Glycosaminoglycan diversity in marine sponge extracellular matrix

Aim of this paper is to report on a screening on sponge ECM glycosaminoglycan (GAG) diversity. To investigate the heterogeneity of sponge extracellular sulphated glycans, we determined their content and distribution in some Mediterranean and Caribbean species. To focus on biological and morpho-functional roles of these molecules in the sponge ECM some selected species were considered as models to investigate the topographic distribution of GAGs in sponge body according with the different architecture of specialized regions

The genetic architecture of membranous nephropathy and its potential to improve non-invasive diagnosis

Membranous Nephropathy (MN) is a rare autoimmune cause of kidney failure. Here we report a genome-wide association study (GWAS) for primary MN in 3,782 cases and 9,038 controls of East Asian and European ancestries. We discover two previously unreported loci, NFKB1 (rs230540, OR = 1.25, P = 3.4 × 10−12) and IRF4 (rs9405192, OR = 1.29, P = 1.4 × 10−14), fine-map the PLA2R1 locus (rs17831251, OR = 2.25, P = 4.7 × 10−103) and report ancestry-specific effects of three classical HLA alleles: DRB1*1501 in East Asians (OR = 3.81, P = 2.0 × 10−49), DQA1*0501 in Europeans (OR = 2.88, P = 5.7 × 10−93), and DRB1*0301 in both ethnicities (OR = 3.50, P = 9.2 × 10−23 and OR = 3.39, P = 5.2 × 10−82, respectively). GWAS loci explain 32% of disease risk in East Asians and 25% in Europeans, and correctly re-classify 20–37% of the cases in validation cohorts that are antibody-negative by the serum anti-PLA2R ELISA diagnostic test. Our findings highlight an unusual genetic architecture of MN, with four loci and their interactions accounting for nearly one-third of the disease risk

Factors affecting <i>S</i>-homocysteinylation of LDL apoprotein B

Background: Hyperhomocysteinemia is an important risk factor for vascular disease and atherosclerosis, but the mechanisms by which homocysteine exerts its deleterious effects are not known. Because oxidation and/or homocysteinylation may increase atherogenicity of LDL, we investigated S-homocysteinylation of LDL as a possible contributor to atherosclerosis pathogenesis. Methods: We used capillary electrophoresis to measure LDL-bound thiols [homocysteine, cysteine (Cys), cysteinylglycine, glutathione, and glutamylcysteine] in 104 healthy study participants We also assessed total plasma thiol concentrations and lipid profiles. Results: Our data suggest that apoprotein B (apoB)-cysteinylglycine (CysGly), apoB-Hcy, and apoB-Cys concentrations are markedly higher in men than in women. The percentage of CysGly and glutathione on apoB was higher than that of the same thiols in plasma, whereas the other thiols were markedly less prevalent in lipoprotein than in plasma. Pearson correlation showed that among all thiols, only total plasma Hcy is related to apoB-Hcy concentrations. Multiple correlation analysis confirmed that total Hcy was the most important determinant of apoB-Hcy. Age and LDL cholesterol also showed positive associations, but Cys and, mainly, CysGly were negatively associated with apoB-Hcy concentrations. Conclusions: apoB-Hcy derivative formation is mainly dependent on total homocysteine concentration. Increased cholesterol concentrations are related to increased apoB-Hcy. CysGly seems to compete with Hcy for binding to LDL apoprotein, suggesting that CysGly may protect against atherosclerosis by decreasing the concentrations of Hcy transferred by LDL from plasma to endothelial and subendothelial spaces

Glycosaminoglycan and transforming growth factor β1 changes in human plasma and urine during the menstrual cycle, in vitro fertilization treatment, and pregnancy

Objective. To evaluate transforming growth factor β1 (TGF-β1) and glycosaminoglycans (GAG) changes in human plasma and urine during the menstrual cycle, IVF-ET, and pregnancy. Design. Prospective clinical study. Setting. University hospital. Patient(s). Thirteen women with apparently normal menstrual cycle (group 1); 18 women undergoing IVF-ET (group 2); and 14 low-risk pregnant women (group 3). Intervention(s). We assayed plasma and urine concentrations of TGF-β1, urine content, and distribution of GAG. Blood and urine samples were collected during days 2 to 3, 12 to 13, and 23 to 24 in group 1; in group 2, samples were obtained at menstrual phase, oocyte pick-up day, and 15 days after ET; in group 3, samples were obtained during gestational weeks 10–12, 22–24, and 30–32 and 1 month after delivery. Main Outcome Measure(s). Changes in TGF-β1 and GAG content. Result(s). The mean value of total urinary trypsin inhibitor/chondroitin sulfate (UTI/CS) showed a distinct peak at day 12 of the menstrual cycle in the fertile women in whom we monitored the ovulatory period. In the IVF-ET group, GAG distribution and TGF-β1 levels showed significant differences during the cycle. We observed increased levels of plasma TGF-β1 15 days after ET. A significant increase of total UTI/CS value with increasing gestation was detected. Conclusion(s). Transforming growth factor β1 and GAG levels could represent an additional tool to monitor reproductive events and could be useful, noninvasive markers of ovulation and ongoing pregnancy

A Novel LIF-CE method for the separation of hyaluronan- and chondroitin sulfate-derived disaccharides: Application to structural and quantitative analyses of human plasma low- and high-charged chondroitin sulfate isomers

The report describes a rapid and simple CE method using LIF detection for the analysis of unsaturated disaccharides obtained from enzymatic depolymerization of plasma chondroitin sulfate (CS) isomers. The disaccharide reducing groups were labeled with 2-aminoacridone (AMAC). The fluorotagged products can be separated by reversed-polarity CE using a sodium acetate buffer, pH 3.8, in the presence of 0.05% methylcellulose. The choice of the appropriate electrophoretic conditions was performed after a deep analysis of the most important parameters affecting analyte separation. In particular, the effect of both run buffer concentration and pH on resolution, efficiency, migration times, and peak area was evaluated. The selected electrophoretic conditions allowed us to separate the CS isomers-derived Δ-disaccharides in less than 12 min, also resolving the nonsulfated disaccharides released from CS isomers from those released from hyaluronan (HA). Moreover, these conditions gave a good reproducibility of both the migration times (CV%, 0.25) and the peak areas (CV%, 1.4). Intra- and interassay CV were 5.37 and 7.23%, respectively, and analytical recovery was about 86%. The applicability of the above method to the quantitative and structural disaccharide analyses of plasma CS isomers was investigated. Data obtained from 44 healthy human subjects were compared with those obtained by a fluorophore-assisted carbohydrate electrophoresis (FACE) reference assay, by using the Passing and Bablok regression and Bland-Altman tests. The developed method could represent a good tool for an ultrasensitive analysis of CS isomers in biological samples from different sources, particularly when samples are available in very low amounts

Evidence for a proinflammatory and proteolytic environment in plaques from endarterectomy segments of human carotid arteries

Objectives—Based on previous observations on apolipoprotein(a), apo(a), in human unstable carotid plaques, we explored whether in the inflammatory environment of human atheroma, proteolytic events affect other hepatic and topically generated proteins in relation to the issue of plaque stability. Methods and Results—Forty unstable and 24 stable plaques from endarterectomy segments of affected human carotid arteries were extracted with buffered saline (PBS) and then 6 mol/L guanidine-hydrochloride (GdHCl) to identify loosely and tightly bound products, respectively. The extracts were studied before and after ultracentrifugation at d 1.21 g/mL. In the extracts, the concentrations of interleukin (IL)-6, -8, and -18 were significantly higher in the unstable plaques and correlated to those of MMP-2 and MMP-9. By Western blots, both apoB and apo(a) were highly fragmented and mostly present in the d 1.21 bottom that also contained fragments of apoE (10 and 22 kDa), decorin, biglycan, and versican. Fragmentation was higher in the unstable plaques. In baseline plasmas, concentrations of lipids, lipoproteins, and ILs did not differ between patients with unstable and stable plaques. Conclusions—In unstable and to a lesser extent in stable plaques, there is a proinflammatory and proteolytic microenvironment with the generation of fragments with potential pathobiological significance that requires investigation

A Proteomic approach to differentiate histologically classified stable and unstable plaques from human carotid arteries

Objectives. By using proteomics we isolated and identified proteins that were expressed/retained in stable and unstable human carotid artery atherosclerotic plaques. Methods. The criteria for plaque instability were the presence of a thin fibrous cap or fissured cap covering the foamy or necrotic core, and the presence of overt, hemorrhagic, ulcerated or thrombotic plaques. Proteins were extracted from finely minced endarterectomy specimens (19 stable and 29 unstable plaques) and separated by two-dimensional gel electrophoresis. Coomassie Blue-stained gels were analysed using PD-Quest software. Results. A total of 57 distinct spots corresponding to 33 different proteins were identified by matrix assisted laser desorption/ionization mass spectrometry using the NCBI database. Most of the spots were present in both types of extracts, although significantly (p &lt; 0.05) differing in abundance between them. Compared to stable plaque, unstable ones showed reduced abundance of: protective enzymes SOD3 and GST, small heat shock proteins HSP27 and HSP20, annexin A10, and Rho GDI. In unstable plaques the more abundant proteins were: ferritin light subunit, SOD 2 and fibrinogen fragment D. For fibrinogen fragment D, the increased levels in unstable versus stable plaques was confirmed by Western blot analysis. Conclusions. Since many of the differentially expressed proteins are known to have a functional role in inflammation and oxidative stress, we speculate that they may be involved in events relating to plaque stability.<br/

Urinary transforming growth factor-β1 in various types of nephropathy

Transforming growth factor-β1 (TGF-β1) is a potent multifunctional polypeptide that is involved in normal renal function and in the development of glomerular sclerosis. It is also an important mediator of the immune and anti-inflammatory responses. The purpose of this study was to examine whether the measurement of urinary TGF-β1 excretion in patients with different types of renal diseases and in newly diagnosed type 1 diabetes mellitus represents a non-invasive tool to evaluate disease activity and to monitor response to therapy. We studied the urinary excretion of TGF-β1 in 57 nephropathic patients divided in different groups according to the underlying disease: 15 had mesangial glomerulonephritis (IgAGN), 9 membranous glomerulonephritis (MGN), 7 rapidly progressive glomerulonephritis (RPGN), 8 systemic lupus erythematosus (SLE), 9 interstitial nephritis (IN), 9 chronic renal failure (CRF). TGF-β1 was also measured in 38 patients with type 1 (insulin-dependent) diabetes mellitus (12 with newly diagnosed diabetes, 26 long-standing diabetes) and 31 healthy controls. Total urinary TGF-β1 concentration was assayed by enzyme-linked immunoassay (ELISA), and expressed as a ratio to urinary creatinine concentration. The urinary TGF-β1 levels were compared with the findings of biopsy and clinical parameters. Urinary TGF-β1 excretion was significantly increased in all groups except MGN, IN and CRF. In non-diabetic patients, urinary TGF-β1 levels correlated with crescent formation, floccular adhesion and mesangial proliferation, but not with the degree of tubulo-interstitial fibrosis. Urinary TGF-β1 levels did not correlate with indices of renal function (serum creatinine, glomerular filtration rate (GFR), albumin excretion rate [AER]). Among diabetic patients, HbA1C significantly correlated with TGF-β1 urinary excretion. Urinary TGF-β1 levels may represent a valid indicator of acute glomerular flogosis associated with mesangial proliferation in glomerulonephrities. In newly diagnosed diabetic patients, hyperglycaemia seems to represent the principal factor leading to TGF-β1 overproduction. Follow-up studies of urinary TGF-β1 levels measured during optimal glycaemic control are necessary to clarify the relationship between hyperglycaemia and TGF-β1 excretion

Age-specific seroprevalence of human herpesvirus 8 in Mediterranean regions

Objective: human herpesvirus 8 (HHV8) is believed to be transmitted mainly by sexual contact; epidemiological data from Africa show, however, that non-sexual transmission routes may also play an important role. To evaluate better the distribution of HHV8 infection in the Mediterranean area, we performed an age-specific seroprevalence study. Methods: Sera were collected from subjects from different geographical areas. The sera were analyzed by immunofluorescence assay (IFA) and enzyme-linked immunosorbent assay (ELISA). A total of 1083 patients were studied, 667 patients from various regions of Italy and 416 from Albania. The patients were stratified into six age groups. Multivariate logistic regression was used to evaluate associations between HHV8 and demographic data. Results: an overall seropositivity rate of 17.6% was observed. The highest rate was observed in Sardinia (25.0%) and the lowest was found in Albania (13.9%). The prevalence rate increased linearly with age, from 9.7% in patients belonging to the 0–14 years age group to 26.3% for patients more than 59 years old. Seropositivity for HHV8 was significantly associated with membership of the 59 years-plus age group. Rates of seropositivity were significantly higher in patients from central southern Italy (OR = 1.7) and Sardinia (OR = 1.8) than in patients from Albania. Conclusions: the data suggest that HHV8 is widespread in the Mediterranean area, including regions like Albania that have not been previously investigated. The statistically significant association between HHV8 seropositivitity and increasing age suggests that non-sexual transmission routes may be involved in the spread of the virus