Role of Amphiregulin in kidney fibrosis

项目摘要 我们研究的总体目标是开发新的治疗和检测/监测策略，以预防 慢性肾病（CKD）和纤维化的进展。缺血再灌注所致急性肾损伤（阿基） 损伤（IRI）或单侧输尿管梗阻（UUO）诱导持续的EGFR活化和肾纤维化 其通过EGFR化学抑制或其在近端小管细胞（PTC）中的基因缺失来预防。的 涉及的特异性EGFR配体仍然未知。我们发现A-去整合素和金属蛋白酶-17 （ADAM 17）近端小管敲除可防止损伤诱导的纤维化，确定促纤维化的来源 PTC的EGFR配体。在损伤的小鼠肾脏中，EGFR配体前双调蛋白（pro-AREG）非常高， 在阿基和CKD患者的尿中sAREG被强烈上调并且在人类中非常显著地升高， 以及纤维化CKD肾活检。在体外，在人近端小管细胞（HPTC）中，sAREG诱导的 持续的EGFR活化和促纤维化靶点。（Kefalogianni JCI Insight 2016）.的目的 应用的目的是确定sAREG在阿基后早期与晚期损伤修复阶段的作用， 人患者样品中的sAREG与肾纤维化和CKD进展的关系。我们的核心假设是， 双调蛋白（sAREG）是表皮生长因子受体（EGFR）的关键配体， 以及在肾损伤中放大促纤维化EGFR信号。我们基于以下初步数据：（1）sAREG 基于sAREG注射和PTC-KO， （2）在患有肾切除术的患者队列的血清样品中，sAREG显著升高， CKD及其水平与肾功能参数呈负相关。该项目的基本原理是， 完成后将确定sAREG作为CKD/纤维化的新型治疗靶点和生物标志物，并确定 涉及的分子机制。我们计划通过两个具体目标来测试我们的中心假设： AREG是否是IRI损伤后早期肾恢复和/或肾纤维化发展所必需的。 目的2：在波士顿肾活检中将sAREG与人类肾纤维化程度和CKD分期联系起来 队列（BKBC，n=770）和慢性肾功能不全队列（CRIC，n=3889）中的CKD进展。这里 我们将sAREG血清/尿液水平与活检时的肾纤维化和CKD进展相关联，并比较 与目前使用的肾脏生物标志物（肌酐、蛋白尿）相关。作为结果，我们希望AREG 近端小管基因敲除可防止损伤诱导的纤维化，sAREG水平与CKD相关 结果和性能优于目前的生物标志物（肌酐，蛋白尿）。这一贡献意义重大 因为它预计将对CKD/纤维化的检测、预防和治疗产生影响。我们的研究是 在我们看来，这是一项创新，因为它将首次确定一种肾纤维化生物标志物， 机械地连接到纤维化过程，并且可以用于治疗靶向和监测。

PROJECT SUMMARY ABSTRACT The overall goal of our research is to develop novel therapeutic and detection/monitoring strategies to prevent progression of chronic kidney disease (CKD) and fibrosis. Acute kidney injury (AKI) due to ischemia reperfusion injury (IRI) or unilateral ureteral obstruction (UUO) in mice induces sustained EGFR activation and kidney fibrosis which is prevented by EGFR chemical inhibition or its genetic deletion in proximal tubule cells (PTCs). The specific EGFR ligand(s) involved remain unknown. We showed that A-disintegrin-and-metalloprotease-17 (ADAM17) proximal tubule knockout protects against injury-induced fibrosis, identifying the source of pro-fibrotic EGFR ligands to PTCs. In the injured mouse kidney, the EGFR ligand pro-Amphiregulin (pro-AREG) was very strongly upregulated and in humans sAREG was very significantly elevated in the urine of AKI and CKD patients, as well as fibrotic CKD kidney biopsies. In vitro, in human proximal tubule cells (HPTCs), sAREG induced sustained EGFR activation and pro-fibrotic targets. (Kefalogianni JCI Insight 2016). The objective of this application is to determine the role of sAREG in early vs. late injury-repair stages after AKI and to firmly link sAREG in human patient samples to kidney fibrosis and to CKD progression. Our central hypothesis is that Amphiregulin (sAREG) is the key epidermal growth factor receptor (EGFR) ligand that is responsible for inducing and amplifying pro-fibrotic EGFR signals in kidney injury. We base this on preliminary data including: (1) sAREG is sufficient and necessary to induce fibrosis after kidney injury in mice, based on sAREG injection and PTC-KO studies in mice (2) sAREG is significantly elevated in serum samples of a nephrectomy cohort of patients with CKD and its levels correlate negatively with kidney function parameters. The rationale for this project is that completion will identify sAREG as a novel therapeutic target and biomarker in CKD/fibrosis, and identify molecular mechanisms involved. We plan to test our central hypothesis with two specific aims: AIM1: Determine whether AREG is necessary for early renal recovery and/or the development of kidney fibrosis after IRI injury. AIM2: To link sAREG to degree of kidney fibrosis and stage of CKD in humans in the Boston Kidney Biopsy Cohort (BKBC, n=770) and to CKD progression in the Chronic-Renal-Insufficiency-Cohort (CRIC, n=3889). Here we will correlate sAREG serum/urine levels to kidney fibrosis on biopsy and CKD progression, and compare this to correlation with currently used kidney biomarkers (Creatinine, proteinuria). As outcomes, we expect that AREG proximal tubule knockout protects against injury-induced fibrosis and that sAREG levels correlate with CKD outcomes and perform better than current biomarkers (creatinine, proteinuria). This contribution is significant because it is expected to have impact on CKD/fibrosis detection, prevention and treatment. Our research is innovative, in our opinion, because, it would for the first time identify a kidney fibrosis biomarker that is directly mechanistically connected to the fibrotic process and could be used for therapeutic targeting and monitoring.