SOX9和VEGF在小儿先天性脊柱侧弯顶椎生长板软骨的表达研究

发布时间：2018-05-20 06:42

本文选题：先天性脊柱侧弯 + 顶椎　；参考：《郑州大学》2012年硕士论文

【摘要】：先天性脊柱侧弯(Congenital scoliosis,CS)是临床上较常见的一种小儿先天性脊柱畸形,其发生发展主要归因于先天存在的异常脊椎导致脊柱纵向不平衡生长。在脊柱纵向发育过程中脊椎生长板软骨是纵向生长的基础,顶椎作为先天性脊柱侧弯的触发点,其生长板软骨的化骨活性直接关系到侧弯的发生发展情况。以往研究已证实脊椎生长板具有类似长骨生长板的特性,生长分化受多种细胞因子的调控。其中SOX9是一个与哺乳动物的性别决定因子SRY(Y染色体上的性别决定区)具有高度同源性的转录因子,表达于除肥大软骨细胞外所有分化型软骨细胞,并贯穿整个软骨形成过程,被认为在软骨内成骨的初始阶段发挥着重要作用。血管内皮生长因子(VEGF)是刺激内皮细胞生长和血管再生的一个重要因素,研究发现在软骨内化骨过程中对于软骨细胞死亡,细胞外基质重塑,血管生成和骨骼形成是一种基本的调节物质。因此通过对侧弯顶椎凸凹两侧生长板软骨VEGF和SOX9表达情况的检测可以判断侧弯两侧软骨板成骨能力的差异,推断小儿先天性脊柱侧弯发病和进展的分子生物学基础,为进一步的分子生物学研究奠定基础。目的通过免疫组化和：mRNA原位杂交方法判定先天性脊柱侧弯患儿凸凹两侧生长板软骨SOX9和VEGF的表达部位和情况,了解两侧生长板软骨生物活性的差异,发现SOX9和VEGF表达的相关性,探讨影响先天性脊柱侧弯发病和进展的分子生物学基础。方法取2010年10月～2011年9月我院收住的20例1～5岁先天性脊柱侧凸患者,男8例,女12例,平均年龄3.2(3.2±0.9)岁,术前订均接受详细的体格检查和影像学检查(包括x线片,全脊柱螺旋CT+三维重建,MRI)。入院后经患者直系亲属的知情同意后,取术中切除的半椎体凸凹两侧生长板软骨,常规HE染色并应用免疫组化和mRNA原位杂交的方法对顶椎生长板的SOX9和VEGF表达进行检测,观察SOX9和VEGF在顶椎生长板软骨的表达位置,以及在顶椎凸凹两侧生长板软骨的表达情况,同时对SOX9、VEGF和VEGFmRNA的表达进行相关性研究。凸凹两侧因子表达结果采用配对t检验,p0.05为差异有统计学意义。相关性分析采用Pearson相关性分析,以p0.05为检验水准。结果 l、凸凹两侧生长板的形态学观察,两侧软骨板HE染色可见生长板有类似长骨生长板的结构,大致可以分为静止层、增殖层和肥大层软骨区。凸侧生长板软骨的增殖区和肥大区厚度更大,细胞排列也较紧密。 2、SOX9主要表达于静止层和增殖层软骨细胞,VEGF主要表达肥大软骨区,少量表达于增殖层,但静止层未见表达。SOX9在凸侧生长板软骨表达量为154.376±8.055,凹侧为121.413±6.751,VEGF在凸凹两侧生长板软骨的表达量分别为134.989±9.731,108.498±8.737,VEGFmRNA在凸凹两侧生长板的表达为116.517±7.536和97.077±6.627,分别经配对t检验p均0.05, SOX9、VEGF、 VEGFmRNA在顶椎凸凹两侧生长板软骨的表达差异有统计学意义。 3.对脊椎生长板软骨SOX9和VEGF的表达进行Pearson相关性分析,r=0.650,p0.05,有统计学意义。对VEGF的表达和VEGFmRNA的表达进行相关性分析,r=0.343,p0.05。无统计计学意义。结论 1、先天性脊柱侧弯患者顶椎生长板软骨,大体观凸侧较凹侧具有较高的生物活性。 2、SOX9、VEGF和VEGFmRNA在先天性脊柱侧弯患儿顶椎凸侧生长板软骨的表达量较凹侧高,这种表达差异可能是小儿先天性脊柱侧弯发病和进展的关键因素。 3、SOX9和VEGF在脊椎生长板不同软骨区表达量不同。整体分析,SoX9和VEGF的表达存在正线性相关。
[Abstract]:Congenital spinal scoliosis (Congenital scoliosis, CS) is a common congenital spinal deformity in children. Its development is mainly attributable to the congenital abnormal spine resulting in the longitudinal uneven growth of the spine. In the longitudinal development of the spine, the vertebral growth plate cartilage is the basis of longitudinal growth and the apex vertebrae as a congenital spinal column. The triggering point of the lateral bending, the osteogenic activity of the growth plate cartilage is directly related to the occurrence and development of the lateral bending. Previous studies have confirmed that the vertebral growth plate has the characteristics similar to the long bone growth plate, and the growth and differentiation is regulated by a variety of cytokines. SOX9 is a sex determinant on the Y chromosome (sex determination on the Y chromosome). A highly homologous transcription factor, expressed in all differentiated chondrocytes except the hypertrophic chondrocytes and throughout the process of cartilage formation, is considered to play an important role in the initial stage of endochondral osteogenesis. Vascular endothelial growth factor (VEGF) is an important factor in stimulating the growth of the inner skin cells and the regeneration of blood vessels. In the process of cartilaginous endochondral bone, the death of cartilage cells, the remodeling of extracellular matrix, angiogenesis and bone formation is a basic regulating substance in the process of endochondral osseous bone. Therefore, the difference in the osteogenesis ability of the cartilaginous plates on both sides of the lateral bending can be judged by the detection of the expression of VEGF and SOX9 on the sides of the growth plate cartilage on both sides of the lateral bending top vertebra. Molecular biological basis for pathogenesis and progression of scoliosis in nature will lay a foundation for further molecular biology research.
objective
The expression of SOX9 and VEGF in the growth plate cartilage on both sides of the children with congenital scoliosis was determined by immunohistochemistry and mRNA in situ hybridization. The differences in the biological activity of the cartilage on both sides of the growth plate were understood, the correlation between the expression of SOX9 and VEGF was found, and the molecular biological basis of the pathogenesis and progression of congenital scoliosis was discussed.
Method
From October 2010 to September 2011, 20 cases of 1~5 year old congenital scoliosis were collected from our hospital, 8 men and 12 women, with an average age of 3.2 (3.2 + 0.9) years old. A detailed physical examination and imaging examination (including X-ray, total spinal spiral CT+ 3D reconstruction, MRI) were accepted before the operation. The expression of SOX9 and VEGF in the apical growth plate was detected by routine HE staining and immunohistochemistry and mRNA in situ hybridization. The expression of SOX9 and VEGF at the apical growth plate cartilage, and the expression of the long plate cartilage on both sides of the apical convex concave were observed, and SOX9, VEGF and VEG were also used for SOX9, VEGF and VEG on both sides of the excised hemivertebra. The expression of FmRNA was studied. The results of both sides of convex and concave factors were detected by paired t, and the difference between P0.05 was statistically significant. The correlation analysis adopted Pearson correlation analysis and P0.05 as the test level.
Result
L, the morphological observation of the growth plates on both sides of the convex concave. The structure of the growth plate similar to the long bone growth plate was observed on both sides of the cartilage plate, which could be divided into the static layer, the proliferating layer and the hypertrophic region. The proliferation area and the hypertrophic area of the protruded growth plate cartilage were more thick and the cells were arranged closely.
2, SOX9 is mainly expressed in the static and proliferating layer cartilage cells. VEGF mainly expresses the hypertrophic cartilage region, which is expressed in a small amount in the proliferation layer, but the expression of.SOX9 in the protruded growth plate cartilage is not 154.376 + 8.055, the concave side is 121.413 + 6.751, and the expression of VEGF on both sides of the convex concave is 134.989 + 9.731108.498 + 8.73, respectively. 7, the expression of VEGFmRNA on both sides of the convex concave is 116.517 + 7.536 and 97.077 + 6.627. The expression of P 0.05, SOX9, VEGF and VEGFmRNA on both sides of the top vertebral convex concave is statistically significant.
3. the expression of SOX9 and VEGF in the spinal growth plate cartilage was analyzed by Pearson correlation, r=0.650, P0.05, and there were statistical significance. The correlation analysis between the expression of VEGF and the expression of VEGFmRNA, r=0.343, p0.05. had no statistical significance.
conclusion
1, the growth plate cartilage of the apical vertebrae in congenital scoliosis has a higher biological activity than the concave side.
2, SOX9, VEGF and VEGFmRNA in children with congenital scoliosis are more expressed in the protruding plate cartilage than the concave side. This difference may be the key factor for the onset and progression of children's congenital scoliosis.
3, the expression levels of SOX9 and VEGF in different cartilage regions of the spinal growth plate were different. Overall analysis showed that there was a positive linear correlation between SoX9 and VEGF expression.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R726.5

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