ABSTRACT
The levels of the MMPs in the biological samples of confirmed patients with gastric cancer are significantly elevated compared to those found in healthy people. Therefore, a novel 3D stochastic microsensor based on graphene oxide, modified with gold nanoparticles and (Z)-N-(pyridin-4-yl-methyl) octadec-9-enamide (namely N2-AuNP/GO), was designed for the determination of MMP-2 in biological samples, and validated for the screening tests of biological samples in order to be used for the early diagnosis of gastric cancer. The proposed sensor presents a low limit of quantification (1.00 × 10-22 g mL-1), high sensitivity (1.84 × 107 s-1 g-1 mL), and a wide working concentration range (1.00 × 10-22-1.00 × 10-7 g mL-1). Recovery values higher than 99.15% were recorded for the assay of MMP-2 in whole blood, gastric tissue tumors, saliva, and urine samples.
Subject(s)
Gold , Graphite , Matrix Metalloproteinase 2 , Metal Nanoparticles , Graphite/chemistry , Humans , Matrix Metalloproteinase 2/blood , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 2/metabolism , Metal Nanoparticles/chemistry , Gold/chemistry , Stomach Neoplasms/diagnosis , Stomach Neoplasms/blood , Biosensing Techniques/methodsABSTRACT
IMPORTANCE: Extracellular matrix proteins and enzymes involved in degradation have been found to be associated with tissue fibrosis and ureteropelvic junction obstruction (UPJO). In this study we developed a promising urinary biomarker model which can identify reduced renal function in UPJ obstruction patients. This can potentially serve as a non-invasive way to enhance surgical decision making for patients and urologists. OBJECTIVE: We sought to develop a predictive model to identify UPJO patients at risk for reduced renal function. DESIGN: Prospective cohort study. SETTING: Pre-operative urine samples were collected in a prospectively enrolled UPJO biomarker registry at our institution. Urinary MMP-2, MMP-7, TIMP-2, and NGAL were measured as well as clinical characteristics including hydronephrosis grade, differential renal function, t1/2, and UPJO etiology. PARTICIPANTS: Children who underwent pyeloplasty for UPJO. MAIN OUTCOME MEASUREMENT: Primary outcome was reduced renal function defined as MAG3 function <40%. Multivariable logistic regression was applied to identify the independent predictive biomarkers in the original Training cohort. Model validation and generalizability were evaluated in a new UPJO Testing cohort. RESULTS: We included 71 patients with UPJO in the original training cohort and 39 in the validation cohort. Median age was 3.3 years (70% male). By univariate analysis, reduced renal function was associated with higher MMP-2 (p = 0.064), MMP-7 (p = 0.047), NGAL (p = 0.001), and lower TIMP-2 (p = 0.033). Combining MMP-7 with TIMP-2, the multivariable logistic regression model predicted reduced renal function with good performance (AUC = 0.830; 95% CI: 0.722-0.938). The independent testing dataset validated the results with good predictive performance (AUC = 0.738). CONCLUSIONS AND RELEVANCE: Combination of urinary MMP-7 and TIMP-2 can identify reduced renal function in UPJO patients. With the high sensitivity cutoffs, patients can be categorized into high risk (aggressive management) versus lower risk (observation).
Subject(s)
Hydronephrosis , Matrix Metalloproteinase 7 , Tissue Inhibitor of Metalloproteinase-2 , Ureteral Obstruction , Biomarkers/urine , Child , Child, Preschool , Female , Humans , Hydronephrosis/etiology , Hydronephrosis/urine , Kidney/physiopathology , Kidney Pelvis/physiopathology , Lipocalin-2/urine , Male , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 7/urine , Prospective Studies , Tissue Inhibitor of Metalloproteinase-2/urine , Ureteral Obstruction/complications , Ureteral Obstruction/surgery , Ureteral Obstruction/urineABSTRACT
Renal tubulointerstitial fibrosis caused by congenital ureteropelvic junction obstruction (UPJO) may lead to the development of obstructive nephropathy (ON) and the impairment of kidney function. Hence, the identification of early biomarkers of this condition might be of assistance in therapeutic decisions. This study evaluates serum and urinary metalloproteinases MMP-1, MMP-2, and MMP-9 and tissue inhibitors of metalloproteinases TIMP-1 and TIMP-2 as potential biomarkers of ON in children with congenital unilateral hydronephrosis (HN) caused by UPJO. Forty-five (45) children with congenital HN of different grades of severity and twenty-one (21) healthy controls were enrolled in the study. Urinary and serum concentrations of MMP-1, MMP-2, MMP-9, TIMP-1 and TIMP-2 were measured using specific ELISA kits. The urinary excretions were expressed as biomarker/creatinine (Cr) ratios. To evaluate the extracellular matrix remodelling process activity, the serum and urinary MMP-1, -2, -9/TIMP-1, -2 ratios were also calculated. In comparison with the controls, patients with HN, independent of the grade, showed significantly increased median serum MMP-9, TIMP-1, and TIMP-2, median urinary MMP-9/Cr, and TIMP-2/Cr ratios. Lower median values of serum MMP-2/TIMP-1, MMP-9/TIMP-1 in patients with HN were also revealed. Additionally, higher urinary MMP-2/Cr, lower urinary MMP-2/TIMP-2, and lower serum MMP-9/TIMP-2 ratios were observed in patients with HN grades 3 and 4. Patients with ON diagnosed by renal scintigraphy had a significantly higher median serum MMP-9 concentration and lower median serum MMP-9/TIMP-1, -2 ratios in comparison with those without this condition. Patients with nonglomerular proteinuria had a significantly higher median serum TIMP-1 concentration, a higher median urinary TIMP-2/Cr ratio, and a lower serum MMP-9/TIMP-1 ratio compared to those without this symptom. The relationship between the measured biomarkers and the relative function of the obstructed kidney showed no correlations. The ROC curve analysis showed a promising diagnostic profile for the detection of ON for serum MMP-9 and the serum MMP-9/TIMP-1 and MMP-9/TIMP-2 ratios. In conclusion, the results of this study suggest that patients with HN, particularly with grades 3 and 4, are at higher risk of renal tubulointerstitial fibrosis. The noninvasive markers of this condition considered are urinary MMP-2/Cr and MMP-9/Cr, serum MMP-9, serum and urinary MMP-2, MMP-9/TIMP-1, -2. Additionally, serum MMP-9 and MMP-9/TIMP-1, -2 may become promising markers of ON.
Subject(s)
Hydronephrosis/congenital , Kidney Tubules/pathology , Matrix Metalloproteinases, Secreted/blood , Matrix Metalloproteinases, Secreted/urine , Tissue Inhibitor of Metalloproteinases/blood , Tissue Inhibitor of Metalloproteinases/urine , Adolescent , Biomarkers/blood , Biomarkers/urine , Case-Control Studies , Child , Child, Preschool , Creatinine/blood , Creatinine/urine , Female , Fibrosis , Humans , Hydronephrosis/blood , Hydronephrosis/urine , Male , Matrix Metalloproteinase 1/blood , Matrix Metalloproteinase 1/urine , Matrix Metalloproteinase 2/blood , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/blood , Matrix Metalloproteinase 9/urine , Tissue Inhibitor of Metalloproteinase-1/blood , Tissue Inhibitor of Metalloproteinase-1/urine , Tissue Inhibitor of Metalloproteinase-2/blood , Tissue Inhibitor of Metalloproteinase-2/urineABSTRACT
AIMS: To assess the predictive values of six urinary markers (nerve growth factor [NGF], brain-derived neurotrophic factor [BDNF], matrix metalloproteinase 2 [MMP-2], tissue inhibitor metalloproteinase 2 [TIMP-2], transformation growth factor ß-1 [TGF-B1], and prostaglandin 2 [PGE2]) for adverse urodynamic features and for upper urinary tract damage in adult patients with spina bifida. MATERIALS AND METHODS: A single-center prospective trial was conducted from March 2015 to March 2017 including all consecutive adult patients with spina bifida seen for urodynamic testing. The urine was collected and stored at -80°C. A urodynamic and an upper urinary tract were systematically performed. At the end of the inclusion period, urines were defrosted and urinary nerve growth factor, BDNF, TIMP-2, and TGF-B1 were assessed using validated ELISA kits. The urinary markers levels were adjusted on the urinary creatinine level. Urinary MMP-2 levels were assessed by zymography. RESULTS: Fourty patients were included. Only TIMP-2 and MMP-2 were significantly associated with poor bladder compliance (P = .043 and P = .039, respectively). TIMP-2 was also the only urinary marker significantly associated with upper urinary tract damage on imaging (OR = 19.81; P = .02). Of all urodynamic parameters, bladder compliance and maximum detrusor pressure were the only ones associated with upper urinary tract damage on imaging (P = .01 and P = .02), The diagnostic performances of urinary TIMP-2 for upper urinary tract damage were slightly superior to PdetMax and bladder compliance with an area under the curve of 0.72. CONCLUSION: Urinary TIMP-2 and MMP-2 were significantly associated with poor bladder compliance and urinary TIMP-2 was significantly associated with upper urinary tract damage. These findings support a pathophysiological role of extracellular matrix remodeling in poor bladder compliance of adult patients with spina bifida.
Subject(s)
Spinal Dysraphism/physiopathology , Urinary Bladder, Neurogenic/urine , Adult , Atrophy , Biomarkers/urine , Brain-Derived Neurotrophic Factor/urine , Compliance/physiology , Dinoprostone/urine , Female , Humans , Hydronephrosis/diagnostic imaging , Kidney/diagnostic imaging , Kidney/pathology , Male , Matrix Metalloproteinase 2/urine , Middle Aged , Nerve Growth Factor/urine , Prospective Studies , Spinal Dysraphism/complications , Tissue Inhibitor of Metalloproteinase-2/urine , Transforming Growth Factor beta1/urine , Urinary Bladder, Neurogenic/etiology , Urinary Bladder, Neurogenic/physiopathology , Urodynamics , Young AdultABSTRACT
The present study assessed protein and gene expression levels of tissue inhibitor of metalloproteinase-2 (TIMP-2), matrix metalloproteinase-2 (MMP-2), and MMP-9 in urine and blood samples of 50 patients with bladder carcinoma. The expression of TIMP-2, MMP-2, and MMP-9 levels with tumor stage and grade was also assessed. Results showed that the expression levels of MMP-2 and MMP-9 in both blood and urine were significantly elevated in group 1 when compared with groups 2 and 3 healthy subjects. The discriminatory ability in the diagnosis of bladder carcinoma of MMP-2 and MMP-9 expression was confirmed by receiver operating characteristic curve analysis that revealed a sensitivity and specificity of 100%. MMP-2 and MMP-9 levels were not correlated with grade or stage of the tumor. With respect to TIMP-2 blood and urine levels, results showed a significant decrease in gene expression levels in bladder carcinoma group, whereas, TIMP-2 protein showed a significant increase in bladder carcinoma.
Subject(s)
Biomarkers, Tumor/metabolism , Early Detection of Cancer , Matrix Metalloproteinase 2/metabolism , Matrix Metalloproteinase 9/metabolism , Tissue Inhibitor of Metalloproteinase-2/metabolism , Urinary Bladder Neoplasms/diagnosis , Adult , Aged , Biomarkers, Tumor/blood , Biomarkers, Tumor/urine , Blotting, Western , Case-Control Studies , Female , Humans , Male , Matrix Metalloproteinase 2/blood , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/blood , Matrix Metalloproteinase 9/genetics , Matrix Metalloproteinase 9/urine , Middle Aged , Prognosis , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity , Tissue Inhibitor of Metalloproteinase-2/blood , Tissue Inhibitor of Metalloproteinase-2/genetics , Tissue Inhibitor of Metalloproteinase-2/urine , Urinary Bladder Neoplasms/blood , Urinary Bladder Neoplasms/enzymology , Urinary Bladder Neoplasms/urineABSTRACT
Objective: Histopathological changes in the kidney in type 1 diabetes mellitus (T1DM) begin before detection of microalbuminuria. Therefore, there is interest in finding a better biomarker for the early detection of diabetic kidney injury. The aim of this present study was to determine whether urinary indicators of fibrosis are detectable early in the development of T1DM in children and if they may predict progressive renal injury. Methods: Urinary matrix metalloproteinase 2 and 9 (MMP2 and MMP9), tissue inhibitor of metalloproteinase 1 and 2 (TIMP1 and TIMP2) and transforming growth factor-ß1 (TGF-ß1) were assessed in 33 patients with T1DM with normal renal functions and in 24 healthy controls. Microalbuminuria was not present in the patient group with the exception of three patients. The results were adjusted to urine creatinine (Cr) and the differences between patients and controls were evaluated. These measurements were repeated after one year and the results were compared with the first year results. Results: Urine MMP2/Cr, MMP9/Cr, TIMP1/Cr, TIMP2/Cr, TGF-ß1/Cr were not different between the patient and control groups (p>0.05). There were also no significant differences between the first and second year results for these biomarkers (p>0.05). None of these parameters were correlated with hemoglobin A1c, body mass index and duration of T1DM. Interestingly, all parameters were negatively correlated to age of onset of T1DM (p<0.05). Conclusion: Our findings suggest that urinary biomarkers of fibrosis do not show an increase in diabetic children without microalbuminuria. The results also indicate that the risk of early fibrosis may increase as age of onset of T1DM decreases.
Subject(s)
Biomarkers/urine , Diabetes Mellitus, Type 1/physiopathology , Diabetes Mellitus, Type 1/urine , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Tissue Inhibitor of Metalloproteinase-1/urine , Tissue Inhibitor of Metalloproteinase-2/urine , Adolescent , Case-Control Studies , Child , Child, Preschool , Female , Follow-Up Studies , Humans , Male , Prognosis , Transforming Growth Factor beta1/urineABSTRACT
BACKGROUND: Preeclampsia, a pregnancy disorder characterized by hypertension and proteinuria, represents the leading cause of fetal and maternal morbidity and mortality in developing countries. The identification of novel and accurate biomarkers that are predictive of preeclampsia is necessary to improve the prognosis of patients with preeclampsia. OBJECTIVE: The objective of this study is to evaluate the usefulness of nine urinary metalloproteinases to predict the risk of preeclampsia development. METHODS: MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-10, MMP-12 and MMP-13 were analyzed in urine (early-pregnancy) from 17 women predicted to develop preeclampsia and 48 controls using the Bio-Plex Pro-Human MMP panel (Bio-Rad, Hercules, CA). RESULTS: Urinary MMP-2 showed differences between groups which allowed us to calculate an increased risk for PE development of up to 20 times among the study population. CONCLUSION: Increased urinary concentration of MMP-2 at 12 and 16 weeks of gestation predicted an increased risk of developing preeclampsia in the study population.
Subject(s)
Biomarkers/urine , Matrix Metalloproteinase 2/urine , Pre-Eclampsia/diagnosis , Pre-Eclampsia/urine , Adolescent , Adult , Female , Gestational Age , Humans , Matrix Metalloproteinases/urine , Predictive Value of Tests , Pregnancy , Prognosis , Risk Factors , Young AdultABSTRACT
Proteinases secreted by the prostate gland have a reproductive function in cleaving proteins in the ejaculate and in the female reproductive tract, but some may have a fundamental role in disease and pathological processes including cancer. The purpose of this study was to determine if there were differences in proteinase activities in urine samples collected following prostate massage of men positive (CaP) or negative (no evidence of malignancy, NEM) for biopsy determined prostate cancer. Matrix metalloproteinase (MMP) and serine proteinase activities were detected using protein substrate zymography. There were no differences in activities of MMP-2, proMMP-9, and MMP-9/NGAL (neutrophil gelatinase associated lipocalin) complex (gelatin substrate) in men with detected prostate cancer, although the latter two were somewhat diminished. A caseinolytic activity of about 75kDa inhibited by calcium did not differ between the NEM and CaP groups. Heparin stimulated calcium sensitive gelatinolytic activities of approximately 22, 42, and 60kDa, but did not affect activities of MMP-2, MMP-9, or the 75kDa caseinolytic activity. The 22, 42, and 60kDa activities appear to be serine proteinases since they were inhibited by benzamidine. There was a significant decrease in the 22kDa heparin-stimulated serine proteinase activity in urines of men with cancer. Proteinase expression and activities, perhaps in combination with other potential markers, may prove useful in urine for detection and evaluation of prostate cancer.
Subject(s)
Biomarkers, Tumor/urine , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Prostatic Neoplasms/urine , Serine Proteases/urine , Aged , Benzamidines/administration & dosage , Calcium/metabolism , Heparin/chemistry , Humans , Lipocalin-2/urine , Male , Middle Aged , Prostatic Neoplasms/enzymology , Prostatic Neoplasms/pathologyABSTRACT
Matrix metalloproteinases (MMPs) are regulated at multiple transcriptional and post-transcriptional levels, among which receptor-mediated endocytic clearance. We previously showed that low-density lipoprotein receptor-related protein-1 (LRP-1) mediates the clearance of a complex between the zymogen form of MMP-2 (proMMP-2) and tissue inhibitor of metalloproteinases, TIMP-2, in HT1080 human fibrosarcoma cells. Here we show that, in BN16 rat yolk sac cells, proMMP-2:TIMP-2 complex is endocytosed through a distinct LRP member, megalin/LRP-2. Addition of receptor-associated protein (RAP), a natural LRP antagonist, caused accumulation of endogenous proMMP-2 and TIMP-2 in conditioned media. Incubation with RAP also inhibited membrane binding and cellular uptake of exogenous iodinated proMMP-2:TIMP-2. Moreover, antibodies against megalin/LRP-2, but not against LRP-1, inhibited binding of proMMP-2:TIMP-2 to BN16 cell surface. BIAcore analysis confirmed direct interaction between the complex and megalin/LRP-2. Conditional renal invalidation of megalin/LRP-2 in mice resulted in accumulation of proMMP-2 and TIMP-2 in their urine, highlighting the physiological relevance of the binding. We conclude that megalin/LRP-2 can efficiently mediate cell-surface binding and endocytosis of proMMP-2:TIMP-2 complex. Therefore megalin/LRP-2 can be considered as a new actor in regulation of MMP-2 activity, an enzyme crucially involved in many pathological processes.
Subject(s)
Enzyme Precursors/metabolism , Gelatinases/metabolism , Matrix Metalloproteinase 2/metabolism , Animals , Cell Line , Cell Membrane/metabolism , Culture Media, Conditioned , Enzyme Precursors/urine , Gelatinases/urine , Kidney/metabolism , Low Density Lipoprotein Receptor-Related Protein-2 , Matrix Metalloproteinase 2/urine , Mice , Mice, Transgenic , Multiprotein Complexes/metabolism , Protein Binding , Protein Transport , Rats , Tissue Inhibitor of Metalloproteinase-2/metabolismABSTRACT
Changes in immune and inflammatory responses may play a crucial role in the development and progression of atherosclerosis, as an autoimmune, chronic and progressive inflammatory disease. Immunological activity and vascular inflammation during atherosclerosis can be modulated by autoimmune responses against self-antigens, according to changeable risk factors (cholesterol, oxidized low-density lipoprotein (ox-LDL) in the vascular wall, fatty acids, etc.), and accompanied by accumulation of leucocytes and proinflammatory cytokines, which stimulate the transcription of matrix metalloproteinases (MMPs), whose concentration are increased in foam cell-rich regions. Regulatory T cells (Tregs) represent a unique subpopulation of T cells specialized in the regulation of immune response and in the suppression of proatherogenic T cells. The aim of our study was to examine the interactions between the concentration of enzyme matrix metalloproteinases 2 and 9 (MMP-2 and 9) in urine and the percentage of Tregs in peripheral blood of two groups of patients: with carotid artery stenosis (CAS), undergoing surgery and with mild atherosclerosis (A) from general practice. The method of enzyme immunoassay (ELISA) was used to determine enzyme MMP expression, and Tregs was examined by flow cytometric analysis. Our data have showed a large increase in the enzyme MMP-2 and 9 in the urine of CAS and A patients in comparison with healthy controls and indicated this method as an easy marker for the monitoring of the development of atherosclerosis. Simultaneously, the diminished number of Tregs in the same patients pointed the importance of these regulatory mechanisms in the etiopathogenesis of atherosclerosis and possible Tregs-mediated therapy.
Subject(s)
Atherosclerosis/immunology , Matrix Metalloproteinase 2/immunology , Matrix Metalloproteinase 9/immunology , T-Lymphocytes, Regulatory/immunology , Aged , Aged, 80 and over , Atherosclerosis/blood , Atherosclerosis/urine , Carotid Stenosis/blood , Carotid Stenosis/immunology , Carotid Stenosis/urine , Cholesterol/immunology , Cholesterol/metabolism , Cytokines/immunology , Cytokines/metabolism , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Global Burden of Disease/statistics & numerical data , Humans , Inflammation Mediators/immunology , Inflammation Mediators/metabolism , Lipoproteins, LDL/immunology , Lipoproteins, LDL/metabolism , Male , Matrix Metalloproteinase 2/metabolism , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/metabolism , Matrix Metalloproteinase 9/urine , Middle Aged , Protein Binding , Risk FactorsABSTRACT
Hernández-Hernández, Magda Elena, Jaime Morales-Romero, Clara Luz Sampieri, Diego Jesús Luna Lozano, Isidra del Carmen Valencia Lezama, Mónica Janett Muñoz Contreras, and Arturo Rodríguez Hernández. Association of urinary activity of MMP-2 with microalbuminuria in an isolated sample of subjects living in high altitude rural locations in México. High Alt Med Biol. 18:209-218, 2017.-Matrix metalloproteinases (MMP) are implicated in remodeling of the renal extracellular matrix. In a cross-sectional study we evaluated renal impairment in general population of high-altitude rural locations in México. Multivariable analysis was performed to identify the association between MMP-2 and MMP-9 and microalbuminuria. Twenty-eight (20.9%) subjects with renal impairment (WRI) and 106 (79.1%) without renal impairment were included. No differences were found relating to sex, location, marital status, current habits, weight, height, body mass index, waist size in males, creatinine in males, and uric acid. In contrast, differences were found among age, level of education, waist size in general and in females, creatinine in general and in females, urinary albumin, urea, glucose, total cholesterol, and triglycerides. Proportions of hypertension, type 2 diabetes mellitus, central abdominal obesity, hypertriglyceridemia, and hypercholesterolemia were greater in the group WRI. Presence of urinary MMP-2 or of both urinary gelatinases and arbitrary unit (AU) values ≥P90 were associated with microalbuminuria. We conclude that AU values ≥P90 of urinary MMP-2 (OR = 20.1, p = 0.002) is associated with microalbuminuria.
Subject(s)
Albuminuria/etiology , Altitude , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Adult , Cross-Sectional Studies , Female , Humans , Male , Mexico , Middle Aged , Multivariate Analysis , Risk Factors , Rural PopulationABSTRACT
BACKGROUND/AIMS: Renal ischemia-reperfusion (I-R) injury (IRI) is an inseparable feature of organ transplantation and may have a negative impact on the graft, its function and survival. Acute tubular necrosis, which is reversible thanks to the regenerative capacity of renal tubular epithelial cells, is the main cause of acute renal failure secondary to IRI. MMP-2 and MMP-9 are proteolytic enzymes involved in digesting proteins that are components of the extracellular matrix (ECM) and the basement membrane of the nephrons. This way post-reperfusion MMP activation allows the inflammatory process to spread. METHODS: In our studies, we focused on identifying whether the concentrations of MMP-2 and MMP-9 and their natural inhibitors TIMP-1 and TIMP-2 in urine sample at day 1 and day 30 as well as after 12 months following renal transplantation are markers of early and long-term renal function during meanly five-years observation. Moreover, in urine sampled at months 6 and 12 after renal transplantation, we determined the content of TGF-ß as a graft fibrosis indicator. RESULTS: MMP-9 concentration in the early post-transplant period is a major marker of early and long-term function of the transplanted kidney. Its increased concentration was correlated with lesions related to tubular atrophy and fibrosis in renal biopsies performed at months 3 and 12 after transplantation. Its concentration is correlated with TGF-ß content in a later period. CONCLUSIONS: TIMP-1 and-2 are primarily markers of an early function of the transplanted kidney. Early post-transplant concentration of MMP-2 is a marker of proteinuria in early and long-term post-transplant periods.
Subject(s)
Graft Survival , Kidney Transplantation , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Tissue Inhibitor of Metalloproteinase-1/urine , Tissue Inhibitor of Metalloproteinase-2/urine , Atrophy , Biomarkers/urine , Fibrosis , Follow-Up Studies , Humans , Time FactorsABSTRACT
CONTEXT: Epithelial-mesenchymal transition (EMT) leads to renal fibrosis and chronic kidney disease (CKD). OBJECTIVE: The aim of this study was to assess the usefulness of survivin, E-cadherin and metalloproteinases (MMPs) as biomarkers of CKD-related complications. MATERIAL AND METHODS: Survivin, E-cadherin, MMP-2, MMP-9 and TGFbeta1 were assessed by ELISA in 41 children with CKD stages 3 to 5 and in 23 controls. RESULTS: The serum and urine values of analyzed parameters were significantly elevated in CKD patients versus controls and correlated with each other. CONCLUSIONS: The observed parameter changes indicate apoptosis, tissue remodeling and fibrosis in CKD children. Urine survivin may become a new biomarker of kidney-specific EMT.
Subject(s)
Cadherins/urine , Inhibitor of Apoptosis Proteins/urine , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Renal Insufficiency, Chronic/urine , Adolescent , Apoptosis , Biomarkers/blood , Biomarkers/urine , Cadherins/blood , Case-Control Studies , Child , Child, Preschool , Cross-Sectional Studies , Enzyme-Linked Immunosorbent Assay , Epithelial-Mesenchymal Transition , Female , Fibrosis , Glomerular Filtration Rate , Humans , Infant , Inhibitor of Apoptosis Proteins/blood , Male , Matrix Metalloproteinase 2/blood , Matrix Metalloproteinase 9/blood , Renal Insufficiency, Chronic/blood , Renal Insufficiency, Chronic/diagnosis , Renal Insufficiency, Chronic/pathology , Survivin , Transforming Growth Factor beta1/blood , Transforming Growth Factor beta1/urineABSTRACT
We have developed a new amplification system for proteinases that is sensitive, simple, and inexpensive to run, exemplified by a horseradish peroxidase (HRP)-conjugated, dual MMP2 (matrix metalloproteinase 2) and ADAM8 (a disintegrin and metalloproteinase 8) peptide substrate assay presented herein. The HRP-conjugated substrate is attached to beads through a 6× histidine tag and then incubated with the target enzyme, cleaving the HRP reporter. This product is subsequently removed from the unreacted bound portions of the substrate by magnetic deposition of the beads. The amount of product is then quantified using a standard HRP color development assay employing 3,3',5,5'-tetramethylbenzidine (TMB) and hydrogen peroxide (H2O2). This HRP amplification system represents a new approach to proteinase assays and could be applied to other enzymes, such as lipases, esterases, and kinases, as long as the unreacted substrate can be physically separated from the product and catalysis by the enzyme to be quantified is not impaired dramatically by steric hindrance from the HRP entity.
Subject(s)
ADAM Proteins/metabolism , Colorimetry/methods , Enzyme Assays/methods , Matrix Metalloproteinase 2/metabolism , Membrane Proteins/metabolism , Amino Acid Sequence , Benzidines/chemistry , Dipeptides/pharmacology , Horseradish Peroxidase/metabolism , Humans , Hydrogen Peroxide/chemistry , Kinetics , Magnets/chemistry , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase Inhibitors/pharmacology , Microspheres , Nitrilotriacetic Acid/analogs & derivatives , Nitrilotriacetic Acid/chemistry , Organometallic Compounds/chemistry , Peptides/chemistry , Peptides/metabolism , Substrate SpecificityABSTRACT
OBJECTIVES: Hepatocellular carcinoma (HCC) is a highly vascularized tumor. In this study, we investigated the prognostic and predictive values of proangiogenic factors in HCC patients receiving radiotherapy. METHODS: Between September 2008 and December 2009, a total of 50 patients treated with radiotherapy were prospectively enrolled in this study. Serum and urine samples were collected <1 week before and after radiotherapy. RESULTS: After completion of radiotherapy, serum vascular endothelial growth factor (VEGF)/platelet (Plt) levels were significantly increased (p < 0.01). Patients who experienced hepatic tumor recurrence outside the radiation field showed higher VEGF-A/Plt levels before and after radiotherapy than patients who did not (p = 0.04), whereas patients who had hepatic tumor recurrence inside the radiation field showed significantly higher matrix metalloproteinase (MMP)-2 levels after radiotherapy (p = 0.04). On multivariate analyses, a high level of either VEGF/Plt or MMP-2 (≥median) before radiotherapy was a significant independent prognostic factor for a worse progression-free survival (p = 0.04). CONCLUSIONS: In HCC patients receiving radiotherapy, levels of VEGF/Plt and MMP-2 before radiotherapy can be useful to predict treatment outcome. This study also suggests the necessity of anti-angiogenic therapy, such as sorafenib, since radiotherapy increases VEGF/Plt levels, and higher levels of VEGF/Plt are associated with a poor outcome.
Subject(s)
Biomarkers, Tumor/analysis , Carcinoma, Hepatocellular/radiotherapy , Liver Neoplasms/radiotherapy , Matrix Metalloproteinase 2/urine , Vascular Endothelial Growth Factor A/blood , Adult , Aged , Analysis of Variance , Carcinoma, Hepatocellular/blood , Carcinoma, Hepatocellular/mortality , Cohort Studies , Disease-Free Survival , Female , Humans , Kaplan-Meier Estimate , Liver Neoplasms/blood , Liver Neoplasms/mortality , Male , Middle Aged , Predictive Value of Tests , Prognosis , Proportional Hazards Models , Prospective Studies , Republic of Korea , Risk Assessment , Survival Analysis , Treatment OutcomeABSTRACT
BACKGROUND: The most frequent cause of kidney allograft loss is chronic allograft injury, often with proteinuria as the clinical feature. Occurrence of proteinuria late after kidney transplantation is associated with worse graft function and patient survival. AIM: The aim of the study was to assess plasma and urine matrix metalloproteinases (MMP-2 and MMP-9) and tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) in proteinuric renal transplant recipients (RTRs). The factors were determined by enzyme-linked immunosorbent assay in 150 RTRs (51 women and 99 men), aged 49.2 ± 11.5 years, at mean 73.4 ± 41.2 months after kidney transplantation (range: 12 to 240 months). RESULTS: Proteinuric RTRs compared with non-proteinuric RTRs had higher median plasma MMP-2 (P = .012), TIMP-1 (P = .0003), and TIMP-2 (P = .0021) concentrations, as well as higher urine MMP-2 (P < .0001) excretion. The presence of proteinuria had no impact on plasma MMP-9 and urine MMP-9, TIMP-1, and TIMP-2. Proteinuria and estimated daily proteinuria (uPr:uCr) correlated positively with plasma MMP-2 (rs = 0.226, P = .0054 and rs = 0.241, P = .003), TIMP-1 (rs = 0.305, P = .00015 and rs = 0.323, P = .000055), TIMP-2 (rs = 0.273, P = .0007 and rs = 0.269, P = .001) and urine MMP-2 (rs = 0.464, P < .0001 and rs = 0.487, P < .0001), respectively. Proteinuric RTRs had impaired graft function with higher median serum creatinine concentrations (1.91 [1.60-2.43] mg/dL versus 1.41 [1.20-1.65] mg/dL, P < .00001) and lower estimated glomerular filtration rate (36 [28-45] mL/min/1.73 m(2) versus 53 [43-61] mL/min/1.73 m(2), P < .00001) than RTRs without proteinuria. CONCLUSIONS: Our research revealed that in RTRs, proteinuria was significantly associated with increased concentrations of enzymes involved in extracellular matrix (ECM) degradation: plasma MMP-2, TIMP-1, TIMP-2, and urine MMP-2. Findings strongly emphasize increased plasma TIMPs in proteinuric RTRs that inhibit degradation of ECM by MMPs and favor excessive deposition of ECM proteins.
Subject(s)
Kidney Transplantation , Matrix Metalloproteinase 2/urine , Tissue Inhibitor of Metalloproteinase-1/blood , Tissue Inhibitor of Metalloproteinase-2/blood , Transplant Recipients , Adult , Enzyme-Linked Immunosorbent Assay , Female , Humans , Male , Matrix Metalloproteinase 9/blood , Middle Aged , Proteinuria/metabolism , Transplantation, HomologousABSTRACT
FDA approval of the first device to use novel biomarkers of kidney damage to assess risk of acute kidney injury (AKI) potentially brings forward diagnosis of moderate-to-severe AKI to a time frame that could enable early intervention. Although the device awaits greater scrutiny, its approval marks the beginning of a new era.
Subject(s)
Acute Kidney Injury/diagnosis , Acute Kidney Injury/urine , Cell Cycle Checkpoints , Early Diagnosis , Insulin-Like Growth Factor Binding Proteins/urine , Matrix Metalloproteinase 2/urine , Acute Kidney Injury/pathology , Acute Kidney Injury/prevention & control , Biomarkers/urine , Female , Fluoroimmunoassay/methods , Humans , Male , Point-of-Care Systems , Sensitivity and Specificity , United States , United States Food and Drug AdministrationABSTRACT
BACKGROUND: A majority of patients with pancreatic malignancies, including both pancreatic ductal adenocarcinoma (PDAC) and pancreatic neuroendocrine tumours (pNETs), present with advanced disease due to a lack of specific symptoms and current diagnostic limitations, making this disease extremely difficult to detect. Our goal was to determine whether urinary matrix metalloproteases (uMMPs) and/or their endogenous inhibitors, urinary tissue inhibitor of metalloproteases (uTIMPs), could be detected in the urine of patients with pancreatic malignancies and whether they may serve as independent predictors of disease status. METHODS: Retrospective analyses of urine samples (n=139) from PDAC and pNET patients as well as age- and sex-matched controls were conducted. Urinary MMP-2 and uTIMP-1 levels were determined using ELISA and zymography. Biomarker expression in tumour and normal pancreatic tissues was analysed via immunohistochemistry (IHC). RESULTS: Multivariable logistic regression analyses indicated that, when controlling for age and sex, uMMP-2 (P<0.0001) and uTIMP-1 (P<0.0001) but not uMMP-9, were significant independent predictors for distinguishing between PDAC patients and healthy controls. Our data also indicated that uMMP-2 was an independent predictor of the presence of pNET. In addition, uTIMP-1 levels could differentiate the two cancer groups, PDAC and pNET, respectively. Immunohistochemistry analysis confirmed that MMP-2 and TIMP-1 protein expression is significantly upregulated in PDAC tissue compared with the normal pancreas. CONCLUSIONS: Taken together, our results suggest that the detection of uMMP-2 and uTIMP-1 may have diagnostic value in the detection of pancreatic malignancies and that uTIMP-1 may be useful in distinguishing between pancreatic adenocarcinoma and neuroendocrine tumours.
Subject(s)
Biomarkers, Tumor/urine , Carcinoma, Pancreatic Ductal/urine , Matrix Metalloproteinase 2/urine , Neuroendocrine Tumors/urine , Pancreatic Neoplasms/urine , Tissue Inhibitor of Metalloproteinase-1/urine , Adult , Carcinoma, Pancreatic Ductal/diagnosis , Case-Control Studies , Enzyme-Linked Immunosorbent Assay , Female , Follow-Up Studies , Humans , Immunoenzyme Techniques , Male , Middle Aged , Neuroendocrine Tumors/diagnosis , Pancreatic Neoplasms/diagnosis , Prognosis , Retrospective StudiesABSTRACT
Sturge-Weber syndrome (SWS) consists of a capillary-venous vascular malformation of the brain, skin and eye. Urine vascular biomarkers have been demonstrated to be abnormal in other vascular anomalies and to correlate with clinical severity and progression. The current study investigated the use of urinary matrix metalloproteinase (MMP)-2, MMP-9, vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) levels to non-invasively monitor the progression of SWS. Fifty-four urine samples were collected from patients seen at the Hunter Nelson Sturge-Weber Center at Kennedy Krieger Institute. Urine was analyzed for MMP-2, MMP-9, VEGF and bFGF levels and correlated with clinical outcome at the time of urine collection (n = 48) and 1 year following urine collection (n = 22). Analysis revealed that MMP-2 (p = 0.033) and MMP-9 (p = 0.010) were significantly more likely to be present in the urine of SWS subjects compared to controls and that bFGF was significantly more likely to be present at abnormal levels (p = 0.005). MMP-2 correlated with a more severe clinical score at the time of urine collection, while both MMP-2 and MMP-9 levels correlated with greater disease severity at time of collection. bFGF levels correlated with improved clinical score 1 year after urine collection. These results suggest that MMP-2 and MMP-9 levels may be useful in assessing SWS progression, as well as indicating which patients might benefit from more aggressive treatment, while bFGF levels may be useful in judging the efficacy of neurologic treatment in SWS.
Subject(s)
Fibroblast Growth Factor 2/urine , Matrix Metalloproteinase 2/urine , Matrix Metalloproteinase 9/urine , Sturge-Weber Syndrome/urine , Vascular Endothelial Growth Factor A/urine , Adolescent , Adult , Biomarkers, Tumor/urine , Child , Child, Preschool , Female , Humans , Infant , Male , Risk Assessment , Sturge-Weber Syndrome/diagnosis , Young AdultABSTRACT
BACKGROUND: The study was undertaken to develop a potential new markers for distinguishing minimal change nephrotic syndrome (MCNS) and focal segmental glomerulosclerosis (FSGS) in children. We hypothesized that matrix metalloproteinase-9/neutrophil gelatinase-associated lipocalin (MMP-9/NGAL) is a better marker of focal sclerosis in the glomerulus then matrix metalloproteinase-9/tissue inhibitor of metalloproteinase-1 (MMP-9/TIMP-1) and matrix metalloproteinase-2/tissue inhibitor of metalloproteinase-2 MMP2/TIMP-2. METHODS: The present study used a sample of 36 children and adolescents subdivided into two groups: I - 20 children with MCNS, subjected to examination twice: A - in relapse of nephrotic syndrome, before treatment and B - after regression of proteinuria; II - 16 children with FSGS. MMPs and TIMPs and NGAL levels were measured in the urine using ELISA kit. MMP-9/TIMP-1, MMP-2/TIMP-2 and MMP-9/NGAL ratios were calculated. RESULTS: Median NGAL/cr. was significantly higher in MCNS and FSGS patients when compared to healthy controls. Both, NGAL and MMP-9 urinary levels were significantly elevated in FSGS subjects, as compared with control subjects. Contrary to FSGS children, in MCNS group, before treatment only NGAL/cr., but not MMP-9/cr. was increased. Urinary concentrations of NGAL and MMP-9 were highly associated with each other (NGAL/cr. vs. MMP-9/cr., r=0.485, p<0.01). Median urine MMP-9/NGAL ratio in FSGS patients was significantly higher than in patients with MCNS. We also found that significant increase in MMP-9/NGAL was associated with FSGS [odds ratio (OR) - 9.0; confidence interval (CI) 1.97-41.07]. CONCLUSION: MMP-9/NGAL ratio may serve as differentiation marker between MCNS and FSGS in nephrotic children.