The role of transport proteins for the renal homoeostasis of arginine derivatives

肾脏的近端小管细胞能够区分结构和化学性质非常相似的物质，从而导致尿液或血浆中的选择性蓄积或消耗。对于许多物质的基本分子机制的研究和理解很少，到目前为止。在我们实验室进行的初步研究表明，身体所需的物质，如半必需氨基酸L-精氨酸或其衍生物高精氨酸在肾脏中被净重吸收，而结构非常相似的尿毒症毒素不对称和对称二甲基精氨酸（ADMA和SDMA）似乎没有被定量重吸收。在慢性肾病患者中，ADMA和SDMA蓄积，而血浆高精氨酸浓度降低。ADMA和SDMA血浆浓度升高以及高精氨酸血浆浓度降低与总死亡率增加相关。据推测，特别是位于近曲小管细胞的不同膜域的转运蛋白负责选择性重吸收或分泌。特别是在具有相反生物功能的化学相似物质的情况下，更好地理解潜在机制可以帮助寻找新的治疗方法以及理解药物的肾脏介导的副作用。该项目的目的是研究L-精氨酸及其衍生物的细胞摄取和载体转运，重组表达近曲小管细胞中表达的转运蛋白的转染细胞模型。此外，我们的目标是分析这些转运过程是否可以被药物修饰，以确定潜在的治疗用途的影响以及介导药物不良反应的新机制。通过应用有针对性的代谢组学和“代谢通量”分析条件永生化近端肾小管细胞（ciPTEC细胞），培养在一个3D细胞模型，我们进一步的目的是分析运输和代谢的相互作用，在肾脏处理不同的L-精氨酸衍生物。将在人肾活检样本中进行参与转运和代谢的相关蛋白的mRNA和蛋白表达分析，以研究慢性肾脏疾病进展期间的表达变化。这些数据也可能有助于确定慢性肾脏疾病患者血浆高精氨酸反常下降的机制。总之，所获得的数据预计将允许识别肾脏对化学相似物质进行差异处理所涉及的关键机制。此外，它的目的是获得数据，看看这些过程是否可以通过药物以潜在有益的方式进行改变，这可能对慢性肾病患者的治疗产生影响。

Proximal tubule cells of the kidney are able to discriminate between structurally and chemically very similar substances leading to selective accumulation or depletion in urine or plasma. For many substances the underlying molecular mechanisms are poorly investigated and understood, so far. Preliminary investigations performed in our laboratory demonstrated, that substances necessary for the body such as the semiessential amino acid L-arginine or its derivative homoarginine are net reabsorbed in the kidney whereas the structurally very similar uremic toxins asymmetric and symmetric dimethylarginine (ADMA and SDMA) appear not to be reabsorbed in a quantitative manner. In Patients with chronic kidney disease ADMA and SDMA accumulate whereas the plasma concentration of homoarginine decreases. Elevated ADMA and SDMA plasma concentrations as well as decreased plasma concentrations of homoarginine are associated with an increased total mortality. It is assumed that especially transport proteins located in the different membrane domains of proximal tubule cells are responsible for the selective reabsorption or secretion. Especially in the case of of chemically similar substances with opposing biological function a better understanding of the underlying mechanisms could aid the search for new therapeutic approaches as well as for the understanding of renally mediated side effects of drugs.Therefore, it is the aim of the proposed project to study the cellular uptake and the vectorial transport of L-arginine and its derivatives using single- and multiple-transfected cell models with recombinant expression of transport proteins expressed in proximal tubule cells. In addition, we aim to analyze whether these transport processes can be modified by drugs, in order to identify effects of potential therapeutic use as well as new mechanisms mediating adverse drug effects. By application of targeted metabolomics and “metabolic-flux” analyses in conditionally immortalized proximal tubular cells (ciPTEC cells), cultured in a 3D cell model, we further aim to analyze the interplay of transport and metabolism in the renal handling of different L-arginine derivatives. Analyses of mRNA and protein expression of relevant proteins involved in transport and metabolism will be conducted in human kidney biopsy samples to study changes in expression during the progression of chronic kidney diseases. These data may also help to identify the mechanisms underlying the paradox decline in plasma homoarginine observed in patients with chronic kidney disease. Taken together, the data obtained are expected to allow the identification of key mechanisms involved in the differential handling of chemically similar substances by the kidney. Moreover, it is aimed to gain data whether these processes can be modified by drugs in a potentially beneficial way, which may have implications for the treatment of patients with chronic kidney disease.