Urine Biomarkers of Kidney Tubule Health, Injury, and Inflammation are Associated with Progression of CKD in Children.

BACKGROUND: Novel urine biomarkers may improve identification of children at greater risk of rapid kidney function decline, and elucidate the pathophysiology of CKD progression. METHODS: We investigated the relationship between urine biomarkers of kidney tubular health (EGF and α-1 microglobulin), tubular injury (kidney injury molecule-1; KIM-1), and inflammation (monocyte chemoattractant protein-1 [MCP-1] and YKL-40) and CKD progression. The prospective CKD in Children Study enrolled children aged 6 months to 16 years with an eGFR of 30-90ml/min per 1.73m2. Urine biomarkers were assayed a median of 5 months [IQR: 4-7] after study enrollment. We indexed the biomarker to urine creatinine by dividing the urine biomarker concentration by the urine creatinine concentration to account for the concentration of the urine. The primary outcome was CKD progression (a composite of a 50% decline in eGFR or kidney failure) during the follow-up period. RESULTS: Overall, 252 of 665 children (38%) reached the composite outcome over a median follow-up of 6.5 years. After adjustment for covariates, children with urine EGF concentrations in the lowest quartile were at a seven-fold higher risk of CKD progression versus those with concentrations in the highest quartile (fully adjusted hazard ratio [aHR], 7.1; 95% confidence interval [95% CI], 3.9 to 20.0). Children with urine KIM-1, MCP-1, and α-1 microglobulin concentrations in the highest quartile were also at significantly higher risk of CKD progression versus those with biomarker concentrations in the lowest quartile. Addition of the five biomarkers to a clinical model increased the discrimination and reclassification for CKD progression. CONCLUSIONS: After multivariable adjustment, a lower urine EGF concentration and higher urine KIM-1, MCP-1, and α-1 microglobulin concentrations were each associated with CKD progression in children.

Urinary phosphate-containing nanoparticle contributes to inflammation and kidney injury in a salt-sensitive hypertension rat model.

Although disturbed phosphate metabolism frequently accompanies chronic kidney disease (CKD), its causal role in CKD progression remains unclear. It is also not fully understood how excess salt induces organ damage. We here show that urinary phosphate-containing nanoparticles promote kidney injury in salt-sensitive hypertension. In Dahl salt-sensitive rats, salt loading resulted in a significant increase in urinary phosphate excretion without altering serum phosphate levels. An intestinal phosphate binder sucroferric oxyhydroxide attenuated renal inflammation and proteinuria in this model, along with the suppression of phosphaturia. Using cultured proximal tubule cells, we confirmed direct pathogenic roles of phosphate-containing nanoparticles in renal tubules. Finally, transcriptome analysis revealed a potential role of complement C1q in renal inflammation associated with altered phosphate metabolism. These data demonstrate that increased phosphate excretion promotes renal inflammation in salt-sensitive hypertension and suggest a role of disturbed phosphate metabolism in the pathophysiology of hypertensive kidney disease and high salt-induced kidney injury.

The association of renal tubular inflammatory and injury markers with uric acid excretion in chronic kidney disease patients.

AIM: To investigate the correlation of renal tubular inflammatory and injury markers with renal uric acid excretion in chronic kidney disease (CKD) patients. METHODS: Seventy-three patients with CKD were enrolled. Fasting blood and morning urine sample were collected for routine laboratory measurements. At the same time, 24 h of urine was collected for urine biochemistry analyses, and 10 ml was extracted from the 24-h urine sample to further detect renal tubular inflammatory and injury markers, including interleukin-18 (IL-18), interleukin 1ß (IL-1ß), neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1). The patients were divided into three tertile groups according to their 24-h urinary uric acid (24-h UUA) levels (UUA1: 24-h UUA ≤ 393.12 mg; UUA2: 393.12 < 24-h UUA ≤ 515.76 mg; UUA3: 24-h UUA > 515.76 mg). The general clinical and biochemical indexes were compared. Multivariable linear regression models were used to test the association of IL-18/Urinary creatinine concentration (IL-18/CR), IL-1ß/CR, NGAL/CR and KIM-1/CR with renal uric acid excretion indicators. RESULTS: All of tested renal tubular inflammation- and injury-related urinary markers were negatively associated with 24-h UUA and UEUA, and the negative correlation still persisted after adjusting for multiple influencing factors including urinary protein and eGFR. Further group analyses showed that these makers were significantly higher in the UUA1 than in the UUA3 group. CONCLUSIONS: Our findings suggest that markers of urinary interstitial inflammation and injury in CKD patients are significantly correlated with 24-h UUA and Urinary excretion of uric acid (UEUA), and those with high 24-h UUA have lower levels of these markers. Renal uric acid excretion may also reflect the inflammation and injury of renal tubules under certain conditions.

Abnormal circadian rhythm of urinary sodium excretion correlates closely with hypertension and target organ damage in Chinese patients with CKD.

Whether the abnormal circadian rhythm of urinary sodium excretion is associated with hypertension in chronic kidney disease (CKD) is poorly understood. In this study, we assessed the relationship between the circadian rhythm of urinary sodium excretion and hypertension. Urinary samples were collected during both the day (07:00 to 22:00) and night (22:00 to 07:00) to estimate night/day urinary sodium excretion ratios. Blood pressure (BP) and clinical data were also measured. A total of 1,099 Chinese CKD patients were recruited, 308 patients were excluded, and 791 patients were final enrolled in this study. Among them, 291 patients were normotensive and 500 were hypertensive CKD patients. A 1:1 propensity score matching (PSM) analysis was performed with age and estimated glomerular filtration rate (eGFR) matched between 190 normotensive and hypertensive patients. In the full cohort and PSM cohort, multivariate regression analysis showed that the night/day urinary sodium excretion ratio was an independent risk factor for clinical hypertension, whereas 24 h urinary sodium excretion, diurnal and nocturnal urinary sodium excretion were not. When the night/day urinary sodium excretion ratios were further divided into tertiles (tertile 1 < 0.47, tertile 2, 0.47-0.84 and tertile 3 > 0.84), multivariate analysis showed that tertile 3 was independently associated with hypertension in the full and PSM cohorts. In addition, tertile 3 was also independently associated with eGFR ≤ 60 mL/min/1.73 m2 and left ventricular hypertrophy. These data suggested that an abnormal circadian rhythm of urinary sodium excretion was independently associated with hypertension and target-organ damage. Individualized salt intake and therapeutic strategies should be used to normalize the natriuretic dipping profile in CKD patients.

Circulating miR-103a-3p contributes to angiotensin II-induced renal inflammation and fibrosis via a SNRK/NF-κB/p65 regulatory axis.

Although angiotensin II (AngII) is known to cause renal injury and fibrosis, the underlying mechanisms remain poorly characterized. Here we show that hypertensive nephropathy (HN) patients and AngII-infused mice exhibit elevated levels of circulating miR103a-3p. We observe a positive correlation between miR-103a-3p levels and AngII-induced renal dysfunction. miR-103a-3p suppresses expression of the sucrose non-fermentable-related serine/threonine-protein kinase SNRK in glomerular endothelial cells, and glomeruli of HN patients and AngII-infused mice show reduced endothelial expression of SNRK. We find that SNRK exerts anti-inflammatory effects by interacting with activated nuclear factor-κB (NF-κB)/p65. Overall, we demonstrate that AngII increases circulating miR-103a-3p levels, which reduces SNRK levels in glomerular endothelial cells, resulting in the over-activation of NF-κB/p65 and, consequently, renal inflammation and fibrosis. Together, our work identifies miR-103a-3p/SNRK/NF-κB/p65 as a regulatory axis of AngII-induced renal inflammation and fibrosis.

NEPHROPATHY IN EARLY RHEUMATOID ARTHRITIS PATIENTS: DOES A SIGNIFICANT RISK EXIST?

Objective of the study layed in assessment of the pathophysiological relation between cell-mediated immunity (tumor necrosis factor-alpha (TNF-α) inflammatory cytokine) activation and renal dysfunction in the patients with early rheumatoid arthritis. We analyzed the data from 35 early rheumatoid arthritis (RA) patients of average age of 50,71±2,25 years (ranged 18-76 years, 80% of women) with 9,21±0,43 months mean duration of the disease by the time of the study initiation. Urine and blood tests were performed to verify the main indicators of kidney function and inflammation cytokines significant interaction. All signs of renal dysfunction at the baseline in the patients with early RA were associated with glomerular filtration rate decrease and excretion of urine protein increase. Dynamics of albuminuria, according to the analysis of variance for one-factor scheme, were significantly determined by the state of disease activity, reflecting the severity of joint damage. High urine ß-2-microglobulin level was significantly associated with the expression rate of main inflammatory cytokines as per binary regression analysis. The obtained dependence showed the dynamics of expression of tubular disorders in early RA with a progressive deterioration which did associate with the levels of TNF-α expression, and variety of the urine miсroglobulin rates in the interval 200-350 µg/L. Reliable correlation (r=0.51, p<0.05) between beta-2-microglobulinuria and TNF-α levels was clearly shown, revealing the relationship described by the formula MGU=- 81+937×log10 (TNF-α) as per regression analysis. The severity of tubular damage in early RA is associated with TNF-α expression, especially in the patients with TNF-α above 250 pg/mL, when microalbuminuria rates were significantly higher (p=0.00043). We identified robust data that in the early RA patients with high TNF-α, the number of reported cases of microalbuminuria was significantly higher than in those with low levels.

Elevated urinary monocyte chemoattractant protein-1 levels in children with Henoch-Schonlein purpura nephritis.

BACKGROUND: Chemokine monocyte chemoattractant protein-1 (MCP-1) has been proved as a potential urinary biomarker in nephropathies. The aim of this study was to investigate the urinary monocyte chemoattractant protein-1 (MCP-1) levels and clinical significance in Henoch-Schonlein purpura (HSP) children with and without nephritis and determine the association of MCP-1 with proteinuria. METHODS: A total of 261 HSP children-with or without nephritis-and 84 healthy control children were enrolled in this study. Of these, 126 HSP nephritis (HSPN) children were subdivided into three groups according to total urine protein in 24 h (TUP): Group A, mild proteinuria group with TUP <25 mg/kg; Group B, moderate proteinuria group with TUP ≥25 mg/kg and <50 mg/kg; Group C, severe proteinuria group with TUP ≥50 mg/kg. Urinary MCP-1 levels were determined by ELISA. Levels of serum creatinine (Cr), blood urea nitrogen (BUN), urinary α1-micro globulin (α1-MG), micro-albumin (mAlb), immunoglobulin G (IgG), transferrin (TRF) and TUP were performed to determine their associations with MCP-1. RESULTS: Urinary MCP-1 was significantly higher in HSPN group in comparison with HSP group and controls (P < 0.05), but no significant difference was found between the HSP group and the healthy group (P > 0.05). The levels of urinary MCP-1 increased in parallel to the enhancement of total urine protein in 24 h in HSPN patients. There were statistically significant differences among these three groups of HSPN children (p < 0.05). Urinary MCP-1 correlated positively with urinary α1-MG, mAlb, IgG, TRF and TUP in HSPN, whereas no correlation was observed with serum Cr and BUN. CONCLUSIONS: MCP-1 was elevated in children with HSPN and correlated with proteinuria. Urinary MCP-1 could be used as a suitable, non-invasive biomarker to provide valuable information not only for the diagnosis of HSPN, but also for evaluation of severity of renal damage.

Urinary metal and metalloid biomarker study of Henoch-Schonlein purpura nephritis using inductively coupled plasma orthogonal acceleration time-of-flight mass spectrometry.

To obtain a better understanding as to whether concentration alterations of metals and metalloids in urine were related to Henoch-Schonlein purpura nephritis (HSPN), the profiles of as many as 29 elements in urine were compared among three groups, the Henoch-Schonlein purpura (HSP), HSPN and a healthy control group. To this end, a reliable method has been developed for the simultaneous quantification of multiple elements including Li, Be, B, Al, Sc, Ti, V, Cr, Mn, Fe, Ni, Co, Cu, Zn, Ga, Ge, As, Se, Rb, Sr, Mo, Cd, Sn, Sb, Cs, Ba, Tl, Pb and Bi in urine using inductively coupled plasma orthogonal acceleration time-of-flight mass spectrometry (ICP-oa-TOF-MS). The process of sample pre-treatment used a direct 20-fold dilution method with centrifuged urine. The internal standard element used for quantification was 103Rh, and 1,4-butanediol was chosen as a matrix matching reagent. The method detection limits of these 29 elements were in the range of 0.04-12ngmL-1. Results of statistical analysis revealed that the concentrations of 15 elements and the element homeostasis were significantly different among these three groups. Our study provides a potential method for HSPN metal and metalloid biomarker discovery.

Urinary Macrophage Migration Inhibitory Factor as a Noninvasive Biomarker in Pediatric Henoch-Schönlein Purpura Nephritis.

PURPOSES: The aims of this study were to investigate urinary macrophage migration inhibitory factor (MIF) levels and their clinical significance in Henoch-Schönlein purpura (HSP) children with or without nephritis (N) and to assess the influence of steroid treatment on the urine MIF levels of HSPN patients. METHODS: Group I comprised 35 children with HSPN who were examined twice (A before treatment and B after steroid treatment). Group II comprised 41 children with HSP. The control group included 32 healthy children. Urinary MIF levels were measured via enzyme linked immunosorbent assay. The levels of serum creatinine, blood urea nitrogen, urinary microalbumin (mAlb), and 24-hour proteinuria were performed to determine their associations with MIF levels. RESULTS: Urinary MIF levels were significantly higher in group I compared with group II and the control group (P < 0.01); however, no significant difference was found between group II and the control group (P > 0.05). Upon examination, albeit urinary MIF concentration was significantly lower in group IB compared with group IA (P < 0.05), these concentrations were statistically higher than that of group II (P < 0.05). In addition, in the HSPN patients, the urinary MIF was positively associated with urinary microalbumin and 24-hour proteinuria but no association with serum creatinine and blood urea nitrogen. CONCLUSIONS: Elevated urinary MIF levels were found to be correlated with proteinuria in pediatric HSPN. An obvious decrease in urinary MIF concentrations among the children with HSPN was associated with steroid treatment. Urinary MIF can be used as a noninvasive biomarker in pediatric HSPN.

A novel in vivo method to quantify slit diaphragm protein abundance in murine proteinuric kidney disease.

Injury of the glomerular filter causes proteinuria by disrupting the sensitive interplay of the glomerular protein network. To date, studies of the expression and trafficking of glomerular proteins have been mostly limited to in vitro or histologic studies. Here, we report a novel in vivo biotinylation assay that allows the quantification of surface expression of glomerular proteins in mice. Kidneys were perfused in situ with biotin before harvest. Afterwards glomeruli were isolated and lyzed. The protein of interest was separated by immunoprecipitation and the amount of surface-expressed protein was quantified by Western blot analysis with streptavidin staining. As proof-of-concept, we examined the presence of nephrin in the slit diaphragm in two well-established murine models of proteinuric kidney disease: nephrotoxic nephritis and adriamycin nephropathy. In proteinuric animals, significantly less nephrin was detected in the slit diaphragm. When proteinuria decreased once again during the course of disease, the amount of surface nephrin returned to the baseline. Our present results suggest that our assay is a valuable tool to study the glomerular filter in proteinuric kidney diseases. Note that the assay is not limited to proteins expressed in the slit diaphragm, and all surface proteins that are accessible to biotin perfusion and immunoprecipitation qualify for this analysis.

Urine Ammonium Predicts Clinical Outcomes in Hypertensive Kidney Disease.

Metabolic acidosis is associated with poor outcomes in CKD. Because impaired renal ammonium excretion is important in the pathogenesis of acidosis, urine ammonium excretion might be a better and perhaps earlier acid-base indicator of risk than serum bicarbonate, particularly in patients without acidosis. We evaluated the association between baseline ammonium excretion and clinical outcomes in African American Study of Kidney Disease and Hypertension participants (n=1044). Median daily ammonium excretion was 19.5 (95% confidence interval [95% CI], 6.5 to 43.2) mEq. In Cox regression models (adjusted for demographics, measured GFR, proteinuria, body mass index, net endogenous acid production, and serum potassium and bicarbonate), hazard ratios of the composite outcome of death or dialysis were 1.46 (95% CI, 1.13 to 1.87) in the low tertile and 1.14 (95% CI, 0.89 to 1.46) in the middle tertile of daily ammonium excretion compared with the high tertile. Among participants without acidosis at baseline, the adjusted hazard ratio for those with ammonium excretion <20 mEq/d was 1.36 (95% CI, 1.09 to 1.71) compared with those with ammonium excretion ≥20 mEq/d. Additionally, compared with participants in the high ammonium tertile, those in the low ammonium tertile had higher adjusted odds of incident acidosis at 1 year (adjusted odds ratio, 2.56; 95% CI, 1.04 to 6.27). In conclusion, low ammonium excretion is associated with death and renal failure in hypertensive kidney disease, even among those without acidosis. Low ammonium excretion could identify patients with CKD and normal bicarbonate levels who might benefit from alkali before acidosis develops.

The ISKDC classification and a new semiquantitative classification for predicting outcomes of Henoch-Schönlein purpura nephritis.

BACKGROUND: Histological findings from primary kidney biopsies were correlated with patient outcomes in a national cohort of paediatric Henoch-Schönlein nephritis (HSN) patients. METHODS: Primary kidney biopsies from 53 HSN patients were re-evaluated using the ISKDC (International Study of Kidney Disease in Children) classification and a modified semiquantitative classification (SQC) that scores renal findings and also takes into account activity, chronicity and tubulointerstitial indices. The ISKDC and SQC classifications were evaluated comparatively in four outcome groups: no signs of renal disease (outcome A, n = 27), minor urinary abnormalities (outcome B, n = 18), active renal disease (outcome C, n = 3) and renal insufficiency, end-stage renal disease or succumbed due to HSN (outcome D, n = 5). For the receiver operating characteristic and logistic regression analyses, outcomes A and B were considered to be favourable and outcomes C and D to be unfavourable. The median follow-up time was 7.3 years. RESULTS: The patients with an unfavourable outcome (C and D), considered together due to low patient numbers, had significantly higher total biopsy SQC scores and activity indices than those who had a favourable one (groups A and B). The chronicity and tubulointerstitial indices differed significantly only between group C + D and group A. The difference in areas under the curve between the total biopsy SQC scores and ISKDC findings was 0.15 [p = 0.04, normal-based 95% confidence interval (CI) 0.007-0.29, bias-controlled 95% CI -0.004 to 0.28]. CONCLUSIONS: Our results suggest that the modified SQC is more sensitive than ISKDC classification for predicting the outcome in HSN cases.

New developments in transplant proteomics.

PURPOSE OF REVIEW: Despite modern immunosuppression, renal allograft rejection remains a major contributor to graft loss. Novel biomarkers may help improve posttransplant outcomes through the early detection and treatment of rejection. Our objective is to provide an overview of proteomics, review recent discovery-based rejection studies, and explore innovative approaches in biomarker development. RECENT FINDINGS: Urine MMP7 was identified as a biomarker of subclinical and clinical rejection using two-dimensional liquid chromatography tandem-mass spectrometry (LC-MS/MS) and improved the overall diagnostic discrimination of urine CXCL10 : Cr alone for renal allograft inflammation. A novel peptide signature to classify stable allografts from acute rejection, chronic allograft injury, and polyoma virus (BKV) nephropathy was identified using isobaric tag for relative and absolute quantitation (TRAQ) and label-free MS, with independent validation by selected reaction monitoring mass spectrometry (SRM-MS). Finally, an in-depth exploration of peripheral blood mononuclear cells identified differential proteoform expression in healthy transplants versus rejection. SUMMARY: There is still much in the human proteome that remains to be explored, and further integration of renal, urinary, and exosomal data may offer deeper insight into the pathophysiology of rejection. Functional proteomics may be more biologically relevant than protein/peptide quantity alone, such as assessment of proteoforms or activity-based protein profiling. Discovery-based studies have identified potential biomarker candidates, but external validation studies are required.

Clinical outcomes in children with Henoch-Schönlein purpura nephritis without crescents.

BACKGROUND: Henoch-Schönlein purpura is the most common vasculitis in children. Its long-term prognosis depends on renal involvement. The management of Henoch-Schönlein purpura nephritis (HSPN) remains controversial. This study reports the prognosis of children with HSPN presenting with class 2 International Study of Kidney Disease in Children (ISKDC) nephritis. METHODS: All children with HSPN class 2 diagnosed between 1995 and 2015 in four pediatric nephrology centers were included, and clinical and biological data were collected from the medical files. The primary endpoint was proteinuria remission defined as a proteinuria <200 mg/L. RESULTS: Ninety-two children were included in the study with a median follow-up of 36 (6-120) months; 28% had nephrotic syndrome, 31% proteinuria >3 g/L, 52% proteinuria between 1 and 3 g/L, and 18% proteinuria <1 g/L. Forty-seven percent of patients received orally treatment with steroids alone, 37% received methylprednisolone pulses followed by steroids orally, 18% received no steroids. Although 85% reached remission during follow-up, 12% did not maintain complete remission over time so that only 75% remained in complete remission by the end of the follow-up. Univariate analysis found a higher likelihood of remission in patients with higher proteinuria at disease onset (p = 0.009). This trend was not found in the multivariate analysis after adjusting for treatments, as patients with higher proteinuria were most often treated with steroids. CONCLUSION: Our study shows that one fourth of patients with HSPN class 2 remain proteinuric and thus carry the risk of developing chronic kidney disease over the long term. This finding, together with the better outcome of patients treated with steroids, is in favor of using high-dose steroids orally or IV in these patients.

Differences in pathological characteristics and laboratory indicators in adult and pediatric patients with Henoch-Schönlein purpura nephritis.

We aimed to investigate the differences in renal histopathological changes and laboratory parameters between adult and pediatric patients with Henoch-Schönlein purpura nephritis (HSPN), and to analyze the correlation between laboratory parameters and renal histopathological grading. A total of 139 patients diagnosed with HSPN between September 2010 and December 2014 at the First Hospital of Jilin University, China, were retrospectively reviewed. The clinical and pathological characteristics were examined and compared between the adult and the pediatric patients. A majority of adult (75.0%) and pediatric (66.2%) patients were categorized as pathological grade III HSPN. Adults having crescent lesions, interstitial fibrosis and renal artery involvement significantly outnumbered child counterparts (all P<0.05). Pathological grading showed a positive correlation with 24-h urine protein (r=0.307, P=0.009), microalbuminuria (r=0.266, P=0.000) and serum globulin (r=0.307, P=0.014), and a negative correlation with serum albumin (r=0.249, P=0.037) in pediatric patients with HSPN. Among adult patients with HSPN, histopathological grading showed a positive correlation with 24-h urine protein (r=0.294, P=0.015), microalbuminuria (r=0.352, P=0.006), α1-microglobulin (r=0.311, P=0.019) and immunoglobulin G (r=0.301, P=0.023) in urine, and serum creatinine (r=0.292, P=0.018). Further, a negative correlation between serum albumin and pathological grading was also observed (r=0.291, P=0.018). In conclusion, the severity of renal pathological lesions in HSPN patients is well reflected by the levels of proteinuria. Adult patients have more severe renal histopathological changes than pediatric patients.

Mining the human urine proteome for monitoring renal transplant injury.

The human urinary proteome provides an assessment of kidney injury with specific biomarkers for different kidney injury phenotypes. In an effort to fully map and decipher changes in the urine proteome and peptidome after kidney transplantation, renal allograft biopsy matched urine samples were collected from 396 kidney transplant recipients. Centralized and blinded histology data from paired graft biopsies was used to classify urine samples into diagnostic categories of acute rejection, chronic allograft nephropathy, BK virus nephritis, and stable graft. A total of 245 urine samples were analyzed by liquid chromatography-mass spectrometry using isobaric Tags for Relative and Absolute Quantitation (iTRAQ) reagents. From a group of over 900 proteins identified in transplant injury, a set of 131 peptides were assessed by selected reaction monitoring for their significance in accurately segregating organ injury causation and pathology in an independent cohort of 151 urine samples. Ultimately, a minimal set of 35 proteins were identified for their ability to segregate the 3 major transplant injury clinical groups, comprising the final panel of 11 urinary peptides for acute rejection (93% area under the curve [AUC]), 12 urinary peptides for chronic allograft nephropathy (99% AUC), and 12 urinary peptides for BK virus nephritis (83% AUC). Thus, urinary proteome discovery and targeted validation can identify urine protein panels for rapid and noninvasive differentiation of different causes of kidney transplant injury, without the requirement of an invasive biopsy.

Elevated Urinary Matrix Metalloproteinase-7 Detects Underlying Renal Allograft Inflammation and Injury.

BACKGROUND: The urinary CXC chemokine ligand (CXCL)10 detects renal transplant inflammation noninvasively, but has limited sensitivity and specificity. In this study, we performed urinary proteomic analysis to identify novel biomarkers that may improve the diagnostic performance of urinary CXCL10 for detecting alloimmune inflammation in renal transplant patients. METHODS: In preliminary studies, adult renal transplant patients with normal histology (n = 5), interstitial fibrosis and tubular atrophy (n = 6), subclinical (n = 6) and clinical rejection (n = 6), underwent in-depth urine protein compositional analysis with LC-MS/MS, and matrix metalloproteinase-7 (MMP7) were identified as a potential candidate for the diagnosis of renal allograft inflammation. Urine MMP7 performance was then studied in a larger, prospective adult renal transplant population (n = 148 urines from n = 133 patients) with matched surveillance/indication biopsies. The diagnostic performance of urinary MMP7 and CXCL10 in combination was next evaluated using concordance (C-) statistics, net reclassification improvement and integrated discrimination improvement indices, to determine whether it was better than CXCL10 alone. RESULTS: Urinary MMP7:creatinine (Cr) was lower in normal transplants compared to those with inflammation: glomerulonephritis (P = 0.009), viral nephropathies (P = 0.002), interstitial fibrosis and tubular atrophy and inflammation (P = 0.04), borderline (P = 0.08), subclinical (P = 0.01) and clinical rejection (P = 0.0006), and acute tubular necrosis (P < 0.0001). Urinary MMP7:Cr and CXCL10:Cr significantly distinguished noninflamed from inflamed biopsies (area under the curve, 0.74 and 0.70, respectively). The addition of urinary MMP7:Cr to CXCL10:Cr improved the diagnostic performance for subclinical and clinical inflammation/injury by integrated discrimination improvement (P = 0.002) and net reclassification improvement (P = 0.006) analyses. CONCLUSIONS: Urinary MMP7:Cr improves the overall diagnostic performance of urinary CXCL10:Cr for distinguishing normal histology from subclinical and clinical inflammation/injury, but not subclinical inflammation alone.

The Expression of miR-192 and Its Significance in Diabetic Nephropathy Patients with Different Urine Albumin Creatinine Ratio.

OBJECTIVE: To investigate the expression of miR-192 and its significance in diabetic nephropathy (DN) patients. METHODS: 464 patients with type 2 diabetes mellitus (T2DM) were divided into normal albuminuria group (NA, n = 157), microalbuminuria group (MA, n = 159), and large amount of albuminuria group (LA, n = 148). 127 healthy persons were selected as the control group (NC, n = 127). The serum miR-192 levels were detected by Real-Time PCR and transforming growth factor-ß1 (TGF-ß1) and fibronectin (FN) were detected by enzyme-linked immunosorbent assay. The relationships among these parameters were analyzed by Pearson correlation analysis and multiple linear regression analysis. RESULTS: The miR-192 in the LA group was significantly lower than other groups, which was lower in the MA group than in the NA group (P < 0.01). The TGF-ß1 and FN in the LA group were significantly higher than other groups, which were higher in the MA group than in the NA group (P < 0.01). The expression of miR-192 was negatively correlated with TGF-ß1, FN, and Ln (UACR) and miR-192, TGF-ß1, and FN were independent relevant factors affecting Ln (UACR) in T2DM (P < 0.01). CONCLUSIONS: These findings indicate that the levels of miR-192 were lower accompanied by the decrease of urine albumin creatinine ratio (UACR) and the association between miR-192 and nephritic fibrosis in DN.