Glucocorticoids are steroid hormones that play diverse roles in numerous normal and pathological processes. They are actively used to treat a wide variety of diseases, including neurodegenerative and inflammatory diseases, cancers, and COVID-19, among others. However, the long-term use of glucocorticoids is associated with numerous side effects. Molecular mechanisms of these negative side effects are not completely understood. Recently, arguments have been made that one such mechanisms may be related to the influence of glucocorticoids on O-glycosylated components of the cell surface and extracellular matrix, in particular on proteoglycans and glycosaminoglycans. The potential toxic effects of glucocorticoids on these glycosylated macromolecules are particularly meaningful for brain physiology because proteoglycans/glycosaminoglycans are the main extracellular components of brain tissue. Here, we aim to review the known effects of glucocorticoids on proteoglycan expression and glycosaminoglycan content in different tissues, with a specific focus on the brain.

Intensive adjuvant radiotherapy (RT) is a standard treatment for glioblastoma multiforme (GBM) patients; however, its effect on the normal brain tissue remains unclear. Here, we investigated the short-term effects of multiple irradiation on the cellular and extracellular glycosylated components of normal brain tissue and their functional significance. Triple irradiation (7 Gy*3 days) of C57Bl/6 mouse brain inhibited the viability, proliferation and biosynthetic activity of normal glial cells, resulting in a fast brain-zone-dependent deregulation of the expression of proteoglycans (PGs) (decorin, biglycan, versican, brevican and CD44). Complex time-point-specific (24–72 h) changes in decorin and brevican protein and chondroitin sulfate (CS) and heparan sulfate (HS) content suggested deterioration of the PGs glycosylation in irradiated brain tissue, while the transcriptional activity of HS-biosynthetic system remained unchanged. The primary glial cultures and organotypic slices from triple-irradiated brain tissue were more susceptible to GBM U87 cells’ adhesion and proliferation in co-culture systems in vitro and ex vivo. In summary, multiple irradiation affects glycosylated components of normal brain extracellular matrix (ECM) through inhibition of the functional activity of normal glial cells. The changed content and pattern of PGs and GAGs in irradiated brain tissues are accompanied by the increased adhesion and proliferation of GBM cells, suggesting a novel molecular mechanism of negative side-effects of anti-GBM radiotherapy.

Glioblastoma (GB) is an aggressive cancer with a high probability of recurrence, despite active chemoradiotherapy with temozolomide (TMZ) and dexamethasone (DXM). These systemic drugs affect the glycosylated components of brain tissue involved in GB development; however, their effects on heparan sulfate (HS) remain unknown. Here, we used an animal model of GB relapse in which SCID mice first received TMZ and/or DXM (simulating postoperative treatment) with a subsequent inoculation of U87 human GB cells. Control, peritumor and U87 xenograft tissues were investigated for HS content, HS biosynthetic system and glucocorticoid receptor (GR, Nr3c1). In normal and peritumor brain tissues, TMZ/DXM administration decreased HS content (5–6-fold) but did not affect HS biosynthetic system or GR expression. However, the xenograft GB tumors grown in the pre-treated animals demonstrated a number of molecular changes, despite the fact that they were not directly exposed to TMZ/DXM. The tumors from DXM pre-treated animals possessed decreased HS content (1.5–2-fold), the inhibition of HS biosynthetic system mainly due to the -3–3.5-fold down-regulation of N-deacetylase/N-sulfotransferases (Ndst1 and Ndst2) and sulfatase 2 (Sulf2) expression and a tendency toward a decreased expression of the GRalpha but not the GRbeta isoform. The GRalpha expression levels in tumors from DXM or TMZ pre-treated mice were positively correlated with the expression of a number of HS biosynthesis-involved genes (Ext1/2, Ndst1/2, Glce, Hs2st1, Hs6st1/2), unlike tumors that have grown in intact SCID mice. The obtained data show that DXM affects HS content in mouse brain tissues, and GB xenografts grown in DXM pre-treated animals demonstrate attenuated HS biosynthesis and decreased HS content.

Introduction. Glucocorticoids are actively used in the treatment of various diseases, however their long-term use leads to numerous negative side-effects, the molecular mechanisms of which remain poorly understood.Aim. Study of the short-term (1–10 days) effects of various doses of dexamethasone (Dex) (0,1–10 mg/kg) on the expression of the glucocorticoid receptor (GR, Nr3c1), core proteins of main proteoglycans and heparan sulfate metabolism-involved genes, as well as the content of carbohydrate macromolecules of glycosaminoglycans in the brain tissue of experimental animals.Materials and methods. In the study, C57Bl/6 mice were used. The expression of GR, proteoglycan core proteins and heparan sulfate metabolism-involved genes was determined by real-time polymerase chain reaction with reverse transcription. The content and localization of GR protein molecule were studied by Western blot and immunohistochemical analysis, and the glycosaminoglycan content was determined by dot-blot analysis and Alcian Blue staining.Results. It was shown that a single Dex administration leads to fast (1–3 days) short-term activation of GR expression (+1.5 times, p 0.05), proteoglycan’s genes (syndecan-3, Sdc3; perlecan, Hspg2; phosphacan, Ptprz1; neurocan, Ncan; +2–3-fold; p 0.05) and heparan sulfate-metabolism-involved genes (Ndst1, Glce, Hs2st1, Hs6st1, Sulf1 / 2; +1.5–2-fold; p 0.05) in the mouse brain, with a return to control values by 7–10 days after Dex administration. At the same time, the effect of Dex on carbohydrate macromolecules of glycosaminoglycans was more delayed and stable, increasing the content of low-sulfated glycosaminoglycans in the brain tissue in a dose-dependent manner starting from day 1 after Dex administration. Highly-sulfated glycosaminoglycans showed more delayed response to Dex administration, and an increase in their content was observed only at higher doses (2.5 and 10 mg/kg) and only on 7–10 days after its administration, apparently, mainly due to an increase in heparan sulfate content.Conclusion. In general, the effect of a single injection of Dex on the transcriptional activity of GR, proteoglycan core proteins and heparan sulfate metabolism-involved genes were short-termed, and the genes expression quickly returned to the normal levels. However, even a single use of Dex significantly increased the content of total as well as highly sulfated glycosaminoglycans in the mouse brain tissue, which can lead to the changes in the composition and structure of the brain tissue, as well as its functional characteristics.

Adjuvant chemoradiotherapy is a standard treatment option for glioblastoma multiforme (GBM). Despite intensive care, recurrent tumors developed during the first year are fatal for the patients. Possibly contributing to this effect, among other causes, is that therapy induces changes of polysaccharide heparan sulfate (HS) chains in the cancer cells and/or tumor microenvironment. The aim of this study was to perform a comparative analysis of heparanase (HPSE) expression and HS content in different normal and GBM brain tissues. Immunohistochemical analysis revealed a significant decrease of HPSE protein content in the tumor (12-15-fold) and paratumorous (2.5-3-fold) GBM tissues compared with normal brain tissue, both in cellular and extracellular compartments. The relapsed GBM tumors demonstrated significantly higher intertumor and/or intratumor heterogeneity of HPSE and HS content and distribution compared with the matched primary ones (from the same patient) (n = 8), although overall expression levels did not show significant differences, suggesting local deterioration of HPSE expression with reference to the control system or by the treatment. Double immunofluorescence staining of various glioblastoma cell lines (U87, U343, LN18, LN71, T406) demonstrated a complex pattern of HPSE expression and HS content with a tendency towards a negative association of these parameters. Taken together, the results demonstrate the increase of intratumor heterogeneity of HPSE protein in relapsed GBM tumors and suggest misbalance of HPSE expression regulation by the adjuvant anti-GBM chemoradiotherapy.

Glucocorticoids are used during glioblastoma treatment to prevent the cerebral edema effect surrounding normal brain tissue. The aim of our study was to investigate the long-term effects of multiple administrations of glucocorticoids onto the glycosylated components (proteoglycans and glycosaminoglycans) of normal brain extracellular matrix and the glucocorticoid receptor (GR, Nr3c1) in an experimental model in vivo. Two-month-old male C57Bl/6 mice (n = 90) were injected intraperitoneally with various doses of dexamethasone (DXM) (1; 2.5 mg/kg) for 10 days. The mRNA levels of the GR, proteoglycans core proteins, and heparan sulfate metabolism-involved genes were determined at the 15th, 30th, 60th, and 90th days by a real-time RT–PCR. The glycosaminoglycans content was studied using dot blot and staining with Alcian blue. A DXM treatment increased total GAG content (2-fold), whereas the content of highly sulfated glycosaminoglycans decreased (1.5–2-fold). The mRNA level of the heparan sulfate metabolism-involved gene Hs3St2 increased 5-fold, the mRNA level of Hs6St2 increased6–7-fold, and the mRNA level of proteoglycan aggrecan increased 2-fold. A correlation analysis revealed an association between the mRNA level of the GR and the mRNA level of 8 of the 14 proteoglycans-coding and 4 of the 13 heparan sulfate metabolism-involved genes supporting GR involvement in the DXM regulation of the expression of these genes. In summary, multiple DXM administrations led to an increase in the total GAG content and reorganized the brain extracellular matrix in terms of its glycosylation pattern.