Biomedical subjects

N K Karamanos

Publications and source records attributed to N K Karamanos.

At least 19 recordsLinked to original sources

PubMed2006

Chondroitin sulfate A chains enhance platelet derived growth factor-mediated signalling in fibrosarcoma cells.

Platelet derived growth factor is involved in the autocrine growth stimulation of malignant cells, the stimulation of angiogenesis and the recruitment and regulation of tumor fibroblasts. PDGF has been shown to physically interact with glycosaminoglycans which are abundant in the fibrosarcoma cell microenvironment. Aim of the present study was to examine the effects of glycosaminoglycans on the mitogenic function of platelet derived growth factor in two human fibrosarcoma cell lines (B6FS, HT1080). For this purpose exogenously added glycosaminoglycans, regulators of endogenous glycosaminoglycan synthesis (sodium chlorate as selective inhibitor and beta-D-xyloside as a stimulator) and specific glycosidases to cleave cell-associated glycosaminoglycans, were utilized. Platelet derived growth factor demonstrated a growth stimulating effect on B6FS, whereas no effect was evident on HT1080 fibrosarcoma cells. Beta-D-xyloside had no effect on the basal level or the platelet derived growth factor-induced cell proliferation, whereas sodium chlorate severely reduced the basal level of proliferation in both cell lines. Significant co-stimulatory effects of chondroitin sulfate A in combination with platelet derived growth factor BB on the growth of HT1080 and B6FS cells were found. The co-stimulatory effect of chondroitin sulfate A was not due to transcriptional up regulation of platelet derived growth factor receptors genes, but rather to more efficient signalling of tyrosine kinase receptors. In conclusion, this study shows that chondroitin sulfate A can enhance the mitogenic activity of platelet-derived growth factor in fibrosarcoma cells utilizing a pathway which involves tyrosine kinases. This result introduces a new modulating role for chondroitin sulfate in signalling pathways critical for cancer growth.

Becaplermin↗

PubMed2006

Evaluation of modal damping factor as a diagnostic tool for osteoporosis and its relation with serum osteocalcin and collagen I N-telopeptide for monitoring the efficacy of alendronate in ovariectomized rats.

Osteoporosis is a metabolic bone disease characterized by reduced bone mass and deterioration of bone microarchitecture. It results from the shift of the osteoblast-osteoclast activity equilibrium in favor of the later. Although, a number of biochemical markers, such as collagen I N-telopeptide (NTx) and osteocalcin (OC), have been used for monitoring bone remodeling, a new, monitoring, non-invasive method, which is based on the measurement of the dynamic characteristic of bone and is known as modal damping factor (MDF), has not been evaluated as a diagnostic tool for osteoporosis. Bisphosphonates, such as alendronate, have an established role in the treatment of osteoporosis. The aim of the present study was, therefore, to evaluate the effects of alendronate on the levels of MDF, serum NTx and OC on osteoporosis induced by ovariectomy in rats. Furthermore, the effects of alendronate on osteoporosis have been histologically evaluated. Fifteen adult female Wistar rats were bilaterally ovariectomized and osteoporosis was histologically confirmed and by the use of peripheral quantitative computerized tomography (pQCT). MDF was applied to assess the bone structural integrity. The serum levels of NTx (37.4+/-0.5 nM bone collagen equivalents, BCE) and OC (111.0+/-8.2 ng/mL) were found to significantly increase following ovariectomy (72.0+/-2.9 nM BCE and 213.5+/-12.1 ng/mL, respectively, p<0.001). As assessed by histology and the levels of NTx and OC in sera, animals treated with alendronate presented a statistically significant deceleration in the progression of the disease in comparison to the no-therapy control group (alendronate group NTx levels: 146.3+/-8.9 nM BCE versus no-therapy control group NTx levels: 265.3+/-14.0 nM BCE, p<0.001, alendronate group OC levels: 205.6+/-18.2 ng/mL versus no-therapy group OC levels: 353.9+/-26.1 ng/mL, p<0.001). Data obtained from the vibration analysis performed illustrated that the change in damping was equal or greater to the change in total and trabecular density, respectively. Damping increased with decreasing bone density, as expected, given that damping accounts for the structural integrity of bone (MDF value before ovariectomy: 0.058+/-0.003 versus MDF value after ovariectomy: 0.098+/-0.003, p<0.001). The higher damping values correspond to more deteriorated structures. In particular, both total and trabecular density were significantly decreased following ovariectomy (total density before ovariectomy: 702.4+/-19.0 versus total density after ovariectomy: 542.2+/-12.8, p<0.001, trabecular density before ovariectomy: 445.3+/-13.0 versus trabecular density after ovariectomy: 396.7+/-8.4, p<0.05). MDF value of the alendronate group (0.07+/-0.002) was significantly lower (p<0.001) as compared to MDF value after ovariectomy (0.098+/-0.003) and that of the no-therapy group (0.1+/-0.004, p<0.001). The administration of alendronate seemed to have no effect on either total or trabecular density, since both parameters continued to decrease (alendronate group total density: 549.4+/-12.3, alendronate group trabecular density: 368.4+/-14.7). However, when this was compared to the no-therapy group, a statistically significant difference of total density at the 0.05 level was observed (no-therapy total density: 464.8+/-9.1). The results of this study suggest that combined measurements of MDF, NTx and OC may be a potential diagnostic tool for osteoporosis and monitoring bone integrity during treatment with bisphosphonates. Furthermore, administration of alendronate showed to offer a critical deceleration in the progression of osteoporosis.

Alendronate↗

PubMed2006

Determination and distribution of N-acetyl- and N-glycolylneuraminic acids in culture media and cell-associated glycoconjugates from human malignant mesothelioma and adenocarcinoma cells.

Sialic acids containing glycoconjugates are very common in human neoplasias and their expression frequently correlates with malignant phenotype and the tumor grade. The majority of tumor markers containing sialic acids in man involve changes in the amount of total sialic acids and in the presence of the two main sialic acid types, Neu5Ac and Neu5Gc, and their derivatives. The aim of the present study was to examine whether malignant mesothelioma cell lines synthesize sialic acid containing glycoconjugates at both the extracellular and cell membrane levels and particularly whether the type and the content of Neu5Ac and Neu5Gc are of biological importance for mesothelioma cell differentiation and evaluation of its prognosis. The study was performed in three human malignant mesothelioma cell lines, two with a fibroblast like phenotype (STAV-FCS and Vester) and one of epithelial differentiation (STAV-AB), which developed from the pleural effusions of patients with malignant mesothelioma and in one human adenocarcinoma cell line (Wart). Neu5Ac and Neu5Gc were determined following a mild hydrolysis step and a sample clean-up procedure. The determination was performed by reversed-phase HPLC after the NeuAc and NeuGc had been converted to per-O-benzoylated derivatives. It was found that Neu5Gc is the major sialic acid in the culture media of all cell lines examined. Molar ratios of Neu5Ac to Neu5Gc showed that Neu5Gc is the predominant sialic acid in the culture medium of the fibroblast-like mesothelioma cells. Neu5Ac is almost undetectable in the cell membrane, whereas Neu5Gc is present in considerable amounts. The obtained results suggest that the type and the content of Neu5Ac and Neu5Gc in culture media are of biological importance for mesothelioma cell differentiation and may be of value in the evaluation of prognosis.

Adenocarcinoma↗

PubMed2006

Identification of the commonest cystic fibrosis transmembrane regulator gene DeltaF508 mutation: evaluation of PCR--single-strand conformational polymorphism and polyacrylamide gel electrophoresis.

In the present study we investigated whether single-strand conformational polymorphism (SSCP) and polyacrylamide gel electrophoresis (PAGE) could be used for the identification of the CFTR DeltaF508 gene mutation, which is commonest in the Greek population. Using DNA from patients carrying this mutation, the appropriate 98 bp region of the CFTR gene was amplified by PCR and the reaction products were analysed by non-radioactive SSCP-electrophoresis using silver staining for band visualization and non-denaturating PAGE to confirm the results. SSCP electrophoretic analysis has been optimized for several parameters in order to achieve the best resolution. Single-strand DNA fragments gave a reproducible pattern of bands, characteristic for the particular mutation. Comparison of the obtain patterns with control samples allowed the detection of the DeltaF508 mutation in the patients studied by SSCP assay and these results were confirmed by the independent method of PAGE. Although SSCP and PAGE can be used for detection of this mutation, PAGE resulted in more distinct patterns than SSCP. It is, therefore, proposed that PAGE can be reliably used for the detection and identification of such a mutation in patients provided that suitable controls are available. The applicability of PAGE to identification of the mutation in carriers, particularly useful for population screening, is also discussed.

Cystic Fibrosis↗

PubMed2006

Immunization with specific polysaccharide antigen reduces alterations in corneal proteoglycans during experimental slime-producing Staphylococcus epidermidis keratitis.

PURPOSE: Staphylococcus epidermidis is a leading cause of bacterial keratitis associated with corneal damage. Corneal integrity is closely associated with matrix macromolecules, such as proteoglycans (PGs) and collagen. The aim of this study was to examine whether active immunization (AI) using a major immunogenic polysaccharide determinant of slime (20-kDa PS) as antigen, and passive immunization (PI) after administration of specific antibodies toward 20-kDa PS affect the distribution of PGs as well as corneal lesions in an experimental model of slime-producing S. epidermidis keratitis. METHODS: For AI, seven rabbits were immunized with 20-kDa PS, whereas for PI, seven rabbits received specific antibodies against 20-kDa PS. Lesions were graded clinically for a 21-day period. Levels of 20-kDa PS antibodies in serum and aqueous humor in both immunization groups were determined by ELISA. The distribution of certain extracellular matrix PGs during corneal healing was analyzed immunohistochemically. RESULTS: Levels of specific anti-20-kDa PS antibodies in serum and aqueous humor obtained after either AI or PI were significantly higher as compared with those in the respective nonimmunized control groups (p<0.001). Clinical grading showed that both AI and PI rabbits had a significantly less corneal damage as compared with infected nontreated rabbits. Immunohistochemical analyses for PGs exhibited significant differences to the wounded regions as compared with noninfected corneal tissue. Accumulation of keratan sulfate PGs and decorin was observed in the corneal stroma of infected rabbits and of heparan sulfate PGs around the new-formed vessels. This phenomenon was significantly reduced in immunized animals in accordance with macroscopically decreased corneal damage observed in these animals. CONCLUSIONS: Results of this study suggest a key role of 20-kDa PS and its antibodies as prophylactic and therapeutic agents in keratitis caused by slime-producing S. epidermidis.

Animals↗

PubMed2006

Transforming growth factor-beta as a key molecule triggering the expression of versican isoforms v0 and v1, hyaluronan synthase-2 and synthesis of hyaluronan in malignant osteosarcoma cells.

Versican, a large sized chondroitin-sulphate proteoglycan (PG), and its binding partner, hyaluronan (HA), are extracellular matrix (ECM) components that play an essential role in transformed cell behavior. Expression of certain versican isoforms has been implicated in cell migration and proliferation of cancer cells and, on the other hand, disruption of HA synthesis by inhibiting hyaluronan synthase-2 (HAS2) expression in osteosarcoma cells by suppressing cell proliferation, invasiveness and motility. Considering that growth factors, such as TGF-beta, bFGF and PDGF-BB, are important regulators for the expression of the ECM macromolecules, in this study we examined the effect of these growth factors on the expression of the various versican isoforms, HA synthases as well as HA synthesis by MG-63 osteosarcoma cells and normal human osteoblastic periodontal ligament cells (hPDL). Real-time PCR and metabolic labelling followed by fine HPLC analysis coupled to radiochemical detection were the methods utilized. It was found that, contrary to normal hPDL cells, osteosarcoma MG-63 cells do not constitutively express the versican isoforms V0 and V1. Exogenous addition of TGF-beta2 stimulated the versican transcript levels mainly by forcing osteosarcoma cells to express V1 and V0 isoforms. PDGF-BB and bFGF had only minor effects in these cells. In hPDL cells a strong stimulation of the V3 transcript by all growth factors was observed. TGF-beta2 was also the major stimulator of HAS2 isoform expression as well as hyaluronan synthesis in osteosarcoma cells, while PDGF-BB exerted dominant influence on HAS2 isoform expression and hyaluronan biosynthesis by osteoblasts. The obtained results show for the first time that TGF-beta2 triggers the malignant phenotype pattern of versican and hyaluronan expression in human osteosarcoma cells and indicate that this growth factor may account for the metastatic potential of these cells.

Becaplermin↗

PubMed2006

Lumican, a small leucine-rich proteoglycan substituted with keratan sulfate chains is expressed and secreted by human melanoma cells and not normal melanocytes.

Melanoma is a frequent and therapy-resistant human disease. Malignant melanocytes modulate their microenvironment in order to penetrate the dermal/epidermal junction and eventually invade the dermis. The small leucine-rich proteoglycans (SLRPs) constitute important constituents of the dermis extracellular matrix (ECM), participating in both the structural and the functional organization of the skin. The role of a keratan sulphate SLRP lumican, has recently been investigated in the growth and metastasis of several cancers. In this study, the expression of lumican was studied in two human melanoma cell lines (WM9, M5) as well as in normal neonatal human melanocytes (HEMN) using real time PCR, western blotting with antibodies against the protein core and keratan sulfate, and treatments with specific enzymes. Both human metastatic melanoma cell lines were found to express lumican mRNA and effectively secrete lumican in a proteoglycan form, characterized to be substituted mostly with keratan sulfate chains. Lumican mRNA was not detected in normal melanocytes. This is the first time that the synthesis and secretion of lumican in human melanoma cell lines is reported. The role of this proteoglycan in the development and progression of malignant melanoma has to be further investigated.

Cell Line, Tumor↗

PubMed2006

Versican but not decorin accumulation is related to metastatic potential and neovascularization in testicular germ cell tumours.

AIMS: To investigate the expression of versican and decorin in patients with testicular germ cell tumours (GCTs) and to correlate this with the clinicopathological findings. Matrix proteoglycans versican and decorin are frequently overexpressed in various malignancies and are involved in the progression of cancer. METHODS AND RESULTS: Overexpression of versican and decorin was detected in GCTs by immunoblotting. Immunohistochemical staining for proteoglycans was performed on 71 cases of paraffin-embedded tissues. In most of the cases increased decorin and versican stromal staining was demonstrated. In both seminomas and non-seminomatous germ cell tumours (NSGCTs) strong staining of decorin was not found to be related to any of the clinicopathological variables. Accumulation of versican was found to be associated with vascular and lymphatic invasion, nodal metastasis and disease stage in seminomas and NSGCTs and, in addition, with tumour size and distant metastasis only in NSGCTs. Additionally, only the deposition of versican was linearly correlated with the number of microvessels in the tumour stroma in GCTs. CONCLUSIONS: Ectopic versican and decorin expression is a frequent feature in GCTs. Versican but not decorin accumulation in GCTs is related to metastatic potential and neovascularization and might be a useful marker for testicular malignancy.

Adolescent↗

PubMed2006

Effects of the natural isoflavonoid genistein on growth, signaling pathways and gene expression of matrix macromolecules by breast cancer cells.

Genistein is a well known protein tyrosine kinase inhibitor. It is structurally similar to 17beta-estradiol and exerts antiestrogenic effects. It also affects the signal transduction components Akt, FAK, ErbB-2 and Bcl-2. Key enzymes implicated in cancer invasion are also affected by genistein. A critical evaluation of the effects of genistein on breast cancer growth, signaling and gene expression is presented in this review.

Antineoplastic Agents↗

PubMed2005

Signal transduction by IL-2 and its receptors as target in treatment of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic and destructive arthropathy with systemic features, the etiopathogenesis of which remains unclear. It is characterized by relapsing and remitting inflammation and hyperplasia of synovial cells. Proinflammatory cytokines, such as interleukin-2 (IL-2), play an important role in maintaining cartilage damage and severe destruction of the joints due to an uncontrolled activation of cellular immunity. An imbalance between proinflammatory and anti-inflammatory mediators is likely to contribute to the chronicity of the disease. Therefore, insight into the activation state of T-cells in different stages of the disease may be important to understand pathogenetic mechanisms underlying RA and could be a lead for the design of future therapeutic strategies. Because of the central role of the IL-2/IL-2 receptor (IL-2R) system in mediation of the immune system, monitoring and manipulation of this system has important diagnostic and therapeutic implications. New approaches in RA therapy with anticytokine agents, which block cytokines and their receptors, are now used as antirheumatic drugs in clinical practice.

Animals↗

PubMed2005

Genistein suppresses the invasive potential of human breast cancer cells through transcriptional regulation of metalloproteinases and their tissue inhibitors.

Progression of breast cancer implicates the degradation of extracellular matrix (ECM) by metallo-proteinases (MMPs), a process with important consequences on the growth and invasiveness of cancer cells in adjacent and distant sites. The isoflavone, genistein--a natural inhibitor of protein tyrosine kinase pathway--inhibits the growth of a wide range of cancer cells in vitro. The aim of this study was to investigate: i) the expression of mRNAs encoded for MMPs and their endogenous inhibitors (TIMPs) associated with pathogenesis and metastatic potential of breast cancer cells; and ii) the effect of genistein on the transcription of MMPs and TIMPs and the invasive potential of breast cancer cells. Gene expression at transcriptional level was examined in cell cultures of two epithelial breast cancer cell lines, the high invasive (ER-negative) MDA-MB-231 and the low invasive (ER-positive) MCF-7, as well as the normal mammary cells (MCF-12A) following RNA isolation and reversed transcriptase polymerase chain reaction (RT-PCR). The inhibitory effect of genistein on functional invasiveness was examined by a cell invasion assay. Cell cycle distribution showed that genistein arrested breast cancer MDA-MB-231, MCF-7 and BT-20 cells in the G2/M phase. Both normal and breast cancer cell lines express the genes of MMP-2, -9, MT1-, MT2-, MT3-MMP and TIMP-1, -2 and -3. MCF-7 express notably less MMPs than MDA-MB-231 cell line. The addition of genistein resulted in down-regulation of the transcription of all MMP genes in MDA-MB-231 and most of MMPs in MCF-7 cells. The inhibitory effect of genistein on MMPs was functionally confirmed, since it significantly reduced the invasion properties of cancer cells in vitro. The obtained results indicate that genistein may be of great value in prevention of breast cancer cell metastasis, since it represents both a transcriptional modulator of genes involved in this pathogenetic process and a suppressor of breast cancer cell invasiveness.

Antineoplastic Agents↗

PubMed2004

Potential use of solid phase immunoassays in the diagnosis of coagulase-negative staphylococcal infections.

Staphylococcus epidermidis is a major nosocomial pathogen, even though it is a member of the normal bacterial flora of skin and the mucous membranes. A major complication is the development of biofilms on implanted medical devices. Diagnosis of coagulase-negative staphylococcal infections relies on the presence of clinical manifestation of infections and on microbiologic evidence, usually obtained after the removal of the biomaterial. Solid-phase immunoassays have not yet been used for routine diagnosis of coagulase-negative staphylococcal infections and distinction between pathogenic and normal cocci. The enzyme immunoassays developed in the last decade are presented in this review article. Serodiagnosis has been attempted by determining antibodies against bacterial cells, mixtures of S. epidermidis slime antigens and discrete slime antigens. Detection or typing of staphylococcal cells has been performed by specific antibodies and lectins. There is still a long way until the application of such assays in the routine clinical laboratory and large clinical studies are necessary.

Antibodies, Bacterial↗

PubMed2004

Determination and biological relevance of serum cross-linked type I collagen N-telopeptide and bone-specific alkaline phosphatase in breast metastatic cancer.

Bone metastasis is a frequent complication of cancer disease. The metastatic spread of cancer to bone is common to many different malignancies, particularly breast (ca. 73%), prostate (ca. 68%) and lung (ca. 36%) cancers. Metastases to bone cause increased bone resorption both from direct effects of the tumor itself and thought osteoclastic activation. The diagnosis and follow-up of bone metastatic cancer patients usually relies on skeletal X-ray and bone scintigraphy. However, the development of biochemical markers, used as indicators of bone metabolism, provides data useful in the clinical practice. The most important markers for bone remodeling process, bone formation and resorption, are bone-specific alkaline phosphatase (BAP) and N-telopeptide of type I collagen (NTx), respectively. In this report, we applied two solid-phase immunoassays used for the determination of BAP and NTx in serum of breast cancer (BC) post-menopausal women with bone metastasis and healthy individuals. BAP level in patients was found to be 45.72 +/- 12.92 U/l, while the normal range for healthy individuals was 14.2 - 42.7 U/l. The respective level of serum NTx was 19.20 +/- 8.87 nM bone collagen equivalents (BCE) for patients and 15.9 +/- 3.8 nM BCE for healthy women. Correlation of the obtained data showed elevated levels for both markers indicating high rate of bone degradation in breast metastatic cancer.

Alkaline Phosphatase↗

PubMed2004

Clara cell secretory protein: determination of serum levels by an enzyme immunoassay and its importance as an indicator of bronchial asthma in children.

Clara cell secretory protein (CC16) is a 16kDa protein secreted by Clara cells in the lining fluid of bronchiolar and bronchial epithelium. CC16 presents several biologic properties, and has been shown to have immunomodulatory and anti-inflammatory activity. It may play a role in controlling inflammation in the airway. There is some evidence that the CC16 level is primarily lower in adult individuals with bronchial asthma, thus contributing to its pathophysiology. This study was designed to examine CC16 serum levels of children, healthy and with asthma. An enzyme solid phase immunoassay utilizing monoclonal antibody to CC16 was the analytical method to determine the protein concentration in blood sera. The method showed excellent linearity, high sensitivity (detection limit: <50 ng/l) and precision. It was found that asthmatic children appear significantly lower levels (P < 0.001) of CC16 in serum as compared to healthy ones. It is, therefore, concluded that CC16 may be a useful diagnostic index of bronchial asthma in the early child-age.

Antibodies, Monoclonal↗

PubMed2004

The presence of a novel extracellular hyaluronidase in squid cranial cartilage.

A new type of hyaluronidase was isolated from squid cranial cartilage. The enzyme seems to be localised extracellularly, since it is extracted from the tissue by 0.5 M sodium acetate, pH 7.0, in the presence of proteinase inhibitors. Degradation studies suggest that the enzyme belongs to the family of endoglycosidases generating oligosaccharides of rather large size. The best activity of the enzyme was observed at pH 7.0 and 37 degrees C and the optimum buffer for digestion was 0.15 M Tris acetate. It is inactive in sodium phosphate, morpholine acetate and HEPES buffers. The enzyme degrades aggrecan, hyaluronan, chondroitin sulphate and oversulphated chondroitin sulphate.

Animals↗

PubMed2003

Capillary electrophoresis and enzyme solid phase assay for examining the purity of a synthetic heparin proteoglycan-like conjugate and identifying binding to basic fibroblast growth factor.

Interaction of basic fibroblast growth factor (bFGF) with heparin/heparan sulfate proteoglycans protects the growth factor against proteolytic degradation and is essential for its cellular activity. Although the structural requirements of heparin and heparan sulfate for the high-affinity binding to bFGF have been extensively examined, studies on intact heparin proteoglycans are limited. In this report, the purity and the binding ability of a heparin proteoglycan-like molecule-the heparin-bovine serum albumin (heparin-BSA) conjugate-was examined using capillary zone electrophoresis (CZE). Furthermore, the affinity of bFGF binding to the heparin-BSA conjugate was studied using an enzyme solid-phase assay. Chondroitin sulfate, dermatan sulfate, hyaluronan, heparan sulfate and variously sulfated disaccharides derived from heparin and heparan sulfate were also studied for their ability to compete with the binding of bFGF to heparin. Heparin-BSA conjugate was synthesized by reductive amination and, following precipitation with 1.5 vols of ethanol-sodium acetate, it was obtained free of contaminating heparin. Heparin-BSA-bFGF conjugate was obtained following incubation of heparin-BSA with bFGF for 2 h at 37 degrees C. Intact heparin, heparin-BSA and heparin-BSA-bFGF conjugates were completely resolved by CZE using 50 mM phosphate, pH 3.5, as operating buffer, reversed polarity (30 kV) and detection at 232 nm. Competitive solid phase assay showed that, among the glycosaminoglycans tested, heparin exhibits the highest affinity binding to bFGF (IC(50) = 6.4 nM). Heparan sulfate showed a lower affinity as compared with that of heparin, whereas all other glycosaminoglycans and heparin/heparan sulfate-derived disaccharides tested showed minute effects. The developed CZE method is rapid and accurate and can be easily used to identify bFGF-interacting heparin preparations of biopharmaceutical importance.

Electrophoresis, Capillary↗

PubMed2003

Versican undergoes specific alterations in the fine molecular structure and organization in human aneurysmal abdominal aortas.

Versican is the major matrix proteoglycan in aortic wall and participates in various biological functions of the tissue. In the present study the molecular characteristics of versican isolated from normal human aorta as well as those of versican expressed in aneurysmal aortic tissue were examined. Versican was isolated by combined anion-exchange and gel permeation chromatography and was further characterized by high-performance liquid chromatography, polyacrylamide gel electrophoresis and immunoblotting. In both tissues versican is exclusively substituted with chondroitin sulfate chains, in contrast to other human tissues where both chondroitin and dermatan sulfate chains are attached onto versican core proteins. Except for the significant decrease in the concentration of versican in the aneurysmal tissue, this PG undergoes specific alterations in the aneurysmal tissue. The molecular size of versican isolated from diseased tissue is decreased with a simultaneous increase in the ratio of glycosaminoglycan to protein in this tissue. The latter reflect the extensive fragmentation of versican in the diseased tissue and most probably the generation of shorter peptides enriched to glycosaminoglycan chains. Although the size of chondroitin sulfate chains is identical in both versican preparations, a significant increase in the percentage of 6-sulfated disaccharides is observed in chondroitin sulfate chains of versican in aneurysmal aortas, which is accompanied by decrease in 4-sulfated and non-sulfated units.

Aged↗

PubMed2002

Advances in analysis of glycosaminoglycans: its application for the assessment of physiological and pathological states of connective tissues.

Glycosaminoglycans are a class of biological macromolecules found mainly in connective tissues as constituents of proteoglycans, covalently linked to their core protein. Hyaluronan is the only glycosaminoglycan present under its single form and possesses the ability to aggregate with the class of proteoglycans termed hyalectans. Proteoglycans are localised both at the extracellular and cellular (cell-surface and intracellular) levels and, via either their glycosaminoglycan chains or their core proteins participate in and regulate several cellular events and (patho)physiological processes. Advances in analytical separational techniques, including high-performance liquid chromatography, capillary electrophoresis and fluorophore assisted carbohydrate electrophoresis, make possible to examine alterations of glycosaminoglycans with respect to their amounts and fine structural features in various pathological conditions, thus becoming applicable for diagnosis. In this review we present the chromatographic and electromigration procedures developed to analyse and characterise glycosaminoglycans. Moreover, a critical evaluation of the biological relevance of the results obtained by the developed methodology is discussed.

Chromatography, Liquid↗