Non-invasive profiling of protease-specific elastin turnover in lung cancer: biomarker potential.

PURPOSE: Elastin is a signature protein of lungs. Increased elastin turnover driven by altered proteolytic activity is an important part of lung tumorigenesis. Elastin-derived fragments have been shown to be pro-tumorigenic, however, little is known regarding the biomarker potential of such elastin fragments. Here, we present an elastin turnover profile by non-invasively quantifying five specific elastin degradation fragments generated by different proteases. METHODS: Elastin fragments were assessed in serum from patients with stage I-IV non-small cell lung cancer (NSCLC) (n = 40) and healthy controls (n = 30) using competitive ELISAs targeting different protease-generated fragments of elastin: ELM12 (generated by matrix metalloproteinase MMP-9 and -12), ELM7 (MMP-7), EL-NE (neutrophil elastase), EL-CG (cathepsin G) and ELP-3 (proteinase 3). RESULTS: ELM12, ELM7, EL-NE and EL-CG were all significantly elevated in NSCLC patients (n = 40) when compared to healthy controls (n = 30) (ELM12, p = 0.0191; ELM7, p < 0.0001; EL-NE, p < 0.0001; EL-CG, p < 0.0001). ELP-3 showed no significant difference between patients and controls (p = 0.8735). All fragments correlated positively (Spearman, r: 0.69-0.81) when compared pairwise, except ELM12 (Spearman, r: 0.042-0.097). In general, all fragments were detectable across all stages of the disease. CONCLUSIONS: Elastin fragments generated by different proteases are elevated in lung cancer patients compared to healthy controls but differ in their presence. This demonstrates non-invasive biomarker potential of elastin fragments in serum from lung cancer patients and suggests that different pathological mechanisms may be responsible for the elastin turnover, warranting further validation in clinical trials.

Production of Elastin-Derived Peptides Contributes to the Development of Nonalcoholic Steatohepatitis.

Affecting more than 30% of the Western population, nonalcoholic fatty liver disease (NAFLD) is the most common liver disease and can lead to multiple complications, including nonalcoholic steatohepatitis (NASH), cancer, hypertension, and atherosclerosis. Insulin resistance and obesity are described as potential causes of NAFLD. However, we surmised that factors such as extracellular matrix remodeling of large blood vessels, skin, or lungs may also participate in the progression of liver diseases. We studied the effects of elastin-derived peptides (EDPs), biomarkers of aging, on NAFLD progression. We evaluated the consequences of EDP accumulation in mice and of elastin receptor complex (ERC) activation on lipid storage in hepatocytes, inflammation, and fibrosis development. The accumulation of EDPs induces hepatic lipogenesis (i.e., SREBP1c and ACC), inflammation (i.e., Kupffer cells, IL-1ß, and TGF-ß), and fibrosis (collagen and elastin expression). These effects are induced by inhibition of the LKB1-AMPK pathway by ERC activation. In addition, pharmacological inhibitors of EDPs demonstrate that this EDP-driven lipogenesis and fibrosis relies on engagement of the ERC. Our data reveal a major role of EDPs in the development of NASH, and they provide new clues for understanding the relationship between NAFLD and vascular aging.

Biomarkers of extracellular matrix turnover are associated with emphysema and eosinophilic-bronchitis in COPD.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by airflow obstruction and loss of lung tissue mainly consisting of extracellular matrix (ECM). Three of the main ECM components are type I collagen, the main constituent in the interstitial matrix, type VI collagen, and elastin, the signature protein of the lungs. During pathological remodeling driven by inflammatory cells and proteases, fragments of these proteins are released into the bloodstream, where they may serve as biomarkers for disease phenotypes. The aim of this study was to investigate the lung ECM remodeling in healthy controls and COPD patients in the COPDGene study. METHODS: The COPDGene study recruited 10,300 COPD patients in 21 centers. A subset of 89 patients from one site (National Jewish Health), including 52 COPD patients, 12 never-smoker controls and 25 smokers without COPD controls, were studied for serum ECM biomarkers reflecting inflammation-driven type I and VI collagen breakdown (C1M and C6M, respectively), type VI collagen formation (Pro-C6), as well as elastin breakdown mediated by neutrophil elastase (EL-NE). Correlation of biomarkers with lung function, the SF-36 quality of life questionnaire, and other clinical characteristics was also performed. RESULTS: The circulating concentrations of biomarkers C6M, Pro-C6, and EL-NE were significantly elevated in COPD patients compared to never-smoking control patients (all p < 0.05). EL-NE was significantly elevated in emphysema patients compared to smoking controls (p < 0.05) and never-smoking controls (p < 0.005), by more than 250%. C1M was inversely associated with forced expiratory volume in 1 s (FEV1) (r = -0.344, p = 0.001), as was EL-NE (r = -0.302, p = 0.004) and Pro-C6 (r = -0.259, p = 0.015). In the patients with COPD, Pro-C6 was correlated with percent predicted Forced Vital Capacity (FVC) (r = 0.281, p = 0.046) and quality of life using SF-36. C6M and Pro-C6, were positively correlated with blood eosinophil numbers in COPD patients (r = 0.382, p = 0.006 and r = 0.351, p = 0.012, respectively). CONCLUSIONS: These data suggest that type VI collagen turnover and elastin degradation by neutrophil elastase are associated with COPD-induced inflammation (eosinophil-bronchitis) and emphysema. Serological assessment of type VI collagen and elastin turnover may assist in identification of phenotypes likely to be associated with progression and amenable to precision medicine for clinical trials.

Circulating desmosine levels do not predict emphysema progression but are associated with cardiovascular risk and mortality in COPD.

Elastin degradation is a key feature of emphysema and may have a role in the pathogenesis of atherosclerosis associated with chronic obstructive pulmonary disease (COPD). Circulating desmosine is a specific biomarker of elastin degradation. We investigated the association between plasma desmosine (pDES) and emphysema severity/progression, coronary artery calcium score (CACS) and mortality.pDES was measured in 1177 COPD patients and 110 healthy control subjects from two independent cohorts. Emphysema was assessed on chest computed tomography scans. Aortic arterial stiffness was measured as the aortic-femoral pulse wave velocity.pDES was elevated in patients with cardiovascular disease (p<0.005) and correlated with age (rho=0.39, p<0.0005), CACS (rho=0.19, p<0.0005) modified Medical Research Council dyspnoea score (rho=0.15, p<0.0005), 6-min walking distance (rho=-0.17, p<0.0005) and body mass index, airflow obstruction, dyspnoea, exercise capacity index (rho=0.10, p<0.01), but not with emphysema, emphysema progression or forced expiratory volume in 1 s decline. pDES predicted all-cause mortality independently of several confounding factors (p<0.005). In an independent cohort of 186 patients with COPD and 110 control subjects, pDES levels were higher in COPD patients with cardiovascular disease and correlated with arterial stiffness (p<0.05).In COPD, excess elastin degradation relates to cardiovascular comorbidities, atherosclerosis, arterial stiffness, systemic inflammation and mortality, but not to emphysema or emphysema progression. pDES is a good biomarker of cardiovascular risk and mortality in COPD.

Neutrophil serine proteases and their endogenous inhibitors in coronary artery ectasia patients.

OBJECTIVE: Proteolytic enzymes possibly contribute to coronary artery ectasia (CAE). This study aimed to determine whether neutrophils, neutrophil serine proteases (NSPs), and their endogenous inhibitors participated in the pathological process of CAE. METHODS: The study consisted of 30 patients with CAE, 30 patients with coronary artery disease (CAD), and 29 subjects with normal coronary arteries (Control). The following circulating items were measured: the main NSPs, including human neutrophil elastase (HNE), cathepsin G (CG), and proteinase 3 (PR3); soluble elastin (sElastin), which was a degradation product of elastin fibres; NSP inhibitors such as α1-protease inhibitor (α1-PI), α2-macroglobulin (α2-MG), secretory leucoprotease inhibitor (SLPI), and elafin; as well as two neutrophil activation markers (myeloperoxidase and lactoferrin) and three classic neutrophil activators [tumor necrosis factor-α (TNF-α), interleukin-8 (IL-8), and bacterial endotoxin]. RESULTS: The levels of HNE, CG, and sElastin were elevated in the CAE group. The levels of α1-PI and α2-MG were also significantly increased in the CAE group. The levels of myeloperoxidase and lactoferrin were higher in the CAE group. The levels of TNF-α, IL-8, and endotoxin were unchanged in the CAE group compared with those in the CAD group. CONCLUSION: Neutrophils may participate in the process of vessel extracellular matrix destruction and coronary ectasia by releasing NSPs in a non-classical manner.

Circulating elastin peptides, role in vascular pathology.

The atherosclerotic process starts with the degradation of elastic fibers. Their presence was demonstrated in the circulation as well as several of their biological properties elucidated. We described years ago a procedure to obtain large elastin peptides by organo-alkaline hydrolysis, κ-elastin. This method enabled also the preparation of specific antibodies used to determine elastin peptides, as well as anti-elastin antibodies in body fluids and tissue extracts. Elastin peptides were determined in a large number of human blood samples. Studies were carried out to explore their pharmacological properties. Similar recent studies by other laboratories confirmed our findings and arose new interest in circulating elastin peptides for their biological activities. This recent trend justified the publication of a review of the biological and pathological activities of elastin peptides demonstrated during our previous studies, subject of this article.

Abnormal levels of serum anti-elastin antibodies in patients with symptomatic carotid stenosis.

BACKGROUND AND OBJECTIVE: A correlation between the levels of antibodies to alpha-elastin (alpha-AEAb) and tropoelastin (tropo-AEAb) and the corresponding peptide concentration is found in human serum in health and disease. Serum elastin peptide and anti-elastin antibodies (AEAb) levels are age-related and vary with the stages of atherosclerotic vascular damage. This study aims to determine if elastin metabolism (assessed by the ratio of tropo-AEAb to alpha-AEAb) differs in patients with symptomatic carotid stenosis versus subjects with asymptomatic stenosis. PATIENTS AND METHODS: Alpha-AEAb and tropo-AEAb were measured by ELISA in blood sera of 65 patients with ultrasound verified high-grade symptomatic carotid stenosis (resulting in stroke 1-7 days before measurement) compared to 51 patients with asymptomatic stenosis. RESULTS: Serum anti-alpha-elastin IgG levels are extremely increased in symptomatic versus asymptomatic carotid stenosis. The ratio of tropo-AEAb (reflecting elastin synthesis) to alpha-AEAb (a function of elastin degradation) was 3.7 in symptomatic stenosis versus 14.2 in asymptomatic stenosis (p<0.001). CONCLUSIONS: There is a significant difference in elastin metabolism in patients with symptomatic carotid stenosis versus asymptomatic stenosis. The ratio of tropo-AEAb to alpha-AEAb as an index of elastin synthesis/degradation proves useful in investigation of atherosclerotic lesions and may represent a new immunologic marker for carotid plaque destabilization.

Acute Myocardial Infarction and Pulmonary Diseases Result in Two Different Degradation Profiles of Elastin as Quantified by Two Novel ELISAs.

BACKGROUND: Elastin is a signature protein of the arteries and lungs, thus it was hypothesized that elastin is subject to enzymatic degradation during cardiovascular and pulmonary diseases. The aim was to investigate if different fragments of the same protein entail different information associated to two different diseases and if these fragments have the potential of being diagnostic biomarkers. METHODS: Monoclonal antibodies were raised against an identified fragment (the ELM-2 neoepitope) generated at the amino acid position '552 in elastin by matrix metalloproteinase (MMP) -9/-12. A newly identified ELM neoepitope was generated by the same proteases but at amino acid position '441. The distribution of ELM-2 and ELM, in human arterial plaques and fibrotic lung tissues were investigated by immunohistochemistry. A competitive ELISA for ELM-2 was developed. The clinical relevance of the ELM and ELM-2 ELISAs was evaluated in patients with acute myocardial infarction (AMI), no AMI, high coronary calcium, or low coronary calcium. The serological release of ELM-2 in patients with chronic obstructive pulmonary disease (COPD) or idiopathic pulmonary fibrosis (IPF) was compared to controls. RESULTS: ELM and ELM-2 neoepitopes were both localized in diseased carotid arteries and fibrotic lungs. In the cardiovascular cohort, ELM-2 levels were 66% higher in serum from AMI patients compared to patients with no AMI (p<0.01). Levels of ELM were not significantly increased in these patients and no correlation was observed between ELM-2 and ELM. ELM-2 was not elevated in the COPD and IPF patients and was not correlated to ELM. ELM was shown to be correlated with smoking habits (p<0.01). CONCLUSIONS: The ELM-2 neoepitope was related to AMI whereas the ELM neoepitope was related to pulmonary diseases. These results indicate that elastin neoepitopes generated by the same proteases but at different amino acid sites provide different tissue-related information depending on the disease in question.

Measurement of MMP-9 and -12 degraded elastin (ELM) provides unique information on lung tissue degradation.

BACKGROUND: Elastin is an essential component of selected connective tissues that provides a unique physiological elasticity. Elastin may be considered a signature protein of lungs where matrix metalloprotease (MMP) -9-and -12, may be considered the signature proteases of the macrophages, which in part are responsible for tissue damage during disease progression. Thus, we hypothesized that a MMP-9/-12 generated fragment of elastin may be a relevant biochemical maker for lung diseases. METHODS: Elastin fragments were identified by mass-spectrometry and one sequence, generated by MMP-9 and -12 (ELN-441), was selected for monoclonal antibody generation and used in the development of an ELISA. Soluble and insoluble elastin from lung was cleaved in vitro and the time-dependent release of fragments was assessed in the ELN-441 assay. The release of ELN-441 in human serum from patients with chronic obstructive pulmonary disease (COPD) (n = 10) and idiopathic pulmonary fibrosis (IPF) (n = 29) were compared to healthy matched controls (n = 11). RESULTS: The sequence ELN-441 was exclusively generated by MMP-9 and -12 and was time-dependently released from soluble lung elastin. ELN-441 levels were 287% higher in patients diagnosed with COPD (p < 0.001) and 124% higher in IPF patients (p < 0.0001) compared with controls. ELN-441 had better diagnostic value in COPD patients (AUC 97%, p = 0.001) than in IPF patients (AUC 90%, p = 0.0001). The odds ratios for differentiating controls from COPD or IPF were 24 [2.06-280] for COPD and 50 [2.64-934] for IPF. CONCLUSIONS: MMP-9 and -12 time-dependently released the ELN-441 epitope from elastin. This fragment was elevated in serum from patients with the lung diseases IPF and COPD, however these data needs to be validated in larger clinical settings.

Serum elastin-derived peptides and anti-elastin antibody in patients with systemic sclerosis.

The elastin metabolism in systemic sclerosis (SSc) has been known to be abnormal. The authors investigated relationship between the clinical manifestations of systemic sclerosis (SSc) and serum levels of soluble elastin-derived peptide (S-EDP) and anti-elastin antibodies. Serum samples were obtained from 79 patients with SSc and 79 age- and sex-matched healthy controls. Concentrations of serum S-EDP and anti-elastin antibodies were measured by ELISA. The serum concentrations of S-EDP in SSc patients were significantly higher than in healthy controls (median, 144.44 ng/mL vs 79.59 ng/mL, P < 0.001). Serum EDP concentrations were found to be correlated with disease duration in SSc (P = 0.002) and particularly in diffuse cutaneous SSc (P = 0.005). Levels of anti-elastin antibodies were found to be more elevated in SSc patients than in healthy controls (median, 0.222 U vs 0.191 U, P = 0.049), more increased in diffuse cutaneous SSc than limited cutaneous SSc (median, 0.368 U vs 0.204 U, P = 0.031). In addition, levels of anti-elastin antibodies were also found to be negatively associated with presence of anti-centromere antibody (P = 0.023). The S-EDP levels were not found to be correlated with levels of anti-elastin antibodies. The increased S-EDP and anti-elastin antibody levels and association with clinical and laboratory characteristics may reflect the abnormal metabolism in SSc.

Potential circulating biomarkers for abdominal aortic aneurysm expansion and rupture--a systematic review.

BACKGROUND: The maximal diameter of abdominal aortic aneurysms (AAAs) is the dominating indication for repair. However half of the AAAs repaired would never have ruptured if left unrepaired, although small AAAs occasionally rupture. Earlier surgery may be associated with a lower mortality. More precise indicators for surgery are warranted. This systematic review identifies potential systemic biomarkers for AAA rupture or expansion. METHODS: MEDLINE/PubMed and EMBASE (from 1985 trough May 2007) were searched with the medical subject heading abdominal aortic aneurysm and keywords "size", "progression" or "growth" or "expansion rate" or "rupture" on the basis of MESH tree and as a text search restricted to English, German, French and Italian. In addition, reference lists were studied and manual searches performed. Observational studies investigating the association of circulating biomarkers with AAA rupture, expansion or size were selected. DATA EXTRACTION: Two reviewers (SU and GU) independently extracted the following data: year of publication, study characteristics, duration of follow-up, circulating biomarker, AAA expansion rate or size or rupture. RESULTS: 699 papers were identified. After exclusion of thoracic aneurysms and cardiac studies (n=118), surgical or medical treatment studies (n=179), case reports and animal studies (n=87), as well as reviews or letters (n=66), 249 articles were selected. Also excluded were 230 papers that did not report AAA size, expansion rate or rupture. 39 papers were included. Several potential biomarkers were identified. The strongest association with AAA was obtained with serum elastin peptides (SEP) and plasmin-antiplasmin (PAP) complexes. Matrix-degrading metalloproteinase 9 (MMP9) and interferon-gamma (IFN-gamma) could have clinical potential while many putative biomarkers showed poor association. CONCLUSIONS: Several circulating agents in peripheral blood may predict AAA size, expansion rate or rupture. Few of them have clinical potential for future use. Confirmative studies and development of multivariate models are needed, together with continuing search for new biomarkers using the discovery based sciences within proteomics and/or genomics.

Increased levels of elastin-derived peptides in cerebrospinal fluid of patients with lacunar stroke.

OBJECTIVE: To investigate whether elastin-derived peptides (EDP) are detectable in the cerebrospinal fluid (CSF) of healthy controls and of patients with acute brain ischemia and if so to assess possible trends in EDP levels in different groups of ischemic stroke patients (small-vessel disease vs. other ischemic strokes; first-ever vs. recurrent stroke). PATIENTS AND METHODS: Levels of EDP were determined by ELISA in blood sera and CSF of 80 patients with acute ischemic stroke (mean age 61.5+/-10.8; age range 47-70; 22 women) and in 15 healthy age- and sex-matched controls (mean age 57.3+/-13.4; age range 50-65). The patients were divided into a group with first ever lacunar stroke (27); first ever non-lacunar ischemic stroke (27) and recurrent stroke (26). EDP were measured early (mean 7 days, range 1-15) after stroke onset. RESULTS: Serum EDP levels were mildly higher in recurrent strokes as compared to first ever lacunar lesion and controls. However, in the CSF the concentrations of EDP in stroke patients were strongly elevated (from 2 up to 30 times depending on subgroup) as compared with healthy subjects. The highest level of EDP in CSF and in the serum was found in recurrent strokes. Subgroup analysis revealed a trend for significantly higher EDP concentrations in CSF in lacunar and recurrent stroke as compared with non-lacunar. CONCLUSIONS: This study is the first application of elastin peptide measurement to human CSF and stroke patients. The increased levels of EDP were detected in CSF of patients with lacunar and recurrent strokes.

Antibodies to elastin peptides in sera of Warmblood horses at different ages.

REASONS FOR PERFORMING STUDY: Early diagnosis and monitoring progression of chronic diseases in elastin-rich tissues, such as chronic progressive lymphoedema in draught horses and chronic obstructive pulmonary disease (COPD) is still a real challenge in the horse. Use of an enzyme-linked immunosorbent assay (ELISA) to detect anti-elastin antibody (AEAb) levels might be useful to assess the status of such diseases. Baseline levels, representing physiological breakdown of elastin in normal horses, are not available at present. HYPOTHESIS: Levels of AEAb in healthy horses are generally low and follow the same age-related pattern as found in man. Therefore, elevation of AEAb levels in serum can be used to evaluate pathological elastin breakdown in elastin-rich tissues. METHODS: Sera of 84 clinically healthy Warmblood horses were evaluated for the presence of AEAbs by means of a modified version of an ELISA technique used in man. The horses were divided in 5 age groups: A) < 4 months; B) 4-23 months; C) 2-3 years; D) 4-10 years; and E) > 11 years. RESULTS: Antibodies to elastin were found in all equine serum samples tested. Their levels were lowest in Group A, low in Groups B and E and highest in animals age 2-10 years. CONCLUSIONS: Measuring AEAbs in serum of horses by an ELISA technique proved to be possible and levels were stable during well-defined life stages. POTENTIAL RELEVANCE: Changes in AEAb levels are expected to be useful for early diagnosis and for monitoring progression of diseases that affect elastin-rich tissues, such as chronic progressive lymphoedema and COPD.

Antibodies to elastin peptides in sera of Belgian Draught horses with chronic progressive lymphoedema.

REASONS FOR PERFORMING STUDY: Chronic progressive lymphoedema (CPL) is a recently recognised disease of the lymphatic system characterised by lesions in the skin of the lower legs in several draught horse breeds, including the Belgian Draught hourse. Clinical signs slowly progress and result in severe disfigurement of the limbs. Ideally, supportive treatment should be started early in the disease process. However early diagnosis and monitoring progression of CPL is still a challenge. HYPOTHESIS: Elastin changes, characterised by morphological alterations as well as increased desmosine levels, in the skin of the distal limbs of horses affected with CPL are probably associated with a marked release of elastin degradation products, which elicit production of circulating anti-elastin antibodies (AEAbs) in the serum. An enzyme-linked immunosorbent assay (ELISA) for detection of serum AEAbs may document elastin breakdown. METHODS: An ELISA technique was used to evaluate levels of AEAbs in sera of 97 affected Belgian Draught horses that were clinically healthy except for possible skin lesions, associated with CPL in their distal limbs. The horses were divided into 5 groups according to the severity of these skin lesions: normal horses (Group 1, n = 36), horses with mild lesions (Group 2, n = 43), horses with moderate lesions (Group 3, n = 8), horses with severe lesions (Group 4, n = 10) and, as a control, healthy Warmblood horses, unaffected by the disease (Group 5, n = 83). RESULTS: Horses with clinical signs of CPL had significantly higher AEAb levels compared to clinically normal Belgian Draught horses and to healthy Warmblood horses. These levels correlated with severity of lesions. CONCLUSIONS: CPL in draught horses is associated with an increase of serum AEAbs. POTENTIAL RELEVANCE: Evaluation of serum levels of AEAbs by ELISA might be a useful diagnostic aid for CPL. Pathological degradation of elastic fibres, resulting in deficient support of the distal lymphatics, is proposed as a contributing factor for CPL in Belgian Draught horses.

Thermal cycling enhances the accumulation of a temperature-sensitive biopolymer in solid tumors.

The delivery of anticancer therapeutics to solid tumors remains a critical problem in the treatment of cancer. This study reports a new methodology to target a temperature-responsive macromolecular drug carrier, an elastin-like polypeptide (ELP) to solid tumors. Using a dorsal skin fold window chamber model and intravital laser scanning confocal microscopy, we show that the ELP forms micron-sized aggregates that adhere to the tumor vasculature only when tumors are heated to 41.5 degrees C. Upon return to normothermia, the vascular particles dissolve into the plasma, increasing the vascular concentration, which drives more ELPs across the tumor blood vessel and significantly increases its extravascular accumulation. These observations suggested that thermal cycling of tumors would increase the exposure of tumor cells to ELP drug carriers. We investigated this hypothesis in this study by thermally cycling an implanted tumor in nude mice from body temperature to 41.5 degrees C thrice within 1.5 h, and showed the repeated formation of adherent microparticles of ELP in the heated tumor vasculature in each thermal cycle. These results suggest that thermal cycling of tumors can be repeated multiple times to further increase the accumulation of a thermally responsive polymeric drug carrier in solid tumors over a single heat-cool cycle. More broadly, this study shows a new approach--tumor thermal cycling--to exploit stimuli-responsive polymers in vivo to target the tumor vasculature or extravascular compartment with high specificity.

Detection of free elastin-derived peptides among diabetic children.

Elastin breakdown products are found in the serum of all human subjects. The presence of these elastin-derived peptides (EDP) and the corresponding antibodies in circulation leads to formation of circulating immune complexes (CIC). The aim of this study was to determine if serum level of free-EDP (unbound in CIC) correlate with the development of microvascular complications in children with Type 1 (insulin-dependent) diabetes mellitus. To this end we used a method for detecting immune complexes (CIF-ELISA) in combination with an ELISA for detection of EDP. The levels of free EDP were studied in sera of 81 diabetic children (mean age 13.46+/-3.51 years, diabetes duration 5.17+/-4.21 years). Forty-two of the children had vascular complications (group 1) and 39 were without vascular complications (group 2). Twenty-one healthy children (mean age 12.6+/-2.47 years) were used as controls. Diabetics showed significantly higher levels of free EDP (68.1+/-25 ng/ml versus 51+/-12.5 ng/ml; p=0.003) compared to the control group. In group 1, free EDP showed significantly higher levels than controls (78.9+/-25.6 ng/ml versus 51+/-12.5 ng/ml; p=0.0001). About 38 of 81 (47%) patients were positive for free EDP (30/42--71% in group 1 and 8/39--21% in group 2). Free EDP levels in all diabetics showed a correlation with insulin dose (r=0.23; p=0.041), and microalbuminuria (r=0.57; p=0.0001). Patients who had vascular pathology showed a correlation of free EDP with microalbuminuria (r=0.41; p=0.0081), retinopathy (r=0.32; p=0.041), insulin dose (r=0.37; p=0.02), HbA1c (r=0.35; p=0.03), systolic blood pressure (r=0.30; p=0.045) and total cholesterol (r=0.36; p=0.02). These findings suggest that elevated levels of free EDP are associated with the development of diabetic vascular complications in children.

Serum elastin-derived peptides in age-related macular degeneration.

PURPOSE: Dysregulation of the extracellular matrix (ECM) plays an important role in the pathogenesis of age-related macular degeneration (AMD). Elastin is a fibrous protein constituent of the ECM, degradation of which may be detected by the presence of serum elastin-derived peptides (S-EDPs) in circulation. This study was undertaken to estimate levels of S-EDPs in patients with AMD compared with age-matched control subjects. METHODS: Fifty-six patients with AMD were classified into two groups: early age-related maculopathy (ARM) and neovascular AMD. The control group consisted of 15 age-matched subjects with no AMD. S-EDP levels in the serum of these subjects was estimated by competitive ELISA, using solubilized alpha-elastin from human aorta and polyclonal antibodies to this antigen. RESULTS: S-EDPs were significantly higher in patients with AMD than in control subjects. In addition, subjects with neovascular AMD had higher levels of S-EDPs than did those with early disease. CONCLUSIONS: The cause of this association between S-EDPs and AMD is unknown, but it suggests that systemic elastin degradation may increase the risk of conversion from early ARM to neovascular AMD. Further studies are needed to confirm whether the serum level of S-EDPs is a useful predictor of conversion from early ARM to neovascular AMD.

[Serum tumor necrosis factor alpha (TNF-alpha) concentration correlates with soluble adhesion molecules and vascular endothelial growth factor (VEGF) in rheumatoid arthritis]. / Stezenie czynnika martwicy nowotworów alpha (TNF-alpha) koreluje ze stezeniem rozpuszczalnych form czasteczek adhezyjnych i naczyniowo-sródblonkowego czynnika wzrostu (VEGF) w reumatoidalnym zapaleniu stawów.

In present study we investigated whether the serum concentration of tumour necrosis factor alpha (TNF-alpha) is associated with soluble adhesion molecules and vascular endothelial growth factor (VEGF) in rheumatoid arthritis (RA) patients. Serum levels of TNF-alpha, soluble intercellular adhesion molecule 1 (sICAM-1), soluble vascular cell adhesion molecule 1 (sVCAM-1), soluble E-selectin (sE-selectin) and VEGF were assessed by ELISA in 38 patients with RA. We demonstrated the TNF-alpha to correlate with sICAM-1 (p < 0.001), sVCAM-1 (p < 0.01) and VEGF (p < 0.01). No association was noticed between TNF-alpha and sE-selectin serum concentrations. Moreover, serum TNF-alpha, sICAM-1, sVCAM-1 and VEGF levels correlated with markers of RA activity such as the erythrocyte sedimentation rate, C reactive protein level and the number of swollen joints. Data presented in this report support the concept of the VEGF synthesis stimulation by TNF-alpha. Analysis of the serum TNF-alpha, sICAM-1, sVCAM-1 and VEGF levels may be useful in the prediction of the RA activity.

Does lipoprotein(a) inhibit elastolysis in abdominal aortic aneurysms?

PURPOSE: to test the hypothesis that there is a negative association between serum levels of lipoprotein(a) (Lp(a)) and elastin-derived peptides (EDP) as well as matrix metalloproteinase (MMP)-9 activation in the aneurysm wall in patients with asymptomatic abdominal aortic aneurysms (AAA). MATERIAL AND METHODS: from 30 patients operated for asymptomatic AAAs, preoperative serum samples and AAA biopsies were collected. Lp(a) (mg/L) and EDP (ng/ml) in serum were measured by enzyme linked immunosorbent assays. MMP-9 activity (arbitrary units) in the AAA wall was measured by gelatin zymography and the ratio: active MMP-9/total MMP-9 were calculated. RESULTS: there was a significant negative correlation (Spearman's rho) between serum levels of Lp(a) and EDP (r= -0.707, p<0.001), as well as the share of activated MMP-9 (r= -0.461, p=0.01) in the AAA wall. CONCLUSION: this preliminary study indicate that Lp(a) inhibit elastolysis in asymptomatic AAA.

Serum elastin peptides in the preoperative evaluation of abdominal aortic aneurysms.

OBJECTIVE: Serum elastin peptides (SEP) have been reported to be associated with the expansion of small abdominal aortic aneurysms (AAA). Consequently, SEP-measurements may predict future rupture, and allow further selection for surgery in cases referred for surgery due to size. MATERIAL AND METHODS: SEP was measured in 90 men and 10 women with AAA, who were considered for surgery as part of the Chichester aneurysm screening programme. Sixty-one patients were electively operated and four because of symptoms. The rest were followed up further. Twelve of these experienced ruptured AAA later. RESULTS: No correlation between last measured AAA-diameter, annual expansion rate and SEP was noticed. However, SEP levels were significantly higher in cases rupturing later, persisting after adjustment for age, sex, and last measured AAA-size. ROC curve analysis concerning SEP as a predictor of rupture later showed an optimal sensitivity and specificity of 67% and 60%, respectively, similar with last measured AAA-size. By combining AAA-size and SEP, the optimal sensitivity and specificity reached 83% and 66%, respectively. CONCLUSION: One sampling of SEP combined with AAA-size in patients referred for AAA surgery may be a clinical useful indicator of high rupture risk.