Abstract P235: Renin and Prorenin Uptake by Human Conditionally Immortalized Proximal Tubule Epithelial Cells Involves Megalin and Mannose 6-phosphate Receptor

Abstract

Renin is filtered by the glomerulus, and subsequently reabsorbed in the proximal tubule in a megalin-dependent manner, as evidenced by a 20-40-fold rise in urinary renin in patients with Dent’s disease or Lowe syndrome, i.e. disorders characterized by defective proximal tubular reabsorption. Remarkably, the reabsorption of filtered prorenin appears to be complete, as it can only be detected in urine of patients with Dent’s disease or Lowe syndrome. To further investigate megalin-mediated renin and prorenin reabsorption, human conditionally immortalized proximal tubule epithelial cells (ciPTEC) were incubated at 37 °C or 4 °C with recombinant human renin or prorenin (1000 U/L), in the absence or presence of 10 mmol/L mannose 6-phosphate (M6P) to block (pro)renin binding and internalization via M6P receptors. Cell lysate (pro)renin levels were measured by immunoradiometric assay both before and after prorenin activation. At 37 °C, cellular renin and prorenin tended to level off at 6 hours, reaching lysate levels of 75±18 and 100±54 pg/mg (mean±SEM), and all prorenin was detected as renin. Incubating ciPTEC at 4 °C diminished the cellular accumulation of renin and prorenin by 90%, and at this temperature prorenin remained in its inactive conformation. M6P blocked renin and prorenin uptake by >80% under all conditions. Importantly, M6P also concentration-dependently blocked the uptake of fluorescently labeled albumin (IC 50 = 3 mmol/L), to the same degree as the megalin inhibitor RAP-GST (IC 50 = 50 μg/mL). Since albumin is internalized in a megalin-dependent manner, these data indicate that M6P, on top of blocking M6P receptors, also interferes with the megalin-dependent uptake process. In conclusion, ciPTEC bind and internalize renin and prorenin in a temperature-dependent manner. This process involves megalin and/or M6P receptors, and results in prorenin activation at 37 °C.