Real-time monitoring techniques and simplistic platelet assays to reduce and refine animal use in cardiovascular and respiratory biomedical research

血小板是血细胞，在凝血以防止失血和在心脏病发作等情况下形成凝块方面很重要。研究血小板以更好地了解疾病并开发新的治疗方法涉及许多小鼠实验，因为凝块的形成受到许多组织的调节，并且不能在动物体外复制。许多研究小组在清醒的动物中进行研究，通过诱导血液凝块，导致动物在肺部血管阻塞后死亡。没有使用麻醉剂，所以动物遭受了相当大的痛苦。我们已经开发出技术，允许在全身麻醉下的一只小鼠中测量凝块形成。这意味着使用更少的小鼠，避免了痛苦的程序。我们现在希望与目前正在进行实验的科学家合作，这些实验涉及死亡的小鼠，以开发和实施替代方案。我们将测量循环血液中的血小板数量来预测凝块的形成，而不是导致死亡。这是因为当血小板在组织中形成凝块时，一般循环中的血小板较少。也可以从每只小鼠中收集一个以上的血液样品，以进一步减少实验所需的小鼠数量。这种技术可以用来代替涉及死亡的模型。这项技术使用简单，因为血液很容易收集，细胞可以在显微镜下计数。该方法将使用现有的用于治疗心脏病发作的药物和来自研究小组的新测试药物进行检查，这些药物目前依赖于涉及死亡的模型。已经有团体同意参与我们的研究，我们将增加参与人数。这种方法将鼓励科学家将使用死亡的研究转向非常简单的替代技术，这些技术使用更少的老鼠，避免痛苦的程序，因为老鼠在整个实验过程中都被麻醉了。血小板也参与了保护我们免受细菌和感染的过程。为了攻击细菌，血小板需要离开血液，穿过组织到达感染处。身体通过一种称为炎症反应的保护机制来做到这一点。在哮喘等疾病中，血小板从血液进入肺部。更好地了解血小板和其他细胞如何进入肺部将导致新的治疗方法。为了研究这一过程，科学家们在小鼠肺部引起炎症，以复制哮喘等疾病。然后在炎症开始后的不同时间杀死小鼠，并通过显微镜检查评估炎症的严重程度以确定组织中的细胞数量。这涉及使用大量小鼠，因为必须在需要研究的每个时间点杀死小鼠。我们的目标是通过使血小板等细胞具有放射性并跟踪它们在单个麻醉动物中的运动来测量细胞进入肺部的运动。我们还将通过从个体麻醉小鼠中采集血液样本来测量循环血液中血小板的下降来评估炎症。新疗法可以通过测量它们对细胞运动的影响来评估。这将涉及使用更少的小鼠，我们预计将减少70%至80%的小鼠使用。总的来说，我们的目标是开发可用于确定可能用于治疗心脏和肺部疾病的药物效果的技术。申请人在这两个领域都有经验。将开发的技术便宜且易于使用，这将增加其他科学家的吸收。

Platelets are blood cells which are important in clotting to prevent blood loss and in the formation of clots during conditions such as heart attack. Studying platelets to better understand disease and to develop new treatments involves many experiments in mice since the formation of clots is regulated by numerous tissues and cannot be replicated outside an animal. A number of research groups conduct studies in conscious animals by inducing blood clots which lead to the death of the animal following blockage of blood vessels in the lungs. Anaesthesia is not used so the animals suffer considerably. We have developed techniques that allow clot formation to be measured in a single mouse under general anaesthesia. This means that fewer mice are used and painful procedures are avoided. We now wish to work with scientists who currently conduct experiments that involve death in concious mice to develop and implement alternatives. Instead of causing death, we shall measure platelet numbers in circulating blood to predict clot formation. This works because when platelets form a clot in a tissue, there are fewer of them in the general circulation. It may also be possible to collect more than one blood sample from each mouse to further reduce the numbers of mice needed for an experiment. This technique may be used instead of models involving death. The technique is simple to use because blood is easy to collect and cells can be counted under a microscope. The method will be checked using existing drugs that are used to treat heart attack and newer test drugs from research groups who are presently dependent upon models involving death. Groups have already agreed to take part in our study and we shall increase the numbers taking part. This approach will encourage scientists to switch studies using death to very simple alternative techniques that use fewer mice and avoid painful procedures because the mice are anaesthetised for the entire experiment.Platelets are also involved in defending us against germs and infection. To attack germs platelets need to leave the blood stream and travel through tissue to reach the infection. The body does this through a protective mechanism called the inflammatory response. In diseases such as asthma, platelets enter the lungs from the bloodstream. Better understanding of how platelets and other cells enter the lung will lead to new treatments. To study this process scientists cause inflammation in the lungs of mice to replicate conditions such as asthma. Mice are then killed at different times after the start of inflammation and the severity of inflammation assessed by microscopic examination to determine the numbers of cells in the tissue. This involves the use of large numbers of mice because a mouse has to be killed at every time point that needs to be studied. We aim to measure cell movement into the lungs by making cells such as platelets radioactive and tracking their movement in an individual anaesthetised animal. We shall also assess inflammation by measuring the fall in platelets in circulating blood by taking blood samples from individual anaesthetised mice. New therapies can be assessed by measuring their effect on cell movement. This will involve the use of fewer mice and we predict reducing mouse use by 70 to 80%.Overall, we aim to develop techniques that can be used to determine the effects of drugs that may be useful in treating diseases of both the heart and lungs. The applicants have experience in both of these areas. The techniques that will be developed are cheap and easy to use which will increase their uptake by other scientists.