【作者】 李小囡；

【导师】 陈刚；

【作者基本信息】 天津医科大学 ， 口腔临床医学， 2007， 硕士

【摘要】 目的骨缺损愈合是一个复杂的过程，涉及多种细胞的增殖、分化和细胞外基质的合成与钙化等。血管的重建贯穿骨修复的全过程，多种细胞因子和生长因子参与新血管的形成。其中血管内皮细胞生长因子（vascular endothelial growth factor，VEGF）是最有力的血管生成因子之一，它通过与血管内皮上的特异性受体结合，发挥强大的促内皮增殖，促血管生成作用。本实验通过建立绝经后骨质疏松（PMO）动物（大鼠）模型，并以该骨质疏松动物模型为平台，进一步以外科方法形成下颌骨骨缺损模型，于不同时间间期检测血清雌二醇（E₂）、碱性磷酸酶（ALP）水平，应用免疫组织化学方法观察实验性骨质疏松症大鼠下颌骨骨缺损愈合过程中骨痂VEGF的动态表达，研究雌激素（E）水平对骨痂VEGF表达的动态影响变化，进一步探讨VEGF在下颌骨骨缺损愈合中的作用。方法健康雌性四月龄Wistar大鼠80只，体重330g左右，大鼠适应环境饲养一周后，将其随机分为3组，分别为：假手术+下颌骨骨缺损组（sham operation+mandible bone defect，简称S组，26只）；去势+下颌骨骨缺损组（OVX+mandible bone defect，简称O组，27只）；去势+雌激素治疗+下颌骨骨缺损组（OVX+estrogen+mandible bone defect，简称E组，下颌骨骨缺损术后第1天开始按1.5mg／kg体重给予尼尔雌醇灌胃，每周2次，作为长期雌激素替代治疗组，27只）。先用去势术（OVX）建立骨质疏松模型，去势术后8周骨质疏松模型成立；然后建立左侧下颌骨骨缺损模型。下颌骨骨缺损术前及术后一、二、三、四周采血分别检测血清雌二醇（E₂）、碱性磷酸酶（ALP）水平。下颌骨骨缺损术后一、二、三、四周分批处死动物，处死取材后制备脱钙骨片，VEGF免疫组织化学染色，病理图文分析检测不同组别不同时间骨痂VEGF的表达。结果1．不同组别不同时间血清生化指标的变化：①血清E₂水平：骨缺损术前，O组和E组均低于S组，差别有统计学意义（P=0.000，P=0.000）；术后一周，三组间均有差别，S组最高，其次为E组，O组最低；术后二、三、四周，S组和E组间均无差别，二者均高于O组，差别有显著性（P＜0.05）。②血清ALP水平：骨缺损术前，O组与E组之间无差别（P=0.967），二者均高于S组（P=0.000，P=0.000）；术后一周，三组间差别无显著性；术后二周，S组和O组差别无统计学意义（P=0.830），但都高于E组（P=0.003，P=0.002）；术后三、四周：三组间均有差别，O组最高，S组其次，E组最低。2．VEGF免疫组织化学染色病理图文分析：术后一周：O组与E组间差别无显著性（P=0.076），但均高于S组（P=0.000，P=0.001）；术后二周：三组间均有差别，O组最高，S组其次，E组最低（P＜0.05）；术后三、四周：S组与E组间无差别，但均低于O组（P＜0.05）；结论1．OVX使O组骨缺损处纤维性混合性骨痂持续时间较S组延长，其骨愈合时间在组织学上较S组延迟两周。表明OP影响大鼠下颌骨骨缺损的早期愈合。2．在骨缺损术后各个时间段O组VEGF的表达均高于S组和E组，差别有统计学意义。说明OVX后VEGF在骨痂中持续高表达，且不同时期VEGF表达的程度不同，参与表达的细胞也不同，术后三周达高峰，此期为骨组织愈合的血管重建阶段，提示VEGF能促内皮增殖、促血管生成，在骨缺损愈合中起重要作用。3．E组与S组的E₂水平及VEGF的表达无差别，说明尼尔雌醇雌激素替代疗法对预防和治疗骨质疏松有作用，对骨质疏松大鼠下颌骨骨缺损愈合有促进作用。更多还原

【Abstract】 ObjectivesTo establish experimental postmenopausal osteoporosis animal model forsecondary mandibular bone defect repairing study. To explore expression levels ofVEGF throughout the bone defect reparation, To study the impact that estrogen onVEGF expression in new formed bone callus, and to discus the function of VEGF inbone defect repairation in mandibula.Materials and Methods80 female Wistar rats were divided randomly into 3 groups: group O（ovariectomy+mandibular bone defect, 27 rats）; group E （OVX+mandibular bonedefect+estrogen replacement therapy）, in this group nilestriol was given 1.5mg/kg,twice a week Postoperatively. A sham operation group as group S was set up forcomparative study. The serum biochemical indexes were detected before theostectomy and in 1,2,3,4 week time intervals postoperatively, and the specimens wereretrieved for decalcified sections. The expression of VEGF was detected byimmunohistochemical technique. All the results were statistically analyzed.Results1.variety of the bio-chemical index sign in different groups of different time①Serous E₂ level: preoperatively, group O and group E is lower than group E, Thedifference was statistically significant （P=0.000, P=0.000）; 1 week postoperatively,there were differences between every two groups, group S is the highest, then group E,the lowest is group O; Through 2 to 4 weeks, there were no difference between groupS and E, that both in a higher level than O, The difference was statistically significant（P＜0.05）。 ②Serous ALP level: preoperatively,there was no difference between group O and E（P=0.967）,that both in higher level than group S （P=0.000, P=0.000）;1 weekpostoperatively, there were no significant difference; in 2 weeks time interval, groupS and O were no significant difference （P=0.830）,but both higher than group E（P=0.003, P=0.002）; in 3 to 4 weeks, there were differences between each twogroups, group O is the highest, followed by group S, the lowest is group E.2. VEGF immunohistochemistry1 week postoperatively: there were no significant difference between group O and E（P=0.076）,and both higher than group S （P=0.000, P=0.001）; there weredifferences between each two groups in 2 weeks, group O is the highest, then group S,the lowest is group E. （P＜0.05）; through 3 to 4 weeks, group S and E were in nosignificant difference, but both higher than group O （P＜0.05）.Conclusions1.OVX made fiber bone callus period longer in bone defect repair, which means OPaffect the early healing of rat’s mandibula bone defect.2.Expression of VEGF in group O was higher than group S and E throughout thewhole study period and the difference was statistically significant, suggested that thatOVX makes VEGF a high durative expression in bone callus, and had a differentdegree and different cells involved in different period, and simultaneously, suggestedthat VEGF can help the propagatation of endothelial cells and formation of vessals,it played an important role in the defect reparing of bone.3. There was no difference of E₂ expression level between group E and S, impliedthat nylestriol works in prevention and therapy of osteoporosis, and helps with defectrepairation of steoporosis rat’s mandibula.更多还原