Biomedical subjects

H Kresse

Publications and source records attributed to H Kresse.

At least 19 recordsLinked to original sources

PubMed1998

Decorin core protein fragment Leu155-Val260 interacts with TGF-beta but does not compete for decorin binding to type I collagen.

It has been shown that small proteoglycans containing leucine-rich repeats in their core proteins can form complexes with TGF-beta. Decorin, a ubiquitously found molecule of the extracellular matrix, is the best-studied example. Therefore, binding domains on its core protein were investigated using recombinant decorin fragments generated as fusion proteins in prokaryotes. The peptide Leu155-Val260 immobilized by the polyhistidine tag on a nickel chelate column bound TGF-beta1 and -beta2 almost as effectively as the largest fragment (Asp45-Lys359) studied. Other peptides were less effective. For the two peptides Asp45-Lys359 and Leu155-Val260 dissociation constants in the nanomolar range for high-affinity binding sites were calculated in a solid-phase assay with immobilized TGF-beta2. Peptide Asp45-Lys359 also contained a lower affinity binding site. Domains with lower affinity were also found in peptides Asp45-Leu155 and Arg63-Gly190. Peptide Leu155-Val260 also formed complexes with TGF-beta in the liquid phase as determined by equilibrium gel filtration. Furthermore, F(ab') fragments of polyclonal antibodies against peptide Leu155-Val260 interfered with TGF-beta binding to peptide Asp45-Lys359 in a dose-dependent manner. Peptide Leu155-Val260, however, is only a weak competitor of the binding of wild-type decorin to reconstituted type I collagen fibrils. Therefore, independent binding sites of decorin for TGF-beta and type I collagen should exist. In support of this hypothesis saturable binding of TGF-beta1 and TGF-beta2 to collagen-bound native decorin could be demonstrated. The bound cytokine could be released in a biologically active form by collagenase treatment. Thus, decorin may play a biological role in storing this cytokine temporarily in the extracellular matrix and in thereby modulating an interaction of TGF-beta with its signaling receptors.

Binding Sites↗

PubMed1998

Developmental regulation of decorin expression in postnatal rat brain.

Here, we report on the expression of the small chondroitin/dermatan sulfate proteoglycan decorin in the developing postnatal rat brain. Northern analysis of brain RNA demonstrated decorin transcripts with peak expression on postnatal day 3 followed by a slow decline to the lower adult level. In situ hybridization and immunohistochemistry revealed postnatal decorin expression in the grey matter of neocortex, hippocampus and thalamus, in myelinated fibre tracts and in several mesenchymal tissues (blood vessels, pia mater and the choroid plexus). In the neocortex, decorin is expressed in a specific laminar pattern with intense staining of the cortical plate and its derivatives, which differs remarkably from the distributions observed for other proteoglycans [B. Miller, A.M. Sheppard, A.R. Bicknese, A.L. Pearlman, Chondroitin sulfate proteoglycans in the developing cerebral cortex: the distribution of neurocan distinguishes forming afferent and efferent axonal pathways, J. Comp. Neurol. 355 (1995) 615-28]. Thus, decorin seems to serve yet unknown functions in the developing rat brain parenchyma in addition to its well-established role as a constituent of the mesenchymal extracellular matrix.

Animals↗

PubMed1998

Receptor-mediated endocytosis of decorin: involvement of leucine-rich repeat structures.

Decorin, a small dermatan sulfate proteoglycan, is characterized by a core protein with central leucine-rich repeat structures and a single glycosaminoglycan chain. It is catabolized by receptor-mediated uptake and subsequent intralysosomal degradation. In the present study, the localization of the receptor binding site(s) along the core protein was investigated. Various recombinant decorin fragments were consistently able to inhibit the endocytosis of wild-type decorin. The most potent inhibitory peptides were those which encompassed the central Leu125-Val230 region, i.e., the fifth to eighth leucine-rich repeat, or at least a part of it. The peptide Leu125-Val230 bound directly to the 51-kDa endocytosis receptor, and Fab fragments of antibodies against this peptide inhibited the endocytosis of decorin in a dose-dependent manner. Decorin constructs expressed in human 293 cells and comprising the full-length coding region or lacking sequences N- and/or C-terminally of the Leu125-Val230 region were all endocytosed with similar clearance rates. These data suggest that the N- and C-terminal domains of the core protein are not required for endocytosis. The receptor binding site is rather represented by contiguous leucine-rich repeat structures of the central part of the core protein. This conclusion is supported by competition experiments with biglycan, a structurally related small proteoglycan.

Binding, Competitive↗

PubMed1998

Decreased activity of acid alpha-glucosidase in a patient with persistent periocular swelling after infusions of hydroxyethyl starch.

After infusion therapy with hydroxyethyl starch (HES) on account of sudden hearing loss, a 68-year-old woman developed a marked and persistent periocular swelling. This extraordinary adverse effect caused us to search for an explanation by means of histopathology, immunohistochemistry, immunoelectron microscopy and biochemistry. In lesional periocular skin and in normal-appearing skin, lysosomal storage of HES could be detected with a specific HES antibody in histiocytes, endothelial cells, basal keratinocytes and small nerves. In the periocular skin, a stronger deposition of HES was found in addition to distinct xanthomatous changes as well as features of lymphoedema. In view of lysosomal HES storage we measured the pH-dependent activity of the lysosomal alpha-glucosidase (GAA) in cultured fibroblasts. We found a 50% decreased activity of the acid GAA, which is consistent with a heterozygous state of glycogenosis type II (Pompe's disease) and potentially of pathogenetic relevance for the intralysosomal accumulation of HES. Xanthomatous changes and lymphoedema are likely to be secondary effects, but contribute considerably to the clinical manifestation of persistent visible swelling. This observation could point to a role for GAA in the elimination of tissue-stored HES. Patients with decreased activities of GAA may be at risk of unusual adverse effects following extraordinary and prolonged tissue storage of HES, especially if it is infused in large quantities.

Aged↗

PubMed1997

Localization of a binding site for the proteoglycan decorin on collagen XIV (undulin).

Through its ability to bind extracellular matrix constituents and growth factors the small leucine-rich chondroitin/dermatan sulfate proteoglycan decorin which is present in many types of connective tissues may play an important biological role in remodeling and maintenance of extracellular matrices during inflammation, fibrosis, and cancer growth. In this study we investigated the known binding of decorin to human collagen XIV. This binding was unaffected when the small collagenous moiety of collagen XIV was removed with collagenase. Therefore, fragments covering the large noncollagenous domain NC3 of collagen XIV were expressed in Escherichia coli, each fused to a 26-kDa fragment of glutathione S-transferase. Using radioiodinated decorin as ligand for the immobilized fusion proteins, a binding site that interacted with the decorin core protein could be assigned to the NH2-terminal fibronectin type III repeat of collagen XIV. In addition, an auxiliary binding site located COOH-terminal to this fibronectin type III repeat interacted with the glycosaminoglycan component of decorin.

Amino Acid Sequence↗

PubMed1997

Critical role of glutamate in a central leucine-rich repeat of decorin for interaction with type I collagen.

The chondroitin/dermatan sulfate proteoglycan decorin is known to interact via its core protein with fibrillar collagens, thereby influencing the kinetics of fibril formation and the final diameter of the fibrils. To define the binding site(s) for type I collagen along the core protein, which is mainly composed of leucine-rich repeat structures, decorin cDNAs were constructed and expressed in human kidney 293 cells. The constructs encoded (i) C-terminally truncated molecules, (ii) core proteins with deletions of selected leucine-rich repeats, or (iii) various point mutations. The deletion of the sixth leucine-rich repeat Met176-Lys201 and the mutation E180K drastically interfered with the binding to reconstituted type I collagen fibrils. In contrast, the deletion of the seventh repeat Leu202-Ser222 led at the most to a marginally impaired binding, although the secretion of this proteoglycan was abnormally low. Decorin with two other point mutations in the sixth leucine-rich repeat, Lys187 --> Gln and Lys200 --> Gln, respectively, bound type I collagen either normally or even better than the normal recombinant proteoglycan. These data suggest that a major collagen-binding site of decorin is located within the sixth leucine-rich repeat and that glutamate-180 within this repeat is of special importance for ionic interactions between the two matrix components.

Amino Acid Sequence↗

PubMed1997

Immunohistochemical localization of the small proteoglycans decorin and biglycan in human intervertebral discs.

Immunohistochemistry was used to study the presence and distribution of the core proteins of the small proteoglycans decorin and biglycan in the various compartments of human intervertebral discs. Both proteoglycans could be found in the outer tendon-like parts of the annulus fibrosus, indicating their potential role in collagen network formation and biomechanical stress resistance. The loss of both proteoglycans in the annulus of individuals older than 50 years reflects a normal age-related change. In the nucleus pulposus, decorin could be found in fibrillar areas of the interterritorial matrix, thereby indicating co-localization of decorin with fibrils containing type II collagen. Biglycan was present in the extracellular matrix of the nucleus pulposus of adults. The pericellular immunoreactive rims observed around nucleus pulposus cells and giant chondrones indicated local biosynthetic activity for these small proteoglycans. The staining patterns in cartilage endplates resembled those found in human hyaline articular cartilage.

Adolescent↗

PubMed1997

Expression of biglycan, decorin and proteoglycan-100/CSF-1 in normal and fibrotic human liver.

The immunohistochemical expression of the three small chondroitin/dermatan sulphate proteoglycans biglycan, decorin and proteoglycan-100 (PG-100), the proteoglycan form of colony-stimulating factor-1 (CSF-1), was studied in normal and fibrotic human liver. In normal liver tissue biglycan and decorin were clearly seen in the space of Disse, which was in contrast to a faint staining of PG-100. Biglycan and decorin were additionally detected around small bile ducts and in vessel walls. In patients with HBs-Ag positive, chronic active hepatitis decorin as well as biglycan showed strong immunoreactivity in fibrotic areas. In contrast to normal liver, PG-100 was visualized in bile duct epithelia. Therefore, PG-100 could serve as an immunohistochemical marker of the involvement of the bile duct system in chronic active hepatitis and progressive liver fibrosis.

Adult↗

PubMed1997

Chondroitin/dermatan sulphate promotes the survival of neurons from rat embryonic neocortex.

Recently we have shown that biglycan, a small chondroitin sulphate proteoglycan of the extracellular matrix, supports the survival of cultured neurons from the developing neocortex of embryonic day 15 rats. Here we investigate the structure-function relationship of this neurotrophic proteoglycan and show that chondroitin/dermatan sulphate chains are the active moieties supporting survival. Heparin, a highly sulphated glucosaminoglycan, is less active than the galactosaminoglycans (chondroitin-4-sulphate, chondroitin-6-sulphate and dermatan sulphate), whereas hyaluronic acid, an unsulphated glucosaminoglycan, does not support neuron survival. Galactosaminoglycans must be in direct contact with neurons to cause survival. Experiments with elevated potassium concentrations and antagonists of voltage-gated calcium channels exclude the involvement of membrane depolarization. However, genistein and an erbstatin analogue, which are inhibitors of tyrosine kinases with low specificity, abolished neuron survival in the presence of chondroitin/dermatan sulphate, whereas a selective inhibitor of neurotrophin receptor kinases (K252a) had no suppressive effect. Thus, yet unidentified tyrosine kinases are involved in the chondroitin/dermatan sulphate-dependent survival of neocortical neurons. In the embryonic stages of rat neocortical development chondroitin sulphate is mainly located in layers I, V and VI and the subplate. Chondroitin sulphate expression is maintained after birth, extends up to cortical layer IV on postnatal day 7, and is down-regulated until postnatal day 21 concomitant with the period of naturally occurring cell death. The latter observation is consistent with a putative role of chondroitin sulphate in the control of neuron survival during cortical histogenesis.

Animals↗

PubMed1997

Differences in decorin and biglycan expression in patients with gastric ulcer healing.

BACKGROUND: The small interstitial proteoglycans decorin and biglycan have been shown to interact with various extracellular matrix molecules and with transforming growth factor-beta. These interactions are proposed to be important for tissue repair, as the former interactions may affect the diameter and spacing of collagen fibrils, and the latter interaction the proliferation and differentiation of cells embedded in the matrix. The aim of this study is to localize these proteoglycans in the stomach and to investigate their suitability as potential markers of extracellular matrix activity in gastric lesions. METHODS: Immunohistochemical techniques and in situ hybridization were used to study the phenotypic expression of these two proteoglycans in routinely processed specimens of human stomach tissue from 8 patients with gastric ulcer and 10 healthy control persons. RESULTS: In normal gastric tissue, immunostaining for both proteoglycans was found in the interstitium, with a more pronounced staining in the pylorus region than in the corpus area. In addition, biglycan showed a strong staining of parietal cells. In specimens of healing gastric ulcers a larger deposition of decorin throughout scar tissue could be shown, and a higher expression of decorin was also found by in situ hybridization. Biglycan was only found at the edges of the lesions. CONCLUSION: This study shows for the first time the presence of decorin and biglycan in human gastric mucosa. We also showed that these proteoglycans may be involved in the gastric ulcer healing processes.

Adult↗

PubMed1997

Increased turnover of small proteoglycans synthesized by human osteoblasts during cultivation with ascorbate and beta-glycerophosphate.

The small proteoglycan decorin had been localized previously at the d-band in the gap zone of collagen fibrils in nonmineralizing tissues. In bone matrix this zone is proteoglycan free and is at least in some species the place where mineralization along collagen fibrils starts. To study the metabolism of the small proteoglycans decorin and biglycan under mineralizing conditions, osteoblasts from human nasal bone were cultured for several weeks in the presence or absence of beta-glycerophosphate and ascorbate. An immediate consequence of the treatment was a reduced expression of decorin, as judged by immune precipitation, whereas the biosynthesis of biglycan was not affected. Pulse-chase experiments were performed with osteoblasts embedded in floating type I collagen gels. In the presence of beta-glycerophosphate and ascorbate, a more rapid turnover of both proteoglycans was noted; the one of biglycan reached statistical significance. Indirect evidence for an enhanced rate of proteoglycan endocytosis was obtained. This effect was not seen in cultured skin fibroblasts. Thus, osteoblasts respond rapidly to mineralizing conditions with alterations of small proteoglycan biosynthesis and turnover.

Adolescent↗

PubMed1996

Cultured astrocytes express biglycan, a chondroitin/dermatan sulfate proteoglycan supporting the survival of neocortical neurons.

Astrocyte-conditioned medium (ACM) supports the survival of rat E15 neocortical neurons. Using a microtiter assay for neuronal survival, we demonstrated that part of the survival activity is associated with a proteoglycan fraction obtained after two chromatographic steps: (1) preparative Q-Sepharose anion-exchange chromatography under non-denaturating conditions and (2) MonoQ chromatography in the presence of 8 M urea. Analytical SDS-polyacrylamide gradient gel electrophoresis of pooled active MonoQ-fractions (MQ-pool) revealed a broad proteoglycan band migrating with an apparent M(r) in the range of 150-400 kDa. Digestion of the MQ-pool with chondroitin-ABC-lyase yielded a major core protein of 50 kDa. In Western blots the high molecular weight (150-400 kDa) material as well as the 50 kDa core protein band were immunoreactive to chicken polyclonal antibodies raised against purified biglycan from rat meningeal fibroblasts. Northern blot analysis of total RNA prepared from highly enriched astrocyte cultures revealed a single 2.9 kb biglycan transcript. By using in situ hybridization we demonstrated that essentially all cells in these cultures expressed biglycan mRNA. Furthermore, highly purified biglycan from bovine cartilage was shown to markedly enhance survival of rat neocortical neurons. In conclusion, we have shown that astrocytes synthesize and release the small chondroitin/dermatan sulfate proteoglycan (CS/DSPG) biglycan, a molecule that was found to support survival of neocortical neurons in vitro.

Animals↗

PubMed1996

Deficient expression of the small proteoglycan decorin in a case of severe/lethal osteogenesis imperfecta.

In osteogenesis imperfecta (OI) the effects of mutations in type I collagen genes generally reflect their nature and localization. Unrelated individuals sharing identical mutations present, in general, similar clinical phenotypes. However, in some such cases the clinical phenotype differs. This variable clinical expression could be the result of abnormalities in other connective tissue proteins. Since decorin is a component of connective tissue, binds to type I collagen fibrils and plays a role in matrix assembly, we studied decorin production in skin fibroblasts from OI patients. Cultured fibroblasts from one patient with extremely severe osteogenesis imperfecta (classified as type II/III) who has an alpha 1(I)gly415ser mutation were found to secrete barely detectable amounts of decorin into culture medium. Western blotting using antibodies raised against decorin confirmed the reduction of the decorin core protein and Northern blot analysis showed decorin mRNA levels below the limit of detection. Cells from a patient, with a less severe phenotype, bearing a mutation in the same position of the triple helix (alpha 1(I)gly415) expressed decorin normally. The different clinical phenotypes could be due to the differing genetic backgrounds of the patients so it is tempting to conclude that in our most severely affected patient the absence of decorin aggravates the clinical phenotype.

Blotting, Northern↗

PubMed1996

Modulation of collagen gel contraction by decorin.

The small dermatan sulphate protein decorin interacts via its core protein with fibrillar collagens, and its glycosaminoglycan chains were proposed to be capable of self-association. It was therefore of interest to study the role of decorin in the contraction of cell-populated collagen lattices. Stable transfection of dihydrofolate reductase-deficient CHO cells with decorin cDNA resulted in impaired collagen lattice contraction. Using normal human skin fibroblasts in serum-free cultures, inclusion of 0.3 microM decorin in the culture medium also led to a delayed collagen gel contraction. Protein-free dermatan sulphate and the dermatan sulphate-degrading enzyme chondroitin ABC lyase were ineffective. Potential interactions between dermatan sulphate chains were studied by gel filtration. A shift in the elution position of [35S]sulphate-labelled decorin-derived glycosaminoglycans by unlabelled decorin could be observed only when the chains were prepared by trypsin. Chains liberated by beta-elimination or by cathepsin C were eluted at identical positions in the presence or absence of decorin. It is therefore unlikely, that the effect of decorin on collagen-gel retraction is brought about solely by glycosaminoglycan-glycosaminoglycan interactions.

Adolescent↗

PubMed1996

Mucopolysaccharidosis type II (Hunter syndrome): mutation "hot spots" in the iduronate-2-sulfatase gene.

Mucopolysaccharidosis type II (MPS II, Hunter syndrome) is an X-chromosomal storage disorder due to deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS). We have identified IDS mutations in a total of 31 families/patients with MPS II, of which 20 are novel and unique and a further 1 is novel but has been found in 3 unrelated patients. One of the mutations detected is of special interest as an AG-->G substitution in an intron, far apart from the coding region, is deleterious by creating a new 5'-splice-donor site that results in the inclusion of a 78-bp intronic sequence. While the distribution of gene rearrangements (deletions, insertions, and duplications) of <20 bp seems to be random over the IDS gene, the analysis of a total of 101 point mutations lying within the coding region shows that they tend to be more frequent in exons III, VIII, and IX. Forty-seven percent of the point mutations are at CpG dinucleotides, of which G:C-to-A:T transitions constitute nearly 80%. Almost all recurrent point mutations involve CpG sites. Analysis of a collective of 50 families studied in our laboratory, to date, revealed that mutations occur more frequently in male meioses (estimated male-to-female ratio between 3.76 and 6.3).

Alternative Splicing↗

PubMed1996

Isolation and cellular localization of the decorin endocytosis receptor.

The small dermatan sulfate proteoglycans decorin and biglycan are efficiently internalized by a variety of cells of mesenchymal origin. This process is modulated, at least under tissue culture conditions, by cell surface-associated heparan sulfate proteoglycans. Receptor proteins of 51 and 26 kDa, respectively, bind to leucine-rich repeat structures of the core proteins of the small proteoglycans but also to highly sulfated domains of heparan sulfate. The 51 kDa protein was purified from rat brain tissue by subcellular fractionation, heparin affinity chromatography and subsequent SDS-PAGE, and was used for raising a polyclonal antiserum. Affinity-purified antibodies also recognize the 26 kDa protein and a few other low molecular weight proteins, suggesting that these proteins represent proteolytic degradation products of the 51 kDa receptor. By confocal laser microscopy, it could be demonstrated that the affinity-purified antibody reacted at 0 degree C with a protein that became internalized and was transported to a perinuclear compartment during 15 min of incubation at 37 degrees C. These findings provide direct evidence that the receptor protein(s) are internalized together with the ligand and reach an endosomal compartment where further sorting can occur.

Animals↗

PubMed1995

Differential expression of the small chondroitin/dermatan sulfate proteoglycans decorin and biglycan after injury of the adult rat brain.

Chondroitin sulfate proteoglycans are widespread extracellular matrix proteins and are specifically upregulated after CNS injury at the lesion site. Many proteoglycan core proteins have been described in the rat brain, but detailed analysis of individual proteoglycans expressed after injury are missing. The present study represents an initial attempt to assess the diversity and timing of lesion-induced expression of proteoglycans in order to elucidate their functional role in CNS injury and repair. Using immunocytochemical methods we analysed the expression of decorin and biglycan in the transected postcommissural fornix of the adult rat. Transection of the fornix induced the upregulation of both decorin and biglycan. However, their expression differed with respect to time course, regional extent and cellular localization. The rapid upregulation of decorin within a wide area around the lesion was followed by a massive appearance of biglycan that remained restricted to the transection site. Three months after lesion, differences of the area size of decorin- and biglycan-immunoreactivities were no longer detectable. Both proteoglycans were restricted to the lesion site and the fornix stumps. While decorin was primarily expressed by astrocytes, biglycan was deposited extracellularly in sheet-like structures. The upregulation of both proteoglycans persisted for at least up to 6 months after lesion. These strong but divergent lesion-induced expression patterns indicate important but different roles of decorin and biglycan in CNS injury.

Animals↗

PubMed1995

Proteoglycan form of colony-stimulating factor-1 (proteoglycan-100). Stimulation of activity by glycosaminoglycan removal and proteolytic processing.

A proteoglycan had been isolated from the conditioned media of a human osteosarcoma cell line and had tentatively been named proteoglycan-100 (PG-100) because of the size of its core glycoprotein. Amino acid sequencing of the purified proteoglycan and cDNA analysis were consistent with the assumption that PG-100 is identical with the proteoglycan form of CSF-1 (or macrophage colony-stimulating factor). PG-100 induced mouse macrophage differentiation. Proliferation of macrophages was stimulated in a dose-dependent manner. On a molar basis, however, about 100- to 300-fold higher doses of PG-100 than of recombinant human (rh)CSF-1 were required for the half-maximal growth-stimulating effect. Upon enzymatic removal of the glycosaminoglycan chain, the purified core protein exhibited higher activity, but was still about 20-fold less active than rhCSF-1. Incubation of the purified proteoglycan for 48 h at 37 degrees C led to the formation of a glycosaminoglycan-free 50-kDa fragment either by autoproteolysis or by the action of a protease not yet identified. The purified fragment exhibited almost the same biologic activity as rhCSF-1. The glycosaminoglycan chain of the growth factor was not only shown to inhibit CSF-1 activity but also to increase the stability of the core protein when the CSF-1-producing osteosarcoma cells were maintained in a collagen lattice. These findings provide a link between a soluble, highly active cytokine and its extracellular matrix storage form of comparatively low activity.

Amino Acid Sequence↗