Understanding the molecular basis of the peritoneal solute transport to improve peritoneal dialysis efficacy and outcome

点击翻译按钮获取中文摘要

Peritoneal dialysis (PD) is a life-saving, cost-effective therapy for an increasing number of patients with end stage chronic kidney disease (CKD5). PD takes advantage of the peritoneum as a semipermeable membrane to remove toxins and water from the patient. The limited purification capacity of PD and the nonphysiological composition of PD fluids, however, compromise technique and patient survival. The molecular structures defining peritoneal solute and water transport are largely unknown.We have established a large biobank of peritoneal and omental tissues providing information on the age related anatomy of the healthy peritoneum, on the PD induced peritoneal transformation process and on associated systemic vasculopathy. The present project aims at a comprehensive understanding of molecular mechanisms of peritoneal solute and water transport across the endothelial and mesothelial cell barrier, which essentially define PD membrane function and their impact on local and systemic vasculopathy. Based on previous studies we identified potential molecular candidates, i.e. junction proteins, which are key components of adjacent epithelial and endothelial cell barriers, regulating sealing and paracellular transport functions. In an untargeted human ex vivo approach we will perform whole exome and proteome analyses of the peritoneum and of omental arterioles from non-CKD patients, from patients with CKD5 and on PD, and from PD patients, who recently underwent renal transplantation and standardized immunosuppression. Key findings will be validated in independent patient cohorts, and related to known pathomechanisms of peritoneal inflammation, angiogenesis and fibrosis, to vasculopathy and PD transport function. In Transwell/Ussing chamber systems human mesothelial cell and endothelial cell monolayers will undergo transepithelial resistance and molecular weight dependent transport measurements of sodium, water and 4-70kDa molecules together with junction abundance and localization studies. Different PD fluid types, respective fluid components, reactive metabolites and well-described uremic toxins as well as established and in silico identified junction modulators and medications will be investigated. CRISPR/Cas9 gene knockout studies will define the functional impact of key junction components. Super resolution microscopy will demonstrate the co-localization of specific junction components and the junction-cytoskeleton-interaction, freeze fracture electron microscopy will visualize the junction strand network continuity and extension and treatment-induced strand breaks. Ex vivo and in vitro findings will then be translated into a uremic mouse model of PD. Modulators of junction function will be tested in order to promote the development of novel PD fluid prototypes and therapeutic concepts improving PD detoxification efficacy and sustainability and thus PD patient outcome. Six national and international cooperating partners will contribute to the study.