HYPERTENSIVE NEPHROPATHY

Abstract

Objective: We aimed at investigating to which degree hypertensive and diabetic nephropathy show evidence of concurrent inflammation and partial epithelial-to-mesenchymal transition (EMT), including elevated expression of EMT promoting AXL receptor tyrosine kinase. Design and method: Formalin-fixed and paraffin-embedded (FFPE) kidney biopsies from adult patients with hypertensive nephropathy (HN, n = 6), type 2 diabetic nephropathy (DN, n = 6), and normal appearing tissue (n = 6) were recruited from the Norwegian Kidney Biopsy Registry. Total RNA was extracted from whole tissue sections using High Pure FFPE-tissue RNA Isolation kit (Roche). Total RNA inputs of 37.8–1324 ng per sample were used for library preparation and sequenced as 75 bp paired end on Illumina NextSeq500. Differentially expressed genes were analysed with Limma-Voom (Bioconductor) and formed the basis for calculation of a generic EMT score, principal component analysis (PCA) and ingenuity pathway analysis (IPA). Soluble AXL (sAXL) in serum samples (n = 14/group) from patients with HN and healthy controls was quantified by ELISA (Human Axl Duoset, R&D). Results: PCA separated diseased from normal tissue with a considerable overlap in expression profiles of HN and DN. The generic EMT score was higher in both HN and DN as compared to normal tissue. Expression of the EMT promoting receptor tyrosine kinase AXL was increased in both diseases. Immunohistochemistry confirmed increased AXL and vimentin, while E-cadherin was decreased in diseased tissues. Accordingly, sAXL levels were elevated in serum samples from HN patients compared to controls. IPA showed all top five canonical pathways to be involved in inflammation. Accordingly, expression deconvolution analyses provided evidence of eosinophils, natural-killer-T-cells, granulocytes and CD8+ effector-memory T-cells and myeloid-dendritic cells in HN. E.g. the CCL5-CCR5 axis represents a therapeutic target for seven existing drugs. Top scoring pathways common for both DN and HN included the iCOS-iCOSL signalling in T helper cells, Th1 & Th2 activation pathway and the Th1/Th2 pathway. Higher presence of the phagosome formation pathway in DN and the T cell receptor signalling pathway in HN differentiated the two pathologies. Conclusions: HN and DN show an increased mesenchymal phenotype and inflammation. AXL receptor tyrosine kinase represents a novel therapeutic target, especially in HN.