Modelling normal and nephrotic axial uptake of albumin and other filtered proteins along the proximal tubule.

Recent studies indicate that filtered albumin is retrieved in the proximal tubule (PT) via three pathways: receptor-mediated endocytosis via cubilin (high affinity) and megalin (low affinity), and fluid-phase uptake. Expression of megalin is required to maintain all three pathways, making it challenging to determine their respective contributions. Moreover, uptake of filtered molecules varies between the sub-segments (S1, S2 and S3) that make up the PT. Here we used new and published data to develop a mathematical model that predicts the rates of albumin uptake in mouse PT sub-segments in normal and nephrotic states, and partially accounts for competition by β2 -microglobulin (β2m) and immunoglobulin G (IgG). Our simulations indicate that receptor-mediated, rather than fluid-phase, uptake accounts for the vast majority of ligand recovery. Our model predicts that ∼75% of normally filtered albumin is reabsorbed via cubilin; however, megalin-mediated uptake predominates under nephrotic conditions. Our results also suggest that ∼80% of albumin is normally recovered in S1, whereas nephrotic conditions or knockout of cubilin shifts the bulk of albumin uptake to S2. The model predicts β2m and IgG axial recovery profiles qualitatively similar to those of albumin under normal conditions. In contrast with albumin, however, the bulk of IgG and β2m uptake still occurs in S1 under nephrotic conditions. Overall, our model provides a kinetic rationale for why tubular proteinuria can occur even though a large excess in potential PT uptake capacity exists, and suggests testable predictions to expand our understanding of the recovery profile of filtered proteins along the PT. KEY POINTS: We used new and published data to develop a mathematical model that predicts the profile of albumin uptake in the mouse proximal tubule in normal and nephrotic states, and partially accounts for competitive inhibition of uptake by normally filtered and pathological ligands. Three pathways, consisting of high-affinity uptake by cubilin receptors, low-affinity uptake by megalin receptors and fluid phase uptake, contribute to the overall retrieval of filtered proteins. The axial profile and efficiency of protein uptake depend on the initial filtrate composition and the individual protein affinities for megalin and cubilin. Under normal conditions, the majority of albumin is retrieved in sub-segment S1 but shifts to sub-segment S2 under nephrotic conditions. Other proteins exhibit different uptake profiles. Our model explains how tubular proteinuria can occur despite a large excess in potential proximal tubule uptake capacity.

Edwards A ，Long KR ，Baty CJ ，Shipman KE ，Weisz OA ... -  《-》

Distinct functions of megalin and cubilin receptors in recovery of normal and nephrotic levels of filtered albumin.

Ren Q ，Weyer K ，Rbaibi Y ，Long KR ，Tan RJ ，Nielsen R ，Christensen EI ，Baty CJ ，Kashlan OB ，Weisz OA ... -  《-》

Endocytic adaptation to functional demand by the kidney proximal tubule.

The kidney proximal tubule (PT) efficiently recovers the low level of albumin and other proteins that normally escape the glomerular filtration barrier. Two large receptors, megalin and cubilin/amnionless (CUBAM), bind to and efficiently retrieve these predominantly low molecular-weight proteins via clathrin-mediated endocytosis. Studies in cell culture models suggest that PT cells may sense changes in shear stress to modulate recovery of filtered proteins in response to normal variations in filtration rate. Impairments in PT endocytic function lead to the excretion of filtered proteins into the urine (tubular proteinuria). Remarkably, when the glomerular filtration barrier is breached, the PT is able to recover excess albumin with a capacity that is orders of magnitude higher than normal. What mediates this excess capacity for albumin uptake under nephrotic conditions, and why doesn't it compensate to prevent tubular proteinuria? Here we propose an integrated new working model to describe the PT recovery of filtered proteins under normal and nephrotic states. We hypothesize that uptake via the fluid phase provides excess capacity to recover high concentrations of filtered proteins under nephrotic conditions. Further, concentration of tubular fluid along the tubule axis will enhance the efficiency of uptake in more distal regions of the PT. By contrast to cells where fluid phase and receptor-mediated uptake are independent pathways, expression of megalin is required to maintain apical endocytic pathway integrity and is essential for both uptake mechanisms. This model accounts for both the high-affinity and the high-capacity responses to filtration load in physiological and pathological states.

Weisz OA 《-》

Receptor-associated protein impairs ligand binding to megalin and megalin-dependent endocytic flux in proximal tubule cells.

Long KR ，Rbaibi Y ，Kashlan OB ，Weisz OA ... -  《-》

Abolishment of proximal tubule albumin endocytosis does not affect plasma albumin during nephrotic syndrome in mice.

The megalin/cubilin receptor complex is required for proximal tubular endocytosis and degradation of filtered albumin. An additional high-capacity retrieval pathway of intact albumin for the recovery of large amounts of filtered albumin has been proposed, possibly involving cooperation between megalin/cubilin and the neonatal Fc receptor. To clarify the potential role of such a pathway, we examined the effects of megalin/cubilin gene inactivation on tubular albumin uptake and plasma albumin levels in nephrotic, podocin knockout mice. Immunofluorescence microscopy of megalin/cubilin/podocin knockout mouse kidneys demonstrated abolishment of proximal tubule albumin uptake, in contrast to the excessive albumin accumulation observed in podocin knockout mice compared to controls. Correspondingly, urinary albumin excretion was increased 1.4 fold in megalin/cubilin/podocin compared to podocin knockout mice (albumin/creatinine: 226 vs. 157 mg/mg). However, no difference in plasma albumin levels was observed between megalin/cubilin/podocin and podocin knockout mice, as both were reduced to approximately 40% of controls. There were no differences in liver albumin synthesis by mRNA levels and protein abundance. Thus, megalin/cubilin knockout efficiently blocks proximal tubular albumin uptake in nephrotic mice but plasma albumin levels did not differ as a result of megalin/cubilin-deficiency, suggesting no significance of the megalin/cubilin-pathway for albumin homeostasis by retrieval of intact albumin.

Weyer K ，Andersen PK ，Schmidt K ，Mollet G ，Antignac C ，Birn H ，Nielsen R ，Christensen EI ... -  《-》