Myeloid cell regulation of adult lung alveologenesis post-pneumonectomy

 描述(申请人提供)：终末期肺部疾病的唯一可用的治疗方法是全肺移植。这一手术受到存活率低和供体肺供应不足的限制。另一种选择是促进内源性祖细胞再生正常肺组织。部分肺切除术(PNX)，即外科手术切除一个或多个肺叶，在许多哺乳动物物种中刺激剩余肺叶的代偿性肺生长。这种成人肺泡形成的模型是通过几种祖细胞的增殖来介导的，包括肺泡上皮2型细胞(AEC2s)，这是一种远端肺上皮干细胞。关于人类肺的再生潜力、人类肺上皮干细胞的身份、控制其激活的分子信号以及微环境对再生的影响等重大问题仍然存在。我们的目标是应用小鼠研究的数据来刺激人类肺的再生。近年来，巨噬细胞参与了组织修复和再生，但对其在肺再生中的作用知之甚少。我们正在使用荧光记者和小鼠遗传功能的获得和丧失来识别在成人肺再生中调节上皮干细胞行为的免疫细胞群。我们将评估巨噬细胞如何有助于远端肺上皮干细胞的再生生态位，以促进PNX术后成人肺泡形成。我们建议首先测量PNX后的免疫细胞群动态和激活状态(目标1)。我们的初步数据显示，在PNX术后，CD115+髓系细胞在再生肺中增加，这些细胞的一部分共表达精氨酸酶-1，这是M2极化的巨噬细胞的标志，促进伤口愈合。我们将进一步描述PNX术后的髓系细胞群动态，并确定这些细胞是通过局部增殖增加，还是来自循环中的单核细胞。然后我们将测试在PNX诱导的肺再生中对巨噬细胞的需求(目标2)。有趣的是，PNX术后肺上皮细胞中趋化因子CCL2表达上调。我们将破坏上皮细胞或其同源受体CCR2中的CCL2，并确定这是如何损害肺再生的。最后，我们的初步数据表明，PNX后的巨噬细胞分泌可能促进血管生成的因子。我们将研究巨噬细胞是直接影响PNX术后AEC2的增殖，还是通过促进血管生成间接影响AEC2，这是PNX诱导肺再生所必需的(AIM 3)。我们将确定CCR2基因敲除小鼠的内皮细胞增殖是否受损，并确定巨噬细胞特异性的促血管生成途径。我们希望确定调节这些效应的机制，这些机制可能被用来刺激肺再生，并可能被开发成终末期肺部疾病患者的新疗法。

 DESCRIPTION (provided by applicant): The only available treatment for end stage lung diseases is a total lung transplant. This procedure is limited by poor survival and an inadequate supply of donor lungs. An alternative is to promote regeneration of normal lung tissue from endogenous progenitor cells. Partial pneumonectomy (PNX), the surgical removal of one or more lobes, stimulates compensatory lung growth in the remaining lobes in many mammalian species. This model of adult alveologenesis is mediated by proliferation of several progenitor populations, including alveolar epithelial type 2 cells (AEC2s), which are distal lung epithelial stem cells. Significant questions remain regarding the regenerative potential of human lungs, the identities of human lung epithelial stem cells, the molecular signals that control their activation, and the influences of the microenvironment on regeneration. Our objective is to apply data from murine studies to stimulate regeneration of human lungs. Recently macrophages have been implicated in tissue repair and regeneration, but little is know about their role in lung regeneration. We are using fluorescent reporters and genetic gain- and loss-of function in mice to identify populations of immune cells that modulate epithelial stem cell behaviors in adult lung regeneration. We will assess how macrophages may contribute to the regenerative niche of distal lung epithelial stem cells to promote adult alveologenesis post-PNX. We propose to first measure immune cell population dynamics and activation states post-PNX (Aim 1). Our preliminary data shows that that CD115+ myeloid cells increase in regenerating lung post-PNX and a subset of these cells co-express Arginase-1, a marker of M2-polarized macrophages shown previously to promote wound healing. We will further characterize the myeloid population dynamics post-PNX and determine if these cells increase by local proliferation or are derived from circulating monocytes. We will then test the requirement for macrophages in PNX-induced lung regeneration (Aim 2). Interestingly, the chemokine CCL2 is upregulated in lung epithelium post-PNX. We will disrupt CCL2 in the epithelium or its cognate receptor, CCR2, and determine how this impairs lung regeneration. Finally, our preliminary data suggests post-PNX macrophages secrete factors that could promote angiogenesis. We will investigate whether macrophages directly influence AEC2 proliferation post-PNX or if they indirectly influence AEC2 by promoting angiogenesis, which is required for PNX-induced lung regeneration (Aim 3). We will determine if endothelial proliferation is impaired in CCR2 knock-out mice and identify macrophage-specific pro-angiogenic pathways. We hope to identify mechanisms that mediate these effects that might be exploited to stimulate lung regeneration and may be developed into novel therapies for patients with end-stage lung disease.