Dexamethasone affects astroglial differentiation in neural stem cells: Role of endogenous interleukin-1 beta

Vishvesh Shende, Francisco Molina-Holgado, Jolanta Opacka-Juffry. University of Roehampton, Department of Life Sciences SW15 4JD, London, UK.

Cytokines in the brain mediate the neural components of the host response to infection, are involved in the neurodegenerative processes resulting for brain trauma, ischaemia and chronic neurodegenerative diseases. However they are also involved in CNS repair and recovery. Interleukin-1β (IL-1β) is an important cytokine involved in the development of brain inflammatory responses during infection, trauma and neurodegenerative processes. However, brain injury is associated with an enhanced release of glucocorticoids, which inhibit IL-1β production and transcription. The purpose of the present study was to understand the role of endogenous IL-1β in the differentiation of neural stem cells (NSC) in the presence of the synthetic glucocorticoid dexamethasone (Dex) and its effects on astroglia, as identified by the glial acidic fibrillary protein (GFAP). NSC were prepared from the cortex of C57BL6/J mice either wild type (WT) or IL-1β knock-out (IL-1β KO) at embryonic day16 (E16). NSCs were exposed to dexamethasone (0.1, 1 and 10µM, t = 48h, n = 4 per group). Cytotoxicity was evaluated by the release of the cytosolic enzyme lactate dehydrogenase (LDH) at 24h. Prior to immunocytochemistry, NSCs underwent differentiation into 24 multi-well plates (Nunc) that had previously been pre-coated with poly-ornithine (50μg/ml) and laminin (5 µg/ml), DMEN-F12 media containing 10% fetal bovine serum and 5ng/ml FGF. NSC cultures were exposed to Dex for 48h and then fixed with 4% paraformaldehyde. GFAP immunocytochemistry was carried out to label astrocytic cells (GFAP primary antibody (1:500) overnight at 4°C, secondary antibody (1:200) for 1 h). Under the conditions tested, Dex showed no toxic effects and it did not affect NSC cell survival as measured by LDH cytotoxicity in WT (F[3,8] = 3.030, P >0.05) and IL-1β KO (F[3,8] = 3.605, P >0.05) (ANOVA - Post Hoc Tukey test, n = 6 per group). Exposure to Dex led to a decrease in GFAP differentiation in WT NSC (F[3,12] = 5.458, P<0.05) (ANOVA - Post Hoc Tukey test, n = 4 per group). However, Dex facilitated the ramification of GFAP-positive cells in NSC derived from IL-1β KO (F[3,12] = 1.517, P >0.05)(ANOVA - Post Hoc Tukey test, n = 4 per group) mice when compared with WT. Cell count analysis indicated that Dex had a pro-gliogenic effect on NSC from IL-1β KO by directing the NSC differentiation to a GFAP-positive lineage. Hence, IL-1β can act as a mediator of astroglial differentiation, which is of relevance to translational research.