Novel mechanisms involving complement cascade in stem cell trafficking

修改后的项目摘要/摘要部分 这种竞争性更新的主要假设是基于新的令人兴奋的数据，这些数据进一步支持了 补体级联(Comc)等天然免疫元件在动员补体中的作用 造血干/祖细胞(HSPC)。COMC由i)经典的、ii)甘露聚糖结合所激活 凝集素(MBL)，和III)替代途径，以及，正如我们已经报道的，缺乏共同末端的小鼠 Comc(C5-/-小鼠)的成分，是较差的动员因子。然而，令我们惊讶的是，我们发现老鼠 不要激活经典的COMC(C1q-/-小鼠)是正常的激动剂。这种“差异”现在被解释为 我们的最新结果表明，MBL依赖的COMC途径而不是经典的COMC途径是关键的参与 动员中的途径。我们最近还发现了两种应激诱导酶，血红素加氧酶1(HO-1)。 和诱导型一氧化氮合酶(INOS)，是细胞迁移的负性调节因子。基于这些发现， 我们已经准备了一份提案，以一种全面的方式，解决了先天因素的参与 HSPC动员中的免疫(Comc，Gr-1粒细胞/单核细胞，自然产生的抗体)。 在三个相互关联的具体目标中，我们解决了关键的：i)启动、ii)扩大和执行阶段 动员。具体目的1.用MBL介导的Comc解释动员的启动阶段 激活。我们认为，经典的Comc激活途径在触发 动员。然而，正如我们最近发现的那样，并不是经典的而是MBL途径在 这一过程。动员剂的主要靶细胞是Gr-1细胞，除了蛋白水解物和 脂解酶，也分泌ROS和危险相关的分子模式分子(DAMP)。而当 蛋白水解酶和脂解酶扰乱了HSPC在ROS暴露的骨髓(BM)壁龛中的保留 与自然产生的IgM抗体和Gr-1细胞释放的DAMs结合的新表位抗原是 由循环模式识别受体(PRR)MBL识别，通过甘露聚糖结合凝集素丝氨酸 蛋白水解酶(MASP)，激活COMC和凝血级联(COAC)。具体目标2.扩增 动员阶段--C5转化酶的产生。由于C5裂解对HSPC的排出至关重要 从BM开始，这个阶段的主要功能是产生C5转换酶活性。这种酶的产生方式是 C3裂解的结果，它提供了经典C5转换酶的成分，另外还通过凝血酶， 具有C5“转化酶样活性”。放大相位也由自发光调制 C3激活的扩增环以及可能参与COMC激活的另一条途径 并将在这一目标上进行详细的研究。具体目标3.通过COMC进行动员的执行阶段。 我们最近的研究结果表明，HSPC从骨髓中流出受到HO-1和iNOS的负调控，这两种基因分别是 细胞迁移的负性调节因子。我们将重点研究这一现象的分子机制。它是 预期这些成果将转化为制定更好的动员方案。 1

Modified Project Summary/Abstract Section The main hypothesis of this competing renewal is based on new exciting data that further supports the pivotal role of the complement cascade (ComC) and other elements of innate immunity in the mobilization of hematopoietic stem/progenitor cells (HSPCs). The ComC is activated by the i) classical, ii) mannan-binding lectin (Mbl), and iii) alternative pathways, and, as we already reported, mice that lack the common distal component of the ComC (C5–/– mice), are poor mobilizers. To our surprise, however, we found that mice that do not activate the classical ComC (C1q–/– mice) are normal mobilizers. This “discrepancy” is now explained by our latest results that show the pivotal involvement of the Mbl-dependent ComC pathway and not the classical pathway in mobilization. We also recently found that two stress-induced enzymes, heme oxygenase 1 (HO-1) and inducible nitric oxide synthetase (iNOS), are negative regulators of cell migration. Based on these findings, we have assembled a proposal that, in a comprehensive way, addresses the involvement of elements of innate immunity (ComC, Gr-1+ granulocytes/monocytes, naturally occurring antibodies) in the mobilization of HSPCs. In three interrelated specific aims, we address the crucial: i) inititation, ii) amplification, and execution phases of mobilization. Specific Aim 1. The initiation phase of mobilization is explained by Mbl-mediated ComC activation. We have maintained that the classical ComC activation pathway plays a crucial role in triggering mobilization. However, as we discovered recently, it is not the classical but the Mbl pathway that is crucial in this process. The main target cells for mobilizing agents are Gr-1+ cells, which, in addition to proteolytic and lipolytic enzymes, also secrete ROS and danger-associated molecular pattern molecules (DAMPs). While proteolyic and lipolytic enzymes perturb the retention of HSPCs in bone marrow (BM) niches, ROS-exposed neoepitope antigens bound by naturally occurring IgM antibodies and DAMPs released from Gr-1+ cells are recognized by the circulating pattern recognition receptor (PRR) Mbl, which, via mannan-binding lectin serine proteases (MASPs), activates the ComC and coagulation cascade (CoaC). Specific Aim 2. The amplification phase of mobilization - the generation of C5 convertase. Since C5 cleavage is crucial for egress of HSPCs from BM, the main function of this phase is generation of C5 convertase activity. This enzyme is generated as a result of C3 cleavage, which provides elements of classical C5 convertase, and additionally by thrombin, which has C5 “convertase-like” activity. The amplification phase is also modulated by a spontaneous amplification loop of C3 activation and by possible involvement of an alternative pathway of ComC activation and will be studied in detail in this aim. Specific Aim 3. Execution phase of ComC-mediated mobilization. Our recent results show that egress of HSPCs from BM is negatively controlled by HO-1 and iNOS, which are negative regulators of cell migration. We will focus on the molecular mechanisms of this phenomenon. It is expected that these results will be translated into the development of better mobilization protocols. 1