Expanded Hemodialysis ameliorates uremia-induced impairment of vasculoprotective KLF2 and concomitant proinflammatory priming of endothelial cells through an ERK/AP1/cFOS-dependent mechanism

Front Immunol. 2023 Sep 19:14:1209464. doi: 10.3389/fimmu.2023.1209464. eCollection 2023.

Abstract

Aims: Expanded hemodialysis (HDx) therapy with improved molecular cut-off dialyzers exerts beneficial effects on lowering uremia-associated chronic systemic microinflammation, a driver of endothelial dysfunction and cardiovascular disease (CVD) in hemodialysis (HD) patients with end-stage renal disease (ESRD). However, studies on the underlying molecular mechanisms are still at an early stage. Here, we identify the (endothelial) transcription factor Krüppel-like factor 2 (KLF2) and its associated molecular signalling pathways as key targets and regulators of uremia-induced endothelial micro-inflammation in the HD/ESRD setting, which is crucial for vascular homeostasis and controlling detrimental vascular inflammation.

Methods and results: First, we found that human microvascular endothelial cells (HMECs) and other typical endothelial and kidney model cell lines (e.g. HUVECs, HREC, and HEK) exposed to uremic serum from patients treated with two different hemodialysis regimens in the Permeability Enhancement to Reduce Chronic Inflammation II (PERCI-II) crossover clinical trial - comparing High-Flux (HF) and Medium Cut-Off (MCO) membranes - exhibited strongly reduced expression of vasculoprotective KLF2 with HF dialyzers, while dialysis with MCO dialyzers led to the maintenance and restoration of physiological KLF2 levels in HMECs. Mechanistic follow-up revealed that the strong downmodulation of KLF2 in HMECs exposed to uremic serum was mediated by a dominant engagement of detrimental ERK instead of beneficial AKT signalling, with subsequent AP1-/c-FOS binding in the KLF2 promoter region, followed by the detrimental triggering of pleiotropic inflammatory mediators, while the introduction of a KLF2 overexpression plasmid could restore physiological KLF2 levels and downmodulate the detrimental vascular inflammation in a mechanistic rescue approach.

Conclusion: Uremia downmodulates vasculoprotective KLF2 in endothelium, leading to detrimental vascular inflammation, while MCO dialysis with the novel improved HDx therapy approach can maintain physiological levels of vasculoprotective KLF2.

Keywords: Krüppel-like factor 2 (KLF2); and expanded hemodialysis therapy (HDx); cardiovascular disease (CVD); chronic kidney disease (CKD); cytokine signaling; end-stage renal disease (ESRD); systemic inflammation; uremic toxins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Endothelial Cells
  • Humans
  • Inflammation / complications
  • Kidney Failure, Chronic* / therapy
  • Kruppel-Like Transcription Factors / genetics
  • Renal Dialysis / adverse effects
  • Renal Dialysis / methods
  • Transcription Factors
  • Uremia* / complications
  • Uremia* / therapy

Substances

  • MCO
  • Transcription Factors
  • KLF2 protein, human
  • Kruppel-Like Transcription Factors

Grants and funding

The results in this paper have not been published previously, but they are in part complementary to a previously published mechanistic follow-up study to the concomitant clinical trial (7). This manuscript is a follow-up of the published clinical trial: PERCI-II-Medium-Cut-Off (MCO), which is registered at ClinicalTrials.gov NCT02084381 https://clinicaltrials.gov/ct2/show/NCT02084381) (7, 17). The current manuscript focuses on the mechanistic side studies that aim to verify promising biological leads identified in the prior clinical trial. The “lab research” was funded by academic grants through BMBF, DFG, BCRT/BSRT and EU-Project Grants and CGS/CSC-Scholarships, as outlined in detail below. The contributions by GM and RC were made possible by funding from the German Federal Ministry for Education and Research (BMBF) and German Research Foundation (DFG project Nephroproteciton #934046635, subproject A03, as part of CRC 1365, and EXPAND-PD; CA2816/1-1) through the Berlin Institute of Health (BIH)-Center for Regenerative Therapies (BCRT) and the Berlin-Brandenburg School for Regenerative Therapies (BSRT, GSC203), respectively, and in part by the European Union’s Horizon 2020 Research and Innovation Program under grant agreements No 733006 (PACE) and 779293 (HIPGEN) and 754995 (EUTRAIN). We also acknowledge financial support from the Open Access Publication Fund of Charité – Universitätsmedizin Berlin and the German Research Foundation (DFG). HZ, DW, and PW were supported by Chinese Scholarship Council (CSC) Stipends (HZ: 202008080251, DW: 202108080082, and PW 202108440113, respectively): https://www.chinesescholarshipcouncil.com/. The 2.1 Million Euro academic financing for the prior PERCI-II clinical study came from the German Ministry for Education and Research (BMBF, Project Codes 1313N11786, 13N11787, 13N11788, and 13N11789) headed on the academic side by RS and DZ at Charité and Prof. Dr. Girndt in Halle, while the clinical study was coordinated by Gambro Dialysatoren GmbH, Hechingen (Gambro AB, including all direct and indirect subsidiaries is now part of Baxter International Inc.) who sponsored parts of the PERCI-II clinical trial (e.g. disposable dialysis materials and standard of care versus newly developed types of MCO dialysis filters from Gambro/Baxter). The subsequent academic research was done independently from Gambro/Baxter, as an academic follow-up study: https://www.gesundheitsindustrie-bw.de/fachbeitrag/pm/dialyseforschung-bmbf-foerdert-von-gambro-koordiniertes-verbundprojekt.