TGFBI Protein Is Increased in the Urine of Patients with High-Grade Urothelial Carcinomas, and Promotes Cell Proliferation and Migration.

Here, we discovered TGFBI as a new urinary biomarker for muscle invasive and high-grade urothelial carcinoma (UC). After biomarker identification using antibody arrays, results were verified in urine samples from a study population consisting of 303 patients with UC, and 128 urological and 58 population controls. The analyses of possible modifying factors (age, sex, smoking status, urinary leukocytes and erythrocytes, and history of UC) were calculated by multiple logistic regression. Additionally, we performed knockdown experiments with TGFBI siRNA in bladder cancer cells and investigated the effects on proliferation and migration by wound closure assays and BrdU cell cycle analysis. TGFBI concentrations in urine are generally increased in patients with UC when compared to urological and population controls (1321.0 versus 701.3 and 475.6 pg/mg creatinine, respectively). However, significantly increased TGFBI was predominantly found in muscle invasive (14,411.7 pg/mg creatinine), high-grade (8190.7 pg/mg) and de novo UC (1856.7 pg/mg; all p < 0.0001). Knockdown experiments in vitro led to a significant decline of cell proliferation and migration. In summary, our results suggest a critical role of TGFBI in UC tumorigenesis and particularly in high-risk UC patients with poor prognosis and an elevated risk of progression on the molecular level.

Calcitriol reduces kidney development disorders in rats provoked by losartan administration during lactation.

Calcitriol has important effects on cellular differentiation and proliferation, as well as on the regulation of the renin gene. Disturbances in renal development can be observed in rats exposed to angiotensin II (AngII) antagonists during lactation period. The lack of tubular differentiation in losartan-treated rats can affect calcitriol uptake. This study evaluated the effect of calcitriol administration in renal development disturbances in rats provoked by losartan (AngII type 1 receptor antagonist) administration during lactation. Animals exposed to losartan presented higher albuminuria, systolic blood pressure, increased sodium and potassium fractional excretion, and decreased glomerular filtration rate compared to controls. These animals also showed a decreased glomerular area and a higher interstitial relative area from the renal cortex, with increased expression of fibronectin, alpha-SM-actin, vimentin, and p-JNK; and an increased number of macrophages, p-p38, PCNA and decreased cubilin expression. Increased urinary excretion of MCP-1 and TGF-ß was also observed. All these alterations were less intense in the losartan + calcitriol group.The animals treated with calcitriol showed an improvement in cellular differentiation, and in renal function and structure. This effect was associated with reduction of cell proliferation and inflammation.

Urine Biomarkers to Predict Response to Lupus Nephritis Therapy in Children and Young Adults.

OBJECTIVE: To delineate urine biomarkers that forecast response to therapy of lupus nephritis (LN). METHODS: Starting from the time of kidney biopsy, patients with childhood-onset systemic lupus erythematosus who were diagnosed with LN were studied serially. Levels of 15 biomarkers were measured in random spot urine samples, including adiponectin, α-1-acid glycoprotein (AGP), ceruloplasmin, hemopexin, hepcidin, kidney injury molecule 1, monocyte chemotactic protein-1, lipocalin-like prostaglandin D synthase (LPGDS), transforming growth factor-ß (TGF-ß), transferrin, and vitamin D binding protein (VDBP). RESULTS: Among 87 patients (mean age 15.6 yrs) with LN, there were 37 treatment responders and 50 nonresponders based on the American College of Rheumatology criteria. At the time of kidney biopsy, levels of TGF-ß (p < 0.0001) and ceruloplasmin (p = 0.006) were significantly lower among responders than nonresponders; less pronounced differences were present for AGP, hepcidin, LPGDS, transferrin, and VDBP (all p < 0.05). By Month 3, responders experienced marked decreases of adiponectin, AGP, transferrin, and VDBP (all p < 0.01) and mean levels of these biomarkers were all outstanding (area under the receiver-operating characteristic curve ≥ 0.9) for discriminating responders from nonresponders. Patient demographics and extrarenal disease did not influence differences in biomarker levels between response groups. CONCLUSION: Low urine levels of TGF-ß and ceruloplasmin at baseline and marked reduction of AGP, LPGDS, transferrin, or VDBP and combinations of other select biomarkers by Month 3 are outstanding predictors for achieving remission of LN. If confirmed, these results can be used to help personalize LN therapy.

Evaluation of Th17 pathway in the diagnosis of autosomal dominant polycystic kidney disease.

INTRODUCTION: Current assessment tools of autosomal dominant polycystic kidney disease (ADPKD) diagnosis are challenging. This study evaluated the possible application of assessment of interleukin (IL)-17-related cytokines and the circulatory T helper 17 cells in the diagnosis of ADPKD. MATERIALS AND METHODS: Enrolling 54 ADPKD patients and 54 healthy individuals, we measured serum and urine levels of IL-6, IL-17, IL-23, and transforming growth factor-ß and the peripheral blood frequency of T helper 17 cells through flowcytometry. We computed sensitivity and specificity of each inflammatory marker as well as their different combinations using the receiver operating characteristic curve and discriminant function analysis. RESULTS: The mean serum and urine levels of IL-17 and IL-23 as well as urine levels of IL-6 were higher in ADPKD patients compared to the healthy controls (P < .001). There was no significant difference in the number of T helper 17 cells between the two groups. Among different combinations of the inflammatory markers, the serum IL-17 was the best factor in the diagnosis of ADPKD with a sensitivity as well as specificity of 100%. CONCLUSIONS: It is likely that T helper 17 pathway is involved in the pathogenesis of ADPKD; therefore, it may be beneficial if such a pathway be considered in its diagnosis.

Changes in urinary angiotensinogen posttreatment in pediatric IgA nephropathy patients.

BACKGROUND: Recently, we demonstrated that urinary angiotensinogen (AGT) levels are increased and reflect intrarenal renin-angiotensin system (RAS) status in pediatric patients with chronic glomerulonephritis. Therefore, this study was performed to test the hypothesis that urinary AGT (UAGT) levels provide a specific index of intrarenal RAS status associated with RAS blockade treatment in pediatric IgA nephropathy (IgAN) patients. METHODS: We measured plasma and UAGT levels and urinary transforming growth factor beta (TGF-ß) levels, after which we performed immunohistochemical analysis of AGT, angiotensin II (Ang II), and TGF-ß in 24 pediatric IgAN patients treated with RAS blockades for 2 years. Paired tests were used to analyze the changes from baseline to study end. RESULTS: Although there was no change in plasma AGT levels, UAGT and TGF-ß levels were significantly decreased after RAS blockade, which was accompanied by the expression levels of AGT, Ang II, and TGF-ß, as well as the magnitude of glomerular injury. Baseline UAGT levels positively correlated with diastolic blood pressure, urinary protein levels, scores for mesangial hypercellularity, and the expression levels of AGT, Ang II, and TGF-ß in renal tissues. CONCLUSIONS: These data indicate that UAGT is a useful biomarker of intrarenal RAS activation, which is associated with glomerular injury during RAS blockade in pediatric IgAN patients.

Mathematical model of renal interstitial fibrosis.

Lupus nephritis (LN) is an autoimmune disease that occurs when autoantibodies complex with self-antigen and form immune complexes that accumulate in the glomeruli. These immune complexes initiate an inflammatory response resulting in glomerular injury. LN often concomitantly affects the tubulointerstitial compartment of the kidney, leading first to interstitial inflammation and subsequently to interstitial fibrosis and atrophy of the renal tubules if not appropriately treated. Presently the only way to assess interstitial inflammation and fibrosis is through kidney biopsy, which is invasive and cannot be repeated frequently. Hence, monitoring of disease progression and response to therapy is suboptimal. In this paper we describe a mathematical model of the progress from tubulointerstitial inflammation to fibrosis. We demonstrate how the model can be used to monitor treatments for interstitial fibrosis in LN with drugs currently being developed or used for nonrenal fibrosis.

Predictive usefulness of urinary biomarkers for the identification of cyclosporine A-induced nephrotoxicity in a rat model.

The main side effect of cyclosporine A (CsA), a widely used immunosuppressive drug, is nephrotoxicity. Early detection of CsA-induced acute nephrotoxicity is essential for stop or minimize kidney injury, and timely detection of chronic nephrotoxicity is critical for halting the drug and preventing irreversible kidney injury. This study aimed to identify urinary biomarkers for the detection of CsA-induced nephrotoxicity. We allocated salt-depleted rats to receive CsA or vehicle for 7, 14 or 21 days and evaluated renal function and hemodynamics, microalbuminuria, renal macrophage infiltration, tubulointerstitial fibrosis and renal tissue and urinary biomarkers for kidney injury. Kidney injury molecule-1 (KIM-1), tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), fibronectin, neutrophil gelatinase-associated lipocalin (NGAL), TGF-ß, osteopontin, and podocin were assessed in urine. TNF-α, IL-6, fibronectin, osteopontin, TGF-ß, collagen IV, alpha smooth muscle actin (α -SMA) and vimentin were assessed in renal tissue. CsA caused early functional renal dysfunction and microalbuminuria, followed by macrophage infiltration and late tubulointerstitial fibrosis. Urinary TNF-α, KIM-1 and fibronectin increased in the early phase, and urinary TGF-ß and osteopontin increased in the late phase of CsA nephrotoxicity. Urinary biomarkers correlated consistently with renal tissue cytokine expression. In conclusion, early increases in urinary KIM-1, TNF-α, and fibronectin and elevated microalbuminuria indicate acute CsA nephrotoxicity. Late increases in urinary osteopontin and TGF-ß indicate chronic CsA nephrotoxicity. These urinary kidney injury biomarkers correlated well with the renal tissue expression of injury markers and with the temporal development of CsA nephrotoxicity.

Brazilian embauba (Cecropia pachystachya) extract reduces renal lesions in 5/6 nephrectomized rats.

INTRODUCTION: Cecropia pachystachya (CP) is a plant rich in polyphenols which inhibits the angiotensin-converting enzyme (ACE) in vitro. Angiotensin II (AII) has an important role in the renal lesion provoked by 5/6 nephrectomy (NE). This study evaluated the CP extract effect on renal lesions provoked by 5/6 NE. MATERIALS AND METHODS: Male Wistar rats submitted to 5/6 NE were treated or not treated with CP extract and followed for 90 days. Systemic blood pressure (SBP), albuminuria, renal functional and structural parameters, ACE activity, urinary levels of monocyte chemoattrant protein-1 (MCP-1) and transforming growth factor ß (TGF-ß) were evaluated. RESULTS: Albuminuria and hypertension were less intense in the treated (NE+CP) group compared to the untreated (NE) group. CP extract treatment reduced the fall in glomerular filtration rate observed in NE rats. Glomerulosclerosis, tubulointerstitial lesions, increase of macrophages and AII positive cells in the renal cortex, as well as increases in renal ACE activity, urinary levels of MCP-1 and TGF-ß were attenuated in NE rats by CP treatment. CONCLUSIONS: The treatment with CP extract reduced the SBP and functional and structural renal changes in 5/6 NE rats. These effects were associated with decreased AII expression, ACE activity and inflammation in the renal cortex.

Expression of CD147, BIGH3 and Stathmin and their potential role as diagnostic marker in patients with urothelial carcinoma of the bladder.

BACKGROUND: Urothelial carcinoma of the bladder is characterised by very high recurrence rate, followed up by cystoscopy which being invasive technique makes the need for non-invasive markers important for Transitional Cell Carcinoma (TCC) detection. CD147 is a transmembrane protein highly expressed in tumour cells which aids in tumour invasion and growth. BIGH3, an Extracellular matrix protein (ECM) which interacts with various ECM component in different tissue system and Stathmin(STMN1) is cytosolic microtubule destabilising protein also called as Oncoprotein18 due to its role in tumour promotion. So far the expression of BIGH3 and STMN1 remains undetermined in cancer subjects including TCC. We therefore studied the levels and molecular expression of these molecules in TCC patients, to evaluate their usefulness as diagnostic markers. METHODS: Thirty consecutive TCC patients and two sets of control- 15 Benign prostatic hyperplasia (BPH) patient and 15 healthy were taken. Serum and urine levels of these molecules were estimated by ELISA and relative mRNA expression by Q-PCR from tumour and normal urothelium. Post-Hoc analysis and ROC curve were determined to evaluate the significance and sensitivity and specificity. RESULTS: The mean concentrations of these molecules were found to be significantly increased (p<0.001) in the serum and urine of TCC patients, with varying significance in each grade for different molecules. The urinary levels of CD147 (67 pg/ml) and serum STMN1 concentration (1.38 ng/ml) showed a specific increase as compared to the controls, while BIGH3 was elevated in both serum and urine samples. Molecular (mRNA) expression was elevated in the high grade (Muscle Invasive) stage of the disease for all the molecules, with a significant 3-fold increase that correlated with disease severity being observed for STMN1. ROC analysis gave optimal combination of sensitivity and specificity for diagnosis of the disease in urine and serum sample for STMN1. CONCLUSION: Of CD147, BIGH3 and STMN1, significant results were obtained for STMN1 and it could serve as the best possible diagnostic marker for TCC detection in future.

The role of the kidney in the systemic elimination of interleukin 6, platelet-derived growth factor and transforming growth factor beta.

STUDY GOAL: The aim of the study was to assess the role of the kidney in systemic elimination of IL-6 and growth factors (PDGF, TGF-ß) by comparison of their concentrations in renal arteries and veins, peripheral veins and urine. MATERIAL AND METHODS: 30 brain-dead kidney donors were included in the study. Samples were obtained during the harvesting procedure. 10 healthy volunteers served as controls. A mathematical model of elimination of investigated proteins from systemic circulation was developed. The elimination ratio (ER) formula indicates the predominance of renal synthesis or degradation and also quantifies the renal uptake (UR) and renal pass (PR) of investigated proteins. Serum levels of investigated proteins were estimated using an immunoenzymatic method (ELISA). RESULTS: Renal IL-6 uptake ratio (UR) amounted to 6.6%, elimination ratio (ER) amounted to 6.4% and pass ratio (PR) amounted to 0.2%. PDGF ratios amounted to 5.1%, 5.0% and 0.1% and TGF-ß ratios amounted to -9%, -9% and 0%, respectively. CONCLUSIONS: The kidney takes part in the elimination of IL-6 and PDGF from systemic circulation. The kidney does not take part in the elimination of TGF-ß.

Increased urinary excretion of nephrin, podocalyxin, and ßig-h3 in women with preeclampsia.

Emerging evidence has shown that podocyte injury and reduced specific podocyte protein expressions contribute to proteinuria in preeclampsia. We collected urine specimens from women with preeclampsia to study whether podocyte-specific protein shedding is associated with renal barrier dysfunction. Urine specimens from women with normal pregnancies and from pregnant women complicated by chronic hypertension were used for comparison. We determined soluble podocyte slit protein nephrin levels in the urine specimens. Podocalyxin, ßig-h3, and VEGF concentrations were also measured. We found that nephrin and podocalyxin were barely detectable in the urine specimens from normal pregnant women and from women with chronic hypertension. In preeclampsia, urinary nephrin and podocalyxin concentrations were significantly increased and highly correlated to each other, r(2) = 0.595. Nephrin and podocalyxin were also correlated with urine protein concentrations. ßig-h3 was detected in the urine specimens from women with preeclampsia, and it is highly correlated with nephrin and podocalyxin concentrations in preeclampsia. ßig-h3 was undetectable in normal pregnancy and pregnancy complicated by chronic hypertension. Elevated VEGF levels were also found in women with preeclampsia compared with those of normal pregnancy and pregnancy complicated by chronic hypertension. These results provide strong evidence that podocyte protein shedding occurs in preeclampsia, and their levels are associated with proteinuria. The finding of urinary ßig-h3 excretion in preeclampsia suggests that increased transforming growth factor activity might also be involved in the kidney lesion in this pregnancy disorder.

The renal transcriptome of db/db mice identifies putative urinary biomarker proteins in patients with type 2 diabetes: a pilot study.

We sought to identify novel urinary biomarkers of kidney function in type 2 diabetes. We screened the renal transcriptome of db/db and db/m mice for differentially expressed mRNA transcripts that encode secreted proteins with human orthologs. Whether elevated urine levels of the orthologous proteins correlated with diminished glomerular filtration rate was tested in a cross-sectional study of n = 56 patients with type 2 diabetes. We identified 36 putative biomarker genes in db/db kidneys: 31 upregulated and 5 downregulated. Urinary protein levels of six selected candidates (endothelin-1, lipocalin-2, transforming growth factor-ß, growth and differentiation factor-15, interleukin-6, and macrophage chemoattractant protein-1) were elevated in type 2 diabetic patients with subnormal glomerular filtration rate (i.e., <90 ml·min(-1)·1.73 m(-2)), independent of microalbuminuria, age, sex, race, and use of angiotensin-converting enzyme inhibitors and angiotensin receptor antagonists. In contrast, urinary levels of fibroblast growth factor were not increased. A composite variable of urine albumin and any of the six candidate markers was associated with subnormal estimated glomerular filtration rate more closely than albumin alone. In addition, urinary endothelin-1, growth and differentiation factor-15, and interleukin-6 were associated with a marker of proximal tubule damage, N-acetyl-ß-d-glucosaminidase activity. These results suggest that gene expression profiling in diabetic mouse kidney can complement existing proteomic-based approaches for renal biomarker discovery in humans.

[Determination of urinary markers of proteolysis/fibrinolysis and fibroangiogenesis in the kidney in hypertensive patients].

AIM: To determine clinical significance of urinary biomarkers of proteolysis/fibrinolysis and fibroangiogenesis in essential hypertension (EH). MATERIAL AND METHODS: Examination of the kidneys was made in 71 patients with EH degree 1-3. Renal function was assessed by 24-h albuminuria, calculated glomerular filtration rate (GFR) by Cockroft-Golt. Early signs of renal damage were microalbuminuria--MAU (diurnal albuminuria 30-300 mg/day), reduction of GFR (< 90 ml/min/1.73 m2). EH patients with hypercreatininemia and GFR under 60 ml/min/1.73m2 corresponding to stage III of chronic kidney disease were not included in the study. An additional nephropathy marker was an elevated index of resistance of interlobular renal arteries (RI > 0.65) as shown by dopplerometry. ELISA examined urinary biomarkers of intercellular and cell-matrix interactions in the kidney in EHpatients and healthy controls (n = 12). RESULTS: MAU was detected in 54 (76%) of 71 EH patients, elevated RI > 0.65--in 37 (52%) patients. Urinary biomarkers of proteolysis/fibrinolysis and fibroangiogenesis were higher in EH patients then in the controls. Urinary excretion of PAI-1, TGF-beta1, VEGF and collagen of type IV in EH patients with MAU was significantly higher than in patients with normoalbuminuria. A strong direct correlation between MAU and the rest above urinary biomarkers was found as well as between urinary excretion of collagen IV and RI. An inverse negative relationship was seen between RI and GFR. CONCLUSION: Renal impairment in EHpatients is a progressive disorder. Each stage of this process has its own clinicodiagnostic markers. Urinary biomarkers ofproteolysis/fibrinolysis and fibroangiogenesis in the kidney are informative for monitoring of early HNP.

Urinary interleukin-6 (IL-6) and transforming growth factor (TGF-ß) levels in corticosteroidtreated patients with IgA nephropathy.

BACKGROUND: Interleukin-6 (IL-6) and transforming growth factor-ß (TGF-ß) are implicated in the progression of IgA nephropathy, which is usually treated with corticosteroids. PATIENTS AND METHODS: Urinary IL-6 and TGF-ß were measured in 21 proteinuric patients with IgA nephropathy, before and after treatment with corticosteroids, to estimate the activity of the disease after remission of proteinuria. RESULTS: Urinary IL-6 and TGF-ß levels at diagnosis were significantly higher in patients with IgA nephropathy compared to healthy subjects. TGF-ß levels, were significantly higher in patients with proteinuria > 1 g/24 h and/or severe mesangial proliferation. Although a significant reduction of proteinuria was observed with corticosteroid treatment, urinary IL-6 and TGF-ß levels remained elevated. Deterioration of renal function over a period of 5 years was observed in 3 patients. High urinary IL-6 levels at diagnosis represent a significant parameter distinguishing patients with progressive course in comparison to those with favorable clinical outcome (p = 0.01). CONCLUSION: Treatment of patients with IgA nephropathy with corticosteroids is followed by remission of proteinuria but still increased urinary IL-6 and TGF-ß excretion. This may be related to an ongoing inflammatory process within the kidney, and further research is required to estimate the value of urinary IL-6 and TGF-ß as markers of activity of the disease.

Blockade of TSP1-dependent TGF-ß activity reduces renal injury and proteinuria in a murine model of diabetic nephropathy.

Transforming growth factor-ß (TGF-ß) is key in the pathogenesis of diabetic nephropathy. Thrombospondin 1 (TSP1) expression is increased in diabetes, and TSP1 regulates latent TGF-ß activation in vitro and in diabetic animal models. Herein, we investigate the effect of blockade of TSP1-dependent TGF-ß activation on progression of renal disease in a mouse model of type 1 diabetes (C57BL/6J-Ins2(Akita)) as a targeted treatment for diabetic nephropathy. Akita and control C57BL/6 mice who underwent uninephrectomy received 15 weeks of thrice-weekly i.p. treatment with 3 or 30 mg/kg LSKL peptide, control SLLK peptide, or saline. The effects of systemic LSKL peptide on dermal wound healing was assessed in type 2 diabetic mice (db/db). Proteinuria (urinary albumin level and albumin/creatinine ratio) was significantly improved in Akita mice treated with 30 mg/kg LSKL peptide. LSKL treatment reduced urinary TGF-ß activity and renal phospho-Smad2/3 levels and improved markers of tubulointerstitial injury (fibronectin) and podocytes (nephrin). However, LSKL did not alter glomerulosclerosis or glomerular structure. LSKL did not increase tumor incidence or inflammation or impair diabetic wound healing. These data suggest that selective targeting of excessive TGF-ß activity through blockade of TSP1-dependent TGF-ß activation represents a therapeutic strategy for treating diabetic nephropathy that preserves the homeostatic functions of TGF-ß.

Renal protective effect of chronic inhibition of COX-2 with SC-58236 in streptozotocin-diabetic rats.

The induction of renal cyclooxygenase-2 (COX-2) in diabetes has been implicated in the renal functional and structural changes in models where hypertension or uninephrectomy was superimposed. We examined the protective effects of 3 mo treatment of streptozotocin-diabetic rats with a highly selective COX-2 inhibitor (SC-58236) in terms of albuminuria, renal hypertrophy, and the excretion of TNF-α and TGF-ß, which have also been implicated in the detrimental renal effects of diabetes. SC-58236 treatment (3 mg·kg(-1)·day(-1)) of diabetic rats resulted in reduced urinary excretion of PGE(2), 6-ketoPGF(1α), and thromboxane B(2), all of which were increased in the diabetic rat compared with age-matched nondiabetic rats. However, serum thromboxane B(2) levels were unchanged, confirming the selectivity of SC-58236 for COX-2. The renal protective effects of treatment of diabetic rats with the COX-2 inhibitor were reflected by a marked reduction in albuminuria, a reduction in kidney weight-to-body weight ratio, and TGF-ß excretion and a marked decrease in the urinary excretion of TNF-α. The protective effects of SC-58236 were independent of changes in plasma glucose levels or serum advanced glycation end-product levels, which were not different from those of untreated diabetic rats. In an additional study, the inhibition of COX-2 with SC-58236 for 4 wk in diabetic rats resulted in creatinine clearance rates not different from those of control rats. These results confirm that the inhibition of COX-2 in the streptozotocin-diabetic rat confers renal protection and suggest that the induction of COX-2 precedes the increases in cytokines, TNF-α, and TGF-ß.

Treatment of diabetic rats with a peroxynitrite decomposition catalyst prevents induction of renal COX-2.

Cyclooxygenase (COX)-2 expression is increased in the kidney of rats made diabetic with streptozotocin and associated with enhanced release of prostaglandins stimulated by arachidonic acid (AA). Treatment of diabetic rats with nitro-L-arginine methyl ester (L-NAME) to inhibit nitric oxide synthase or with tempol to reduce superoxide prevented these changes, suggesting the possibility that peroxynitrite (ONOO) may be the stimulus for the induction of renal COX-2 in diabetes. Consequently, we tested the effects of an ONOO decomposition catalyst, 5,10,15,20-tetrakis(N-methyl-4'-pyridyl)porphyrinato iron(III) (FeTMPyP), which was administered for 3-4 wk after the induction of diabetes. FeTMPyP treatment normalized the twofold increase in the expression of nitrotyrosine, a marker for ONOO formation, in the diabetic rat and prevented the increase in renal COX-2 expression without modifying the two- to threefold increases in renal release of prostaglandins PGE(2) and 6-ketoPGF(1α) in response to AA. FeTMPyP treatment of diabetic rats reduced the elevated creatinine clearance and urinary excretion of TNF-α and transforming growth factor (TGF)-ß, suggesting a renoprotective effect. Double immunostaining of renal sections and immunoprecipitation of COX-2 and nitrotyrosine suggested nitration of COX-2 in diabetic rats. In cultured human umbilical vein endothelial cells (HUVECs) exposed to elevated glucose (450 mg/dl) or ONOO derived from 3-morpholinosydnonimine (SIN-1), expression of COX-2 was increased and was prevented when endothelial cells were treated with FeTMPyP. These results indicate that elevated glucose increases the formation of ONOO, which contributes to the induction of renal COX-2 in the diabetic rat.

Urinary transforming growth factor beta-1 as a marker of renal dysfunction in sickle cell disease.

Renal dysfunction affects 5-18% of patients with sickle cell disease (SCD). To date, no studies have described urinary levels of transforming growth factor ß-1 (TGF-ß1), a marker of fibrosis, and neutrophil gelatinase-associated lipocalin (NGAL), a marker of acute/chronic kidney disease, as biomarkers in identifying patients at risk of developing renal disease in SCD. We hypothesized that SCD subjects will have increased urinary excretion of TGF-ß1 and NGAL compared with healthy controls (CTR). We examined 51 SCD subjects: 42 HbSS, 8 HbSC, and 1 HbSD. Sixteen out of 42 patients with HbSS were on hydroxyurea (HU). Urinary excretion of TGF-ß1 was 26.4 ± 1.5 pg/mgCr in SCD subjects vs 15.0 ± 2.4 pg/mgCr in CTR (p<0.00001). SCD patients with hemoglobin < 9 g/dl had higher urinary TGF-ß1 than patients with milder anemia (p=0.002). Urinary TGF-ß1 trended lower in HbSS patients treated with HU (23.61 ± 2.6 pg/mgCr), vs patients not on HU (27.69 ± 1.8 pg/mgCr; p=0.055). There was no correlation between urinary TGF-ß1 and microalbuminuria or estimated glomerular function. There was no difference in urinary NGAL in SCD patients vs CTR. We suggest that urinary TGF-ß1 may serve as a marker of early renal injury in SCD.

[Correlation between urinary biomarkers and pathological lesions in drug-induced tubulointerstitial nephritis].

OBJECTIVE: To find some urinary biomarkers for differential diagnosis in drug-induced tubulointerstitial nephritis (DTIN) by accessing the relationship between their changes and the pathological processes of DTIN. METHODS: Forty patients biopsy-proven DTIN and 24 healthy controls were enrolled in this study. The urine samples of DTIN patients were collected on the day of biopsy and were measured for the following biomarkers: urinary TGF-ß with enzyme-linked immunosorbent assay (ELISA); urinary IL-6 with radio-immunoassay; N-acetyl-ß-D-glucosaminidase (NAG) with enzyme-substrate colorimetric assay; α1-microglobulin (α1-MG) with immune transmission turbidity method. Meanwhile, the pathological changes on renal biopsy were analyzed semi-quantitatively and scored. The relationship between these pathological changes and the urinary biomarkers were analyzed and receiver operating characteristic curve (ROC curve) of those biomarkers in distinguishing different pathological lesions were constructed. RESULTS: Urinary α1-MG levels were positively correlated to interstitial edema, inflammatory cell infiltration and tubular atrophy, while urinary IL-6 were negatively correlated to the above parameters. Urinary NAG had positive relationship with inflammatory cell infiltration and tubular atrophy, while urinary TGF-ß had positive relationship with tubular atrophy. ROC analysis results indicated that the area under curve (AUC) of α1-MG was larger than that of NAG (0.797 vs 0.734, P<0.05). Combined measurement of α1-MG and NAG could enhance the sensitivity of detecting interstitial inflammatory cell infiltration. For detecting interstitial edema and tubular atrophy, only α1-MG had statistical significance (AUC=0.723, P=0.027; AUC=0.774, P=0.008, respectively). CONCLUSIONS: Urinary α1-MG and NAG levels can reflect the severity of interstitial inflammatory infiltration, combined measurement of both can enhance sensitivity. Urinary α1-MG level can also reflect the degree of interstitial edema and tubular atrophy.

[Urinary excretion of proinflammatory cytokines and transforming growth factor beta at early stages of diabetic nephropathy].

AIM: To examine correlations between urine excretion of proinflammatory cytokines, transforming growth factor beta (TGF-b) and changes in renal structure and function, quality of glycemia control in patients with type 1 diabetes mellitus. MATERIAL AND METHODS: Urinary excretion of interleukine 1-beta (IL-1b), monocytic chemoattractive protein-1 (MCP-1), RANTES and TGF-b was measured with enzyme immunoassay in 57 patients including 22 patients with normal albuminuria, 23--with microalbuminuria, 12--with macroalbuminuria. Creatinine clearance was subnormal in 8 patients with macroalbuminuria. The control group consisted of 10 healthy persons. Morphological examination of renal biopsies was performed in 8 patients with normoalbuminuria and 10 patients with microalbuminuria. RESULTS: Patients with normoalbuminuria had excretion of MCP-1 significantly higher than in controls. Microalbuminuria patients showed high excretion of IL-1b, MCP-1 and TGF-b. Excretion of IL-1b, MCP-1, RANTES and TGF-b in patients with macroalbuminuria was higher than in controls and other groups of patients. Excretion of cytokines and TGFb correlated inversely with glomerular filtration rate and hemoglobin level. Positive correlations were detected between excretion of IL-1b, MCP-1, TGFb and glycated hemoglobin A(1c). In patients with normo- and microalbuminuria cytokine and TGFb excretion correlated with thickness of glomerular and glomerular basal membrane. CD68-positive macrophages were detected in the intersticium of 1 patient with normoalbuminuria and 6 patients with microalbuminuria. CONCLUSION: Urinary excretion of proinflammatory cytokines and TGF-b was elevated in patients with DM-1 having micro- and macroalbuminuria suggesting participation of inflammation in development of diabetic nephropathy.