MicroRNAs Predict and Regulate Heme Oxygenase-1 Expression

可供捐赠的器官短缺，导致更多人使用已故捐赠者的器官。这些器官在没有足够的血液供应的情况下承受相对较长的时间，随后会受到损伤(缺血再灌注损伤(IRI))。IRI与移植后立即移植肾功能减退有关，长期与移植失败率增加有关。药物血红素精氨酸(HA)增加了天然保护酶血红素加氧酶-1(HO-1)，在检查炎症和IRI的研究中产生了广泛的有益效果。特别是，在白细胞免疫细胞“单核细胞”及其衍生组织细胞“巨噬细胞”中HO-1的增加似乎对保护很重要。现在有一些注册的临床试验评估HA在治疗炎症性疾病中的使用，包括胰腺炎、糖尿病和肾移植(HOT1研究)。在即将到来的多中心HOT2试验中，将确定HA在术后早期对肾移植功能的影响。在HOT1研究中，我们注意到个体对HA治疗的反应各不相同，一些个体在血液单核细胞中产生更高水平的HO-1。我们发现，多个小RNA分子的microRNAs(MiRs)与单个HO-1对治疗的反应密切相关。由于在治疗前血液MIR的差异是明显的，我们认为这种“MIR信号”可以预测治疗反应，因此可以用来对个体进行个性化治疗。我们还假设，这些MIR可能通过干扰单核细胞和巨噬细胞中HO-1的分子通路来调节药物效应。通过取消这些MIR的作用，我们可以在HO-1的调节中识别新的治疗靶点。在这项研究中，我们计划研究在“HOT2”临床试验中招募的肾移植患者中MIR的作用。我们将确定我们的miR信号是否可以预测肾移植患者单核细胞在HA治疗后HO-1的上调。我们将通过检查之前两个临床试验的储存样本来确定我们的miR特征是否适用于其他人群，在这两个临床试验中，HA被给予接受肝脏和心脏手术的患者。确定这一点可能会使我们能够对那些未来可能有反应的人进行个性化的HA治疗。众所周知，MIR可以抑制基因表达。我们将使用人巨噬细胞进行细胞培养实验，以研究这些miRs在HA治疗后调节HO-1表达的机制。我们将在HA治疗前增加和阻断关键miRs的表达，看看这是否会影响HO-1的反应。我们还将测试可能调节这些miRs表达的因素。这可能使我们能够确定HO-1的新调节因子，这些调节因子可能在未来被用作治疗靶点。这项研究将在爱丁堡国际著名的玛格丽特女王研究所(QMRI)进行，该研究所毗邻爱丁堡皇家医院(RIE)。RIE是HOT2研究的主要中心，并得到爱丁堡临床试验单位的支持。这将使我们能够招募患者并在同一地点分析样本。QMRI是巨噬细胞、microRNAs、HO-1专家的大本营，之前已经支持了3项使用HA完成的临床试验。当地现有的专业知识将帮助我们提交这项建议。这是一个新的和现有的研究领域，对肾移植患者和更广泛的人群都有潜在的好处。确定这个miR信号是否可以用于个性化药物对患者具有潜在的直接好处，同时取消miR调节HO-1的机制可能允许开发新的治疗方法，可以在所有患者中上调HO-1。

The shortage of organs available for donation has resulted in the increased use of organs from deceased donors. These organs endure a relatively prolonged period without an adequate blood supply and subsequently become injured (ischaemia reperfusion injury (IRI)). IRI is associated with reduced function of the transplanted kidney immediately following transplantation and long term is associated with increased rates of transplant loss. The drug heme arginate (HA) increases the natural protective enzyme heme oxygenase-1 (HO-1), resulting in a wide range of beneficial effects in studies examining inflammation and IRI. In particular, increases in HO-1 in the white blood immune cells 'monocytes' and their derived tissue cells 'macrophages' appear to be important for protection. There are now a number of registered clinical trials assessing the use of HA in the treatment of inflammatory conditions including pancreatitis, diabetes and renal transplantation (HOT1 study). In the upcoming multicentre 'HOT2' trial, the effect of HA on kidney transplant function in the early post-operative period will be determined.In the HOT1 study, we noted that individuals responded variably to HA treatment, with some individuals generating higher levels of in HO-1 in blood monocytes. We have found that multiple small RNA molecules 'microRNAs' (miRs) are strongly associated with individual HO-1 response to treatment. As differences in blood miRs are evident prior to treatment, we believe that this 'miR signature' may predict treatment response and therefore could be used to personalise treatment to the individual. We also hypothesise that these miRs may regulate drug effects by interfering with the molecular pathway of HO-1 in monocytes and macrophages. By unpicking the action of these miRs, we may identify novel therapeutic targets in the regulation of HO-1. In this study, we plan to investigate the role of miRs in a subset of kidney transplant patients recruited to the 'HOT2' clinical trial. We will determine whether our miR signature can predict HO-1 upregulation following HA treatment in the monocytes of kidney transplant patients. We will determine whether our miR signature is applicable to other populations by examining stored samples from 2 previous clinical trials where HA was given to patients undergoing liver and cardiac surgery. Determining this may allow us to personalise HA treatment to those likely to respond in future. MiRs are known to suppress gene expression. We will use cell culture experiments using human macrophages to examine the mechanism in which these miRs regulate HO-1 expression following HA treatment. We will both increase and block the expression of key miRs before HA treatment, to see if this influences HO-1 response. We will also test factors which may regulate the expression of these miRs. This may allow us to identify novel regulators of HO-1 which could potentially be exploited as therapeutic targets in future. This research will be conducted in the internationally renowned Queens Margaret Research Institute (QMRI) in Edinburgh which is adjacent to the Royal Infirmary of Edinburgh (RIE). The RIE is the lead centre for the HOT2 study and is supported by the Edinburgh Clinical Trials Unit. This will allow us to recruit patients and analyse samples on the same site. The QMRI is home to experts in macrophages, microRNAs, HO-1 and has supported 3 clinical trials using HA to completion previously. The local expertise available, will assist us in delivering this proposal. This is a novel and exiting area of research with potential benefits to both renal transplant patients and the wider population. Determining if this miR signature can be used to personalise medicine is of potential immediate benefit to patients, whilst unpicking the mechanism in which miRs regulate HO-1 may allow the development new treatments that can upregulate HO-1 in all patients.