Analysis of the uptake of native versus modified albumin by renal tubular epithelial cells

Ly, D 2015, Analysis of the uptake of native versus modified albumin by renal tubular epithelial cells, Doctor of Philosophy (PhD), Medical Sciences, RMIT University.


Document type: Thesis
Collection: Theses

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Title Analysis of the uptake of native versus modified albumin by renal tubular epithelial cells
Author(s) Ly, D
Year 2015
Abstract Albumin is the most abundant protein in blood and plays important homeostatic roles in the regulation of oncotic pressure and as a carrier of many small proteins, peptides and hormones. Albumin in the circulation can be modified and thus the circulation contains a mixture of native albumin (i.e. not modified) and modified albumin. A significant amount of albumin is lost from the blood each day as a result of the filtration. This filtered albumin in the early filtrate is thought to be mostly taken up by, and degraded in, the early renal proximal tubular epithelial cells, while a small proportion is excreted in the final urine. This degradation of albumin by proximal tubules operates via the megalin scavenger receptor and an endocytic pathway.

Experimental measurement of renal albumin re-uptake and excretion remains a controversial issue as the test probes used to measure these parameters rely on modified albumin probes. These modifications can potentially change the surface charge and tertiary structure of the albumin molecule and therefore alter the way albumin is filtered and absorbed by the kidney. Hence, there still exists an overall paucity of data in relation to albumin handling and how albumin is processed and excreted in the urine. This is not only a complicating factor for diagnosis, but since these probes are often used to study renal function, it also obscures our basic understanding of molecular renal physiology and how albumin is processed. A major gap in our knowledge is whether the megalin-based albumin uptake is a non-selective process, or whether megalin has a significantly different affinity for modified versus native albumin.

This project developed a uniformly labelled “native” albumin to provide a better diagnostic understanding of urinary albumin excretion. The project sought to examine how proximal tubules take up native versus modified albumin. All previous work in this area has used modified albumin probes. The results indicate that the megalin pathway has a much greater affinity for modified compared to native albumin. Based upon common conserved structural motifs between albumin and known megalin ligands, a series of 6 albumin peptides were designed as potential inhibitors of the megalin-albumin interaction. The peptide was identified as a good tool for future investigation of albumin uptake by megalin in vitro and in vivo.

A common sign of kidney injury is albuminuria. This can occur in chronic diseases such as diabetic nephropathy, but it can also occur in acute kidney injury such as in patients with bacterial sepsis. Animal models have shown that the bacterial cell component, lipopolysaccharide (LPS), can induce albuminuria. However, whether LPS-induced albuminuria operates via a direct effect on albumin uptake by tubular epithelial cells is not known. Therefore, this project examined the effect of LPS on OK cells and found this to inhibit the uptake of Alexa488-BSA following 2 or 24 hours stimulation. However, blockade of LPS induced c-Jun amino terminal kinase (JNK) signalling prevented the effect of LPS on albumin uptake, suggesting that this may explain the ability of JNK inhibition to prevent LPS-induced albuminuria in mice.
Degree Doctor of Philosophy (PhD)
Institution RMIT University
School, Department or Centre Medical Sciences
Keyword(s) albumin
megalin
recombinant albumin
auto-quenching
proximal tubular cells
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Created: Fri, 11 Sep 2015, 10:21:32 EST by Denise Paciocco
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