血管平滑肌细胞5-羟色胺受体1B差异表达影响手臂振动病发生的核因子-κB信号机制研究

Hand-transmitted vibration widely exists in occupational activities,which brings severe hazards to human health and may lead to Handarm Vibration Syndrome (HAVS),which is hard to remedy and almost irreversible.Previous epidemiological and in vivo animal experiment studies focused on exploration of the influencing factors,biomarkers and analysis of elementary mechanisms of HAVS.We have found that several Single Nucleotide Polymorphisms (SNPs) which are in the 5'flanking and coding region of 5-hydroxytryptamine receptor 1B (HTR1B) gene are associated with HAVS susceptibility.Based on what we found previously, we intend to analyze and screen haplotypes in the 5'flanking region that differ in regulating HTR1B expression,with which we will further construct different restructuring plasmids and then achieve cell models of which the 5-HT1B expression could be regulated.This project consists of animal experiments,epidemiological studies and cell models studies.How and to what extent the effects of diversity levels of 5-HT1B expression on vascular smooth muscle cells(SMC) proliferation,differentiation and the ability of withstand oxidant stress will be studied.Whether serotonin could activate NF-κB in SMC will be confirmed and We will unveil whether nuclear factor-κB pathway plays a crucial role in the progression of HAVS,confirm and evaluate its contribution in the process of HAVS.Besides,whether hand transmitted vibration plays a role in regulating the expression of α2C-adrenocepter and endothelin receptors through the NF-κB mechanisms will also be analyzed and confirmed both in cell models and populations exposed to vibration. All these mentioned above will acquire mechanisms explaining the progressivity of HAVS even after abstaining from vibration and provide evidences for HAVS mechanisms study at the cellular and molecular level.

手传振动接触广泛、危害严重，手臂振动病(HAVS)缺乏有效治疗手段，预后差。现有研究主要通过流行病学和动物实验探讨其影响因素和生物标志物，尚缺乏深入探讨其发生机制的细胞模型和方法。我们前期发现5-羟色胺1B受体(5-HT1B)基因5'端(2000bp)和编码区单核苷酸位点多态性与手传振动的损伤效应关联。本研拟进一步增加样本量以确认上述关联性，并分析人群中不同频率5-HT1B编码基因HTR1B 5'端(2000bp)单体型，研究其调控转录的功能活性差异，进而构建具有不同5-HT1B受控表达水平的人血管平滑肌细胞模型。通过这一细胞模型探讨5-羟色胺对平滑肌细胞增殖、分化、迁移，表面α2C肾上腺素和内皮素受体表达的影响以及氧化应激损伤差异的核因子-κB机制，并通过振动动物模型和人群血样分析相关指标验证上述机制。这将为在细胞和分子水平明确手臂振动病发生机制提供基础和依据。

5-HTR1B由HTR1B基因所编码，在神经元、血小板和VSMC表面高表达，是原发雷诺病的易感基因。研究表明，VSMC 是振动损伤微循环的靶点。本项目采取病例-对照研究的方法,收集广东某运动器材厂从事手传振动作业 1 年以上的 77 名男性打磨工人作为接振组，以 80 名不从事手传振动作业的男性健康人为对照组。进行职业流行病学调查，计算手传振动作业工人的累计暴露指数（CEI）；并采集血样，测定血浆中sICAM-1、ET、TGF-β水平，采用spss软件二分类非条件Logistic回归分析估计各因素与血浆因子水平的关系，发现振动暴露组中 CVEL 高水平亚组和有 VWF 亚组与 TGF-β 水平相关（P<0.05），而与sICAM-1、ET的水平无关（P均>0.05）。HTR1B基因多态性分析，检测某运动器材加工厂振动作业工人5-羟色胺受体1B基因(HTR1B)多态性位点，Logistic回归分析其与VWF的关联性，Haploview软件进行其遗传特征和连锁不平衡(LD)分析。结果共筛查到7个已知单核苷酸多态性位点(SNP)，其中rs17273700位点突变杂合型(AG)，rs11568817突变杂合型(AC)，rs6298突变杂合(AG)与纯合型(GG)和rs6296突变杂合(GC)与纯合(CC)型相对其各自纯合型均有增加VWF发生风险的趋势(P<0.05)；但在校正年龄、吸烟、饮酒和工龄差异后均无统计学意义(P>0.05)；各位点基因型频率均符合Hardy-Weinberg(H-W)平衡(P>0.05)。rs17273700和rs11568817与rs130058、rs6296与rs6297之间分别呈强LD，rs17273700与rs11568817为完全LD。项目组扩大了样本量进行了某运动器材厂接振工人的整群抽样检测。结果人群各位点基因型频率均符合Hardy-Weinberg(H-W)平衡(P>0.05)。在校正年龄、吸烟、饮酒和工龄差异后，对接触手臂振动小于10年的工人，rs6298(GG)和rs6298(AG)比rs6298(AA)患白指风险高，OR值和95%CI分别为4.78(1.01,22.64)和5.84(1.53,22.32)。初步验证了HTR1B基因多态性与振动性白指发生的相关性。研究表明，HTR1B基因多态性与振动性白指发生的相关性。