Correlation analysis of Trial of Org 10172 in acute stroke treatment classification and National Institutes of Health Stroke Scale score in acute cerebral infarction with risk factors

SUMMARY OBJECTIVE: The aim of this study was to investigate the correlation between the Trial of Org 10172 in acute stroke treatment classification and the National Institutes of Health Stroke Scale score of acute cerebral infarction as well as acute cerebral infarction's risk factors. METHODS: The clinical data of 3,996 patients with acute cerebral infarction hospitalized in Hebei Renqiu Kangjixintu Hospital from January 2014 to November 2018 were analyzed retrospectively. According to Trial of Org 10172 in acute stroke treatment, they were divided into five groups: arteriosclerosis, cardio cerebral embolism, arterial occlusion, other causes, and unknown causes. Through questionnaire design, routine physical examination, and physical and chemical analysis of fasting venous blood samples, the risk factors were evaluated, and the correlation between Trial of Org 10172 in acute stroke treatment classification and National Institutes of Health Stroke Scale classification was analyzed using multivariate logistic regression. In addition, the relationship between National Institutes of Health Stroke Scale score and risk factors in different groups was compared, and the correlation between Trial of Org 10172 in acute stroke treatment classification and National Institutes of Health Stroke Scale score was analyzed. RESULTS: Multivariate logistic regression analysis showed that diabetes, atrial fibrillation or stroke history, age, and education level were related to Trial of Org 10172 in acute stroke treatment classification. In the National Institutes of Health Stroke Scale comparison, the scores of the cardio cerebral embolism group were significantly higher than those of the other four groups, and patients with diabetes, atrial fibrillation, or stroke history had a high share, especially atrial fibrillation (33.06%). CONCLUSIONS: The nerve function defect is more serious after acute cerebral infarction with cardiogenic cerebral embolism, indicating a poor prognosis.

A new heparan sulfate from the mollusk Nodipecten nodosus inhibits merozoite invasion and disrupts rosetting and cytoadherence of Plasmodium falciparum

BACKGROUND Despite treatment with effective antimalarial drugs, the mortality rate is still high in severe cases of the disease, highlighting the need to find adjunct therapies that can inhibit the adhesion of Plasmodium falciparum-infected erythrocytes (Pf-iEs). OBJECTIVES In this context, we evaluated a new heparan sulfate (HS) from Nodipecten nodosus for antimalarial activity and inhibition of P. falciparum cytoadhesion and rosetting. METHODS Parasite inhibition was measured by SYBR green using a cytometer. HS was assessed in rosetting and cytoadhesion assays under static and flow conditions using Chinese hamster ovary (CHO) and human lymphatic endothelial cell (HLEC) cells expressing intercellular adhesion molecule-1 (ICAM1) and chondroitin sulfate A (CSA), respectively. FINDINGS This HS inhibited merozoite invasion similar to heparin. Moreover, mollusk HS decreased cytoadherence of P. falciparum to CSA and ICAM-1 on the surface of endothelial cells under static and flow conditions. In addition, this glycan efficiently disrupted rosettes. CONCLUSIONS These findings support a potential use for mollusk HS as adjunct therapy for severe malaria.

Prospective evaluation of Chondroitin sulfate, Heparan sulfate and Hyaluronic acid in prostate cancer

ABSTRACT Purpose: The present study evaluates chondroitin sulfate (CS) and heparan sulfate (HS) in the urine and hyaluronic acid (HA) in the plasma of patients with prostate cancer before and after treatment compared to a control group. Materials and Methods: Plasma samples were used for HA dosage and urine for quantification of CS and HS from forty-four cancer patients and fourteen controls. Clinical, laboratory and radiological information were correlated with glycosaminoglycan quantification by statistical analysis. Results: Serum HA was significantly increased in cancer patients (39.68 ± 30.00 ng/ mL) compared to control group (15.04 ± 7.11 ng/mL; p=0.004) and was further increased in high-risk prostate cancer patients when compared to lower risk patients (p = 0.0214). Also, surgically treated individuals had a significant decrease in seric levels of heparan sulfate after surgical treatment, 31.05 ± 21.01 μg/mL (before surgery) and 23.14 ± 11.1 μg/mL (after surgery; p=0.029). There was no difference in the urinary CS and HS between prostate cancer patients and control group. Urinary CS in cancer patients was 27.32 ± 25.99 μg/mg creatinine while in the men unaffected by cancer it was 31.37 ± 28.37 μg/mg creatinine (p=0.4768). Urinary HS was 39.58 ± 32.81 μg/ mg creatinine and 35.29 ± 28.11 μg/mg creatinine, respectively, in cancer patients and control group (p=0.6252). Conclusions: Serum HA may be a useful biomarker for the diagnosis and prognosis of prostate cancer. However, urinary CS and HS did not altered in the present evaluation. Further studies are necessary to confirm these preliminary findings.

Glycomics expression analysis of sulfated glycosaminoglycans of human colorectal cancer tissues and non-neoplastic mucosa by electrospray ionization mass spectrometry / Análise da expressão de glicosaminoglicanos sulfatados no tecido humano neoplásico colorretal e na mucosa não neoplásica por espectrometria de massa por ionização por electrospray

ABSTRACT Objective To determine the presence of glycosaminoglycans in the extracellular matrix of connective tissue from neoplastic and non-neoplastic colorectal tissues, since it has a central role in tumor development and progression. Methods Tissue samples from neoplastic and non-neoplastic colorectal tissues were obtained from 64 operated patients who had colorectal carcinoma with no distant metastases. Expressions of heparan sulphate, chondroitin sulphate, dermatan sulphate and their fragments were analyzed by electrospray ionization mass spectrometry, with the technique for extraction and quantification of glycosaminoglycans after proteolysis and electrophoresis. The statistical analysis included mean, standard deviation, and Student’st test. Results The glycosaminoglycans extracted from colorectal tissue showed three electrophoretic bands in agarose gel. Electrospray ionization mass spectrometry showed characteristic disaccharide fragments from glycosaminoglycans, indicating their structural characterization in the tissues analyzed. Some peaks in the electrospray ionization mass spectrometry were not characterized as fragments of sugars, indicating the presence of fragments of the protein structure of proteoglycans generated during the glycosaminoglycan purification. The average amount of chondroitin and dermatan increased in the neoplastic tissue compared to normal tissue (p=0.01). On the other hand, the average amount of heparan decreased in the neoplastic tissue compared to normal tissue (p= 0.03). Conclusion The method allowed the determination of the glycosaminoglycans structural profile in colorectal tissue from neoplastic and non-neoplastic colorectal tissue. Neoplastic tissues showed greater amounts of chondroitin sulphate and dermatan sulphate compared to non-neoplastic tissues, while heparan sulphate was decreased in neoplastic tissues.

RESUMO Objetivo Determinar a presença de glicosaminoglicanos na matriz extracelular do tecido conjuntivo colorretal neoplásico e não neoplásico, tendo em vista seu papel central no desenvolvimento e na progressão dos tumores. Métodos Amostras de tecidos colorretais neoplásicos e não neoplásicos foram obtidas de 64 pacientes operados com carcinoma colorretal sem metástases a distância. As expressões de heparan sulfato, sulfato de condroitina e sulfato de dermatan e seus fragmentos foram analisadas por espectrometria de massa por ionização por electrospray, com técnica de extração e quantificação de glicosaminoglicanos após proteólise e eletroforese. Para análise estatística, utilizaram-se média, desvio padrão e teste t de Student. Resultados Em gel de agarose, os glicosaminoglicanos extraídos de tecido colorretal mostraram três bandas eletroforéticas. A espectrometria de massa por ionização por electrospray mostrou fragmentos de dissacarídeos característicos de glicosaminoglicanos e indicou sua característica estrutural. Alguns picos na espectrometria de massa por ionização por electrospray não foram caracterizados como fragmentos de açúcares, sugerindo a presença de fragmentos de proteínas estruturais dos proteoglicanos, formadas durante a purificação dos glicosaminoglicanos. A quantidade média de condroitina e dermatan aumentou no tecido neoplástico em relação ao tecido normal (p=0,01). Por outro lado, a quantidade média de heparan foi menor no tecido neoplásico em relação ao tecido normal (p=0,03). Conclusão O método empregado permitiu determinar o perfil estrutural dos glicosaminoglicanos nas amostras. Tecidos neoplásicos apresentaram maiores quantidades de sulfato de condroitina e sulfato de dermatan em comparação com os não neoplásicos, enquanto o sulfato de heparan foi encontrado em menores quantidades nos tecidos neoplásicos.

Effect of castration on renal glycosaminoglycans and their urinary excretion in male and female rats with chronic renal failure

Glycosaminoglycans (GAGs) participate in a variety of processes in the kidney, and evidence suggests that gender-related hormones participate in renal function. The aim of this study was to analyze the relationship of GAGs, gender, and proteinuria in male and female rats with chronic renal failure (CRF). GAGs were analyzed in total kidney tissue and 24-h urine of castrated (c), male (M), and female (F) Wistar control (C) rats (CM, CMc, CF, CFc) and after 30 days of CRF induced by 5/6 nephrectomy (CRFM, CRFMc, CRFF, CRFFc). Total GAG quantification and composition were determined using agarose and polyacrylamide gel electrophoresis, respectively. Renal GAGs were higher in CF compared to CM. CRFM presented an increase in renal GAGs, heparan sulfate (HS), and proteinuria, while castration reduced these parameters. However, CRFF and CRFFc groups showed a decrease in renal GAGs concomitant with an increase in proteinuria. Our results suggest that, in CRFM, sex hormones quantitatively alter GAGs, mainly HS, and possibly the glomerular filtration barrier, leading to proteinuria. The lack of this response in CRFMc, where HS did not increase, corroborates this theory. This pattern was not observed in females. Further studies of CRF are needed to clarify gender-dependent differences in HS synthesis.

Heparan sulphate, its derivatives and analogues share structural characteristics that can be exploited, particularly in inhibiting microbial attachment

Heparan sulphate (HS) and the related polysaccharide, heparin, exhibit conformational and charge arrangement properties, which provide a degree of redundancy allowing several seemingly distinct sequences to exhibit the same activity. This can also be mimicked by other sulphated polysaccharides, both in overall effect and in the details of interactions and structural consequences of interactions with proteins. Together, these provide a source of active compounds suitable for further development as potential drugs. These polysaccharides also possess considerable size, which bestows upon them an additional useful property: the capability of disrupting processes comprising many individual interactions, such as those characterising the attachment of microbial pathogens to host cells. The range of involvement of HS in microbial attachment is reviewed and examples, which include viral, bacterial and parasitic infections and which, in many cases, are now being investigated as potential targets for intervention, are identified.

Glycosaminoglycan distribution in the rat uterine cervix during the estrous cycle

OBJECTIVE: To analyze the amount of glycosaminoglycans in the uterine cervix during each phase of the rat estrous cycle. DESIGN: Based on vaginal smears, forty female, regularly cycling rats were divided into four groups (n = 10 for each group): GI - proestrous, GII - estrous, GIII - metaestrous and GIV - diestrous. Animals were sacrificed at each phase of the cycle, and the cervix was immediately removed and submitted to biochemical extraction and determination of sulfated glycosaminoglycans and hyaluronic acid. The results were analyzed by ANOVA followed by the Bonferroni post-hoc test. RESULTS: The uterine cervix had the highest amount of total sulfated glycosaminoglycans and dermatan sulfate during the estrous phase (8.90 ± 0.55 mg/g of cetonic extract, p<0.001; and 8.86 ± 0.57 mg/g of cetonic extract, p<0.001). In addition, there was more heparan sulfate at the cervix during the proestrous phase (0.185 ± 0.03 mg/g of cetonic extract) than during any other phase (p<0.001). There were no significant changes in the concentration of hyaluronic acid in the uterine cervix during the estrous cycle. CONCLUSION: Our data suggest that the amount of total sulfated glycosaminoglycans may be influenced by hormonal fluctuations related to the estrous cycle, with dermatan sulfate and heparan sulfate being the glycosaminoglycans most sensitive to hormonal change.

O papel da matriz extracelular na cicatrização de feridas bucais / Extracellular matrix role on the wound healing of oral lesions

A matriz extracelular (MEC) é uma estrutura complexa que proporciona o arcabouço que sustenta as células nos tecidos de todos os mamíferos. Uma contínua reconstrução e mudança nos constituintes da MEC ocorrem normalmente durante o processo de reparo de uma ferida. Assim, pode-se afirmar que a cicatrização é uma integração dinâmica que envolve células, MEC e mediadores solúveis. Dessa maneira, a cicatrização depende não só das células e polipeptídeos disponíveis, mas do microambiente da matriz extracelular (MEC). Devido à imensa gama de fatores de origem local ou sistêmica, exógena ou endógena, certamente ocorrem modificações muito mais complexas e profundas nos componentes da MEC e suas inter-relações com as células envolvidas no processo de reparação de feridas bucais, o que influencia a qualidade e quantidade de tecido reparador em cada tipo de ferimento.

Extracellular matrix (ECM) is a complex structure which supports and maintains all the cells organized in tissues in mammalians. A continuous reconstruction and change of the ECM components normally occur during the repair of injured tissue. Thus, it can be affirmed that wound healing is a dynamic integration among cells, ECM and soluble mediators. So it depends not only on the available polypeptides and cells, but especially on the micro environment of the ECM. Because of the enormous endogenous and exogenous, local and systemic factors related, many complex and profound modifications certainly occur in the components of the ECM and also in their relation with all the cells involved in oral lesion repair. These factors influence the quality and quantity of the repairing tissue in each type of lesion.

Perfil de glicosaminoglicanos sulfatados no útero de camundongas durante o ciclo estral / Profile of sulphated glycosaminoglycans content in the murine uterus during the different phases of the estrous cycle

OBJETIVOS: Quantificar glicosaminoglicanos sulfatados (GAGs) no útero de camundongas durante o ciclo estral. MÉTODOS: Utilizaram-se quatro grupos de camundongas virgens com 100 dias de idade (n= 10 cada) conforme a fase ciclo estral: proestro, estro, metaestro e diestro. Amostras da porção média dos cornos uterinos foram preparadas para observação em microscopia de luz (H/E e Alcian blue + PAS). Os GAGs foram extraídos e caracterizados por eletroforese em gel de agarose. Os dados foram analisados pelo teste t de Student não pareado. RESULTADOS: A microscopia de luz, os GAGs sulfatados apresentam-se em todas as camadas do útero, em especial no endométrio, entre as fibras colágenas, na membrana basal e ao redor dos fibroblastos. A análise bioquímica mostrou haver dermatam sulfato (DS), condroitim sulfato (CS) e heparam sulfato (HS) durante todas as fases do ciclo estral. Não houve separação eletroforética clara entre DS e CS, de modo que estes dois GAGs foram considerados em conjunto (DS+CS) (proestro = 0,854 ± 0,192; estro = 1,073 ± 0,254; metaestro = 1,003 ± 0,255; e diestro = 0,632 ± 0,443 μg/mg). Os resultados de HS foram: proestro = 0,092 ± 0,097; estro = 0,180 ± 0,141; metaestro = 0,091 ± 0,046; e diestro = 0,233 ± 0,147 μg/mg. A concentração DS+CS apresentou-se maior no estro (ação estrogênica) e a do HS no diestro (ação progestagênica). CONCLUSÃO: Os GAGs no útero de camundongas sofrem alterações durante as fases do ciclo estral, refletindo o constante processo de renovação, sendo modulados pelos hormônios sexuais.

OBJECTIVE: Identification and quantitation of sulphated glycosaminoglycans (GAGs) in the uterus of female mice during the estrous cycle. METHODS: Four groups (n = 10 each) of virgin, 100-day old female mice were assembled according to the estrous cycle phase: proestrus, estrus, metaestrus and diestrus. Samples of the median portion of uterine horns were processed for light microscopy examination (H/E and Alcian blue + PAS). The GAGs were extracted and characterized by agarose gel electrophoresis. Data were analyzed by the unpaired Student's t-test. RESULTS: At light microscopy GAGs appear in all layers of the uterus, especially in the endometrium, between collagen fibers, in the basal membrane and around fibroblasts. Biochemical analyses disclosed presence of dermatan sulphate (DS), chondroitin sulphate (CS and heparan sulphate (HS) during all estral cycle phases. There was no clear electrophoretic separation between DS and CS, thus these two GAGs were considered together (DS+CS) (proestrus = 0.854 ± 0.192; estrus = 1.073 ± 0.254; metaestrus = 1.003 ± 0.255; diestrus = 0.632 ± 0.443 μg/mg). HS was as follows: proestrus = 0.092 ± 0.097; estrus = 0.180 ± 0.141; metaestrus = 0.091 ± 0.046; diestrus = 0.233 ± 0.147 μg/mg. The uterine content of DS+CS peaked at estrus (estrogenic action) and that of HS at diestrus (progestagen action). CONCLUSION: Due to a constant turnover process, there are definite alterations in the uterine profile of GAGs content during the estrous cycle in mice, which may be modulated by female sex hormones.

Effects of sulfated polysaccharides on tumour biology / Efectos de los polisacáridos sulfatados en la biología tumoral

Sulfated polysaccharides can act not only as anticoagulants but also as tumour inhibitors. Recent studies suggest that sulfated polysaccharides could affect tumour cells directly. Sulfated polysaccharides could inhibit the metastasis and proliferation of tumour cells by binding to growth factors and cell adhesion molecules. Moreover, sulfated polysaccharides could inhibit heparanase, which cleaves heparan sulfate chains of heparan sulfate proteoglycans and cause release of growth factors sequestered by heparan sulfate chains. Some sulfated polysaccharides can induce apoptosis and differentiation of tumour cells, but the mechanism is uncertain. In addition, sulfated polysaccharides can enhance the innate and adaptive immune response for tumour cells. Thus, the anti-tumour mechanism of sulfated polysaccharides can be explained, at least partly, through the effects on tumour biology directly.

Los polisacáridos sulfatados podrían actuar no solamente como anticoagulantes, sino también como inhibidores del tumor. Estudios recientes sugieren que los polisacáridos sulfatados podrían afectar directamente las células tumorales. Los polisacáridos tumorales podrían inhibir la metástasis y la proliferación de las células tumorales por medio de la unión con los factores de crecimiento y las moléculas de adhesión celular. Además, los polisacáridos sulfatados podrían inhibir la heparanasa, que rompe las cadenas de heparán-sulfato del proteoglicano de heparán-sulfato, dando lugar a la liberación de los factores de crecimiento secuestrados por las cadenas de heparán-sulfato. Algunos polisacáridos sulfatados podrían inducir la apoptosis y diferenciación de las células tumorales, pero el mecanismo es incierto. Además, los polisacáridos sulfatados podrían mejorar la respuesta inmunológica innata y adaptativa frente a las células tumorales. De este modo, el mecanismo antitumoral de los polisacáridos sulfatados pudiera explicarse – al menos parcialmente – a partir de los efectos sobre la biología tumoral directamente.

Heparinóides do crustáceo Goniopsis cruentata: estrutura, atividades farmacológicas e interação com células endoteliais / Heparinoids the crab Goniopsis cruentas: structura, pharmacological activities and interation with endothelial cells

Heparinóides A e B foram isolados dos tecidos do caranguejo Goniopsis cruentata após proteólise, cromatografia de troca iônica, fracionamento com acetona caracterizados por eletroforeses (géis de agarose e pofiacrilamida), análises uimicas, enzimáticas e de Ressonância Nuclear Magnética. Essas análises 10straram que os heparinóides A e B (12,8-13,6 kDa) apresentam pequenas iferenças estruturais o que sugere que eles tenham estruturas primárias emelhantes. Análises enzimáticas e de RNM revelaram que os heparinóides do aranguejo (Hepn) estão enriquecidos em dissacarídeos dissulfatados contendo lucosaminas N-sulfatadas ligadas a unidades de ácido D-glucurônico 2-0ulfatado. Além disso, Hepn mostraram baixa atividade anticoagulante in vitro pelos létodos de aPTI, PT, HEPTES TI. Por outro lado, apresentam alta atividade ntitrombótica in vivo e menor efeito antihemostático quando comparados com a eparina de mamíferos. O efeito dos Hepn nas células endoteliais de aorta de coelho (RAEC) foi também investigado. Hepn estimularam a síntese de heparam ulfato antitrombótico de forma semelhante a heparina de mamíferos (Hep). Ensaios e competição entre Hepn e Hep biotinilada mostraram que hepn competem arcialmente pelos mesmos sítios de ligação da heparina de mamíferos em RAEC. epn foram biotinilados e a ligação às RAEC foi analisada usando técnicas ítoquímicas e microscopia confocaf. A ligação dos heparinóides A e 8 biotinilados s RAEC mostrou ser dose dependente com Ko de 87,5 nM e 119,4 nM para ligação s células e ligação de maior afinidade à matriz extracelular com Ko de 28 nM e 7,5 nM, respectivamente. Por outro lado, esses compostos foram detectados apenas na matriz extracelular (MEC) das células endoteliais e colocalizaram )m a fibronectina. Esses resultados mostram que embora os heparinóides presentem diferenças estruturais quando comparados com heparinas de lamiferos, mantiveram atividade antitrombótica e capacidade de interagir com a IEC das células endoteliais. Além disso, uma vez que hepn podem deslocar a Jação de Mhep, pode-se concluir que Hepn e Hep interagem com os mesmos sítios ligação na MEC de RAEC quando estimulam a síntese de heparam sulfato antitrombótico. Essa é a primeira descrição de um composto rico em resíduos de ácido glucurônico 2-O-sulfato que é potente antitrombótico embora com baixa atividade anticoagulante in vitro.

Mucopolisacaridosis / Mucopolysaccharidosis

Dentro de las innumerables enfermedades consideradas como errores innatos del metabolismo, se encuentran las mucopolisacaridosis (MPS), pertenecientes al grupo de las enfermedades de almacenamiento o depósito lisosomal, que siendo muy variadas obedecen a deficiencias enzimáticas específicas y se generan por alteración en la función de las enzimas encargadas de degradar los glucosaminoglicanos (GAGs) Dermatán Sulfato, Queratán Sulfato, Heparán Sulfato, Condroitín Sulfato y Acido Hialurónico. Estas moléculas son polisacáridos que hacen parte estructural del tejido conectivo o bien de algunos fluidos corporales como el líquido sinovial y el humor vítreo. La alteración en la actividad de las enzimas encargadas de su catabolismo tiene consecuencias graves para el organismo humano. Hacemos a continuación una revisión del estado del arte en cada uno de los tipos de MPS, haciendo énfasis en aspectos como son la estructura y función de los GAGs en nuestro organismo, la genética involucrada en la génesis de estas patologías, su enfoque diagnóstico y su posible tratamiento.

The mucopolysaccharidoses (MPS) are a group of disorders caused by deficience of lysosomal enzymes needed for the stepwise degradation of glycosaminoglycans (mucopolysaccharides). Depending on the enzyme deficiency, the catabolism of dermatan sulfate, heparan sulfate, keratan sulfate and chondroitin sulfate may be blocked, singly or in combination. Undegraded glycosaminoglycan molecules are stored in lysosomes. Their accumulation eventually results in cell, tissue, or organ dysfunction. To date, 10 enzymes deficiences that give rise to MPS have been identificated. This review gives some information over this molecules, their function, clinical features, diagnostic and therapeutic posibilities.

Sulfated proteoglycans as modulators of neuronal migration and axonal decussation in the developing midbrain

Proteoglycans are abundant in the developing brain and there is much circumstantial evidence for their roles in directional neuronal movements such as cell body migration and axonal growth. We have developed an in vitro model of astrocyte cultures of the lateral and medial sectors of the embryonic mouse midbrain, that differ in their ability to support neuritic growth of young midbrain neurons, and we have searched for the role of interactive proteins and proteoglycans in this model. Neurite production in co-cultures reveals that, irrespective of the previous location of neurons in the midbrain, medial astrocytes exert an inhibitory or nonpermissive effect on neuritic growth that is correlated to a higher content of both heparan and chondroitin sulfates (HS and CS). Treatment of astrocytes with chondroitinase ABC revealed a growth-promoting effect of CS on lateral glia but treatment with exogenous CS-4 indicated a U-shaped dose-response curve for CS. In contrast, the growth-inhibitory action of medial astrocytes was reversed by exogenous CS-4. Treatment of astrocytes with heparitinase indicated that the growth-inhibitory action of medial astrocytes may depend heavily on HS by an as yet unknown mechanism. The results are discussed in terms of available knowledge on the binding of HS proteoglycans to interactive proteins, with emphasis on the importance of unraveling the physiological functions of glial glycoconjugates for a better understanding of neuron-glial interactions

A avaliaçäo da adesäo, migraçäo e proliferaçäo da linhagem celular CHO=K1 frente à mutante CHO-745 deficiente na biossíntese de glicosaminoglicanos sulfatados: amostra que a integrina alfa-5beta-1 é um proteoglicano envolvido na migraçäo celular / The role of cell surface glycosaminoglycans in cell division: adhesion and proliferation of CHO cells and CHO-K1 cells and a mutant CHO-745 which is deficient in syntesis of proteoglycans due to lack of activity of xylosyl transferase

Na presente tese examinamos o papel dos proteoglicanos e glicosaminoglicanos na divisao celular: adesao e proliferaçao de células em cultura CHO ("chinese hamster ovary") e sua mutante CHO-745 que é deficiente na síntese de glicosaminoglicanos devido a falta da enzima xilosiltransferase. Usando diferentes quantidades de células (selvagem e mutante) pouca adesao pode ser observada na presença de laminina e colágeno tipo I. Entretanto quando fibronectina e vitronectina foram usadas como substratos houve um aumento de adesao dos dois tipos celulares. Sómente a CHO selvagem mostrou adesao em funçao do tempo sobre colágeno tipo IV. Estes resultados sugerem que as duas linhagens celulares possuem diferentes propriedades de adesao. Ensaios de adesao usando colágeno tipo IV com células CHO cultivadas na presença de xilosídeo ou clorato, mostraram reduçao nos níveis de adesao, confirmando a importancia dos glicosaminoglicanos neste fenômeno. Várias evidências experimentais sugerem que os proteoglicanos de hepara sulfato estao envolvidos na adesao celular como moduladores positivos da proliferaça celular e como composto chave no processo de divisao celular. Ensaios de proliferaçao e ciclo celular demonstraram que uma diminuiçao das quantidades do proteoglicano nao inibem a proliferaçao da mutante CHO-745 quando comparadas com a célula selvagem pois os dois tipos celulares entram na fase S ao redor das 8 horas. A inteiraçoes celulas-matriz estao implicadas em uma grande variedade d funçoes. Estas inteiraçoes sao principalmente mediadas por integrinas que sa receptores da superfície celular e estao aparentemente envolvidas em adesao, migraçao e diferenciaçao. Por exemplo, a asj3, integrina está envolvida na migraçao e adesao da células sobre fibronectina. Também neste estudo, usando marcaçao radioativa com sulfato e imunoprecipitaçao...(au)

Development of new heparin-like compounds and other antithrombotic drugs and their interaction with vascular endothelial cells

The anticlotting and antithrombotic activities of heparin, heparan sulfate, low molecular weight heparins, heparin and heparin-like compounds from various sources used in clinical practice or under development are briefly reviewed. Heparin isolated from shrimp mimics the pharmacological activities of low molecular weight heparins. A heparan sulfate from Artemia franciscana and a dermatan sulfate from tuna fish show a potent heparin cofactor II activity. A heparan sulfate derived from bovine pancreas has a potent antithrombotic activity in an arterial and venous thrombosis model with a negligible activity upon the serine proteases of the coagulation cascade. It is suggested that the antithrombotic activity of heparin and other antithrombotic agents is due at least in part to their action on endothelial cells stimulating the synthesis of an antithrombotic heparan sulfate.

Effect of epithelial debridement on human cornea proteoglycans

Corneal transparency is attributed to the regular spacing and diameter of collagen fibrils, and proteoglycans may play a role in fibrillogenesis and matrix assembly. Corneal scar tissue is opaque and this opacity is explained by decreased ultrastructural order that may be related to proteoglycan composition. Thus, the objectives of the present study were to characterize the proteoglycans synthesized by human corneal explants and to investigate the effect of mechanical epithelial debridement. Human corneas unsuitable for transplants were immersed in F-12 culture medium and maintained under tissue culture conditions. The proteoglycans synthesized in 24 h were labeled metabolically by the addition of 35S-sulfate to the medium. These compounds were extracted by 4 M GuHCl and identified by a combination of agarose gel electrophoresis, enzymatic degradation with protease and mucopolysaccharidases, and immunoblotting. Decorin was identified as the main dermatan sulfate proteoglycan and keratan sulfate proteoglycans were also prominent components. When the glycosaminoglycan side chains were analyzed, only keratan sulfate and dermatan sulfate were detected (~50 percent each). Nevertheless, when these compounds were 35S-labeled metabolically, the label in dermatan sulfate was greater than in keratan sulfate, suggesting a lower synthesis rate for keratan sulfate. 35S-Heparan sulfate also appeared. The removal of the epithelial layer caused a decrease in heparan sulfate labeling and induced the synthesis of dermatan sulfate by the stroma. The increased deposit of dermatan sulfate proteoglycans in the stroma suggests a functional relationship between epithelium and stroma that could be related to the corneal opacity that may appear after epithelial cell debridement

Astroglial cells derived from lateral and medial midbrain sectors differ in their synthesis and secretion of sulfated glycosaminoglycans

Astroglial cells derived from lateral and medial midbrain sectors differ in their abilities to support neuritic growth of midbrain neurons in cocultures. These different properties of the two types of cells may be related to the composition of their extracellular matrix. We have studied the synthesis and secretion of sulfated glycosaminoglycans (GAGs) by the two cell types under control conditions and ß-D-xyloside-stimulated conditions, that stimulate the ability to synthesize and release GAGs. We have confirmed that both cell types synthesize and secrete heparan sulfate and chondroitin sulfate. Only slight differences were observed between the proportions of the two GAGs produced by the two types of cells after a 24-h labeling period. However, a marked difference was observed between the GAGs produced by the astroglial cells derived from lateral and medial midbrain sectors. The medial cells, which contain derivatives of the tectal and tegmental midline radial glia, synthesized and secreted ~2.3 times more chondroitin sulfate than lateral cells. The synthesis of heparan sulfate was only slightly modified by the addition of ß-D-xyloside. Overall, these results indicate that astroglial cells derived from the two midbrain sectors have marked differences in their capacity to synthesize chondroitin sulfate. Under in vivo conditions or a long period of in vitro culture, they may produce extracellular matrix at concentrations which may differentially affect neuritic growth

Efeito da desepitelizaçäo na síntese de glicosaminoglicanos em córneas humanas mantidas em cultura / Effect of epithelial debridement on glycosaminoglycan synthesis by human corneal explants

Os objetivos do presente trabalho foram identificar os glicosaminoglicanos presentes na córnea humana, analisar a síntese dos glicosaminoglicanos em córneas humanas mantidas em cultura e avaliar o efeito da remoção do epitélio sobre a síntese destes compostos em córneas mantidas em cultura. Para tal, foram utilizadas córneas do Banco de Olhos do Hospital São Paulo que foram rejeitadas para transplantes. Os glicosaminoglicanos foram extraídos após proteólise dos tecidos com papaína e identificados por uma combinação de eletroforese em gel de agarose e degradação enzimática com mucopolisacaridases bacterianas específicas. Os principais glicosaminoglicanos extraídos da córnea humana foram o queratam sulfato e o dermatam sulfato, cada um correspondendo a cerca de 50 por cento do total. Para analisar os glicosaminoglicanos sintetizados em cultura, as córneas foram mantidas em meio Fl2, por 24 horas, a 37§C, em atmosfera contendo 2,5 por cento de gás carbônico. Ao meio de cultura foi adicionado 35S-sulfato, de forma que os glicosaminoglicanos sintetizados em 24 horas fossem, então, metabolicamente marcados. OS 35S-glicosaminoglicanos foram isolados da mesma forma descrita acima, exceto que foram vistos por radioautograma e quantificados em um contador de cintilações. O principal glicosaminoglicano sintetizado foi o dermatam sulfato (72 por cento do total), seguido pelo queratam sulfato (l7 por cento). Além desses dois glicosaminoglicanos, apareceu heparam sulfato (11 por cento), que não foi identificado como constituinte da córnea por estar em pequena quantidade (abaixo do limite de detecção do método). Esses resultados sugerem que o dermatam sulfato e o heparam sulfato apresentem uma taxa de turnover maior que a do queratam sulfato. A desepitelização corneana levou a uma alteração no padrão de síntese de glicosaminoglicanos, com uma maior marcação de dermatam sulfato e uma diminuição no heparam sulfato, em relação ao controle. Este efeito foi proporcional à área da córnea desepitelizada. Para verificar quais Os glicosaminoglicanos sintetizados em cada camada celular, o epitélio, o endotélio e o estroma foram separados e os glicosaminoglicanos sintetizados em cada uma delas foram identificados. Verificou-se que os ceratócitos foram os principais responsáveis pela síntese de dermatam sulfato e queratam sulfato, enquanto as células epiteliais produziram basicamente heparam sulfato. Esta observação explica a ...(au)

Urinary heparan sulphate is increased in normoalbuminuric diabetic patients

Forty-nine normoalbuminuric diabetic patients were studied: 22 males and 27 females, in whom urinary heparan sulphate (HS), albuminuria, creatininemia, creatininuria, creatinine clearance, HbA1c and arterial pressure (AP) were determined. Two groups were discerned: group 1, Type 1 DM, diabetic cases (n = 16); and group 2, Type 2 DM diabetic cases (n = 33). Patients were compared with 24 healthy controls: 12 men and 12 women, who showed a mean value + SD of 0.36 + 0.18 mg/24 h HS with significant differences between males and females (0.43 + 0.15 versus 0.28 + 0.17, respectively; p = 0.02). The total population of diabetic cases rendered a mean of 0.68 + 0.44 and comparison with controls proved highly significant (p < 0.001). Globally, male patients had a mean of 0.82 + 0.48 and females 0.54 + 0.35, with p < 0.02. Group 1 and 2 values of HS were not significantly different. HS levels falled to correlate either with age, body mass index (BM), time since onset of diabetes, albuminuria, creatininemia, creatininuria, creatinine clearance, HbA1c or arterial hypertension. To conclude: both normal and diabetic males eliminate a greater quantity of HS than females. Normoalbuminuric diabetic patients of both types eliminate a greater quantity of HS regardiess of arterial pressure and time since onset of diabetes.

Glicosaminoglicanas / Glicosaminoglicanas

Este trabalho tem por objetivo, através do levantamento bibliográfico, a insvestigação das glicosaminoglicanas (GAGs) da substância fundamental amorfa do tecido conjuntivo e seu relacionamento com a polpa dental. Na polpa dental normal existem 4 tipos principais de GAGs: ácido hialurônico, condrointin sulfato (os principais), dermatan sulfato e heparan sulfato, sendo que o condoitin sulfato é responsável, durante a dentinogênese, pela mineralização pois pode agir como ponto de captura de íons cálcio e parece influenciar nos processos de fluorose e anquilose. O condroitin sulfato está presente na polpa dental, gengiva, ligamento periodontal, dentina e pré-dentina, osso alveolar, cemento; no tecido mesenquimal, saco dentário e papila dental. As GAGs são macromoléculas, altamente hidrofílicas, então uma das principais funções é regular a capacidadde de reter água o que auxilia a manter o equilíbrio de sal e água na polpa dental, regula fenômenos inflamatórios, transporte de íons (nutrição celular), e desempenham importante função nas interações celulares determinando as características físicas da polpa