及疼痛、炎症与血管增生的关系。方法：(1)将20只SD大鼠(雌雄不限,体重300g左右)编码,利用电脑产生随机数,进而将实验动物分为实验组(16只)、假手术组(4只)。实验组行左膝前交叉韧带切除、左膝半月板切除造模,假手术组只打开关节囊,不切除前交叉韧带、半月板,对照组不做任何处理,以亚纳米级硫酸钡与实验用明胶为原料,在65℃-70℃的水浴锅中配置成所需造影剂,以大鼠的腹主动脉为注入段,下腔静脉为流出段,在用肝素钠注射液与生理盐水混合置换出下肢血液,然后利用甲醛溶液固定血管形态,进行造影剂的灌注。标本处死后冷冻一夜固定,次日进行标本取材,实验组分别于术后3周、6周、9周、12周取材。取材部位位于大鼠膝关节标本内侧胫骨平台骨-软骨单元,进行大体观察与组织学评分、Mirco-CT血管形态计量学分析、增生血管的三维重建、血管增生相关基因的PCR等检测、VEGF因子的免疫组化染色。(2)取2013年5月-2014年5月行全膝人工关节置换术后的OA患者的胫骨平台标本50例,于-20°下保存,所取标本均与患者签署自愿捐赠协议。根据软骨评分将OA标本共分为4组：A组(Ⅰ级)、B组(Ⅱ级)、C组(Ⅲ级)、D组(Ⅳ级)。进行VEGF免疫组化染色、组织染色、PCR基因扩增等检测,比较不同程度病变的血管增生情况。结果：(1)显微CT显示：造影剂充填胫骨平台、股骨髁的微血管,显影效果良好。手术组在切除前交叉韧带与半月板三周后即出现胫骨平台的软骨退变,随着时间的进行,关节炎病变加重。根据大体观察与组织学评分,术后3周：关节软骨点状损伤。术后6周：关节软骨呈片状退变,破坏范围增大。术后9周：关节软骨呈全层缺损。术后12周：有骨赘形成。利用显微CT对胫骨平台与股骨髁进行扫描,通过设定阈值对增生的微血管进行三维成像,手术组各期增生的微血管与假手术组相比,均有显著差异(P0.05),手术各组增生的微血管与时间呈相关性变化,趋势为先增加后减少,在术后9周达到高峰。(2)四组标本中,C组标本的VEGF免疫组化染色的阳性率最高,各组进行血管生成相关的VEGF基因扩增检测,C组标本高于其他各组。结论：(1)利用明胶配比硫酸钡可以对大鼠进行充分的血管充盈灌注；显微CT可以对关节内微血管进行定性、定量的分析。在大鼠骨关节炎模型中,关节内增生的微血管与时间呈相关性变化,先升高再降低,在关节炎中期,增生的微血管达到峰值(3)血管增生在人骨关节炎的发病的机制中至关重要,在病变早期,随着关节炎的症状加重,软骨内增生的血管增多,在病变晚期,软骨全层缺损,骨赘形成,血管增生减少。
[Abstract]:Background: osteoarthritis of the knee (knee osteoarthritis KOA) is one of the most common variety of elderly degenerative diseases, pain of knee joint activity, osteophyma, varus deformity is a common symptom. There are many factors involved in KOA lesions of the weight exceed the standard, severe trauma, excessive activity is related. Cartilage of knee osteoarthritis in the visible microvascular invasion, angiogenesis mechanism is not clear, various factors regulating mechanism, further study the level of molecular biology. The lack of image, intuitive, specificity observation observation method of OA blood vessel increase position and quantity of students. This research is based on the micro CT detection of human and rat knee osteoarthritis (OA) in different stages of cartilage, subchondral bone vascular hyperplasia, to explore the different stages of osteoarthritis, vascular proliferation of different parts, to further elucidate the mechanism of cartilage vascular invasion, explore arthritis The incidence and mechanism of pain, inflammation and vascular proliferation. Methods: (1) 20 SD rats (male or female, weight about 300g) encoding, random numbers generated by computer, and then the experimental animal were divided into experimental group (16 rats), sham operation group (4 rats) in experimental group. Left knee anterior cruciate ligament resection, left knee meniscectomy model, sham operation group only open the joint capsule, without resection of anterior cruciate ligament, meniscus, the control group without any treatment, with sub nanometer barium sulfate and experiment with gelatin as raw materials, into the desired contrast agent configuration in a water bath at 65 -70 C. in the rat abdominal aorta for injection, inferior vena cava is the outflow segment in Heparin Sodium Injection, and injected with the replacement of blood, and then use Formaldehyde Solution for fixed vascular morphology, contrast agent perfusion. Specimens were frozen overnight and fixed, the specimens were drawn, experimental group 鍒�浜庢湳鍚�3鍛，

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