Dataset Information

The effect of beta-D-xylosides on the proliferation and proteoglycan biosynthesis of monoblastic U-937 cells.

ABSTRACT: The monoblastic cell line U-937 was cultured in the presence of C-ethyl beta-D-xyloside (E-xyl), hexyl beta-D-thioxyloside (HX-xyl), p-nitrophenyl beta-D-xyloside, phenyl beta-D-xyloside or phenyl alpha-D-xyloside. All of the beta-D-xylosides inhibited proliferation, but HX-xyl was by far the most efficient, and had a maximum effect at 1 mM concentration. The inhibitory effect of HX-xyl could be reversed; after washing, the HX-xyl-treated cells proliferated with a pattern similar to that of control cells. For more detailed analysis of the effects of beta-D-xylosides on cell proliferation and chondroitin sulphate (CS)/chondroitin sulphate proteoglycan (CSPG) structure, a comparison between the effects of E-xyl and HX-xyl was made. Treating the cells with 1 mM-HX-xyl resulted in a large increase in CS synthesis, whereas 1 mM-E-xyl had only minor effects on the rate of PG/glycosaminoglycan synthesis. Sepharose CL-6B gel chromatography of medium and cell fractions from 35S-labelled cells revealed that HX-xyl treatment resulted in the expression of only free CS chains, whereas E-xyl exposure leads to the synthesis of both large and small CSPGs, as well as some free CS chains. The expression of elevated levels of free CS chains was clearly correlated to the inhibition of proliferation. The proliferation of U-937-4, a clone of U-937 synthesizing ten times more CSPG/CS than the parent line, was equally inhibited by HX-xyl treatment. With this clone, however, there was no stimulation of CS synthesis after xyloside exposure, indicating that the elevated level of CS evident after xyloside treatment of the parent cell line is not causing the inhibition of proliferation. Furthermore, the biosynthesis of hyaluronate was shown not to be implicated in the xyloside-induced decrease in proliferation. The inhibition of proliferation observed in the presence of 1 mM-HX-xyl did not lead to differentiation of the cells into macrophage-like cells, as is observed when the cells are cultured in the presence of phorbol esters, agents also known to inhibit proliferation of U-937 cells.

SUBMITTER: Kolset SO

PROVIDER: S-EPMC1133682 | biostudies-other | 1990 Feb

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description:Incorporation of [35S]]sulphate, [3H]glucose and [3H]serine into glycosaminoglycans and proteoglycans of embryonic-chicken sternum was measured in vitro in incubation medium containing 4-methylumbelliferyl beta-D-xyloside or p-nitrophenyl beta-D-xyloside at low concentrations, and in the absence of inhibitors of protein synthesis. Incorporation of sulphate was decreased by 80% in incubations in which 1mM-4-methylumbelliferyl beta-xyloside or 2.5 mM-p-nitrophenyl beta-xyloside was present; under these conditions, serum factors stimulated incorporation to only a small extent. When the concentration of the xyloside was decreased tenfold, incorporation of sulphate was inhibited by 60-70%, but when normal human serum or L-3,3',5-tri-iodothyronine or both were also added to the incubation medium, incorporation was markedly stimulated. Experiments in which [35S]sulphate and [3H]glucose were incorporated simultaneously, and enzymic analysis of glycosaminoglycans formed in such experiments, indicated that chondroitin sulphate formed in the presence of 0.1 mM-4-methylumbelliferyl beta-xyloside contained 30-40% less sulphate than did chondrotin sulphate synthesized in the absence of xylosides. Similar experiments, with [3H]serine instead of [3H]glucose, suggested also a 20-30% decrease in chain length of the chondroitin sulphate; this was confirmed by direct gel filtration of labelled glycosaminoglycans on a calibrated column. Incorporation of [3H]glucose or [3H]serine was stimulated by serum and tri-iodothyronine in parallel with incorporation of sulphate. The changes seen in the total chondroitin sulphate were mirrored in the major proteoglycan fraction, purified by isopycnic centrifugation of salt-extracted proteoglycans. The labelling pattern of chondroitin sulphate from this proteoglycan indicated that decreased sulphation of chondroitin sulphate was largely due to the inferior ability of short polysaccharide chains to accept sulphate, with some direct interference with transfer of sulphate to all chains. The results also suggested that the action of serum factors on synthesis of proteochondroitin sulphate is exercised at the level of either protein synthesis or transport to the sites of initiation of polysaccharide synthesis.

Project description:Glycosaminoglycans (GAGs) play a critical role in binding and activation of growth factors involved in cell signaling critical for developmental biology. The biosynthetic pathways for GAGs have been elucidated over the past decade and now analytical methodology makes it possible to determine GAG composition in as few as 10 million cells. A glycomics approach was used to examine GAG content, composition, and the level of transcripts encoding for GAG biosynthetic enzymes as murine embryonic stem cells (mESCs) differentiate to embryoid bodies (EBs) and to extraembryonic endodermal cells (ExE) to better understand the role of GAGs in stem cell differentiation. Hyaluronan synthesis was enhanced by 13- and 24-fold, most likely due to increased expression of hyaluronan synthase-2. Chondroitin sulfate (CS)/dermatan sulfate (DS) synthesis was enhanced by 4- and 6-fold, and heparan sulfate (HS) synthesis was enhanced by 5- and 8-fold following the transition from mESC to EB and ExE. Transcripts associated with the synthesis of the early precursors were largely unaltered, suggesting other factors account for enhanced GAG synthesis. The composition of both CS/DS and HS also changed upon differentiation. Interestingly, CS type E and highly sulfated HS both increase as mESCs differentiate to EBs and ExE. Differentiation was also accompanied by enhanced 2-sulfation in both CS/DS and HS families. Transcript levels for core proteins generally showed increases or remained constant upon mESC differentiation. Finally, transcripts encoding selected enzymes and isoforms, including GlcNAc-4,6-O-sulfotransferase, C5-epimerases, and 3-O-sulfotransferases involved in late GAG biosynthesis, were also enriched. These biosynthetic enzymes are particularly important in introducing GAG fine structure, essential for intercellular communication, cell adhesion, and outside-in signaling. Knowing the changes in GAG fine structure should improve our understanding the biological properties of differentiated stem cells.

Dataset Information

The effect of beta-D-xylosides on the proliferation and proteoglycan biosynthesis of monoblastic U-937 cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets