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Robert Stern

Publications and source records attributed to Robert Stern.

At least 19 recordsLinked to original sources
PubMed2006

A case of Churg-Strauss syndrome associated with antiphospholipid antibodies.

Churg-Strauss syndrome (CSS) is a systemic vasculitis affecting both small- and medium-sized blood vessels, almost invariably affecting the lung, and frequently associated with cutaneous involvement. Microvascular vaso-occlusion leading to digital gangrene is not a feature of CSS. We report an unusual case of a patient with CSS with antiphospholipid antibodies who developed severe digital gangrene in addition to cutaneous vasculitis. The presence of antiphospholipid antibodies is not a feature usually seen in association with CSS. While the full clinical spectrum of CSS is still being defined, the identification of additional features associated with this syndrome might help to better understand the pathogenesis of the disease and to have an impact on both management and prognosis.

Antibodies, Antiphospholipid↗
PubMed2006

Hyaluronic acid: a natural biopolymer with a broad range of biomedical and industrial applications.

Hyaluronic acid (hyaluronan, HA) is a linear polysaccharide formed from disaccharide units containing N-acetyl-D-glucosamine and glucuronic acid. It has a high molecular mass, usually in the order of millions of Daltons, and interesting viscoelastic properties influenced by its polymeric and polyelectrolyte characteristics. HA is present in almost all biological fluids and tissues. In clinical medicine, it is used as a diagnostic marker for many diseases including cancer, rheumatoid arthritis and liver pathologies, as well as for supplementation of impaired synovial fluid in arthritic patients by means of intra-articular injections. It is also used in certain ophthalmological and otological surgeries and cosmetic regeneration and reconstruction of soft tissue. Herein we present an overview of the occurrence and physiological properties of HA, as well as of the recent advances in production biotechnology and preparation of the HA-based materials for medical application.

Biocompatible Materials↗
PubMed2006

Hyaluronan fragments: an information-rich system.

Hyaluronan is a straight chain, glycosaminoglycan polymer of the extracellular matrix composed of repeating units of the disaccharide [-D-glucuronic acid-beta1,3-N-acetyl-D-glucosamine-beta1,4-]n. Hyaluronan is synthesized in mammals by at least three synthases with products of varying chain lengths. It has an extraordinary high rate of turnover with polymers being funneled through three catabolic pathways. At the cellular level, it is degraded progressively by a series of enzymatic reactions that generate polymers of decreasing sizes. Despite their exceedingly simple primary structure, hyaluronan fragments have extraordinarily wide-ranging and often opposing biological functions. There are large hyaluronan polymers that are space-filling, anti-angiogenic, immunosuppressive, and that impede differentiation, possibly by suppressing cell-cell interactions, or ligand access to cell surface receptors. Hyaluronan chains, which can reach 2 x 10(4) kDa in size, are involved in ovulation, embryogenesis, protection of epithelial layer integrity, wound repair, and regeneration. Smaller polysaccharide fragments are inflammatory, immuno-stimulatory and angiogenic. They can also compete with larger hyaluronan polymers for receptors. Low-molecular-size polymers appear to function as endogenous "danger signals", while even smaller fragments can ameliorate these effects. Tetrasaccharides, for example, are anti-apoptotic and inducers of heat shock proteins. Various fragments trigger different signal transduction pathways. Particular hyaluronan polysaccharides are also generated by malignant cells in order to co-opt normal cellular functions. How the small hyaluronan fragments are generated is unknown, nor is it established whether the enzymes of hyaluronan synthesis and degradation are involved in maintaining proper polymer sizes and concentration. The vast range of activities of hyaluronan polymers is reviewed here, in order to determine if patterns can be detected that would provide insight into their production and regulation.

Animals↗
PubMed2005

Tumor cells enhance their own CD44 cleavage and motility by generating hyaluronan fragments.

Hyaluronan (HA) is an extracellular matrix glycosaminoglycan that interacts with cell-surface receptors, including CD44. Although HA usually exists as a high molecular mass polymer, HA of a much lower molecular mass that shows a variety of biological activities can be detected under certain pathological conditions, particularly in tumors. We previously reported that low molecular weight HAs (LMW-HAs) of a certain size range induce the proteolytic cleavage of CD44 from the surface of tumor cells and promote tumor cell migration in a CD44-dependent manner. Here, we show that MIA PaCa-2, a human pancreatic carcinoma cell line, secreted hyaluronidases abundantly and generated readily detectable levels of LMW-HAs ranging from approximately 10- to 40-mers. This occurred in the absence of any exogenous stimulation. The tumor-derived HA oligosaccharides were able to enhance CD44 cleavage and tumor cell motility. Inhibition of the CD44-HA interaction resulted in the complete abrogation of these cellular events. These results are consistent with the concept that tumor cells generate HA oligosaccha-rides that bind to tumor cell CD44 through the expression of their own constitutive hyaluronidases. This enhances their own CD44 cleavage and cell motility, which would subsequently promote tumor progression. Such an autocrine/paracrine-like process may represent a novel activation mechanism that would facilitate and promote the malignant potential of tumor cells.

Animals↗
PubMed2005

Magnetic resonance imaging visualization of hyaluronidase in ovarian carcinoma.

Hyaluronan, a high molecular weight, negatively charged polysaccharide, is a major constituent of the extracellular matrix. High molecular weight hyaluronan is antiangiogenic, but its degradation by hyaluronidase generates proangiogenic breakdown products. Thus, by expression of hyaluronidase, cancer cells can tilt the angiogenic balance of their microenvironment. Indeed, hyaluronidase-mediated breakdown of hyaluronan correlates with aggressiveness and invasiveness of ovarian cancer metastasis and with tumor angiogenesis. The goal of this work was to develop a novel smart contrast material for detection of hyaluronidase activity by magnetic resonance imaging (MRI). Gadolinium-diethylenetriaminepentaacetic acid (GdDTPA) covalently linked to hyaluronan on the surface of agarose beads showed attenuated relaxivity. Hyaluronidase, either purified from bovine testes or secreted by ES-2 and OVCAR-3 human epithelial ovarian carcinoma cells, activated the hyaluronan-GdDTPA-beads by rapidly altering the R1 and R2 relaxation rates. The change in relaxation rates was consistent with the different levels of biologically active hyaluronidase secreted by those cells. Hyaluronan-GdDTPA-beads were further used for demonstration of MRI detection of hyaluronidase activity in the proximity of s.c. ES-2 ovarian carcinoma tumors in nude mice. Thus, hyaluronan-GdDTPA-beads could allow noninvasive molecular imaging of hyaluronidase-mediated tilt of the peritumor angiogenic balance.

Animals↗
PubMed2005

Structures of vertebrate hyaluronidases and their unique enzymatic mechanism of hydrolysis.

Human hyaluronidases (Hyals) are a group of five endo-beta-acetyl-hexosaminidase enzymes, Hyal-1, -2, -3, -4, and PH-20, which degrade hyaluronan using a hydrolytic mechanism of action. Catalysis by these Hyals has been shown to follow a double-displacement scheme. This involves a single Glu residue within the enzyme, the only catalytic residue, as the proton donor (acid). Also involved is a carbonyl group of the hyaluronan (HA) N-acetyl-D-glucosamine as a unique type of nucleophile. Thus the substrate participates in the mechanism of action of its own catalysis. An oxocarbonium ion transition state is postulated, but there is no formation of a covalent enzyme-glycan intermediate, as found in most such reactions. The major domain is catalytic and has a distorted (beta/alpha)8 triose phosphate isomerase (TIM) barrel fold. The C-terminal domain is separated by a peptide linker. Each Hyal has a different C-terminal sequence and structure, the function of which is unknown. These unique C-termini may participate in the additional function(s) associated with these multifunctional enzymes.

Amino Acid Sequence↗
PubMed2005

An assay for bacterial and eukaryotic chondroitinases using a chondroitin sulfate-binding protein.

A simple, rapid, and reproducible microtiter-based chondroitinase (CSase) assay is reported here, based on the competition of chondroitin sulfate (CS) with immobilized hyaluronan (HA) for the binding of TSG-6 protein, the product of TNF-inducible gene 6. Although the catabolic reaction of bacterial and other prokaryotic CSase enzymes, often referred to as the chondroitin lyases, can be followed by tracking the generation of unsaturated bonds by the spectrophotometrical determination of the absorbance at 232 nm, no rapid, sensitive, and simple assay has been devised to date for measuring the activity of the vertebrate enzymes that cleave their substrate exclusively by hydrolysis. We provide data demonstrating that the CSase assay described here is suitable for the determination of the activities of both classes of enzymes. For the bacterial enzyme CSase ABC, both the determination of the absorbance at 232 nm and the assay based on TSG-6 binding are suitable using the same range of enzyme activities. However, for testicular hyaluronidase, considerably higher enzyme activities were needed to cleave CS than to cleave HA. Using the HA-binding domain of aggrecan for a comparison, we determined that the interaction between TSG-6 and chondroitin sulfate is uniquely suited for this CSase assay.

Animals↗
PubMed2005

Hyaluronan metabolism: a major paradox in cancer biology.

Paradoxically, both hyaluronan (HA) and hyaluronidases, the enzymes that eliminate HA, can correlate with cancer progression. Levels of HA on the surface of tumor cells are indicators of poor outcome. Certain hyaluronidases, products of tumor suppressor genes eliminated in the course of tumor spread, are used clinically in anti-cancer chemotherapy regimens. Such information would indicate that cancer progression is inhibited by hyaluronidase. Yet progression of certain cancers correlates with levels of hyaluronidase activity. An attempt is made here to understand such apparent contradictions by examining details of HA metabolism. Anabolic and catabolic pathways are comprised of the HA synthases and hyaluronidases, respectively. There are several enzymes that synthesize HA, each under a different control mechanism, generating products of differing polymer size. The hyaluronidases degrade HA in step-wise fashion, the polymer decreasing in size in quantum steps, each size-specific polymer having a different biological activity. Superimposed on these are the potent hyaluronidase inhibitors, about which very little is known. These components of HA metabolism are reviewed here for possible roles in supporting or suppressing malignant transformation, growth, invasion and metastatic spread of tumors. Such a systematic approach may reveal mechanisms used in the course of cancer progression, resolve some of the apparent disparities, render new prognostic markers, and provide new targets for therapeutic intervention.

Glucuronosyltransferase↗
PubMed2005

Contribution of oxidative-reductive reactions to high-molecular-weight hyaluronan catabolism.

Since the content of hyaluronan (HA)-degrading enzymes in synovial fluid (SF), if any, is extremely low, the high rate of HA turnover in SF is to result from a cause different from enzymatic catabolism. An alternative and plausible mechanism is that of oxidative-reductive degradation of HA chains by a combined action of oxygen and transition metal cations maintained in a reduced oxidation state by ascorbate.

Hyaluronic Acid↗
PubMed2005

Differentiation of mesothelioma from adenocarcinoma in serous effusions: the role of hyaluronic acid and CD44 localization.

Differentiating cells of mesothelial origin from adenocarcinoma (ACA) based on morphology alone can be a diagnostic challenge, especially in cytological specimens. Malignant mesothelioma (MM) is characterized by accumulation of abundant intracellular hyaluronic acid (HA), a feature that is not reported in ACA. The purpose of this study was to evaluate the significance of cellular HA using an HA-specific binding peptide (HABP) and the expression of its principal receptor, the standard CD44 molecule (CD44S). Archival paraffin-embedded cell blocks of serous fluids from 28 cases of reactive mesothelial cells, 14 cases of MM, 20 cases of metastatic ovarian carcinomas, 17 cases of metastatic breast carcinomas, 12 cases of metastatic lung ACA, and 12 cases of metastatic gastrointestinal ACA were stained with HA using a biotinylated HABP and CD44S. Positive staining was defined as droplet to diffuse cytoplasmic staining for HA and uniform membranous staining for CD44S. All MMs and 93% (26/28) of the benign mesothelial cells were positive for intracytoplasmic HA vs. none of ACAs. CD44S was expressed in 100% (28/28) of mesothelial hyperplesia, 86% (12/14) of MMs, 70% (14/20) of ovarian carcinomas, 29% (5/17) of breast carcinomas, 25% (3/12) of gastrointestinal ACAs, and 8% (1/12) of lung ACAs. In MM and reactive mesothelial cells, CD44S stained cell membranes diffusely with highlights on the villous surfaces and in ACA it was focal and confined to cell membranes. Immunostaining with HA is a reliable marker that can distinguish between cells of mesothelial origin (reactive mesothelial cells and MM) and ACA. The CD44S staining pattern of cells of mesothelial origin is of diagnostic significance. CD44 may prove useful in conjunction with other stains in the differential diagnosis of mesothelioma and ADA.

Adenocarcinoma↗
PubMed2005

Expression of hyaluronic acid and its receptors, CD44s and CD44v6, in normal, hyperplastic, and neoplastic endometrium.

The interaction between epithelial tumor cells and their surrounding stroma is important in tumor progression and metastasis. This is accomplished through a number of transmembrane receptors that interact with stromal extracellular matrix molecules. One of these receptors, CD44, binds to extracellular matrix component hyaluronic acid (HA). The purpose of this study was to evaluate the significance of HA, CD44s, and CD44v6 in benign, hyperplastic, atypical, and malignant endometrial epithelia. Archival paraffin-embedded cell blocks from proliferative endometrium (n = 11), secretory endometrium (n = 12), simple hyperplasia (n = 13), complex hyperplasia without atypia (n = 9), complex hyperplasia with atypia (n = 17), and adenocarcinoma (n = 21) were stained for HA, CD44s, and CD44v6. HA was detected throughout the normal menstrual cycle but was more intense during the secretory phase. Only during the secretory phase was CD44s expressed in the stromal cells in 11 cases (92%), whereas CD44v6 was detected in glandular epithelium in 9 (75%). CD44s was expressed in the glandular epithelium in 2 (15%) cases of simple hyperplasia, 4 (44%) of complex hyperplasia without atypia, 14 (82%) of complex hyperplasia with atypia, and in 16 (76%) of adenocarcinoma. CD44v6 was expressed in the glandular epithelium in 1 (11%) case of complex hyperplasia without atypia, 17 (100%) cases of complex hyperplasia with atypia, and in 18 (86%) cases of adenocarcinoma, but in none of the cases of simple hyperplasia. The endometrial stromal cells expressed CD44v6 in 1 (8%) case of simple hyperplasia, 6 (67%) of complex hyperplasia without atypia, 8 (47%) of complex hyperplasia with atypia, and in 3 (14%) of adenocarcinoma. We concluded that in the normal menstrual cycle, the timing of peak staining of HA and CD44s in the stroma and the up-regulation of CD44v6 in secretory glands are coincident with the period in which the endometrium is most receptive to embryo implantation. HA is more abundant in the stroma adjacent to the tumor, suggesting that interactions between tumor cells and stromal HA promote tumorigenesis. With progression from hyperplasia and with increasing atypia to adenocarcinoma, levels of stromal HA, glandular CD44v6, and glandular and stromal CD44s all increase. Thus, HA and CD44 are both involved in the development and progression of endometrial cancer.

Adenocarcinoma↗
PubMed2005

Scleromyxedema-like lesions of patients in renal failure contain hyaluronan: a possible pathophysiological mechanism.

BACKGROUND: Patients with renal failure have been identified recently, some on dialysis, others with renal transplants, who have scleromyxedema-like skin changes. These lesions are characterized grossly by extensive thickening of skin, brawny pigmentation, papules, and subcutaneous nodules. Mucinous deposits are observed histologically that resemble those in scleromyxedema. METHODS: Biopsies of these lesions were stained with a biotinylated hyaluronan (HA)-binding protein coupled to an avidin-peroxidase reaction. RESULTS: These lesions are associated with marked deposition of HA in the papillary dermis. CONCLUSIONS: HA turnover is cleared rapidly in the circulation by both liver and kidney. Evidence suggests that high molecular size HA chains, which are anti-inflammatory, antiangiogenic, and immuno-suppressive are cleared by the liver. By contrast, intermediate-size fragments, which are highly angiogenic, inflammatory, and a stimulus for fibrous deposition, are cleared by the kidney. The accumulation of such fragments in renal failure can account for HA deposition in the dermis and may be a mechanism for the nephrogenic fibrosing dermopathy that can accompany these lesions.

Female↗
PubMed2004

CD44 interaction with Na+-H+ exchanger (NHE1) creates acidic microenvironments leading to hyaluronidase-2 and cathepsin B activation and breast tumor cell invasion.

We have explored CD44 (a hyaluronan (HA) receptor) interaction with a Na(+)-H(+) exchanger (NHE1) and hyaluronidase-2 (Hyal-2) during HA-induced cellular signaling in human breast tumor cells (MDA-MB-231 cell line). Immunological analyses demonstrate that CD44s (standard form) and two signaling molecules (NHE1 and Hyal-2) are closely associated in a complex in MDA-MB-231 cells. These three proteins are also significantly enriched in cholesterol and ganglioside-containing lipid rafts, characterized as caveolin and flotillin-rich plasma membrane microdomains. The binding of HA to CD44 activates Na(+)-H(+) exchange activity which, in turn, promotes intracellular acidification and creates an acidic extracellular matrix environment. This leads to Hyal-2-mediated HA catabolism, HA modification, and cysteine proteinase (cathepsin B) activation resulting in breast tumor cell invasion. In addition, we have observed the following: (i) HA/CD44-activated Rho kinase (ROK) mediates NHE1 phosphorylation and activity, and (ii) inhibition of ROK or NHE1 activity (by treating cells with a ROK inhibitor, Y27632, or NHE1 blocker, S-(N-ethyl-N-isopropyl) amiloride, respectively) blocks NHE1 phosphorylation/Na(+)-H(+) exchange activity, reduces intracellular acidification, eliminates the acidic environment in the extracellular matrix, and suppresses breast tumor-specific behaviors (e.g. Hyal-2-mediated HA modification, cathepsin B activation, and tumor cell invasion). Finally, down-regulation of CD44 or Hyal-2 expression (by treating cells with CD44 or Hyal-2-specific small interfering RNAs) not only inhibits HA-mediated CD44 signaling (e.g. ROK-mediated Na(+)-H(+) exchanger reaction and cellular pH changes) but also impairs oncogenic events (e.g. Hyal-2 activity, hyaluronan modification, cathepsin B activation, and tumor cell invasion). Taken together, our results suggest that CD44 interaction with a ROK-activated NHE1 (a Na(+)-H(+) exchanger) in cholesterol/ganglioside-containing lipid rafts plays a pivotal role in promoting intracellular/extracellular acidification required for Hyal-2 and cysteine proteinase-mediated matrix degradation and breast cancer progression.

Amides↗
PubMed2004

Serum hyaluronan and hyaluronidase: very early markers of toxic liver injury.

BACKGROUND: Dimethylnitrosamine (DMN), a potent hepatotoxin, administered to rats, provides a convenient model for toxic liver injury. Indicators of early liver injury are important clinically, for surveillance, for screening new drugs that are potentially hepatotoxic and for identifying drugs that protect against liver injury. Both cirrhosis and wound healing culminate in deposition of fibrous connective tissue and scarring. Increased hyaluronan (HA) occurs in the earliest stage of wound healing. Hyaluronidase, the enzyme that degrades hyaluronan, is also elevated whenever rapid turnover of hyaluronan occurs. We test the hypothesis that elevated levels of circulating hyaluronan and hyaluronidase could provide indicators of very early liver damage. METHODS: Dimethylnitrosamine was administered to adult male albino rats by intraperitoneal injections for 7 consecutive days. RESULTS: Increased serum hyaluronan levels observed on day 2 reached a maximum on day 4. Hyaluronidase was elevated on the first day and reached a maximum on day 2 that was 30-times control levels. Hyaluronan-specific staining in liver sections was maximal on day 7, occurring predominantly in portal triads and in sinusoidal spaces. Individual hepatocytes were slightly enlarged and contained intracellular hyaluronan, which was not evident in control sections. Though circulating hyaluronan levels had decreased after day 4, continued hyaluronan staining persisted in liver sections through day 21. Conventional indicators of liver injury, such as serum aminotransferase enzymes, did not reach a peak until day 7. Conventional gross and histopathological changes, including severe centrilobular congestion and hemorrhagic necrosis, were observed only after day 7. Both hyaluronan and hyaluronidase are indicators of very early liver damage in the dimethylnitrosamine-treated rat, occurring well before conventional indicators appear, or before overt histopathologic changes of liver damage can be seen. However, levels are increased only transiently, indicating that serial assays are necessary. CONCLUSIONS: Measures of circulating hyaluronidase activity may be used to assess liver damage.

Alanine Transaminase↗
PubMed2004

Inhibition of hyaluronan degradation by dextran sulphate facilitates characterisation of hyaluronan synthesis: an in vitro and in vivo study.

The concentration and molecular weight of hyaluronan often dictates its physiological function. Consequently full characterisation of the anabolic products and turnover rates of HA could facilitate understanding of the role that HA metabolism plays in disease processes. In order to achieve this it is necessary to interrupt the dynamic balance between concurrent HA synthesis and degradation, achievable through the inhibition of the hyaluronidases, a group of enzymes which degrade HA. The sulphated polysaccharide, dextran sulphate has been demonstrated to competitively inhibit testicular hyaluronidase in a non-biological system, but its application to in vitro biological systems had yet to be developed and evaluated. This study determined the inhibitory concentrations of dextran sulphate against both testicular and Streptomyces hyaluronidase in a cell-free and breast cancer model followed by characterisation of the effect that hyaluronidase inhibition exerted on HA synthesis and degradation. The IC(100) of dextran sulphate for both hyaluronidases in a cell-free and biological system was determined to be >or=400 microg/ml. At concentrations up to 10 mg/ml the dextran sulphate did not effect breast cancer cell proliferation or morphology, while at 400 microg/ml HA degradation was totally inhibited, enabling an accurate quantitation of HA production as well as characterisation of the cell-associated and liberated HA. FACS quantitation of the HA receptor CD44, HA synthase and the hyaluronidases HYAL 1 and HYAL 2 demonstrated that dextran sulphate down-regulated CD44 and HA synthase while upregulating the hyaluronidases. These results suggest dynamic feedback signalling and complex mechanisms occur in the net deposition of HA in vivo.

Animals↗
PubMed2004

Hyaluronan catabolism: a new metabolic pathway.

A new pathway of intermediary metabolism is described involving the catabolism of hyaluronan. The cell surface hyaluronan receptor, CD44, two hyaluronidases, Hyal-1 and Hyal-2, and two lysosomal enzymes, beta-glucuronidase and beta-N-acetylglucosaminidase, are involved. This metabolic cascade begins in lipid raft invaginations at the cell membrane surface. Degradation of the high-molecular-weight extracellular hyaluronan occurs in a series of discreet steps generating hyaluronan chains of decreasing sizes. The biological functions of the oligomers at each quantum step differ widely, from the space-filling, hydrating, anti-angiogenic, immunosuppressive 10(4)-kDa extracellular polymer, to 20-kDa intermediate polymers that are highly angiogenic, immuno-stimulatory, and inflammatory. This is followed by degradation to small oligomers that can induce heat shock proteins and that are anti-apoptotic. The single sugar products, glucuronic acid and a glucosamine derivative are released from lysosomes to the cytoplasm, where they become available for other metabolic cycles. There are 15 g of hyaluronan in the 70-kg individual, of which 5 g are cycled daily through this pathway. Some of the steps in this catabolic cascade can be commandeered by cancer cells in the process of growth, invasion, and metastatic spread.

Animals↗