BACKGROUND: Glomerular hyperfiltration (GH) is an important mechanism in the development of albuminuria in hypertension. Upregulation of COX2 (cyclooxygenase 2) and prostaglandin E 2 (PGE 2 ) was linked to podocyte damage in GH. We explored the potential renoprotective effects of either separate or combined pharmacological blockade of EP2 (PGE 2 receptor type 2) and EP4 (PGE 2 receptor type 4) in GH. METHODS: We conducted in vivo studies in a transgenic zebra fish model ( Tg[fabp10a:gc-EGFP] ) suitable for analysis of glomerular filtration barrier function and a genetic rat model with GH, albuminuria, and upregulation of PGE 2 . Similar pharmacological interventions and primary outcome analysis on albuminuria phenotype development were conducted in both model systems. RESULTS: Stimulation of zebra fish embryos with PGE 2 induced an albuminuria-like phenotype, thus mimicking the suggested PGE 2 effects on glomerular filtration barrier dysfunction. Both separate and combined blockade of EP2 and EP4 reduced albuminuria phenotypes in zebra fish and rat models. A significant correlation between albuminuria and podocyte damage in electron microscopy imaging was identified in the rat model. Dual blockade of both receptors showed a pronounced synergistic suppression of albuminuria. Importantly, this occurred without changes in arterial blood pressure, glomerular filtration rate, or tissue oxygenation in magnetic resonance imaging, while RNA sequencing analysis implicated a potential role of circadian clock genes. CONCLUSIONS: Our findings confirm a role of PGE 2 in the development of albuminuria in GH and support the renoprotective potential of combined pharmacological blockade of EP2 and EP4 receptors. These data support further translational research to explore this therapeutic option and a possible role of circadian clock genes.

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Objective: Glomerular hyperfiltration associates with increased fluid flow shear stress (FFSS) on podocytes. This can lead to irreversible podocyte damage, which in turn negatively affects the permeability and integrity of the glomerular filtration barrier. AngiotensinII (AngII) is an important mediator in chronic kidney disease. The negative effects of AngII are at least partially opposed by the alternative signaling pathways within the renin-angiotensin system (RAS) consisting of angiotensin-converting enzyme 2 (ACE2), angiotensin-(1-7) (Ang-[1-7]) and the MAS1 receptor. This counteracting pathway may exert renoprotective effects and support anti-inflammatory processes. As the pathomechanisms and signaling pathways underlying podocyte damage induced by FFSS are not fully understood, our aim is to elucidate the potential renoprotective impact of the alternative RAS axis in human podocytes in the context of glomerular hyperfiltration. Design and method: Conditionally immortalized human podocytes (hPCs) were used for mechanistic studies. Cells seeded on collagen IV coated Culture Slips® were inserted in a Streamer® Shear Stress Device and exposed to FFSS of 1dyne/cm2 for 2h. Control cells were not exposed to FFSS. Afterwards, the formation rate of AngII to Ang-[1-7] was measured in whole cell lysates (1 experiment, n = 6 samples per condition) using spiked AngII as a substrate with and without addition of Z-Pro-Prolinal and MLN-4760. Additionally, gene expression analyses of the enzymes involved in Ang-[1-7] formation were performed. Results: Preliminary data show an increased AngII to Ang-[1-7] formation rate in hPCs following FFSS exposure compared to control. Ang-[1-7] formation under both conditions is reduced by addition of Z-Pro-Prolinal, whereas MLN-4760 has no effect. RNA-expression levels of prolylcarboxypeptidase and prolylendopeptidase are not significantly affected upon FFSS compared to control. In contrast, ACE2 is significantly upregulated by FFSS although expression levels are substantially lower under both conditions compared to prolylcarboxypeptidase and prolylendopeptidase. Conclusions: Our results suggest an elevated AngII to Ang-[1-7] formation rate upon FFSS in hPCs compared to no-flow control. Under both settings, Ang-[1-7] formation in hPCs seems to be mediated by prolylcarboxypeptidase and/or prolylendopeptidase but not ACE2. These results support further studies to confirm the potential role and mechanisms of Ang-[1-7] upregulation in response to glomerular hyperfiltration.