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Proteases and protease inhibitors of urinary extracellular vesicles in diabetic nephropathy.

Musante L, Tataruch D, Gu D, Liu X, Forsblom C, Groop PH, Holthofer H - J Diabetes Res (2015)

Bottom Line: Urine samples were divided into groups based on the level of albuminuria and UEVs isolated by hydrostatic dialysis and screened for relative changes of 34 different proteases and 32 protease inhibitors, respectively.Interestingly, myeloblastin and its natural inhibitor elafin showed an increase in the normo- and microalbuminuric groups.Similarly, a characteristic pattern was observed in the array of protease inhibitors, with a marked increase of cystatin B, natural inhibitor of cathepsins L, H, and B as well as of neutrophil gelatinase-associated Lipocalin (NGAL) in the normoalbuminuric group.

View Article: PubMed Central - PubMed

Affiliation: Centre for Bioanalytical Sciences (CBAS), Dublin City University, Dublin 9, Ireland.

ABSTRACT
Diabetic nephropathy (DN) is one of the major complications of diabetes mellitus (DM), leads to chronic kidney disease (CKD), and, ultimately, is the main cause for end-stage kidney disease (ESKD). Beyond urinary albumin, no reliable biomarkers are available for accurate early diagnostics. Urinary extracellular vesicles (UEVs) have recently emerged as an interesting source of diagnostic and prognostic disease biomarkers. Here we used a protease and respective protease inhibitor array to profile urines of type 1 diabetes patients at different stages of kidney involvement. Urine samples were divided into groups based on the level of albuminuria and UEVs isolated by hydrostatic dialysis and screened for relative changes of 34 different proteases and 32 protease inhibitors, respectively. Interestingly, myeloblastin and its natural inhibitor elafin showed an increase in the normo- and microalbuminuric groups. Similarly, a characteristic pattern was observed in the array of protease inhibitors, with a marked increase of cystatin B, natural inhibitor of cathepsins L, H, and B as well as of neutrophil gelatinase-associated Lipocalin (NGAL) in the normoalbuminuric group. This study shows for the first time the distinctive alterations in comprehensive protease profiles of UEVs in diabetic nephropathy and uncovers intriguing mechanistic, prognostic, and diagnostic features of kidney damage in diabetes.

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Related in: MedlinePlus

Protease inhibitors array. (a) Nitrocellulose membrane slides were acquired at the same time as fluorescent intensity. (b) Relative quantification of positive protease (signal-to-noise ratio ≥3) normalised by a pool made of all the samples included in the analysis. P: pool, H: healthy control, N: normoalbuminuric, M: microalbuminuric, and Ma: macroalbuminuric.
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fig6: Protease inhibitors array. (a) Nitrocellulose membrane slides were acquired at the same time as fluorescent intensity. (b) Relative quantification of positive protease (signal-to-noise ratio ≥3) normalised by a pool made of all the samples included in the analysis. P: pool, H: healthy control, N: normoalbuminuric, M: microalbuminuric, and Ma: macroalbuminuric.

Mentions: Based on the same assay, a protease inhibitor array to screen the expression of protease inhibitors was performed. Out of 32 inhibitors 19 were present in the same pool, and 15 showed more than 1.5-fold change (Table 2 and Figure 6). Also in this array we can identify different trends. Firstly, 6 inhibitors appeared to be more abundant (cystatin B, fetuin B, angiotensinogen, serpin A8, serpin F1, and elafin) progressively in the 3 diabetic groups. Those with decreased amounts showed either a progressive decrease which becomes significant in the micro- and/or macroalbuminuric groups (e.g., serpin B5 and TIMP-2) or very low amount in all the 3 DN groups (cystatin C E/M, EMMPRIN/CD147, HA-2, and HE4/WFDC2). Finally, two protease inhibitors (Lipocalin-2/NGAL and Protease Nexin II) showed a peculiar increase in the normoalbuminuric group and a sharp decrease in the macroalbuminuric cohort.


Proteases and protease inhibitors of urinary extracellular vesicles in diabetic nephropathy.

Musante L, Tataruch D, Gu D, Liu X, Forsblom C, Groop PH, Holthofer H - J Diabetes Res (2015)

Protease inhibitors array. (a) Nitrocellulose membrane slides were acquired at the same time as fluorescent intensity. (b) Relative quantification of positive protease (signal-to-noise ratio ≥3) normalised by a pool made of all the samples included in the analysis. P: pool, H: healthy control, N: normoalbuminuric, M: microalbuminuric, and Ma: macroalbuminuric.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4383158&req=5

fig6: Protease inhibitors array. (a) Nitrocellulose membrane slides were acquired at the same time as fluorescent intensity. (b) Relative quantification of positive protease (signal-to-noise ratio ≥3) normalised by a pool made of all the samples included in the analysis. P: pool, H: healthy control, N: normoalbuminuric, M: microalbuminuric, and Ma: macroalbuminuric.
Mentions: Based on the same assay, a protease inhibitor array to screen the expression of protease inhibitors was performed. Out of 32 inhibitors 19 were present in the same pool, and 15 showed more than 1.5-fold change (Table 2 and Figure 6). Also in this array we can identify different trends. Firstly, 6 inhibitors appeared to be more abundant (cystatin B, fetuin B, angiotensinogen, serpin A8, serpin F1, and elafin) progressively in the 3 diabetic groups. Those with decreased amounts showed either a progressive decrease which becomes significant in the micro- and/or macroalbuminuric groups (e.g., serpin B5 and TIMP-2) or very low amount in all the 3 DN groups (cystatin C E/M, EMMPRIN/CD147, HA-2, and HE4/WFDC2). Finally, two protease inhibitors (Lipocalin-2/NGAL and Protease Nexin II) showed a peculiar increase in the normoalbuminuric group and a sharp decrease in the macroalbuminuric cohort.

Bottom Line: Urine samples were divided into groups based on the level of albuminuria and UEVs isolated by hydrostatic dialysis and screened for relative changes of 34 different proteases and 32 protease inhibitors, respectively.Interestingly, myeloblastin and its natural inhibitor elafin showed an increase in the normo- and microalbuminuric groups.Similarly, a characteristic pattern was observed in the array of protease inhibitors, with a marked increase of cystatin B, natural inhibitor of cathepsins L, H, and B as well as of neutrophil gelatinase-associated Lipocalin (NGAL) in the normoalbuminuric group.

View Article: PubMed Central - PubMed

Affiliation: Centre for Bioanalytical Sciences (CBAS), Dublin City University, Dublin 9, Ireland.

ABSTRACT
Diabetic nephropathy (DN) is one of the major complications of diabetes mellitus (DM), leads to chronic kidney disease (CKD), and, ultimately, is the main cause for end-stage kidney disease (ESKD). Beyond urinary albumin, no reliable biomarkers are available for accurate early diagnostics. Urinary extracellular vesicles (UEVs) have recently emerged as an interesting source of diagnostic and prognostic disease biomarkers. Here we used a protease and respective protease inhibitor array to profile urines of type 1 diabetes patients at different stages of kidney involvement. Urine samples were divided into groups based on the level of albuminuria and UEVs isolated by hydrostatic dialysis and screened for relative changes of 34 different proteases and 32 protease inhibitors, respectively. Interestingly, myeloblastin and its natural inhibitor elafin showed an increase in the normo- and microalbuminuric groups. Similarly, a characteristic pattern was observed in the array of protease inhibitors, with a marked increase of cystatin B, natural inhibitor of cathepsins L, H, and B as well as of neutrophil gelatinase-associated Lipocalin (NGAL) in the normoalbuminuric group. This study shows for the first time the distinctive alterations in comprehensive protease profiles of UEVs in diabetic nephropathy and uncovers intriguing mechanistic, prognostic, and diagnostic features of kidney damage in diabetes.

Show MeSH
Related in: MedlinePlus