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Proteoglycans (PGs) are important constituents of the plasma membrane and of the basement membrane supporting the endothelial cell layer. Changes in the amounts or the structures of PGs in the endothelium may affect important functions such as turnover of lipoproteins, filtration properties, and regulation of chemokines during inflammation, which are all relevant in diabetes. The purpose of this study was to investigate if hyperglycemic conditions would affect the biosynthesis and secretion of PGs in cultured primary human endothelial cells. Primary human umbilical cord vein endothelial cells were established and cultured in vitro. The cells were cultured either in medium with low glucose (LG) (1 g/l) or high glucose (HG) (4.5 g/l). From day 3–4 cells were labeled with 35S-sulfate for 24 h. 35S-Labeled macromolecules (medium) were purified by gel chromatography, and isolated macromolecules were analyzed by gel chromatography after different types of treatment, electrophoresis, and immunoprecipitation. Lower levels of secreted PGs were found in human endothelial cells exposed to HG. The major part of the PGs released was of the heparan sulfate (HS) type, and immunoprecipitation experiments showed that one such PG was syndecan-1. However, there was no difference in the ratio between HS and chondroitin sulfate (CS) under the different experimental conditions. Further, the PGs expressed neither differ with regard to molecular size of the glycosaminoglycan (GAG) chains, nor were their polyanionic properties affected by the different experimental conditions. The results obtained suggest that treatment of primary human endothelial cells with hyperglycemia leads to a decrease in PG secretion in primary cultures of human endothelial cells. [ABSTRACT FROM AUTHOR]

Background Hyperglycaemia may contribute to endothelial dysfunction. Disturbances in endothelial functions include changes in the extracellular matrix underneath the cells. This may result from altered biosynthesis of matrix molecules or from modified biosynthesis and secretion of enzymes involved in the turnover of extracellular matrix. One important class of such enzymes are the matrix metalloproteinases (MMPs). Aim of the study The aim of this study was to investigate whether the condition of high glucose concentration relevant both to diabetes type 1 and 2 and metabolic syndrome, would affect the synthesis and release of MMPs in human umbilical cord endothelial cells (HUVEC) in vitro. Methods The HUVEC were isolated and cultured in vitro. The cells were exposed to medium with either low glucose (LG, 1 g/l) or high glucose (HG, 4.5 g/l) or the advanced glycation end product (AGE) Nε-(carboxymethyl) lysine bovine serum albumin (CMLBSA), at a concentration of 10 µg/ml. The HUVEC-conditioned media were harvested and subjected to gelatin zymography and Western blotting. Results When HUVEC were incubated with HG or CML-BSA under serum free conditions a decreased secretion of pro MMP-2 was observed, both with gelatin zymography and Western blotting. The HUVEC also secreted MMP-9, but at lower levels, and effects of HG treatment were not significant. When HUVEC were stimulated with phorbol 12-myristate 13-acetate (PMA) secretion of pro MMP-2 was not increased, but the activation of pro MMP-2 into lower molecular forms increased, irrespective of culturing in LG, HG or CML-BSA. Conclusion The HUVEC exposed to high glucose or AGE exhibit decreased secretion of MMP-2. These findings may be relevant in understanding the altered turnover of the endothelial extracellular matrix observed in the diabetic state. [ABSTRACT FROM AUTHOR]