Urinary peptidomics in youths with and without type 1 diabetes

Although the early mechanisms responsible for diabetic kidney disease remain unclear, it is widely believed that chronic hyperglycemia disrupts the proteolytic milieu in the diabetic kidney and may contribute to early kidney injury. We thus performed mechanistic peptidomics in type 1 diabetes before the onset of microalbuminuria. In the discovery cohort of 30 participants, we identified 6550 peptides from 753 proteins. After removing false hits and potential contaminants, there were 6323 quantified peptides: 5708 peptides can be found in youths with type 1 diabetes, 5011 in healthy controls, and 4396 common to both groups. After applying a cut-off of 100%, there were 162 peptides found in each and every urine sample. From this subset, fifteen peptides were differentially excreted between youths with type 1 diabetes and their non-diabetic peers (P < 0.05, t-test); five remained significant after Benjamini-Hochberg adjustment (Q < 0.05). Excretion rates of six peptides were validated in a second cohort of thirty participants using parallel reaction monitoring (separate PRIDE file). Interestingly, these peptides originated from a small region near the C-terminus of uromodulin, a kidney-specific protein. In silico analyses of cleavage sites implicated several metallo and serine proteases involved in wound healing. Two of the uromodulin peptides activated NFκB in a TLR4-dependent manner in cultured kidney tubular cells and also promoted neutrophil migration in vitro. In summary, the differential excretion of uromodulin peptides may shed light onto early inflammatory processes in the diabetic kidney.