The role of cilia in inflammasome activation and their role in polycystic kidney disease

主要的肾脏纤毛病变是成人常染色体显性多囊肾病（ADPKD）和儿童隐性肾单位营养不良。这些实体在囊肿的数量和大小方面不同，但共享间质性炎症和纤维化。在ADPKD中，炎症驱动囊肿生长，在这两种情况下，纤维化导致肾功能丧失并导致终末期肾病。我们的小组正在研究ADPKD中哪些免疫途径失调及其细胞和分子机制。我们已经发现肾小管上皮细胞的纤毛调节肾脏炎症。这是由激酶LKB 1的纤毛复合物以及多囊蛋白1（PC 1）和NPHP 1（ADPKD和NPH中最常突变的基因产物）协调的。在肾脏蛋白质组学筛选中，我们鉴定了激酶NEK 7作为LKB 1的相互作用物，并发现NEK 7在功能上与LKB 1/PC 1/NPHP 1复合物相互作用。NEK 7是一种重要的调节剂的类受体NLRP 3，并发挥了至关重要的作用，在激活炎性小体。这些大复合物是先天免疫系统的一部分，对危险分子作出反应，并引起半胱天冬酶介导的白细胞介素-10和-18的活化、细胞损伤和炎症。在小鼠中早期ADPKD中的表达筛选揭示了炎性小体相关转录物如半胱天冬酶1和IL-18的上调。在蛋白质水平上，在早期ADPKD中观察到半胱天冬酶1的活化，并且其定位于早期囊肿。MDCK细胞表达所有炎性体组分，并在刺激时显示NEK 7依赖性IL-18释放。我们假设LKB 1、PC 1、NPHP 1和NEK 7的纤毛复合物调节肾上皮细胞中的炎性小体激活。PC 1的缺乏导致炎性小体活化和肾脏炎症。在目标1中，我们将研究睫状体炎性小体调节的细胞和分子机制。在目标2中，我们将使用遗传动物模型来测试通过草酸盐晶体激活炎性小体是否会加剧表型，或者通过ASC敲除系统性灭活炎性小体是否会改善疾病。在目标3中，我们将在小鼠的ADPKD中使用转基因策略，以区分免疫细胞与肾上皮中炎性小体的作用。在目标4中，我们将用炎性小体抑制剂治疗ADPKD小鼠，以确定该途径中的药物干预是否改善疾病。

The main renal ciliopathies are autosomal dominant polycystic kidney disease (ADPKD) in adults and recessive nephronophthisis in children. These entities differ with respect to number and size of cysts but share interstitial inflammation and fibrosis. In ADPKD inflammation drives cyst growth and in both fibrosis causes loss of renal function and leads to end stage renal disease. Our group is working on which immunological pathways are deregulated in ADPKD and the cellular and molecular mechanisms thereof. We have found that cilia of renal tubular epithelial cells regulate renal inflammation. This is orchestrated by a ciliary complex of the kinase LKB1, together with polycystin 1 (PC1) and NPHP1, the gene products most commonly mutated in ADPKD and NPH. In a proteomic screen from kidney we identified the kinase NEK7 as an interactor of LKB1 and find that NEK7 functionally interacts with the LKB1/PC1/NPHP1 complex. NEK7 is an essential regulator of the nod-like-receptor NLRP3 and plays a crucial role in the activation of inflammasomes. These macro-complexes are part of the innate immune system, responding to danger molecules and causing caspase mediated activation of the interleukins – 1ß and -18, cell injury and inflammation. Expression screens in early ADPKD in the mouse reveal upregulation of inflammasome associated transscripts such as caspase 1 and IL-18. On the protein level activation of caspase 1 is seen in early ADPKD and is localized to early cysts. MDCK cells express all inflammasome components and reveal NEK7 dependent IL-18 release upon stimulation. We hypothesize that a ciliary complex of LKB1, PC1, NPHP1 and NEK7 regulates inflammasome activation in renal epithelial cells. The lack of PC1 leads to inflammsome activation and renal inflammation. In aim 1 we will examine the cellular and molecular mechanisms underlying ciliary inflammasome regulation. In aim 2 we will use genetic animal models to test if activation of inflammasomes through oxalate crystals will aggravate the phenotype, or if systemic inactivation of inflammasomes through ASC knock-out will ameliorate the disease. In aim 3 we will use a transgenic strategy in ADPKD in the mouse to differentiate between the role of inflammasomes in immune cells vs. renal epithelia. In aim 4 we will treat ADPKD mice with an inflammasome inhibitor to ascertain if pharmacological intervention in this pathway ameliorates the disease.