Albuminuria in patients with type 1 diabetes is directly linked to changes in the lysosome-mediated degradation of albumin during renal passage.

Abstract

Previous studies by our group have shown that albumin is metabolized in rodents during renal passage and excreted in the urine as a mixture of intact protein and albumin-derived fragments. The aim of this study was to examine whether albumin is metabolized during renal passage in nondiabetic volunteers and in type 1 diabetic patients with varying levels of albuminuria. Nine nondiabetic normoalbuminuric volunteers and 11 type 1 diabetic patients with albumin excretion rates varying from normoalbuminuria to macroalbuminuria were studied. Each subject received an intravenous injection of tritium-labeled albumin ([ 3 H]-albumin). Urine was collected at 4 h and 24 h after injection and analyzed by size exclusion chromatography. The amount of intact and fragmented albumin was quantified, and each fraction was analyzed by radioimmunoassay (RIA) for albumin. [ 3 H]-albumin in nondiabetic volunteers was metabolized during renal passage to small peptide fragments not detectable by conventional RIA (only 0.05-3.8% of the total urinary radioactivity was associated with intact albumin). The process responsible for albumin fragmentation was similar in diabetic patients with normoalbuminuria (intact albumin represented 0.01-4.0% of total urinary radioactivity). However, there was a reduction in the fragmentation ratio (fragmented:intact) in diabetic patients with micro-or macroalbuminuria (intact albumin represented 2.7-55.5%, P = 0.048). This change in the fragmentation ratio was directly related to the degree of albuminuria. These results have important implications for understanding the mechanisms underlying albuminuria in nondiabetic volunteers and type 1 diabetic patients. In nondiabetic volunteers, the renal processing of albumin involves a relatively rapid and comprehensive degradation of albumin to small fragments (range 1-15 kDa). The degradation process is inhibited in diabetic nephropathy in proportion to the level of albuminuria detected by RIA. Diabetes 49:1579-1584, 2000 M icroalbuminuria, as measured by radioimmunoassay (RIA), is generally equated with the development of diabetic nephropathy in type 1 diabetes (1,2). The change in albumin excretion rate (AER) has traditionally been explained as being due to changes in the glomerular permselectivity barrier and intraglomerular pressure (3,4) with the assumption that albumin remains intact during filtration and renal passage. However, recent studies by our group using tritium-labeled albumin ([ 3 H]-albumin) in a rodent model have demonstrated that albumin excretion could be significantly influenced by the metabolism of albumin to small peptide fragments during its renal passage (5-9). The fragmentation is extensive, with 90-95% of the urinary albumin representing a complex fragment population of >30 different fragments with molecular weights in the range of 1-15 kDa. These albumin-derived fragments are not detected by standard immunochemical assays (6,8). A small number of studies have previously identified albumin fragments in human urine, although the source of...

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Autophagy-Lysosome Pathway in Renal Tubular Epithelial Cells Is Disrupted by Advanced Glycation End Products in Diabetic Nephropathy

“…Diabetes-induced decrease in the lysosomal enzyme activity (25,26) and reduction in the renal lysosome-mediated degradation of albumin (27) have been implicated in several studies. We found that AGEs-RAGE interaction, but not endocytosis of AGEs, induced a significant decline of lysosomal activity and degradative potential for DQ-ovalbumin.…”

Glomerular Permselectivity Factors Are Not Responsible for the Increase in Fractional Clearance of Albumin in Rat Glomerulonephritis

“…Recent studies have clearly demonstrated that the RIA is specific for intact/macromolecular albumin and this work has shown (Figure 4) that the RIA only detects macromolecular IgG and transferrin. 11,13 As the RIA fails to detect the protein fragments that are excreted, its use results in a severe underestimation of the FC of proteins. It is apparent that normal functioning of the kidney entails almost complete degradation of filtered protein before excretion.…”

“…Albumin RIAs and many of the nonspecific total protein assays fail to detect fragmented albumin. 11,[13][14][15] Thus, there is in fact a much larger excretion of albumin by the normal kidney than previously assumed and this will have a profound influence on urinary excretion measurements. IgG and transferrin undergo similar processing to albumin as studied in healthy rats by the degradation pathway, in that they are subject to extensive fragmentation before excretion.…”