退行性脊柱侧凸的血清比较蛋白质组学研究
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摘要
研究背景：
退行性脊柱侧凸(degenerative scoliosis, DS)不仅导致外观畸形,还可引起腰痛、下肢放射痛、间歇性跛行等,甚至损害心肺功能。随着人口老龄化趋势及老年人生活方式的转变,DS逐渐成为导致脊柱功能障碍、影响老年人生活质量的一种重要的腰椎退行性疾病。DS的病因不明,可能与椎间盘、关节突关节的不对称退变、骨质疏松和遗传学因素有关,但这些并不是发生退行性脊柱侧凸的直接原因。目前关于DS的研究绝大多数都是针对该病的治疗,涉及病因学方面的基础研究很少,而且还没有分子生物学方面的报道。因此,弄清DS的病因,确立DS发病及进展相关的分子标志物,从而找到有效预测和防治侧凸加重的方法对于该病的临床诊治具有重大意义。2D-DIGE是近年来发展的新蛋白质组技术,是目前定量蛋白质组学研究中可信度和准确性最高的技术之一。对DS进行比较蛋白质组学研究,寻找患者血清的蛋白质差异,有助于从分子水平认识DS的发生发展机制,也可以为DS的诊断和防治提供有效的分子标志物。
研究目的：
1.构建DS患者与对照间血清差异蛋白质组图谱。
2.应用质谱鉴定技术,对差异蛋白质点进行鉴定。
3.结合蛋白功能网络研究成果,分析所鉴定的蛋白质在DS发生发展中的作用。
研究方法：
1.采集12例退行性脊柱侧凸患者和12例年龄、性别匹配的腰椎管狭窄患者的血清。经去高丰度蛋白、CyDye染料交叉标记,然后进行荧光双向差异凝胶电泳、图像扫描,并用DeCyder v.5.02图像分析软件对DIGE图像进行分析和寻找差异点。
2.对筛选出的差异点进行基质辅助激光解吸／电离飞行时间质谱(MALDI-TOF-MS)
分析,检索IPI-HUMAN数据库鉴定蛋白。
研究结果：
1.经过荧光双向差异凝胶电泳、DeCyder软件分析发现,退行性脊柱侧凸患者和腰椎管狭窄患者血清中有11个蛋白质差异点,两者之间的差异有统计学意义。其中3个点在DS血清中表达上调,8个点表达下调。
2.11个蛋白质差异点经质谱分析,IPI-HUMAN数据库鉴定出7个差异蛋白。
结论：
1.首次对DS患者进行蛋白质组研究,成功构建DS患者与腰椎管狭窄患者血清差异蛋白质组资料库。
2.应用质谱鉴定技术,成功鉴定出7个差异表达蛋白质。这种蛋白质的差异表达究竟是DS发生的始动因素还是畸形引起的继发改变尚不能明确。其具体意义、具体作用及其是否具有DS特异性尚需进一步研究以阐明。研究结果为DS的发病和进展机制研究,筛选可作为DS诊断标记的血清标志物及药物作用靶点奠定了理论基础。
Background:
Degenerative scoliosis (DS) not only leads to the appearance of deformity, but also can lead to low back pain, sciatica, intermittent claudication, and even damages the cardiovascular system. With the population aging trends and lifestyle changes of the elderly, DS has become an important degenerative lumbar disease causing spinal dysfunction and affecting quality of life of the elderly. The pathogenesis of DS is still unknown. Although osteoporosis, asymmetric degenerative disc disease, facet tropism and inheritance have been implicated as factors in the development of degenerative scoliosis, none has been shown to be directly related. Most of current research on the DS is about the treatment of the disease, and the basic research on the etiology is few. Furthermore, there has no molecular biology study been reported. Therefore, It is very important to ascertain the etiology of degenerative scoliosis and establish related molecular markers predicting and controlling the scoliosis. The recently introduced differential in gel electrophoresis(DIGE) technology is believed to be a highly reliable and accurate tool for quantitative analysis of differentially expressed proteins. Comparative Analysis of Serum Proteomes of degenerative scoliosis contribute to understand the molecular mechanism of pathogenesis of DS and can provide effective molecular markers to diagnosis and control this disease.
Objectives:
1. To establish two dimensional difference in gel electrophoresis(2D-DIGE) of serum of degenerative scoliosis and control.
2. To identify the differently expressed proteins in sernm of patients using mass spectrometry techniques.
3. To analyze the role of the identified proteins in the development of degenerative scoliosis through the present protein functional network of research results.
Methods:
1. Serum was collected from 12 degenerative scoliosis patients and 12 age-and gender-matched patients with lumbar spinal stenosis. The samples were labeled with different CyDye, followed by 2D-DIGE, map scanning, and then the maps of protein spots were compared using specific software DeCyder v5.02.
2. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and IPI-HUMAN data base.
Results:
1.11 spots that were differently expressed in the sera of DS patients were found and identified. There were 3 proteins significantly up-regulated and 8 spots significantly down-regulated in serum of DS patients.
2. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS). Seven proteins were identified from samples of DS.
Conclusions:
1. This is the first time to study comparative serum proteomes of degenerative scoliosis. And the serum 2D-DIGE of DS patients and lumbar spinal stenosis were successfully settled.
2. There were 7 protein spots of significance between patients with DS and lumbar spinal stenosis analyzed by DeCyder.Though the result is preliminary, and it is still not clear that these differences are original factors of DS or secondary cellular responses to deformity. reactions. Further investigations are necessary to illustrate the functioning pathway, the specificity and the mechanism of how these proteins contribute to pathogenesis of DS. The information obtained with this proteomic analysis will be very useful in understanding the pathophysiology of DS as well as in finding candidates as new diagnostic biomarkers or drug targets of DS.
退行性脊柱侧凸(degenerative scoliosis, DS)不仅导致外观畸形,还可引起腰痛、下肢放射痛、间歇性跛行等,甚至损害心肺功能。随着人口老龄化趋势及老年人生活方式的转变,DS逐渐成为导致脊柱功能障碍、影响老年人生活质量的一种重要的腰椎退行性疾病。DS的病因不明,可能与椎间盘、关节突关节的不对称退变、骨质疏松和遗传学因素有关,但这些并不是发生退行性脊柱侧凸的直接原因。目前关于DS的研究绝大多数都是针对该病的治疗,涉及病因学方面的基础研究很少,而且还没有分子生物学方面的报道。因此,弄清DS的病因,确立DS发病及进展相关的分子标志物,从而找到有效预测和防治侧凸加重的方法对于该病的临床诊治具有重大意义。2D-DIGE是近年来发展的新蛋白质组技术,是目前定量蛋白质组学研究中可信度和准确性最高的技术之一。对DS进行比较蛋白质组学研究,寻找患者血清的蛋白质差异,有助于从分子水平认识DS的发生发展机制,也可以为DS的诊断和防治提供有效的分子标志物。
研究目的：
1.构建DS患者与对照间血清差异蛋白质组图谱。
2.应用质谱鉴定技术,对差异蛋白质点进行鉴定。
3.结合蛋白功能网络研究成果,分析所鉴定的蛋白质在DS发生发展中的作用。
研究方法：
1.采集12例退行性脊柱侧凸患者和12例年龄、性别匹配的腰椎管狭窄患者的血清。经去高丰度蛋白、CyDye染料交叉标记,然后进行荧光双向差异凝胶电泳、图像扫描,并用DeCyder v.5.02图像分析软件对DIGE图像进行分析和寻找差异点。
2.对筛选出的差异点进行基质辅助激光解吸／电离飞行时间质谱(MALDI-TOF-MS)
分析,检索IPI-HUMAN数据库鉴定蛋白。
研究结果：
1.经过荧光双向差异凝胶电泳、DeCyder软件分析发现,退行性脊柱侧凸患者和腰椎管狭窄患者血清中有11个蛋白质差异点,两者之间的差异有统计学意义。其中3个点在DS血清中表达上调,8个点表达下调。
2.11个蛋白质差异点经质谱分析,IPI-HUMAN数据库鉴定出7个差异蛋白。
结论：
1.首次对DS患者进行蛋白质组研究,成功构建DS患者与腰椎管狭窄患者血清差异蛋白质组资料库。
2.应用质谱鉴定技术,成功鉴定出7个差异表达蛋白质。这种蛋白质的差异表达究竟是DS发生的始动因素还是畸形引起的继发改变尚不能明确。其具体意义、具体作用及其是否具有DS特异性尚需进一步研究以阐明。研究结果为DS的发病和进展机制研究,筛选可作为DS诊断标记的血清标志物及药物作用靶点奠定了理论基础。
Background:
Degenerative scoliosis (DS) not only leads to the appearance of deformity, but also can lead to low back pain, sciatica, intermittent claudication, and even damages the cardiovascular system. With the population aging trends and lifestyle changes of the elderly, DS has become an important degenerative lumbar disease causing spinal dysfunction and affecting quality of life of the elderly. The pathogenesis of DS is still unknown. Although osteoporosis, asymmetric degenerative disc disease, facet tropism and inheritance have been implicated as factors in the development of degenerative scoliosis, none has been shown to be directly related. Most of current research on the DS is about the treatment of the disease, and the basic research on the etiology is few. Furthermore, there has no molecular biology study been reported. Therefore, It is very important to ascertain the etiology of degenerative scoliosis and establish related molecular markers predicting and controlling the scoliosis. The recently introduced differential in gel electrophoresis(DIGE) technology is believed to be a highly reliable and accurate tool for quantitative analysis of differentially expressed proteins. Comparative Analysis of Serum Proteomes of degenerative scoliosis contribute to understand the molecular mechanism of pathogenesis of DS and can provide effective molecular markers to diagnosis and control this disease.
Objectives:
1. To establish two dimensional difference in gel electrophoresis(2D-DIGE) of serum of degenerative scoliosis and control.
2. To identify the differently expressed proteins in sernm of patients using mass spectrometry techniques.
3. To analyze the role of the identified proteins in the development of degenerative scoliosis through the present protein functional network of research results.
Methods:
1. Serum was collected from 12 degenerative scoliosis patients and 12 age-and gender-matched patients with lumbar spinal stenosis. The samples were labeled with different CyDye, followed by 2D-DIGE, map scanning, and then the maps of protein spots were compared using specific software DeCyder v5.02.
2. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and IPI-HUMAN data base.
Results:
1.11 spots that were differently expressed in the sera of DS patients were found and identified. There were 3 proteins significantly up-regulated and 8 spots significantly down-regulated in serum of DS patients.
2. The differentially expressed spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS). Seven proteins were identified from samples of DS.
Conclusions:
1. This is the first time to study comparative serum proteomes of degenerative scoliosis. And the serum 2D-DIGE of DS patients and lumbar spinal stenosis were successfully settled.
2. There were 7 protein spots of significance between patients with DS and lumbar spinal stenosis analyzed by DeCyder.Though the result is preliminary, and it is still not clear that these differences are original factors of DS or secondary cellular responses to deformity. reactions. Further investigations are necessary to illustrate the functioning pathway, the specificity and the mechanism of how these proteins contribute to pathogenesis of DS. The information obtained with this proteomic analysis will be very useful in understanding the pathophysiology of DS as well as in finding candidates as new diagnostic biomarkers or drug targets of DS.
引文
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