Fibrocyte Contribution to Systemic Fibrosis in Chronic Kidney Disease

暴露于基于Gd的磁共振成像(MRI)对比剂是导致 获得性肾源性系统性纤维化(NSF)，这是一种首次在患者中发现的严重衰弱疾病 有急性或慢性肾功能损害。到目前为止，核磁共振造影剂在这些患者中被认为是安全的。 因此不分青红皂白地使用。基于Gd的造影剂仍然是磁共振成像和 目前仍缺乏其他选择。申请人的初步数据显示，MRI对比剂引起直接的增生性病变 体外细胞外基质合成增加对人成纤维细胞的影响。肾衰大鼠 核磁共振成像检查显示皮肤明显异常，包括骨吸收。 骨髓来源的细胞。MRI对比剂处理的小鼠同样表现为皮肤纤维化，真皮细胞增多。 此外，在受影响的大鼠和小鼠皮肤以及其他器官(如 肾脏和肝脏)来自MRI对比剂处理的动物。这项提议的总体目标是 确定暴露于空气中后触发纤维化异常模式的候选机制 肾功能不全背景下基于Gd的mri对比研究。中心假设是，在 设置肾功能不全，MRI对比剂暴露会触发靶器官的促纤维化状态，如 皮肤。后续产生的特异性趋化因子激活和招募骨髓源性和循环 间充质前体细胞，或纤维细胞，到受影响的区域。这些细胞使纤维化过程复杂化。 通过合成和减少细胞外基质的产生，诱导细胞的增殖 细胞。实验将解决以下具体目标：1.确定 基于Gd的对比剂会导致NSF。申请人已证明骨髓来源的细胞 约占皮损真皮细胞的44%。皮肤中的Gd沉积是否会引起 促纤维化细胞因子的产生，导致纤维细胞募集或如果在其他 目前尚不清楚骨髓等部位是否对成纤维细胞前体细胞进行编程以磨练到特定部位。这个 申请者将通过将骨髓供体(转基因、标记的)大鼠暴露在核磁共振中来进行测试 进行对比，然后将这些骨髓移植到受到致命性辐射的动物体内。1B：证明 基于Gd的造影剂会导致循环中纤维细胞的增加。磁共振成像对比度的影响 大鼠在特定时间点的循环纤维细胞数量将被量化。2a：确定 趋化因子和受体负责向特定组织募集骨髓来源的细胞。 假设单核细胞趋化蛋白-(MCP-)1/趋化因子受体2(CCR2)轴 在MRI造影剂诱导的系统性纤维化中调节纤维细胞的募集。化学媒介物 导致循环细胞募集到NSF皮损的机制尚未得到很好的研究。对NSF的抵抗力将是 在具有遗传缺陷的动物身上进行检查，将探索CCR2。这种蛋白质的作用将得到验证 通过将CCR2缺陷的骨髓移植到正常骨髓的骨髓移植实验 (野生型)小鼠，反之亦然。2B：检验Gd的生物分布不同的假设 在小鼠和大鼠肾功能正常和肾功能不全之间。即使之前的出版物 明确地表明，肝脏是MRI对比剂治疗的大鼠和小鼠中Gd的主要储存库， 皮肤总是与NSF有关，而且在实验动物中比任何肝脏病理都要严重得多。 申请人发现的日期。如果Gd的沉积是引发这种疾病的核心，也许 Gd在肾功能不全和肾功能不全状态下的生物分布不同。Gd NSF中可能受影响的器官(皮肤、肺、心脏、横隔膜、肝脏、脾、骨骼、肌肉)中的蓄积 将在肾功能不全(肾次全切除)的动物和假治疗的对照组中进行比较。这个 研究结果将有助于更好地理解系统性纤维化是如何发生的，以及为什么某些器官是靶向的。

Exposure to gadolinium-based magnetic resonance imaging (MRI) contrast is a major risk factor in the acquisition of nephrogenic systemic fibrosis (NSF), a severely debilitating disease first recognized in patients with acute or chronic renal impairment. MRI contrast agents heretofore were considered safe in these patients and thus indiscriminately used. Gadolinium-based contrast agents remain a mainstay for MR imaging and alternatives are still lacking. The applicant's preliminary data show that MRI contrast elicits a direct proliferative effect on human fibroblasts with an increase in extracellular matrix synthesis in vitro. Rats with renal insufficiency exposed to MRI contrast demonstrate significant skin abnormalities, including recruitment of bone marrow-derived cells. MRI contrast-treated mice similarly demonstrate skin fibrosis with dermal hypercellularity. Furthermore, gadolinium has been detected in affected rat and mouse skin as well as other organs (such as the kidneys and livers) from MRI contrast-treated animals. The overall objective of this proposal is to identify candidate mechanisms that trigger the aberrant pattern of fibrosis after exposure to gadolinium-based MRI contrast in the setting of renal insufficiency. The central hypothesis is that in the setting of renal insufficiency, MRI contrast exposure triggers a pro-fibrotic state in target organs, such as the skin. Subsequent generation of specific chemokines activate and recruit bone marrow-derived and circulating mesenchymal precursor cells, or fibrocytes, to the affected areas. These cells compound the fibrotic process by synthesizing and reducing the generation of extracellular matrix and inducing the proliferation of resident cells. Experiments will address the following Specific Objectives: 1a: Determine the mechanism by which gadolinium-based contrast induces NSF. The applicant has proven that bone marrow-derived cells constitute around 44% of the dermal cellularity in skin lesions. Whether gadolinium deposition in the skin elicits the generation of pro-fibrotic cytokines that lead to fibrocyte recruitment or if gadolinium stimulation in other sites, such as the bone marrow, programs fibroblast precursors to hone to specific sites is unknown. The applicant will test this by "priming" bone marrow donor (transgenic, "tagged") rats with exposure to MRI contrast and then transplanting this bone marrow into lethally-irradiated animals. 1b: Demonstrate that gadolinium-based contrast induces an increase in circulating fibrocytes. The effect of MRI contrast on the number of circulating fibrocytes at specific time points will be quantified in rats. 2a: Determine the chemokines and receptors responsible for recruitment of bone marrow-derived cells to specific tissues. The hypothesis is that the monocyte chemoattractant protein- (MCP-) 1/chemokine receptor 2 (CCR2) axis modulates the recruitment of fibrocytes in MRI contrast-induced systemic fibrosis. The chemical mediators that lead to the recruitment of circulating cells to NSF lesions is not well-explored. Resistance to NSF will be examined in animals with a genetic deficiency CCR2 will be explored. The role of this protein will be validated by using bone marrow transplant experiments where CCR2-deficient bone marrow is transplanted to normal (wild-type) mice and vice-versa. 2b: Test the hypothesis that biodistribution of gadolinium differs between normal renal function and renal insufficiency in mice and rats. Even though prior publications definitively indicate that the liver is a major reservoir for gadolinium in MRI-contrast treated rats and mice, the skin is invariably involved in NSF and to a far greater extent than any liver pathology in experimental animals found to date by the applicant. If the deposition of gadolinium is the nidus for triggering the disease, perhaps biodistribution of gadolinium differs between the states of renal sufficiency and insufficiency. Gadolinium accumulation in organs potentially affected in NSF (skin, lungs, heart, diaphragm, liver, spleen, bone, muscle) will be compared in animals with renal insufficiency (subtotal nephrectomies) and sham-treated controls. The findings will lead to a better understanding of how systemic fibrosis occurs and why certain organs are targeted.