Fibrinogen Mechanisms on Perivascular Fibroblast activation after Stroke

瘢痕形成是中枢神经系统(CNS)损伤和疾病的常见反应，也是阻碍CNS功能再生的主要障碍。中枢性非神经性“纤维化”病变核心由基质细胞和中枢神经系统相关巨噬细胞组成，基质细胞局限于血管和脑膜壁龛，包括血管周围巨噬细胞和外周来源巨噬细胞，它们共同产生过量的抑制性细胞外基质成分，如胶原蛋白。先前的研究发现，血管周围成纤维细胞(PVF)是中枢神经系统疾病中“纤维化”的非神经性病变瘢痕的新来源。调控纤维化瘢痕形成的因素，更具体地说，在中枢神经系统疾病中，PVF的激活在很大程度上仍不清楚。我们发现血源性凝血因子纤维蛋白原作为潜在的初级成纤维细胞激活信号具有意想不到的作用，指出纤维蛋白原是血管通透性和纤维性瘢痕形成之间的分子联系。我们将通过利用药理学和遗传学工具抑制/耗尽PVF细胞表面受体来确定纤维蛋白原对PVF激活和迁移影响瘢痕成分的作用，并研究纤维蛋白原诱导的血管周围巨噬细胞激活对PVF激活的影响。我们将通过单细胞RNA测序、RNAScope和免疫组织化学染色来确定纤维蛋白原诱导的PVF分子特性的变化及其与髓系细胞的交叉通讯。最后，我们将操纵纤维蛋白原诱导的PVF激活，以改善大脑修复和功能。总体而言，这项研究将使我们能够为开发新的治疗方法找到新的分子靶点，以促进中风患者的功能恢复，这可能适用于其他中枢神经系统损伤和疾病。

Scar formation is a common response to central nervous system (CNS) injury and disease and a major obstacle blocking CNS functional regeneration. The central non-neural “fibrotic” lesion core is composed of stromal cells, which are restricted to vascular and meningeal niches, and CNS-associated macrophages, including perivascular macrophages and peripherally derived macrophages, which together produce an excess of inhibitory extracellular matrix components, such as collagens. Previous work identified perivascular fibroblasts (PVF) as a novel source of the “fibrotic” non-neural lesion scar in CNS disease. Factors regulating fibrotic scar formation and, more specifically, PVF activation in CNS disease remain largely unknown. We identified an unexpected role for the blood-derived coagulation factor fibrinogen as a potential primary fibroblast activation signal, pointing to fibrinogen as a molecular link between vascular permeability and fibrotic scar formation. We will identify the effect of fibrinogen on PVF activation and migration affecting scar composition by utilizing pharmacologic and genetic tools to inhibit/deplete PVF cell surface receptors and to investigate the effect of fibrinogen-induced perivascular macrophage activation on PVF activation. We will determine the fibrinogen-induced changes in the molecular identity of PVF and their cross-communication with myeloid cells by single-cell RNA-Sequencing, RNAscope and immunohistochemical staining. Finally, we will manipulate fibrinogen-induced PVF activation towards improved brain repair and function. Overall, this study will enable us to pinpoint new molecular targets for the development of novel therapeutic approaches to promote functional recovery in stroke, which might then be applicable for other CNS injuries and diseases.