Single Cell Phosphoprofiling of Diabetic Immune Cells

描述(由申请人提供)：免疫系统通过许多不同细胞群之间的多种相互作用发挥作用。这些相互作用导致细胞内蛋白水平、亚细胞定位和激活状态的变化，从而对正常和病理免疫系统功能产生深远影响。初步数据表明，使用抗CD3抗体的全身治疗可以成功地治疗最近高血糖的非肥胖糖尿病小鼠，从而抑制β细胞的破坏和受损组织的再生。我们推测，抗CD3抗体可能直接作用于胰岛浸润的CD3淋巴细胞，或可能激活调节性T细胞。传统的生化技术在使我们能够观察这些小鼠胰腺和淋巴组织中稀有细胞群中的瞬时信号事件方面几乎毫无用处。使用我们最新开发的11色多参数流式细胞术分析免疫谱系中的活性激酶和磷蛋白，我们可以连续拍摄多个细胞群体中的信号格局快照。使用我们最近开发的贝叶斯网络推理算法分析这些流式细胞仪数据将允许前所未有的快速确定自身免疫细胞亚群中的信号事件，并指出新的治疗方式。我们的实验室开发了一套荧光分子探针，可以让我们看到单个细胞对环境的反应和反应。我们将使用这些探针分析从糖尿病小鼠身上提取的细胞，以确定免疫系统对抗胰腺产生胰岛素的细胞的机制。这将为治疗糖尿病的药物提供新的靶点。

DESCRIPTION (provided by applicant): The immune system functions via multiple interactions between many different cell populations. These interactions lead to changes in intracellular protein levels, subcellular localization and activation state, which can have profound effects on both normal and pathological immune system function. Preliminary data has shown that systemic treatment with an anti-CD3 antibody can successfully treat recently hyperglycemic non- obese diabetic mice, resulting in inhibition of beta cell destruction and regeneration of damaged tissue. We hypothesize that the anti-CD3 antibody may act directly on islet infiltrating CD3 lymphocytes, or may activate regulatory T cells. Conventional biochemical techniques are virtually useless in enabling us to look at transient signaling events in rare cell populations in the pancreatic and lymphoid tissues of these mice. With our newly develped 11-color multiparameter flow cytometric analysis for active kinases and phosphoproteins in immune lineages we can take sequential snapshots of the signaling landscape in multiple cell populations. Analysis of this flow cytometry data using our recently developed Bayesian Network Inference algorithms will allow for an unprecedented and rapid determination of the signaling events in autoimmune cell subsets and point to new therapeutic modalities. Our laboratory has developed a set of fluorescent molecular probes that allow us to see how individual cells respond and react to their environment. We will use these probes to analyse cells taken from diabetic mice in order to determine the mechanisms by which the immune system turns against the insulin-producing cells of the pancreas. This will provide information that could point to new targets for drugs to treat diabetes.