Has-miR-335和siRNA-BMP4在脊柱后纵韧带钙化疾病中干预BMP4基因突变引起的异位骨化机制研究

Ossification of the posterior longitudinal ligament (OPLL) disease opress human spinal cord to acroparalysis, our pre-research had sequenced and identified missense mutation of rs17563 of BMP4 coding area was associated with the occurrence of OPLL and increased severity from 450 OPLL patients and 550 matched controls,because of OPLL has characteristics of genetic susceptibility. And further, we proved that missense mutation of rs17563 could induce the overexpression of BMP4 to promote the heterotopic ossification. Hence, in this project, we first enlarge the sample numbers to 1000 cases and detect and confirm the more SNPs of BMP4 from chinese north area OPLL patients, based on the results of our pre-research, we want to detect the microRNAs for regulation of BMP4 expression, and confirm the mechanism that has-miR-335 could induce heterotopic ossification via Smads and p38MAPK cell signal pathway by using of molecular biology analysis methods, which provide a basis for early prognosis of OPLL; next, we want to observe the ossification change with injection of siRNA-BMP4 in the hererotopic ossification rat model, and discuss the possibility of intervention or amelioration of OPLL, provide the experimental foundation for early prognosis and intervention of OPLL translation research.

颈胸段后纵韧带异位骨化压迫人的脊髓导致肢体瘫痪，本课题的前期研究针对脊柱后纵韧带骨化（OPLL）的遗传易感性特点，对北方地区450例OPLL患者和550例正常者的骨形态发生蛋白4（BMP4）进行了测序，发现编码区rs17563位点的错意突变与OPLL的发生和严重程度密切相关，并证实了BMP4基因rs17563位点突变造成的BMP4过表达具有促进OPLL异位骨化的作用。为此，本课题通过扩大OPLL病例至1000例并在前期研究的基础上利用分子生物学等手段在细胞及动物模型基础上进一步分析、检测具有调控BMP4表达作用的microRNAs，确认miR-335通过Smads和p38MAPK细胞通路诱导异位成骨的机制，为OPLL早期预警提供依据；并通过干扰BMP4基因表达，观察大鼠异位成骨组织在siRNA-BMP4作用下的成骨变化，探讨干预或改善OPLL异位骨化的可能性，为临床转化医学提供理论依据。

颈胸段后纵韧带异位骨化，可压迫人的脊髓导致不同程度肢体瘫痪，其发病率在中国北方地区逐年上升。本课题针对脊柱后纵韧带骨化（OPLL）的遗传易感性特点，对北方地区1000例OPLL患者和550例正常者的骨形态发生蛋白4（BMP4）进行了测序，发现编码区rs17563位点的错意突变与OPLL的发生和严重程度密切相关，并证实了BMP4基因rs17563位点突变造成的BMP4过表达具有促进OPLL异位骨化的作用。并使用分子生物学等手段在细胞及动物模型基础上进一步分析、检测了具有调控BMP4表达作用的microRNAs，确认miR-335通过Smads和p38MAPK细胞通路干预异位成骨的机制，为OPLL早期预警提供依据；并通过干扰BMP4基因表达，验证siRNA-BMP4对大鼠异位成骨组织的抑制作用，深入探讨了干预或改善OPLL异位骨化的可能性，不仅为临床转化医学提供理论依据，也为接下来开展家族性OPLL遗传易感基因的功能研究奠定了实验基础。