环状RNA参与高血压及其并发缺血性脑卒中机制的分子流行病学研究

Our previous study used microRNA (miRNA) chip to screen relevant miRNA for hypertension and vivificated 3 kinds of miRNA (miR-511, miR-520a-3p and miR-1274a) abnormal expression comparing to health control subjects. Further study showed that miR-511 polymorphism rs4748424 was significantly associated with essential hypertension. The results indicated that miRNA and encoding gene variation may be involved in the pathogenesis of hypertension. Recent studies supported that circular non-coding RNA (circRNA) played an important role in the pathomechanism of atherosclerosis with the interaction of miRNAs but there was rare study investigating the relationship between the circRNA and hypertension as well as stroke. The project would conduct a large sample size case-control study to screen the serum abnormal expressed circRNA relevant to hypertension and stroke and evaluate the association of the encoding genes of candidate circRNAs with hypertension and stroke by combining with the significant miRNA identified previously. In addition, the generalized multifactor dimensionality reduction (GMDR) analysis will be used to evaluate gene-gene and gene-environment interaction. The expected results of this study would be helpful to illuminate the epigenetic mechanism of circRNA involved in the pathogenesis of hypertension and stroke and provide sensitive biomarkers for evaluation of clinical prognosis of hypertension and stroke and theoretical basis for the screening of drug targets and disease prevention.

课题组前期通过微小RNA（miRNA）筛选并验证高血压患者有3种miRNA异常表达，其中miR-511编码基因变异与原发性高血压存在显著关联，结果表明miRNA及其基因变异可能共同参与了高血压发病机制。最近国外研究提示，环状非编码RNA（circRNA）通过与miRNA交互作用参与了动脉粥样硬化病理机制，但circRNA与高血压、脑卒中关系的研究少见报道。本课题采用病例对照研究设计,宏观人群研究与分子遗传学研究相结合，通过筛选高血压及缺血性脑卒中与正常对照差异表达的circRNA，分析circRNA和编码基因变异与前期发现异常表达miRNA的关系及其对高血压、脑卒中发病的作用，应用广义多因子降维法（GMDR）分析基因-基因、基因-环境交互作用，结果将有助于系统阐明高血压和缺血性脑卒中的表观遗传学机制，为其临床治疗药物设计靶点提供思路和线索，并可为疾病预后评价和人群防制提供敏感指标理论依据。

最近国外研究提示circRNA可能参与了心脑血管疾病的发病机制。本课题在第一阶段选择有代表性高血压、脑卒中新发病例和健康对照各10例，通过高通量测序筛选出高血压及缺血性脑卒中差异表达的circRNA；第二阶段选择性别年龄匹配的原发性高血压病例、缺血性脑卒中病例和健康对照，经qRT-PCR定量检测加以初步筛选和重复验证。在高血压病例对照中，按照|Log2FoldChange|>1和P<0.05检验水准，从数据库中筛选出355个差异表达circRNA，其中有150个circRNA上调，205个circRNA下调。进一步在同地区102对高血压比例对照中对部分差异表达circRNA进行复制验证。结果发现，6个circRNAs表达水平在病例和对照中具有统计学差异（P值均<0.05）。hsa_circ_0020915，hsa_circ_0072544，hsa_circ_0008638在病例组中高表达；hsa_circ_0000246，hsa_circ_0003363在病例组中低表达；hsa_circ_0137803在病例组中低表达。在缺血性脑卒中病例对照中，筛选出734个差异表达circRNA，其中430个上调circRNA，304个下调（|Log2FoldChange|>1和P<0.05）。进一步在缺血性脑卒中病例145例和对照150例中对部分差异表达circRNA进行qRT-PCR验证。结果发现，hsa_circ_0064555、hsa_circ_0118748、hsa_circ_0095288、hsa_circ_0125195的表达水平在病例对照组中，差异具有统计学意义（P值均<0.05）；缺血性脑卒中外周血样本中hsa_circ_0118748、hsa_circ_0095288、hsa_circ_0125195的表达水平显著降低，hsa_circ_0064555表达水平在卒中病例中升高。并且功能预测表明，经验证差异表达的circRNA可能参与高血压、缺血性脑卒中的病理生理过程，对疾病的诊断、治疗及预后具有一定的价值，可能成为疾病干预的新的靶点。结果将有助于深入探讨高血压及缺血性脑卒中的分子机制，并可为疾病预后评价和人群防制提供敏感指标理论依据。

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