研究背景
高血压是世界范围内一个严重的公共卫生问题，影响超过30%的人口和70%的老年人。原发性高血压（essential hypertension，EH）指没有确切病因的高血压。低级别炎症已被认为在高血压和心血管疾病中起着重要的病理生理作用。包括单核细胞在内的炎症细胞在高血压的发生中起着重要作用。免疫细胞可以渗透到多个器官中，从而导致其功能紊乱并导致血压升高。事实上，实验性高血压动物模型和高血压患者的肾脏和血管系统中单核细胞/巨噬细胞和淋巴细胞增加，提示PBMCs的炎症作用在血管功能障碍和高血压中起着关键作用。
研究目的
1) 通过以微量泵泵注Ang II（1500 ng/kg/min），诱导小鼠建立高血压模型，在此基础上，应用基因表达谱芯片检测血管组织中有明显差异表达的基因，筛选与炎症反应、细胞增殖和凋亡相关的基因。
2) 采用病例对照实验，原发性高血压组纳入166人，健康对照组纳入306人。收集外周血标本，用LUMINEX的方法，在血清中检测在动物组织中差异表达的蛋白。
3) 用实时荧光定量qPCR的方法，在外周血单个核细胞中检测差异蛋白及其受体的mRNA的含量，并初步探讨信号通路和发病机制，为临床诊断和治疗，提供新的帮助。
研究方法
1）小鼠高血压模型的构建及差异基因的筛选与验证
我们用微量泵灌注Ang II（1500 ng/kg/min）分别持续1、3、7天，每组3只小鼠，诱导小鼠建立高血压模型，健康对照组用微量泵灌注等量生理盐水。取小鼠冠状动脉组织构建基因表达谱芯片，筛选高血压1、3、7天组的差异表达基因，用生物信息学技术GO和KEGG富集分析，分析差异基因富集的基因功能和信号通路。用qPCR的方法，在两组动物组织中检测mRNA的相对表达量，对基因表达谱芯片的结果进行验证。
2） 高血压病人血清中的差异蛋白含量变化研究
我们采用病例对照实验，共收取原发性高血压病人166人，健康对照者306人。收集外周血标本，用LUMINEX的方法，在血清中检测在动物组织中差异表达的蛋白的含量，并用多变量逻辑回归校正其他变量的影响。
3） 汉族原发性高血压病人外周血单个核细胞中差异蛋白基因表达水平变化研究
我们采用病例对照实验，共收取原发性高血压病人166人，健康对照者306人。收集外周血单个核细胞标本，提取细胞总mRNA，在外周血单个核细胞上检测在动物组织中差异表达基因及其受体的mRNA含量。初步探讨信号通路和发病机制，为临床诊断和治疗，提供新的帮助。
结果及结论
Ang II 诱导的小鼠高血压模型中，我们筛选出了9个差异表达上调的基因，包括：IGFBP2、IL-6、IL-15、IL-18、IL-33、CCL2、CXCL1、OPN、OPG。结合以往已发表的文章，在心血管疾病和炎症状态下，游离VCAM-1升高。在原发性高血压病人外周血清中，我们发现IL-18、CCL2、OPG、OPN以及IL-33的可溶性受体sST2升高。在原发性高血压组和健康对照组中，我们发现IL-6、IL-15、IL-33、IGFBP2、CXCL1、VCAM-1在两组间没有统计学差异，可能与组织特异性有关。与对照组比较，原发性高血压组外周血单个核细胞上的IL-6ST mRNA显著下降，IL-6ST表达下降，STAT的磷酸化和二聚化减少，引起下游信号通路的改变，会促进高血压的发生。IL-18 mRNA显著升高，无论是直接或通过氧化应激途径和基质金属蛋白，都能改变内皮功能或诱导血管平滑肌细胞迁移和/或增殖，产生伴随高血压发生的血管变化。OPG mRNA显著升高，OPG与RANKL和TRAIL的结合变化影响了下游信号通路。在原发性高血压组外周血单个核细胞上，IL-33未见明显变化，sST2显著升高，ST2L显著降低，影响了IL-33/ST2信号通路，可能导致高血压的发生。
 

background
hypertension is a serious public health problem worldwide, affecting more than 30% of the population and 70% of the elderly. essential hypertension refers to hypertension without a definite cause. low-grade inflammation has been considered to play an important pathophysiological role in hypertension and cardiovascular diseases. inflammatory cells, including monocytes, play an important role in the development of hypertension. immune cells can penetrate multiple organs, leading to dysfunction and elevated blood pressure. in fact, the increase of monocyte/macrophage and lymphocyte in the kidney and vascular system of experimental hypertensive animal models and hypertensive patients suggests that the inflammation of pbmcs plays a key role in vascular dysfunction and hypertension.
ive
1) ang ii (1500 ng/kg/min) was injected by micropump to induce hypertension in mice. on this basis, differentially expressed genes were detected by gene expression microarray，and genes related to inflammation, cell proliferation and apoptosis were screened.
2) a case-control study was conducted in which 166 people were enrolled in the primary hypertension group and 306 in the healthy control group. peripheral blood samples were collected and the differentially expressed proteins in animal tissues were detected by luminex.
3) real-time qpcr was used to detect the content of differentially expressed proteins and their receptors in peripheral blood mononuclear cells, and to explore the signaling pathway and pathogenesis preliminarily, so as to provide new help for clinical diagnosis and treatment.
methods
1) establishment of a mouse model of hypertension and screening and validation of differentially expressed genes
ang ii (1500 ng/kg/min) was infused with micropump for 1, 3 and 7 days, respectively, to establish hypertensive model. three mice were in each group. the healthy control group was infused with normal saline by micropump. gene expression profiles were constructed from mouse coronary artery tissues, and differentially expressed genes were screened from hypertensive groups. the gene functions and signaling pathways of differentially expressed genes were analyzed by bioinformatics technology including go and kegg enrichment analysis. the relative expression of rna in two groups of animal tissues was detected by qpcr, and the results of gene expression microarray were verified.
2) study on the changes of serum differential protein content in patients with hypertension
a total of 166 patients with essential hypertension and 306 healthy controls were enrolled in a case-control study. peripheral blood samples were collected and the protein content differentially expressed in animal tissues was detected by luminex method. the influence of other variables was corrected by multivariate logistic regression.
3) changes of differential protein gene expression in peripheral blood mononuclear cells of patients with essential hypertension
a total of 166 patients with essential hypertension and 306 healthy controls were enrolled in the case-control study. peripheral blood mononuclear cells (pbmcs) were collected to extract the total cell rna and detect the differentially expressed genes and their receptors in animal tissues on pbmcs. to explore the signaling pathway and pathogenesis in order to provide new help for clinical diagnosis and treatment.
results and conclusion
in ang ii-induced hypertension mouse model, 10 differentially expressed genes were screened, including igfbp2, il-6, il-15, il-18, il-33, ccl2, cxcl1, opn and opg. combined with previous published articles, free vcam-1 increased in patients with cardiovascular disease and inflammation. in peripheral serum of patients with essential hypertension, we found elevated levels of il-18, ccl2, opg, opn and soluble receptor sst2 of il-33. in essential hypertension group and healthy control group, we found no significant difference in il-6, il-15, il-33, igfbp2, cxcl1, vcam-1 between the two groups, which may be related to tissue specificity. compared with the control group, the expression of il-6st and the phosphorylation and dimerization of stat in peripheral blood mononuclear cells of essential hypertension group decreased significantly, which led to the change of downstream signaling pathway and promoted the occurrence of hypertension. interleukin-18 (il-18) mrna increased significantly. both direct or peroxidative stress pathways and matrix metalloproteinases could alter endothelial function or induce migration and/or proliferation of vascular smooth muscle cells, resulting in vascular changes associated with hypertension. opg mrna increased significantly, which was related to the combination of opg with rankl and trail affecting the downstream quotation pathway. in the peripheral blood mononuclear cells of essential hypertension group, il-33 did not change significantly, sst2 increased significantly, st2l decreased significantly, affecting the il-33/st2 signaling pathway, which may lead to hypertension.