Skin-Intrinsic Immunosuppressive Mesenchymal Stem Cells and aGVHD

项目摘要/摘要 急性移植物抗宿主病(Agvhd)是异基因干细胞的主要限制性并发症。 移植治疗白血病/淋巴瘤和某些实体肿瘤。AGVHD发生在T 供者骨髓中的淋巴细胞攻击受者体内对至关重要的功能至关重要的细胞 器官，即皮肤、肝脏和肠道。在过去30年的研究中，墨菲博士的实验室 研究发现，在皮肤和鳞状粘膜中，效应器-靶细胞相互作用的第一个部位由 次要组织相容性抗原的差异，涉及携带VB特异性标记的T细胞 选择性地对小鼠毛囊的上皮网脊以及富含上皮干细胞的隆起区域进行检测。 这种关联通过表达干细胞标记物的相邻上皮细胞的凋亡而导致死亡， 细胞角蛋白15.对这些效应器肢体的全身治疗抑制充满了问题，包括 对免疫重建和钝化移植物抗肿瘤效应的间接影响。然而，我们现在知道 通过与该提案的合作PI马库斯博士和娜塔莎·弗兰克博士合作的发现， 皮肤和鳞状粘膜含有能够有效抑制免疫的固有细胞通路。 具体地说，我们发现了一种具有干细胞特征的真皮树突状细胞，表达 多药耐药转运蛋白ABCB5，也表达PD-1。这个细胞具有抑制T细胞的能力 反应，包括同种异体反应性，因此像皮肤一样具有重大但尚未被探索的前景- 内源性途径可能被用来抑制aGVHD中的组织损伤。在这项提案中，墨菲博士和 弗兰克博士联合起来，致力于解决利用皮肤固有的 免疫抑制能力，以阻止移植物抗宿主病。该提案的具体目标是确定影响 表达PD-1免疫抑制的真皮间充质干细胞对最早和最特异的影响 效应器-靶细胞相互作用导致移植物抗宿主病。为此，我们提出了三个具体目标： 1)将时空多重免疫图谱与疾病发病率和恢复率的各个方面联系起来 在aGVHD中；2)剖析aGVHD中ABCB5皮肤干细胞的功能并确定PD-1/PD-L1为中心 皮肤内源性免疫抑制策略；3)应用ABCB5+皮肤干细胞移植 以时空多重免疫模式作为效应器-靶点影响aGVHD严重程度的策略 生物化验。这些方法包括最先进的效应-靶细胞的多重免疫表型 敏感和特异地代表aGVHD中组织损伤的相互作用，并利用ABCB5基因敲除 以及PD-L1免疫球蛋白注射和ABCB5+干细胞植入背景下的谱系追踪模型。这个 基本原理是，在这项工作成功完成后，我们将确定新的治疗方法 运用皮肤手法治疗和预防aGVHD皮肤和鳞状黏膜病变 内源性免疫抑制途径。

Project Summary/Abstract Acute graft-versus-host disease (aGVHD) is a major limiting complication of allogeneic stem cell transplantation for the treatment of leukemia/lymphoma and certain solid tumors. aGVHD occurs when T lymphocytes from donor bone marrow attack cells within the recipient that are critical to the function vital organs, namely skin, liver and gut. During the past three decades of research, Dr. Murphy’s laboratory has found that in skin and squamous mucosae, the first sites of effector-target cell interaction provoked by differences in minor histocompatibility antigens, involve T cells bearing Vb-specific markers that home selectively to epithelial rete ridges as well as the epithelial stem cell-rich bulge regions of murine hair follicles. This association results in death by apoptosis of adjacent epithelial cells that express the stem cell marker, cytokeratin 15. Systemic therapeutic inhibition of these effector limbs is riddled with problems that include collateral impact on immune reconstitution and blunting of graft-versus-tumor effects. However, we now know through discoveries made in collaboration with co-PIs of this proposal, Drs. Markus and Natasha Frank, that the skin and squamous mucosae harbor intrinsic cellular pathways capable of potent immunosuppression. Specifically, we have discovered a dermal dendritic cell that has stem cell characteristics, expresses the multidrug resistance transporter ABCB5, and also expresses PD-1. This cell has the ability to thwart T cell responses, including alloreactivity, and therefore holds significant but as yet unexplored promise as a skin- intrinsic pathway that may be leveraged to inhibit the tissue injury in aGVHD. In this proposal, Dr. Murphy and the Drs. Frank have combined forces to address the long-term objective of harnessing the skin’s intrinsic immunosuppressive capabilities to thwart aGVHD. The proposal’s specific objective is to determine the impact of PD-1-expressing immunosuppressive dermal mesenchymal stem cells on the earliest and most specific effector-target cell interactions responsible for aGVHD. To this end, we have proposed three specific aims: 1) to correlate spatiotemporal multiplex immune profiling with all aspects of disease morbidity and recovery in aGVHD; 2) to dissect ABCB5 dermal stem cell function in aGVHD and define PD-1/PD-L1-centered strategies for skin-intrinsic immunosuppression; and 3) to apply ABCB5+ dermal stem cell transplant strategies to influence aGVHD severity using spatiotemporal multiplex immune profiling as effector-target bioassay. The approaches involve state-of-the-art multiplex immunophenotyping of effector-target cell interactions that sensitively and specifically typify tissue injury in aGVHD, and leverage ABCB5 gene knockout and lineage tracing models in context of PD-L1 Ig administration and ABCB5+ stem cell engraftment. The rationale is that upon successful completion of this work, we will identify novel therapeutic approaches to the treatment and prevention of aGVHD skin and squamous mucosal pathology through manipulation of skin- intrinsic immunosuppressive pathways.