The Role of Fibrocytes in Liver Fibrosis

描述（由申请人提供）：慢性肝损伤导致肝纤维化，其特征在于细胞外基质蛋白（ECM）的广泛沉积、TGF-β 1和循环内毒素（脂多糖，LPS）水平的增加以及产生胶原的肌成纤维细胞的活化。肝纤维化的发展伴随着骨髓（BM）来源的纤维细胞的募集，所述纤维细胞是表达CD 45和胶原蛋白-α 1（I）的细胞，先前涉及肺、皮肤和肾中纤维化的发病机制。我们已经证明，纤维细胞通过在纤维化肝脏中包含4-6%的胶原蛋白产生细胞并分化成纤维原性肌成纤维细胞而促成肝纤维化。尽管数量较少，但我们的初步数据表明，天然抑制剂血清淀粉样蛋白P（SAP）对纤维细胞功能的抑制导致纤维细胞流入纤维化肝脏的阻断，并显著减弱肝纤维化。目前的建议调查SAP对肝纤维化的影响，纤维细胞募集和分化成肌成纤维细胞，并在肝纤维化过程中使用两种模型，四氯化碳（CCl 4）和胆管结扎（BDL）的炎症信号的调解。我们预计纤维细胞在肝纤维化的发病机制中发挥重要作用（AIM 1）。因此，纤维细胞可能是抗纤维化的有吸引力的靶点。 疗法为了支持我们的假设，我们开发了一种互补的方法来研究纤维细胞在肝纤维化中的作用。利用Cre-loxP重组技术，我们获得了无纤维细胞的转基因小鼠（AIM 2）。体内纤维细胞的消融通过在胶原蛋白-α 1（I）+CD 45+纤维细胞中过表达白喉毒素-α（DTA）或受体（DTR）来实现。纤维细胞的缺失还没有报道，这将为纤维细胞在肝纤维化发病机制中的作用提供宝贵的见解。我们相信，纤维细胞可能成为一个新的抗纤维化治疗的目标。纤维细胞抑制剂SAP用于肺和皮肤纤维化的临床试验，可能为肝纤维化患者提供一种潜在的新疗法。在这里，我们确定了骨髓来源的纤维细胞（造血或间充质）的起源，并确定了它们与共同髓系祖细胞（CMP）和单核细胞（AIM 3）的关系。我们预测纤维细胞起源于c-kit+Sca-1+造血干细胞，但在造血过程中发育先于CMP。一致地，纤维细胞具有高可塑性，并且响应于肝损伤而迁移到纤维化肝，但也迁移到脾，在那里它们介导抗微生物应答。胶原蛋白的表达如何影响纤维细胞的功能尚不清楚。我们通过产生胶原突变型纤维细胞来解决这个问题，这些纤维细胞下调胶原-a1（I）的产生（5 'SL），或表达更稳定的不可切割的Col 1a 1 rr蛋白（AIM 4）。在体内研究突变纤维细胞的功能，并与野生型纤维细胞进行比较。我们预计，突变胶原缺陷的纤维细胞表现出的缺陷，在迁移，分化成肌成纤维细胞，而Col 1a 1 rr突变的纤维细胞预计具有高度纤维化的表型。

DESCRIPTION (provided by applicant): Chronic liver injury results in hepatic fibrosis, which is characterized by extensive deposition of extracellular matrix proteins (ECM), increased levels of TGF-¿1 and circulating endotoxin (lipopolysaccharide, LPS), and activation of collagen producing myofibroblasts. Development of liver fibrosis is accompanied by recruitment of bone marrow (BM)-derived fibrocytes, that are CD45 and collagen-a1(I) expressing cells, previously implicated in the pathogenesis of fibrosis in lungs, skin, and kidneys. We have demonstrated that fibrocytes contribute to liver fibrosis by comprising 4-6% of collagen producing cells in the fibrotic liver and differentiating into fibrogenic myofibroblasts. Despite of low numbers, our preliminary data suggest that inhibition of fibrocyte function(s) by the natural inhibitor Serum Amyloid P (SAP), results in the blockage of fibrocyte flux into fibrotic liver, and significantly attenuates liver fibrosis. Current proposal investigates the effect of SAP on liver fibrosis, fibrocyte recruitment and differentiation into myofibroblasts, and mediation of inflammatory signals in the course of liver fibrosis using two models of liver injury, carbontetrachloride (CCl4 and bile duct ligation (BDL). We anticipate that fibrocytes play an important role in pathogenesis of liver fibrosis (AIM 1). Therefore, fibrocytes may present an attractive target for anti-fibrotic therapy. To support our hypothesis, we developed a complimentary approach to study the role of fibrocytes in liver fibrosis. Using Cre-loxP recombination technique, we generate transgenic mice devoid of fibrocytes (AIM 2). Ablation of fibrocytes in vivo is achieved by overexpression of Diphtheria toxin-a (DTA) or receptor (DTR) in Collagen-a1(I)+CD45+ fibrocytes. Deletion of fibrocytes has not been reported, and will provide the invaluable insight into the role of fibrocytes in pathogenesis of liver fibrosis. We believe that fibrocytes may become a novel target for anti-fibrotic therapy. Futhermore, fibrocyte inhibitor SAP, which is used in clinical trals for lung and skin fibrosis, may present a potentially new treatment for patients with liver fibrosi. Here we determine the origin of BM-derived fibrocytes (hematopoietic or mesenchymal), and define their relationship to common myeloid progenitors (CMP) and monocytes (AIM 3). We predict that fibrocytes originate from c-kit+Sca-1+ hematopoietic stem cells, but developmentally precede CMP during hematopoiesis. In concordance, fibrocytes possess high plasticity, and in response to liver injury migrate to fibrotic liver, but also to spleen where they mediate antimicrobial response. It remains unclear how expression of collagen affects functions of fibrocytes. We address this question by generation of collagen-mutant fibrocytes, which downregulate collagen-a1(I) production (5'SL), or express a more stable uncleavable Col1a1rr protein (AIM 4). The functions of mutant fibrocytes are studied in vivo and compared to the wild type fibrocytes. We expect that mutant collagen-deficient fibrocytes exhibit a defect in transmigration, differentiation into myofibroblasts, while Col1a1rr-mutant fibrocytes are expected to possess a highly fibrogenic phenotype.