RESUMO
Sepsis represents a syndromic response to infection and frequently acts as a common pathway leading to fatality in the context of various infectious diseases globally. The pathology of severe sepsis is marked by an excess of inflammation and activated coagulation. A substantial contributor to mortality in sepsis patients is widespread microvascular thrombosis-induced organ dysfunction. Multiple lines of evidence support the notion that sepsis induces endothelial damage, leading to the release of glycosaminoglycans, potentially causing microvascular dysfunction. This review aims to initially elucidate the relationship among endothelial damage, excessive inflammation, and thrombosis in sepsis. Following this, we present a summary of the involvement of glycosaminoglycans in coagulation, elucidating interactions among glycosaminoglycans, platelets, and inflammatory cells. In this section, we also introduce a reasoned generalization of potential signal pathways wherein glycosaminoglycans play a role in clotting. Finally, we discuss current methods for detecting microvascular conditions in sepsis patients from the perspective of glycosaminoglycans. In conclusion, it is imperative to pay closer attention to the role of glycosaminoglycans in the mechanism of microvascular thrombosis in sepsis. Dynamically assessing glycosaminoglycan levels in patients may aid in predicting microvascular conditions, enabling the monitoring of disease progression, adjustment of clinical treatment schemes, and mitigation of both acute and long-term adverse outcomes associated with sepsis.
Assuntos
Coagulação Sanguínea , Glicosaminoglicanos , Sepse , Humanos , Sepse/sangue , Sepse/complicações , Glicosaminoglicanos/sangue , Animais , Plaquetas/metabolismo , Microvasos , Transdução de Sinais , Inflamação/sangue , Microcirculação , Trombose/sangueRESUMO
Plasma and urine glycosaminoglycans (GAGs) are long, linear sulfated polysaccharides that have been proposed as potential noninvasive biomarkers for several diseases. However, owing to the analytical complexity associated with the measurement of GAG concentration and disaccharide composition (the so-called GAGome), a reference study of the normal healthy GAGome is currently missing. Here, we prospectively enrolled 308 healthy adults and analyzed their free GAGomes in urine and plasma using a standardized ultra-high-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry method together with comprehensive demographic and blood chemistry biomarker data. Of 25 blood chemistry biomarkers, we mainly observed weak correlations between the free GAGome and creatinine in urine and hemoglobin or erythrocyte counts in plasma. We found a higher free GAGome concentration - but not a more diverse composition - in males. Partitioned by gender, we also established reference intervals for all detectable free GAGome features in urine and plasma. Finally, we carried out a transference analysis in healthy individuals from two distinct geographical sites, including data from the Lifelines Cohort Study, which validated the reference intervals in urine. Our study is the first large-scale determination of normal free GAGomes reference intervals in plasma and urine and represents a critical resource for future physiology and biomarker research.
Assuntos
Glicosaminoglicanos , Adulto , Biomarcadores/sangue , Biomarcadores/urina , Cromatografia Líquida de Alta Pressão , Estudos de Coortes , Glicosaminoglicanos/sangue , Glicosaminoglicanos/química , Glicosaminoglicanos/urina , Humanos , Masculino , Espectrometria de Massas em Tandem/métodosRESUMO
Glycosaminoglycans (GAGs) are sulfated, negatively charged polysaccharides produced in almost every cell of the human body. As GAGs are extracellularly localized, the changes in body fluids such as blood and urine may reflect pathological changes in the urinary system as observed in other pathologies. In this review, we determined the potential of urinary and/or serum GAG levels as a marker for kidney and urothelial system diseases. We performed a search in the PubMed, MEDLINE, and ScienceDirect databases until September 30, 2019. A number of studies reported changes in the urinary and/or plasma GAG levels or composition in urological diseases and conditions, such as renal cell carcinoma, kidney stone, bladder carcinoma, and overactive bladder. GAGs were found to have a predictive biomarker potential that could be limited by generalizability concerns.
Assuntos
Glicosaminoglicanos/sangue , Glicosaminoglicanos/urina , Doenças Urológicas/sangue , Doenças Urológicas/urina , Humanos , Doenças Urológicas/diagnósticoRESUMO
BACKGROUND: Portal vein thrombosis (PVT) is the most common thrombotic complication in cirrhosis; however, local risk factors involved in its pathogenesis are still not fully investigated. The aim of the study was to evaluate hemostasis and endothelial damage in the portal vein in patients with cirrhosis and portal hypertension. METHODS: Adult cirrhotics undergoing transjugular intrahepatic portosystemic shunt were consecutively enrolled. Rotational thromboelastometry (ROTEM), dosage of total circulating glycosaminoglycans (GAGs), and endotoxemia levels (lipopolysaccharide [LPS]), along with evaluation of endothelial dysfunction by quantification of circulating endothelial microparticles (MPs), were performed on citrated peripheric and portal venous blood samples from each enrolled patient. RESULTS: Forty-five cirrhotics were enrolled. ROTEM analysis revealed the presence of a significant heparin-like effect in portal blood (median É angle NATEM 50° vs. HEPTEM 55°, p = 0.027; median coagulation time NATEM 665 s vs. HEPTEM 585 s, p = 0.006), which was not detected in peripheral blood, and was associated with a higher concentration of circulating GAGs. Even though total annexin V-MP circulating MPs were less concentrated in the splanchnic district, the proportion of MPs of endothelial origin, with respect to annexin V-MP, was significantly increased in the portal district (p = 0.036). LPS concentration was higher in portal (197 pg/mL) compared with peripheral blood (165 pg/mL) (p < 0.001). CONCLUSION: Evidences of a damage of glycocalyx along with increased concentration of endothelial MPs suggest the presence of a significant endothelial alteration in the portal vein with respect to peripheral veins. Portal site-specific endothelial damage could hamper its antithrombotic properties and may represent an important local risk factor in the pathogenesis of PVT.
Assuntos
Endotélio Vascular/patologia , Cirrose Hepática/patologia , Veia Porta/patologia , Adulto , Coagulação Sanguínea , Micropartículas Derivadas de Células , Endotélio Vascular/fisiopatologia , Endotoxemia/sangue , Feminino , Glicocálix/patologia , Glicosaminoglicanos/sangue , Heparina Liase/farmacologia , Humanos , Hipertensão Portal/etiologia , Hipertensão Portal/cirurgia , Masculino , Pessoa de Meia-Idade , Derivação Portossistêmica Cirúrgica , Tromboelastografia , Trombose Venosa/prevenção & controleRESUMO
Objectives Mucopolysaccharidosis type I (MPS I) was added to our expanded screening panel in 2015. Since then, 127,869 newborns were screened by measuring α-L-iduronidase (IDUA) enzyme activity with liquid chromatography tandem mass spectrometry (LC-MS/MS). High false positives due to frequent pseudodeficiency alleles prompted us to develop a second-tier test to quantify glycosaminoglycan (GAG) levels in dried blood spot (DBS). Methods Heparan-sulfate (HS) and dermatan-sulfate (DS) were measured with LC-MS/MS after methanolysis. DBSs were incubated with methanolic-HCl 3 N at 65 °C for 45 min. Chromatographic separation used an amide column with a gradient of acetonitrile and water with 10 mM ammonium acetate in a 9-min run. The method was validated for specificity, linearity, lower limit of quantification (LOQ), accuracy and precision. Results Intra- and inter-day coefficients of variation were <15% for both metabolites. Reference values in 40 healthy newborns were: HS mean 1.0 mg/L, 0-3.2; DS mean 1.5 mg/L, 0.5-2.7). The two confirmed newborn MPS I patients had elevated HS (4.9-10.4 mg/L, n.v. <3.2) and DS (7.4-8.8 mg/L, n.v. <2.7). Since its introduction in February 2019, the second-tier test reduced the recall rate from 0.046% to 0.006%. Among 127,869 specimens screened, the incidence was 1:63,935 live births. Both patients started enzyme replacement therapy (ERT) within 15 days of birth and one of them received allogenic hematopoietic stem cell transplantation (HSCT) at ht age of 6 months. Conclusions GAGs in DBS increased the specificity of newborn screening for MPS I by reducing false-positives due to heterozygosity or pseudodeficiency. Early diagnosis and therapeutical approach has improved the outcome of our patients with MPS I.
Assuntos
Glicosaminoglicanos/análise , Iduronidase/análise , Mucopolissacaridose I/diagnóstico , Cromatografia Líquida/métodos , Glicosaminoglicanos/sangue , Humanos , Iduronidase/sangue , Recém-Nascido , Mucopolissacaridose I/sangue , Triagem Neonatal/métodos , Valores de Referência , Espectrometria de Massas em Tandem/métodosRESUMO
The aim of the study was to determine whether plasma levels of total glycosaminoglycans (GAGs), matrix metalloproteinases (MMPs) (MMP-3, MMP-10), and their tissue inhibitors (TIMPs) (TIMP-1, TIMP-2) as well as transforming growth factor ß (TGF-ß) differ in the patients with systemic sclerosis (SSc) in relation to the healthy subjects. Plasma samples were obtained from 106 people (64 patients with SSc and 42 healthy individuals) and measured for MMP-3, MMP-10, TIMP-1, TIMP-2, and TGF-ß levels using ELISA methods. GAGs isolated from plasma samples were quantified using a hexuronic acid assay. The plasma levels of total GAGs, TIMP-1, TIMP-2, and TGF-ß were significantly higher, while MMP-3 was significantly decreased in SSc patients compared to the controls. We have revealed a significant correlation between plasma GAGs and TGF-ß (r = -0.47) and TIMP-2 (r = 0.38), respectively. The results of this study revealed that remodeling of the extracellular matrix, reflected by quantitative changes in plasma glycosaminoglycans, occurs during systemic sclerosis. Thus, the alterations in GAG metabolism connected with SSc may lead to systemic changes in the properties of the connective tissue extracellular matrix.
Assuntos
Glicosaminoglicanos/sangue , Metaloproteinase 10 da Matriz/sangue , Metaloproteinase 3 da Matriz/sangue , Escleroderma Sistêmico/sangue , Inibidor Tecidual de Metaloproteinase-1/sangue , Inibidor Tecidual de Metaloproteinase-2/sangue , Fator de Crescimento Transformador beta/sangue , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Severe malaria is mostly caused by Plasmodium falciparum, resulting in considerable, systemic inflammation and pronounced endothelial activation. The endothelium forms an interface between blood and tissue, and vasculopathy has previously been linked with malaria severity. We studied the extent to which the endothelial glycocalyx that normally maintains endothelial function is involved in falciparum malaria pathogenesis by using incident dark-field imaging in the buccal mucosa. This enabled calculation of the perfused boundary region, which indicates to what extent erythrocytes can permeate the endothelial glycocalyx. The perfused boundary region was significantly increased in severe malaria patients and mirrored by an increase of soluble glycocalyx components in plasma. This is suggestive of a substantial endothelial glycocalyx loss. Patients with severe malaria had significantly higher plasma levels of sulfated glycosaminoglycans than patients with uncomplicated malaria, whereas other measured glycocalyx markers were raised to a comparable extent in both groups. In severe malaria, the plasma level of the glycosaminoglycan hyaluronic acid was positively correlated with the perfused boundary region in the buccal cavity. Plasma hyaluronic acid and heparan sulfate were particularly high in severe malaria patients with a low Blantyre coma score, suggesting involvement in its pathogenesis. In vivo imaging also detected perivascular hemorrhages and sequestering late-stage parasites. In line with this, plasma angiopoietin-1 was decreased while angiopoietin-2 was increased, suggesting vascular instability. The density of hemorrhages correlated negatively with plasma levels of angiopoietin-1. Our findings indicate that as with experimental malaria, the loss of endothelial glycocalyx is associated with vascular dysfunction in human malaria and is related to severity.
Assuntos
Endotélio Vascular/patologia , Glicocálix/patologia , Malária Falciparum/patologia , Mucosa Bucal/patologia , Hemorragia Bucal/patologia , Angiopoietina-1/sangue , Angiopoietina-2/sangue , Biomarcadores/sangue , Criança , Pré-Escolar , Endotélio Vascular/fisiopatologia , Feminino , Glicosaminoglicanos/sangue , Humanos , Lactente , Malária Falciparum/sangue , Malária Falciparum/diagnóstico por imagem , Malária Falciparum/fisiopatologia , Masculino , Mucosa Bucal/irrigação sanguínea , Mucosa Bucal/diagnóstico por imagem , Mucosa Bucal/fisiopatologia , Hemorragia Bucal/sangue , Hemorragia Bucal/diagnóstico por imagem , Hemorragia Bucal/fisiopatologiaRESUMO
BACKGROUND: The endothelial glycocalyx controls vascular permeability, cellular signaling, blood-endothelial cell adhesion, extravasation, and transmission of shear stress signals. Burn injury compromises integrity of this layer increasing vascular permeability, which is further exacerbated by large volumes of (intravenous) crystalloids. We have shown that enteral resuscitation is able to reverse burn-induced acute kidney injury (AKI), and herein, we present a follow-up examination of the integrity of the glycocalyx layer and its relationship with renal dysfunction after burn injury. MATERIALS AND METHODS: Anesthetized Yorkshire pigs sustained 40% total body surface area full-thickness contact burns and recovered in metabolic cages for one of three treatments: no fluids (oral or intravenous); (n = 6), ad libitum water (n = 6), or volume-matched oral rehydration solution (ORS; n = 6) for 48 h. Urine and blood were collected at baseline (BL), 6, 12, 24, 32, and 48 h after burn at which point kidneys were harvested. RESULTS: In no fluid and water groups (but not ORS), plasma levels of glycosaminoglycans (GAGs) were elevated after burn (P ≤ 0.031). Syndecan-1 was elevated by 6 h after burn in all animals, but levels declined by 24 h with enteral fluids. Urinary GAGs in the no-fluid group were elevated after burn. No differences among treatments were detected in syndecan-1 levels, or glomerular lectin within the kidney. CONCLUSIONS: Collectively, these data demonstrate that ORS prevented increases in circulating GAGs. Furthermore, an inexpensive and simple method for detecting GAGs provides a sensitive measure of endotheliopathy after burn.
Assuntos
Queimaduras/metabolismo , Glicocálix/fisiologia , Glicosaminoglicanos/análise , Injúria Renal Aguda/diagnóstico , Injúria Renal Aguda/terapia , Animais , Modelos Animais de Doenças , Células Endoteliais/fisiologia , Glicosaminoglicanos/sangue , Glicosaminoglicanos/urina , Túbulos Renais/patologia , Lectinas/análise , Soluções para Reidratação/uso terapêutico , Suínos , Sindecana-1/análiseRESUMO
Therapeutic development and monitoring require demonstration of effects on disease phenotype. However, due to the complexity of measuring clinically-relevant effects in rare multisystem diseases, robust biomarkers are essential. For the mucopolysaccharidoses (MPS), the measurement of glycosaminoglycan levels is relevant as glycosaminoglycan accumulation is the primary event that occurs due to reduced lysosomal enzyme activity. Traditional dye-based assays that measure total glycosaminoglycan levels have a high background, due to a normal, baseline glycosaminoglycan content in unaffected individuals. An assay that selectively detects the disease-specific non-reducing ends of heparan sulfate glycosaminoglycans that remain undegraded due to deficiency of a specific enzyme in the catabolic pathway avoids the normal background, increasing sensitivity and specificity. We evaluated glycosaminoglycan content by dye-based and non-reducing end methods using urine, serum, and cerebrospinal fluid from MPS I human samples before and after treatment with intravenous recombinant human alpha-l-iduronidase. We found that both urine total glycosaminoglycans and serum heparan sulfate derived non-reducing end levels were markedly decreased compared to baseline after 26â¯weeks and 52â¯weeks of therapy, with a significantly greater percentage reduction in serum non-reducing end (89.8% at 26â¯weeks and 81.3% at 52â¯weeks) compared to urine total glycosaminoglycans (68.3% at 26â¯weeks and 62.4% at 52â¯weeks, pâ¯<â¯0.001). Unexpectedly, we also observed a decrease in non-reducing end levels in cerebrospinal fluid in all five subjects for whom samples were collected (mean 41.8% reduction, pâ¯=â¯0.01). The non-reducing ends in cerebrospinal fluid showed a positive correlation with serum non-reducing end levels in the subjects (r2â¯=â¯0.65, pâ¯=â¯0.005). Results suggest utility of the non-reducing end assay in evaluating a therapeutic response in MPS I.
Assuntos
Terapia de Reposição de Enzimas , Glicosaminoglicanos/sangue , Glicosaminoglicanos/urina , Mucopolissacaridose I/tratamento farmacológico , Biomarcadores/sangue , Técnicas de Laboratório Clínico , Monitoramento de Medicamentos/métodos , Glicosaminoglicanos/líquido cefalorraquidiano , Humanos , Iduronidase/genética , Iduronidase/uso terapêuticoRESUMO
In some patients diabetic ketoacidosis (DKA) causes acute endothelial injury and multiorgan failure. Measurement of glycosaminoglycan (GAG) and advanced glycation end products (AGE) could provide information to help understand the biochemical events associated with poor outcomes in these patients. This study included 37 patients with DKA admitted to an intensive care unit. Blood was collected from these patients during the first day of hospitalization, 24 hours after the first sample, and 72 hours after the first sample when possible. ELISA-based assays were used to measure glucose, hemoglobin A1c, AGE, glycated albumin, and GAG levels in serum. Twenty healthy control subjects with no history of diabetes donated 1 blood sample. Control subjects had a mean age of 36.3±12.1 years; patients with DKA had a mean age of 38.1±18.5 years. Admission laboratory tests in patients with DKA included glucose 546±296 mg/dL, bicarbonate 10.1±5.5 mEq/L, anion gap 31.8±7.8 mEq/L, and creatinine 1.1±1.0 mg/dL. Patients with DKA had significantly higher level glucose and free glycated hemoglobin. Control subjects had significantly higher levels of AGE and glycated albumin. There were no differences in soluble receptor for AGE levels or GAG levels between the control subjects and patients with DKA. Patients with DKA had lower circulating levels of AGE and glycated albumin than control subjects. These results may reflect absorption of these proteins to damaged capillary surfaces or loss of proteins into interstitial spaces secondary to increased endothelial permeability.
Assuntos
Cetoacidose Diabética/sangue , Hemoglobinas Glicadas/análise , Produtos Finais de Glicação Avançada/sangue , Glicosaminoglicanos/sangue , Adulto , Biomarcadores/sangue , Glicemia/análise , Feminino , Humanos , MasculinoRESUMO
INTRODUCTION: The prevalence of cardiovascular (CV) comorbidity in patients with chronic kidney disease (CKD) is high, particularly in endstage renal disease (ESRD). There is an ongoing search for novel biomarkers of CV disease in this population. OBJECTIVES: We aimed to investigate the associations of matrix proteoglycans (PGs) and glycosaminoglycans (GAGs), collagen, and arterial calcifications with selected serum and plasma markers of endothelial dysfunction, inflammation, oxidative stress, and bone turnover in patients with ESRD. PATIENTS AND METHODS: We enrolled 47 adult patients (32 men) with stage 5 CKD. The following parameters were investigated: fibrinogen, soluble thrombomodulin (sTM), plasminogen activator inhibitor 1 (PAI1), stromal cellderived factor 1α (SDF1α), calcium (Ca), phosphate (Pi), intact parathormone, interleukin 6, highsensitivity Creactive protein (hsCRP), ferric reducing ability of plasma, 2,2diphenyl1picrylhydrazyl scavenging, ferric reducing ability of ascorbate in plasma, fetuinA, fibroblast growth factor 23, osteopontin, osteoprotegerin, osteocalcin, transforming growth factor ß (TGFß), hepatocyte growth factor, secreted protein acidic and rich in cysteine, as well as matrix metalloproteinase 2. Radial artery specimens were stained with alizarin red for calcifications, alcian blue for PGs and GAGs, and sirius red for collagen. RESULTS: We observed positive correlations between PG and GAG, collagen, and calcification staining. The most intense (grade 3) alcian blue staining was significantly correlated with diabetes as well as higher levels of Ca × Pi product, hsCRP, fibrinogen, SDF1α, PAI1, and sTM. However, PAI1 was the only significant predictor of grade 3 alcian blue staining in a multiple logistic regression model adjusted for hemodialysis, Ca× Pi product, and hsCRP levels. CONCLUSIONS: Coagulation disorders and endothelial dysfunction are the hallmarks of ESRD. The levels of SDF1α, PAI1, sTM, and fibrinogen may be novel predictors of early vascular wall alterations and may serve as CV risk markers.
Assuntos
Biomarcadores/sangue , Doenças Cardiovasculares/sangue , Colágeno/sangue , Glicosaminoglicanos/sangue , Falência Renal Crônica/sangue , Falência Renal Crônica/complicações , Proteoglicanas/sangue , Artéria Radial/química , Idoso , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/fisiopatologia , Comorbidade , Feminino , Humanos , Falência Renal Crônica/epidemiologia , Falência Renal Crônica/fisiopatologia , Masculino , Pessoa de Meia-Idade , Polônia/epidemiologiaRESUMO
The discrimination of glycosaminoglycans (GAGs) is a challenging task but of great importance to ensure their safe use in clinics. Herein, four supramolecular AIE nanoassemblies denoted as PDDA-TPE100, PDDA-TPE75, PDDA-TPE50, and PDDA-TPE25 were synthesized by loading different amounts of the negatively charged AIEgen TPE onto the surface of the positively charged polymer PDDA. These AIE nanoassemblies were utilized for the construction of a fluorescent sensor array, which was able to discriminate various GAGs on the basis of a compaction to displacement reaction mechanism. LDA and HCA results revealed that GAGs including Hep, Chs, HA, CTS, DS, and OSCS can be discriminated with 100% accuracy. The four-sensor array was then simplified to a three-sensor array using PDDA-TPE100, PDDA-TPE75, and PDDA-TPE50 as the sensors, which was determined to be still powerful in discrimination of various GAGs, accurate identification of unknown GAG samples, sensitive detection of trace OSCS contaminant in Hep, and identification of Hep and other biologically abundant anions. Moreover, the three-sensor array was even successfully applied for differentiating GAGs in serum media.
Assuntos
Técnicas Biossensoriais/métodos , Glicosaminoglicanos/sangue , Heparina/sangue , Nanocompostos/química , Polímeros/química , Controle de Qualidade , Glicosaminoglicanos/química , Heparina/química , HumanosRESUMO
Effective regeneration of damaged soft orbital tissues in Graves' ophthalmopathy (GO) requires coordinated remodeling of the extracellular matrix. Matrix metalloproteinases (MMPs) play an important role in the synthesis and degradation homeostasis of extracellular matrix components in various physiological and pathological conditions. Their proteolytic activity is inhibited by tissue inhibitors of metalloproteinases (TIMP). The biochemical processes taking place in extraocular muscles and retrobulbar tissue fibrogenesis in GO are not fully understood. Aims - to assess some biochemical mechanisms of extraocular muscles and retrobulbar tissue fibrogenesis in GO patients. MATERIAL AND METHODS: The study included 65 people (130 eyes) at the age of 43 (35-50) years. Three groups of subjects were formed: 32 patients with a moderate GO severity (clinical group), 18 patients with autoimmune thyroid pathology without GO (comparison group), and 15 healthy persons (control). The diagnosis was based on clinical, laboratory, and instrumental data. A comprehensive ophthalmologic examination and blood sampling for determination of MMP-1, -13, TIMP-1, sulfated glycosaminoglycans (sGAG) and antibodies to thyroid-stimulating hormone receptor (TSHRAbs) were conducted. The data were statistically processed using the program Statistica 10.0. RESULTS: An elevated level of MMP-13, observed in all GO patients (p<0.05). For the active phase of GOP, the comparison with the control group showed a 3.5-fold increase in MMP-13 (p<0.001) and 1.17-fold rise in TIMP-1 (p>0.05). Pulse glucocorticoid therapy reduced MMP-13 by 48.6% (p<0.001), TIMP-1 by 2.7% (p<0.001), and TSHRAbs - by 93% (p<0.001) compared with active GO, but these indicators were higher than the reference limits of control (p>0.05). In inactive GO, despite increased MMP-13, TIMP-1 decreased to the reference values (p=0.533). There were no significant differences in MMP-1 in groups of subjects (p=0.865). CONCLUSIONS: We have found imbalance between MMP-13 and TIMP-1 production in different activity phases of GO. Active GO is characterized by an increase in serum MMP-13 and TIMP-1. Dysregulation of intercellular matrix remodeling, possibly, underlies the development of extraocular muscles and retrobulbar tissue fibrosis in GO.
Assuntos
Oftalmopatia de Graves/sangue , Metaloproteinase 13 da Matriz/sangue , Metaloproteinase 1 da Matriz/sangue , Inibidor Tecidual de Metaloproteinase-1/sangue , Adulto , Biomarcadores/sangue , Feminino , Glicosaminoglicanos/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Receptores da Tireotropina/metabolismoRESUMO
BACKGROUND: The homologous plasma proteins prekallikrein and factor XI (FXI) circulate as complexes with high molecular weight kininogen. Although evidence supports an interaction between the prekallikrein-kininogen complexes and vascular endothelium, there is conflicting information regarding FXI binding to endothelium. OBJECTIVE: To study the interaction between FXI and blood vessels in mice. METHODS: C57Bl/6 wild-type or F11-/- mice in which variants of FXI were expressed by hydrodynamic tail vein injection, received intravenous infusions of saline, heparin, polyphosphates, protamine, or enzymes that digest glycosaminoglycans (GAGs). Blood was collected after infusion and plasma was analyzed by western blot for FXI. RESULTS AND CONCLUSIONS: Plasma FXI increased 5- to 10-fold in wild-type mice after infusion of heparin, polyphosphates, protamine, or GAG-digesting enzymes, but not saline. Similar treatments resulted in a much smaller change in plasma FXI levels in rats, and infusions of large boluses of heparin did not change FXI levels appreciably in baboons or humans. The releasable FXI fraction was reconstituted in F11-/- mice by expressing murine FXI, but not human FXI. We identified a cluster of basic residues on the apple 4 domain of mouse FXI that is not present in other species. Replacing the basic residues with alanine prevented the interaction of mouse FXI with blood vessels, whereas introducing the basic residues into human FXI allowed it to bind to blood vessels. Most FXI in mice is noncovalently associated with GAGs on blood vessel endothelium and does not circulate in plasma.
Assuntos
Endotélio Vascular/metabolismo , Fator XI/metabolismo , Glicosaminoglicanos/sangue , Animais , Sítios de Ligação , Trombose das Artérias Carótidas/sangue , Trombose das Artérias Carótidas/induzido quimicamente , Cloretos/toxicidade , Fator XI/química , Deficiência do Fator XI/sangue , Compostos Férricos/toxicidade , Heparina/farmacologia , Humanos , Cininogênios/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Moleculares , Papio , Pré-Calicreína/metabolismo , Ligação Proteica , Conformação Proteica , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Especificidade da Espécie , Eletricidade EstáticaRESUMO
BACKGROUND: In this study, the effect of 15-month anti-tumor necrosis factor alpha (TNF-α) treatment on circulating levels of plasma sulfated glycosaminoglycans (GAGs) and the nonsulfated GAG hyaluronic acid (HA) in female rheumatoid arthritis (RA) patients was assessed. METHODS: Plasma was obtained from healthy subjects and RA women treated with TNF-α antagonists (etanercept or adalimumab or certolizumab pegol) in combination with methotrexate. GAGs were isolated from plasma samples using ion exchange low-pressure liquid chromatography. Total sulfated GAGs were quantified using a hexuronic acid assay. Plasma levels of keratan sulfate (KS) and HA were measured using immunoassay kits. RESULTS: Total sulfated GAGs and HA levels were higher in female RA patients before treatment in comparison to healthy subjects. KS levels did not differ between RA women and controls. Anti-TNF-α treatment resulted in normalization of plasma total GAG and HA levels in RA patients, without any effect on KS levels. CONCLUSIONS: Our results suggest that anti-TNF-α therapy has a beneficial effect on extracellular matrix remodeling in the course of RA.
Assuntos
Antirreumáticos/administração & dosagem , Artrite Reumatoide/sangue , Artrite Reumatoide/tratamento farmacológico , Glicosaminoglicanos/sangue , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Adalimumab/administração & dosagem , Adulto , Biomarcadores/sangue , Certolizumab Pegol/administração & dosagem , Quimioterapia Combinada , Etanercepte/administração & dosagem , Feminino , Humanos , Metotrexato/administração & dosagem , Pessoa de Meia-Idade , Fatores de Tempo , Resultado do Tratamento , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Dried blood spot (DBS) technology is a cheap and easy method largely applied in newborn screening. Mucopolysaccharidoses (MPS) are characterized by the deficit of enzymes that degrade glycosaminoglycans (GAGs) characterized by progressive worsening of the conditions. For a possible early diagnosis of MPS, we developed a method of uronic acid (UA)-GAGs determination in DBS of 600 healthy newborns and from a small group of MPS subjects matched for age. Spotted blood UA-GAGs of the normal newborns are composed of 67.2% chondroitin sulfate (CS), 28.6% heparan sulfate (HS) and 4.4% hyaluronic acid with a CS/HS ratio of 2.35 and a total GAGs content of 0.43 µg/DBS. A chemical evaluation of CS and HS structure was performed by measuring their disaccharide composition, sulfation and the overall charge density. The DBS of four different MPS types presented an increase of total or single UA-GAGs content and/or modifications of the CS and HS disaccharide composition as well as chemical signature also related to the MPS enzymatic defect. The modifications of the UA-GAGs composition, parameters and structure of healthy newborns determined in DBS would be useful for a possible early diagnosis of various MPS types.
Assuntos
Teste em Amostras de Sangue Seco , Glicosaminoglicanos/sangue , Glicosaminoglicanos/química , Mucopolissacaridoses/sangue , Mucopolissacaridoses/diagnóstico , Configuração de Carboidratos , Humanos , Recém-NascidoRESUMO
To explore the correlation between glycosaminoglycan (GAG) levels and mucopolysaccharidosis (MPS) type, we have evaluated the GAG levels in blood of MPS II, III, IVA, and IVB and urine of MPS IVA, IVB, and VI by tandem mass spectrometry. Dermatan sulfate (DS), heparan sulfate (HS), keratan sulfate (KS; mono-sulfated KS, di-sulfated KS), and the ratio of di-sulfated KS in total KS were measured. Patients with untreated MPS II had higher levels of DS and HS in blood while untreated MPS III had higher levels of HS in blood than age-matched controls. Untreated MPS IVA had higher levels of KS in blood and urine than age-matched controls. The ratio of blood di-sulfated KS/total KS in untreated MPS IVA was constant and higher than that in controls for children up to 10â¯years of age. The ratio of urine di-sulfated KS/total KS in untreated MPS IVA was also higher than that in age-matched controls, but the ratio in untreated MPS IVB was lower than controls. ERT reduced blood DS and HS in MPS II, and urine KS in MPS IVA patients, although GAGs levels remained higher than the observed in age-matched controls. ERT did not change blood KS levels in MPS IVA. MPS VI under ERT still had an elevation of urine DS level compared to age-matched controls. There was a positive correlation between blood and urine KS in untreated MPS IVA patients but not in MPS IVA patients treated with ERT. Blood and urine KS levels were secondarily elevated in MPS II and VI, respectively. Overall, measurement of GAG levels in blood and urine is useful for diagnosis of MPS, while urine KS is not a useful biomarker for monitoring therapeutic efficacy in MPS IVA.
Assuntos
Glicosaminoglicanos/sangue , Glicosaminoglicanos/urina , Mucopolissacaridoses/sangue , Mucopolissacaridoses/urina , Adolescente , Adulto , Biomarcadores/sangue , Biomarcadores/urina , Criança , Pré-Escolar , Dermatan Sulfato/sangue , Dermatan Sulfato/urina , Feminino , Glicosaminoglicanos/isolamento & purificação , Heparitina Sulfato/sangue , Heparitina Sulfato/urina , Humanos , Sulfato de Queratano/sangue , Sulfato de Queratano/urina , Masculino , Mucopolissacaridoses/classificação , Mucopolissacaridoses/patologia , Mucopolissacaridose II/sangue , Mucopolissacaridose II/patologia , Mucopolissacaridose II/urina , Mucopolissacaridose III/sangue , Mucopolissacaridose III/patologia , Mucopolissacaridose III/urina , Mucopolissacaridose IV/sangue , Mucopolissacaridose IV/patologia , Mucopolissacaridose IV/urina , Mucopolissacaridose VI/sangue , Mucopolissacaridose VI/patologia , Mucopolissacaridose VI/urina , Espectrometria de Massas em Tandem , Adulto JovemRESUMO
Mucopolysaccharidosis IVA (MPS IVA, Morquio A syndrome) is an autosomal recessive disorder caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase. Deficiency of this enzyme leads to the accumulation of specific glycosaminoglycans (GAGs), chondroitin-6-sulfate (C6S) and keratan sulfate (KS), which are mainly synthesized in the cartilage. Therefore, the substrates are stored primarily in the cartilage and its extracellular matrix (ECM), leading to a direct impact on bone development and successive systemic skeletal spondylepiphyseal dysplasia. The skeletal-related symptoms for MPS IVA include short stature with short neck and trunk, odontoid hypoplasia, spinal cord compression, tracheal obstruction, obstructive airway, pectus carinatum, restrictive lung, kyphoscoliosis, platyspondyly, coxa valga, genu valgum, waddling gait, and laxity of joints. The degree of imbalance of growth in bone and other organs and tissues largely contributes to unique skeletal dysplasia and clinical severity. Diagnosis of MPS IVA needs clinical, radiographic, and laboratory testing to make a complete conclusion. To diagnose MPS IVA, total urinary GAG analysis which has been used is problematic since the values overlap with those in age-matched controls. Currently, urinary and blood KS and C6S, the enzyme activity of GALNS, and GALNS molecular analysis are used for diagnosis and prognosis of clinical phenotype in MPS IVA. MPS IVA can be diagnosed with unique characters although this disorder relates closely to other disorders in some characteristics. In this review article, we comprehensively describe clinical, radiographic, biochemical, and molecular diagnosis and clinical assessment tests for MPS IVA. We also compare MPS IVA to other closely related disorders to differentiate MPS IVA. Overall, imbalance of growth in MPS IVA patients underlies unique skeletal manifestations leading to a critical indicator for diagnosis.
Assuntos
Condroitina Sulfatases/genética , Mucopolissacaridose IV/genética , Prognóstico , Cartilagem/metabolismo , Cartilagem/patologia , Sulfatos de Condroitina/sangue , Sulfatos de Condroitina/urina , Terapia de Reposição de Enzimas , Glicosaminoglicanos/sangue , Glicosaminoglicanos/urina , Humanos , Sulfato de Queratano/sangue , Sulfato de Queratano/urina , Mucopolissacaridose IV/sangue , Mucopolissacaridose IV/tratamento farmacológico , Mucopolissacaridose IV/urina , FenótipoRESUMO
The glycosaminoglycans (GAGs) heparan sulfate, dermatan sulfate, and heparin are important anticoagulants that inhibit clot formation through interactions with antithrombin and heparin cofactor II. Unfractionated heparin, low-molecular-weight heparin, and heparin-derived drugs are often the main treatments used clinically to handle coagulatory disorders. A wide range of proteins have been reported to bind and neutralize these GAGs to promote clot formation. Such neutralizing proteins are involved in a variety of other physiological processes, including inflammation, transport, and signaling. It is clear that these interactions are important for the control of normal coagulation and influence the efficacy of heparin and heparin-based therapeutics. In addition to neutralization, the anticoagulant activities of GAGs may also be regulated through reduced synthesis or by degradation. In this review, we describe GAG neutralization, the proteins involved, and the molecular processes that contribute to the regulation of anticoagulant GAG activity.
Assuntos
Anticoagulantes/uso terapêutico , Coagulação Sanguínea/efeitos dos fármacos , Glicosaminoglicanos/antagonistas & inibidores , Antagonistas de Heparina/uso terapêutico , Heparina/uso terapêutico , Animais , Anticoagulantes/efeitos adversos , Sítios de Ligação , Dermatan Sulfato/antagonistas & inibidores , Dermatan Sulfato/sangue , Glicosaminoglicanos/sangue , Heparina/efeitos adversos , Antagonistas de Heparina/efeitos adversos , Heparitina Sulfato/antagonistas & inibidores , Heparitina Sulfato/sangue , Humanos , Ligação ProteicaRESUMO
Certain recessively inherited diseases result from an enzyme deficiency within lysosomes. In mucopolysaccharidoses (MPS), a defect in glycosaminoglycan (GAG) degradation leads to GAG accumulation followed by progressive organ and multiple system dysfunctions. Current methods of GAG analysis used to diagnose and monitor the diseases lack sensitivity and throughput. Here we report a LC-MS method with accurate metabolite mass analysis for identifying and quantifying biomarkers for MPS type I without the need for extensive sample preparation. The method revealed 225 LC-MS features that were >1000-fold enriched in urine, plasma and tissue extracts from untreated MPS I mice compared to MPS I mice treated with iduronidase to correct the disorder. Levels of several trisaccharides were elevated >10000-fold. To validate the clinical relevance of our method, we confirmed the presence of these biomarkers in urine, plasma and cerebrospinal fluid from MPS I patients and assessed changes in their levels after treatment.