Erythrocyte Glycocalyx Reports Glomerular Endothelial Glycocalyx Damage in Disease and Predicts Glomerular Albumin Permeability

Background: The endothelial glycocalyx (EnGlx) is damaged in acute and chronic kidney diseases contributing to the development albuminuria and vascular damage. However, directly detecting EnGlx loss has proven difficult, limiting research and slowing the development of targeted therapeutics. The red blood cell (RBC) and endothelial glycocalicies contain shared components. Both are exposed to circulating sheddases. We hypothesised therefore that the RBCGlx could provide a surrogate measure of EnGlx damage. Methods: RBC from male Wistar rats and human trials were labelled with fluorescently tagged Glx-binding lectins and rhodamine 18. Images were obtained using confocal microscopy. Analysis was ‘blinded’, using machine learning to identify RBC and custom analysis software to measure the distance between Gaussian modelled fluorescence profile peaks to provide a measure of RBCGlx ‘depth’. Results: In rats (control, STZ (diabetic), STZ spironolactone (treated)) we confirmed a linear corelation between RBCGlx and glomerular EnGlx depth(r2 0.72, p<0.0001, n=19). RBCGlx thickness also corelated with cardiac capillary EnGlx thickness measured using electron microscopy (r2 0.78, p=0.0016, n=8). Serial blood samples confirmed significant RBCGlx damage 4 weeks after STZ, however a significant therapeutic response to spironolactone was detected 2 weeks after treatment. In all groups RBCGlx thickness corelated with ex vivo glomerular albumin permeability (MOA lectin, r2 0.95, p<0.0001, n=9,) suggesting RBCGlx damage can be used to predict glomerular damage. On human kidney biopsies (minimal change nephropathy and thin basement membrane disease) RBCGlx thickness corelated (r2 0.9226, p <0.0001, n=12) with cortical capillary EnGlx thickness. Measuring the RBCGlx in healthy pregnant women (venous blood) we confirmed a corelation with sublingual GlycoCheck™ readings (r2 0.25, p=0.007, n=27). In patients with COVID-19 we rapidly confirmed significant RBCGlx damage (t-test, p=0.0002, n=26,) supporting published work suggesting EnCGlx damage. Conclusion: Our test provides a reliable surrogate marker of EnGlx damage in kidney disease and predicts endothelial function. Larger human trials are underway to investigate the full potential of this major discovery in renal and cardiovascular disease monitoring and therapeutics. Funding: Government Support – Non-U.S.